| 2020 | Endothelial dysfunction is an early indicator of sepsis and neutrophil degranulation of septic shock in surgical patients. | BJS Open | Stratification of the severity of infection is currently based on the Sequential Organ Failure Assessment (SOFA) score, which is difficult to calculate outside the ICU. Biomarkers could help to stratify the severity of infection in surgical patients.Levels of ten biomarkers indicating endothelial dysfunction, 22 indicating emergency granulopoiesis, and six denoting neutrophil degranulation were compared in three groups of patients in the first 12 h after diagnosis at three Spanish hospitals.There were 100 patients with infection, 95 with sepsis and 57 with septic shock. Seven biomarkers indicating endothelial dysfunction (mid-regional proadrenomedullin (MR-ProADM), syndecan 1, thrombomodulin, angiopoietin 2, endothelial cell-specific molecule 1, vascular cell adhesion molecule 1 and E-selectin) had stronger associations with sepsis than infection alone. MR-ProADM had the highest odds ratio (OR) in multivariable analysis (OR 11·53, 95 per cent c.i. 4·15 to 32·08; P = 0·006) and the best area under the curve (AUC) for detecting sepsis (0·86, 95 per cent c.i. 0·80 to 0·91; P < 0·001). In a comparison of sepsis with septic shock, two biomarkers of neutrophil degranulation, proteinase 3 (OR 8·09, 1·34 to 48·91; P = 0·028) and lipocalin 2 (OR 6·62, 2·47 to 17·77; P = 0·002), had the strongest association with septic shock, but lipocalin 2 exhibited the highest AUC (0·81, 0·73 to 0·90; P < 0·001).MR-ProADM and lipocalin 2 could be alternatives to the SOFA score in the detection of sepsis and septic shock respectively in surgical patients with infection. |
| 2019 | Triggering Receptor Expressed on Myeloid Cells-1 Inhibitor Targeted to Endothelium Decreases Cell Activation. | Front Immunol | TREM-1 (Triggering Receptor Expressed on Myeloid cells-1) is an immunoreceptor expressed on neutrophils, monocytes/macrophages, and endothelial cells. It amplifies the inflammatory response driven by Toll-Like Receptors (TLR) engagement. The pharmacological inhibition of TREM-1 confers protection in several pre-clinical models of acute inflammation. In this study, we aimed to investigate the role of TREM-1 in endothelial cells using a sneaking ligand construct (SLC) inhibiting TREM-1 in the endothelium. The SLC was made of 3 modules: an E-selectin targeting domain, a exotoxin a translocation domain, and a 7 aa peptide (LSKSLVF) that contains the interaction site between TREM-1 and its adaptor protein DAP-12. SLC peptide was effectively picked up by endothelial cells following LPS stimulation. It decreased LPS induced TREM-1 up-regulation and cell activation, neutrophils extravasation, and improved median survival time during experimental peritonitis in mice. We reported that a targeted endothelial TREM-1 inhibition is able to dampen cell activation and to confer protection during septic shock in mice. The use of such cell-specific, ligand- independent TREM-1 inhibitors deserve further investigations during acute or chronic inflammatory disorders. |
| 2019 | Pathogenic role of palatine tonsils in palmoplantar pustulosis: A review. | J Dermatol | Palmoplantar pustulosis (PPP) is characterized by symmetrical, erythematous, scaly plaques, with numerous, sterile, non-bacterial, pinpoint pustules, which are restricted to the palms and soles. Because several reports have described the efficacy of tonsillectomy for improvement in PPP skin lesions, we consider that PPP is tonsil-induced autoimmune/inflammatory syndrome (TIAS) while other factors are also involved in the pathogenesis of PPP. Here, the association between PPP pathogenesis and TIAS was examined, with a focus on results of previous studies. PPP patients show a hyperimmune response to indigenous bacteria such as α-streptococci, due to impaired immunological tolerance towards such organisms. Such a novel immune response leads to T-cell activation through the abnormal expression of secondary stimulation molecules, including cytotoxic T-lymphocyte-associated antigen 4, inducible T-cell co-stimulator and Smad7, in the tonsils of PPP patients. Activated tonsillar T cells express cutaneous lymphocyte antigen (CLA), CCR6 and β1-integrin, enter the blood circulation and are recruited to PPP skin lesions. Within lesions, T cells roll onto endothelial cells through the interaction between CLA and E-selectin, migrate into the extravascular area through β1-integrin-vascular cell adhesion molecule 1 binding, and assemble in the skin through CCL20-CCR6 binding. Hyperimmune responses to autoantigens such as keratin and heat shock proteins could also be involved in PPP pathogenesis, through the stimulation of the T-helper 17 reaction. |
| 2020 | Critical Role of Mortalin/GRP75 in Endothelial Cell Dysfunction Associated with Acute Lung Injury. | Shock | Mortalin/GRP75 (glucose regulated protein 75), a member of heat shock protein 70 family of chaperones, is involved in several cellular processes including proliferation and signaling, and plays a pivotal role in cancer and neurodegenerative disorders. In this study, we sought to determine the role of mortalin/GRP75 in mediating vascular inflammation and permeability linked to the pathogenesis of acute lung injury (ALI). In an aerosolized bacterial lipopolysaccharide inhalation mouse model of ALI, we found that administration of mortalin/GRP75 inhibitor mean kinetic temperature-077, both prophylactically and therapeutically, protected against polymorphonuclear leukocytes influx into alveolar airspaces, microvascular leakage, and expression of pro-inflammatory mediators such as interleukin-1β, E-selectin, and tumor necrosis factor TNFα. Consistent with this, thrombin-induced inflammation in cultured human endothelial cells (EC) was also protected upon before and after treatment with mean kinetic temperature-077. Similar to pharmacological inhibition of mortalin/GRP75, siRNA-mediated depletion of mortalin/GRP75 also blocked thrombin-induced expression of proinflammatory mediators such as intercellular adhesion molecule-1 and vascular adhesion molecule-1. Mechanistic analysis in EC revealed that inactivation of mortalin/GRP75 interfered with the binding of the liberated NF-κB to the DNA, thereby leading to inhibition of downstream expression of adhesion molecules, cytokines, and chemokines. Importantly, thrombin-induced Ca signaling and EC permeability were also prevented upon mortalin/GRP75 inactivation/depletion. Thus, this study provides evidence for a novel role of mortalin/GRP75 in mediating EC inflammation and permeability associated with ALI. |
| 2019 | Sepsis causes right ventricular myocardial inflammation independent of pulmonary hypertension in a porcine sepsis model. | PLoS One | Right ventricular (RV) myocardial dysfunction is a common feature in septic shock. It can worsen outcome, but the etiology is poorly understood. Pulmonary artery hypertension (PAH) plays a part in the pathogenesis of the right heart dysfunction in sepsis but its importance is unknown. In pigs, PAH in sepsis is substantial and the translational value of porcine sepsis models therefore questioned. We hypothesized that porcine sepsis causes a myocardial inflammatory response which leads to myocardial dysfunction independent of PAH.Sepsis was induced by Escherichia coli-infusion in 10 pigs resulting in PAH and increased right ventricular pressure (RVP). The same degree of RVP was achieved by external pulmonary artery banding (PAB) in a consecutive series of 6 animals.Sepsis, but not PAB, led to increase in endothelial damage marker PAI-1 and cytokines TNF and IL-6 (all p<0.05) in plasma. In myocardium, TNF and IL-6 were significantly elevated in sepsis, TNF in both ventricles and IL-6 mostly in RV, while IL-1β, IL-18 and C5a were significantly higher in RV compared to LV after PAB (all p<0.05). Myocardial mRNA levels of IL-1β, IL-6, IL-18, IP-10, E-selectin and PAI-1 were significantly elevated in RV and LV during sepsis compared to PAB, while Caspase-1 was decreased in septic compared to PAB animals (all p<0.05). Cathepsin L activity was increased in RV by PAB, while sepsis inhibited this response. Escherichia coli-induced sepsis caused myocardial inflammation independent of PAH.Prominent PAH should therefore not exclude porcine sepsis models to further our understanding of human sepsis. |
| 2019 | Partial Deletion of Tie2 Affects Microvascular Endothelial Responses to Critical Illness in A Vascular Bed and Organ-Specific Way. | Shock | Tyrosine kinase receptor (Tie2) is mainly expressed by endothelial cells. In animal models mimicking critical illness, Tie2 levels in organs are temporarily reduced. Functional consequences of these reduced Tie2 levels on microvascular endothelial behavior are unknown. We investigated the effect of partial deletion of Tie2 on the inflammatory status of endothelial cells in different organs. Newly generated heterozygous Tie2 knockout mice (exon 9 deletion, ΔE9/Tie2) exhibiting 50% reduction in Tie2 mRNA and protein, and wild-type littermate controls (Tie2), were subjected to hemorrhagic shock and resuscitation (HS + R), or challenged with i.p. lipopolysaccharide (LPS). Kidney, liver, lung, heart, brain, and intestine were analyzed for mRNA levels of adhesion molecules E-selectin, vascular cell adhesion molecule 1 (VCAM-1), and intercellular cell adhesion molecule 1 (ICAM-1), and CD45. Exposure to HS + R did not result in different expression responses of these molecules between organs from Tie2 or Tie2 mice and sham-operated mice. In contrast, the LPS-induced mRNA expression levels of E-selectin, VCAM-1, and ICAM-1, and CD45 in organs were attenuated in Tie2 mice when compared with Tie2 mice in kidney and liver, but not in the other organs studied. Furthermore, reduced expression of E-selectin and VCAM-1 protein, and reduced influx of CD45 cells upon LPS exposure, was visible in a microvascular bed-specific pattern in kidney and liver of Tie2 mice compared with controls. In contrast to the hypothesis that a disbalance in the Ang/Tie2 system leads to increased microvascular inflammation, heterozygous deletion of Tie2 is associated with an organ-restricted, microvascular bed-specific attenuation of endothelial inflammatory response to LPS. |
| 2019 | Levels of circulating microparticles in septic shock and sepsis-related complications: a case-control study. | Minerva Anestesiol | Microparticles (MP) have been largely studied as potential biomarkers in septic shock (SS) though their biological and clinical relevance is still unclear. This case-control study describes the trend of various MP subtypes during SS to evaluate their possible association with severity of illness and sepsis-related complications (disseminated intravascular coagulation [DIC] and acute kidney injury [AKI]).Forty patients admitted to the Intensive Care Unit with SS and 40 matched healthy volunteers were recruited. AnnexinV+, E-selectin+, thrombomodulin (TM+), leukocyte-derived (CD45+, CD36+) and platelet-derived MP (PMP-expressed as PMP/platelets ratio) were measured by flow-cytometry at baseline, on day 1, 3 and 7 after diagnosis. Severity of illness was assessed by Sequential Organ Failure Assessment Score, duration of vasoactive support and mechanical ventilation. Sepsis-related complications were considered.Overall, septic patients showed higher levels of all MP considered compared to controls. TM+MP were significantly lower in more severe sepsis, while CD36+MP and PMP/platelets ratio were significantly increased in patients requiring longer vasoactive support and mechanical ventilation. As for sepsis-related complications, a higher PMP/platelets ratio in patients who developed DIC and increased E-selectin+MP in subjects who developed AKI were observed. PMP/platelets ratio at baseline was significantly associated with longer vasoactive support (OR=1.59 [1.05-2.42]), longer mechanical ventilation (OR=1.6 [1.06-2.42]) and DIC occurrence (OR=1.45 [1.08-1.96]).A global response through extra-vesiculation of endothelial cells, leukocytes and platelets during the early stages of SS was confirmed. The cellular activation was detected until day 3 after diagnosis. PMP/platelets ratio at diagnosis may be useful to evaluate SS severity and DIC occurrence. |
| 2018 | Clinical characteristics and prognosis of serous body cavity effusions in patients with sepsis: a retrospective observational study. | BMC Anesthesiol | Cavity effusion is common in patients with infectious diseases. However, the incidence rate and characteristics of serous cavity effusions (SCE) in septic patients are not clear to date. The objective of this study was to investigate the incidence and characteristics of SCE in septic patients and to explore the correlations between the bloody effusions and the illness severity/prognosis in septic patients.From January 2010 to January 2015, a total of 214 patients with severe sepsis and septic shock were enrolled in this retrospective observational study. Thoracentesis or abdominal paracentesis was performed in 45 septic patients because of massive pleural effusions or ascites. The serum concentrations of VEGF, VEGFR, Ang, sICAM-1, sVCAM-1, E-selectin, Serpine1 and VE-cadherin in 45 septic patients underwent paracentesis were measured by enzyme-linked immunosorbent assay (ELISA).Of the 214 septic patients, 155 (72.4%) had SCE according to imaging or ultrasound manifestations. 45 subjects with SCE underwent therapeutic thoracentesis or abdominal paracentesis. Effusion laboratory analysis showed that exudates were predominant when compared with transudates (95.6% vs. 4.4%), and 16 (35.6%) patients suffered bloody effusions. Compared with patients with non-bloody effusions, those with bloody effusions showed higher critical illness scores (13 vs. 17 for APACHE II; 7 vs. 9 for SOFA), and higher mortality (6.9% vs. 62.5%). Moreover, patients with bloody effusions had delayed TT and APTT, increased D-dimer concentration, and higher serum levels of CRP and PCT (P < 0.05). In addition, the serum levels of Ang2, sVCAM-1 and E-selectin were significantly higher in patients with bloody effusions than in those with non-bloody effusions (P < 0.05). However, the serum level of VEGFR2 was lower in patients with bloody fluids (P = 0.025).The incidence of serous cavity effusion is high in patients with sepsis. The septic patients with bloody effusions suffer a more inflammatory burden and a worse prognosis compared to septic patients with non-bloody effusions. |
| 2018 | Lower serum kallistatin level is associated with 28-day mortality in patients with septic shock. | J Crit Care | Investigation for whether serum levels of kallistatin, vascular cell adhesion molecule-1 (VCAM-1), and E-selectin are associated with outcomes in patients with septic shock MATERIAL AND METHODS: Biomarker levels were measured using blood samples from patients with septic shock at admission, 24 h, and 72 h and from healthy volunteers. The primary outcome was 28-day mortality.Fifty-eight survivors, fourteen non-survivors, and six healthy volunteers were enrolled. Serum kallistatin level was lower and serum VCAM-1 and E-selectin levels were higher in patients at admission compared with healthy volunteers. Serum kallistatin levels were higher in survivors compared with non-survivors at all time points (4.4 μg/mL [2.9-6.1] vs. 2.5 μg/mL [2.1-5.0], P = 0.019 at admission; 4.3 μg/mL [3.3-5.2] vs. 3.2 μg/mL [2.2-3.8], P = 0.004 at 24 h; 3.1 μg/mL [2.5-4.2] vs. 2.3 μg/mL [1.7-3.1], P = 0.012 at 72 h), while VCAM-1 and E-selectin levels showed no difference. In the multivariable analysis, serum kallistatin level at 24 h was independently associated with 28-day mortality (OR, 0.29; 95% CI, 0.08-0.69, P = 0.024).Lower serum kallistatin level at 24 h was independently associated with 28-day mortality in patients with septic shock. |
| 2018 | Relationships between markers of neurologic and endothelial injury during critical illness and long-term cognitive impairment and disability. | Intensive Care Med | Neurologic and endothelial injury biomarkers are associated with prolonged delirium during critical illness and may reflect injury pathways that lead to poor long-term outcomes. We hypothesized that blood-brain barrier (BBB), neuronal, and endothelial injury biomarkers measured during critical illness are associated with cognitive impairment and disability after discharge.We enrolled adults with respiratory failure and/or shock and measured plasma concentrations of BBB (S100B), neuronal (UCHL1, BDNF), and endothelial (E-selectin, PAI-1) injury markers within 72 h of ICU admission. At 3 and 12 months post-discharge, we assessed participants' global cognition, executive function, and activities of daily living (ADL). We used multivariable regression to determine whether biomarkers were associated with outcomes after adjusting for relevant demographic and acute illness covariates.Our study included 419 survivors of critical illness with median age 59 years and APACHE II score 25. Higher S100B was associated with worse global cognition at 3 and 12 months (P = 0.008; P = 0.01). UCHL1 was nonlinearly associated with global cognition at 3 months (P = 0.02). Higher E-selectin was associated with worse global cognition (P = 0.006 at 3 months; P = 0.06 at 12 months). BDNF and PAI-1 were not associated with global cognition. No biomarkers were associated with executive function. Higher S100B (P = 0.05) and E-selectin (P = 0.02) were associated with increased disability in ADLs at 3 months.S100B, a marker of BBB and/or astrocyte injury, and E-selectin, an adhesion molecule and marker of endothelial injury, are associated with long-term cognitive impairment after critical illness, findings that may reflect mechanisms of critical illness brain injury. |
| 2017 | A Mouse Model of Anaphylaxis and Atopic Dermatitis to Salt-Soluble Wheat Protein Extract. | Int Arch Allergy Immunol | Wheat allergy and other immune-mediated disorders triggered by wheat proteins are growing at an alarming rate for reasons not well understood. A mouse model to study hypersensitivity responses to salt-soluble wheat protein (SSWP) extract is currently unavailable. Here we tested the hypothesis that SSWP extract from wheat will induce sensitization as well as allergic disease in mice.Female BALB/cJ mice were weaned onto a plant protein-free diet. The mice were injected a total of 4 times with an SSWP (0.01 mg/mouse) fraction extracted from durum wheat along with alum as an adjuvant. Blood was collected biweekly and SSWP-specific IgE (SIgE) and total IgE (TIgE) levels were measured using ELISA. Systemic anaphylaxis upon intraperitoneal injection with SSWP was quantified by hypothermia shock response (HSR). Mucosal mast cell degranulation was measured by the elevation of mMCP-1 in the blood. The mice were monitored for dermatitis. Skin tissues were used in histopathology and for measuring cytokine/chemokine/adhesion molecule levels using a protein microarray system.Injection with SSWP resulted in time-dependent SIgE antibody responses associated with the elevation of TIgE concentration. Challenge with SSWP elicited severe HSR that correlated with a significant elevation of plasma mMCP-1 levels. Sensitized mice developed facial dermatitis associated with mast cell degranulation. Lesions expressed significant elevation of Th2/Th17/Th1 cytokines and chemokines and E-selectin adhesion molecule.Here we report a mouse model of anaphylaxis and atopic dermatitis to SSWP extract that may be used for further basic and applied research on wheat allergy. |
| 2017 | Markers of acute kidney injury in patients with sepsis: the role of soluble thrombomodulin. | Crit Care | Endothelial activation and damage occur early during sepsis, with activated coagulopathy and playing a major role in the pathophysiology of sepsis-induced acute kidney injury (AKI). The aim of this study was to compare the various biomarkers of endothelial injury with the biomarkers of coagulation and inflammation and to determine a significant predictor of AKI in patients with sepsis.We conducted a single-center, retrospective, observational study on patients with sepsis fulfilling the Third International Consensus Definitions for Sepsis and Septic Shock criteria admitted to an adult intensive care unit (ICU) at a university hospital from June 2011 to December 2016. Levels of 13 biomarkers were measured on ICU admission, including markers of endothelial injury (soluble thrombomodulin [sTM], E-selectin, protein C, and plasminogen activator inhibitor-1 [PAI-1]) and markers of coagulation derangement (platelet count, fibrin degradation product [FDP], prothrombin time [PT], fibrinogen, α-plasminogen inhibitor [α-PI], antithrombin III [AT III], plasminogen, thrombin-antithrombin complex, and plasmin-α-plasmin inhibitor complex). All patients with sepsis were reviewed, and the development of AKI was evaluated. Multivariate logistic regression analysis was performed to identify significant independent predictive factors for AKI.Of the 514 patients admitted with sepsis, 351 (68.3%) developed AKI. Compared with the non-AKI group, all the endothelial biomarkers were significantly different in the AKI group (sTM [23.6 vs. 15.6 U/ml, P < 0.0001], E-selectin [65.5 vs. 46.2 ng/ml, P = 0.0497], PAI-1 [180.4 vs. 75.3 ng/ml, P = 0.018], and protein C [45.9 vs. 58.7 ng/ml, P < 0.0001]). Biomarkers of coagulopathy and inflammation, platelet counts, FDP, PT, α-PI, AT III, plasminogen, and C-reactive protein were significantly different between the two groups. Multivariable logistic regression analysis showed that sTM was an independent predictive factor of AKI, with an AUROC of 0.758 (P < 0.0001).Endothelial biomarkers were significantly changed in the sepsis patients with AKI. Particularly, sTM was an independent predictive biomarker for the development of AKI that outperformed other coagulation and inflammation biomarkers as well as organ function in patients with sepsis. |
| 2017 | The Effects of Selective Hematopoietic Expression of Human IL-37 on Systemic Inflammation and Atherosclerosis in LDLr-Deficient Mice. | Int J Mol Sci | The human cytokine interleukin (IL)-37 has potent anti-inflammatory capacities, and hematopoietic cell-specific transgenic overexpression of IL-37 in mice protects against septic shock and colitis. In the present study we investigated the effect of hematopoietic expression of IL-37 on atherosclerosis development under low-grade inflammatory conditions. Low-density lipoprotein receptor (LDLr)-deficient mice were lethally irradiated and transplanted with bone marrow from IL-37-transgenic or control wild-type mice and fed a Western-type diet (WTD; 1% cholesterol) for eight weeks. Metabolic and inflammatory parameters were monitored and atherosclerosis was assessed in the aortic valve area. Hematopoietic IL-37 expression did not influence body weight, food intake and plasma cholesterol levels during the study. Plasma soluble E-selectin levels were increased with WTD-feeding as compared to chow-feeding, but were not influenced by IL-37 expression. IL-37 expression reduced the inflammatory state as indicated by reduced white blood cell counts and by reduced basal and lipopolysaccharide-induced cytokine response by peritoneal macrophages ex vivo. IL-37 expression did not influence the atherosclerotic lesion area. Lesion composition was marginally affected. Smooth muscle cell content was decreased, but macrophage and collagen content were not different. We conclude that under low-grade inflammatory conditions, hematopoietic IL-37 expression reduces the inflammatory state, but does not influence atherosclerosis development in hyperlipidemic LDLr-deficient mice. |
| 2017 | Sirtuin 2 Regulates Microvascular Inflammation during Sepsis. | J Immunol Res | . Sepsis and septic shock, the leading causes of death in noncoronary intensive care units, kill more than 200,000/year in the US alone. Circulating cell-endothelial cell interactions are the rate determining factor in sepsis inflammation. Sirtuin, a seven-member family of proteins (SIRT1-7), epigenetically controls inflammation. We have studied the roles of SIRTs 1, 3, and 6 in sepsis previously. In this project, we studied the role of SIRT2 on sepsis-related inflammation. . Sepsis was induced in C57Bl/6 (WT), SIRT2 knockout (SIRT2KO), and SIRT2 overexpressing (SIRT2KI) mice by cecal ligation and puncture (CLP). We studied leukocyte/platelet adhesion using intravital microscopy and E-selectin/ICAM-1 adhesion molecule expression in the small intestine with immunohistochemistry (IHC) six hours post-CLP/sham surgery. We also studied 7-day survival rates in WT, SIRT2KO, and SIRT2KI sepsis mice. . Compared to WT mice, SIRT2KO mice show exaggeration while SIRT2KI mice show attenuation of cellular adhesion with sepsis in the small intestine. We also show that the small intestinal E-selectin and ICAM-1 expressions increased in SIRT2KO and decreased in SIRT2KI mice versus those in WT sepsis mice. We show that the 7-day survival rate is decreased in SIRT2KO and increased in SIRT2KI sepsis mice. . SIRT2 modulates microvascular inflammation in sepsis and affects survival. |
| 2017 | Xuebijing Injection () increases early survival rate by alleviating pulmonary vasopermeability in rats subjected to severe burns. | Chin J Integr Med | To investigate the effects of Xuebijing Injection (, XBJ) on survival rate and pulmonary vasopermeability in a rat model of severe scald injury.Rats were divided into two experiments: experiment 1 was monitored for 12 h post-injury for survival analysis after severe burns; in experiment 2, rats were killed for determination of pulmonary vascular permeability and pro-inflflammatory mediators. In both experiments, rats were subject to third-degree 50% total body surface area (TBSA) burns or sham injury followed by XBJ or normal saline (NS) treatment. In addition, rat pulmonary microvascular endothelium cells (PMECs) were pretreated with either XBJ or phosphate buffer saline (PBS), and then subjected to sham serum or scald serum stimulation for 2 or 6 h, followed by transwell examination for the permeability of PMECs. Meanwhile, pro-inflflammatory mediators in PMECs culture supernatant were also investigated.The average survival time in the scald+XBJ group was 582.1±21.2 min, which was signifificantly longer than that in the scald + NS group (345.8±25.4 min, P<0.01). Plasma levels of tumor necrosis factor-alpha (TNF-α), E-selectin, interleukin-6 (IL-6), vascular permeability and water content of lung tissues were signifificantly increased in animals after severe burns (P<0.01). However, administration of XBJ signifificantly decreased these levels in plasma and lung tissue. In in vitro cell experiments, XBJ markedly attenuated permeability in PMECs monolayer and reduced the levels of TNF-α, IL-6 and soluble E-selectin after stimulation with scald serum (P<0.01).XBJ increases early survival rate by alleviating pulmonary vasopermeability and inhibiting pro-inflflammatory mediators in rats subjected to lethal scald injury. XBJ may be a potent drug in treatment of severe burns. |
| 2016 | Effects of Early Continuous Venovenous Hemofiltration on E-Selectin, Hemodynamic Stability, and Ventilatory Function in Patients with Septic-Shock-Induced Acute Respiratory Distress Syndrome. | Biomed Res Int | . To investigate the effects of 72-hour early-initiated continuous venovenous hemofiltration (ECVVH) treatment in patients with septic-shock-induced acute respiratory distress syndrome (ARDS) (not acute kidney injury, AKI) with regard to serum E-selectin and measurements of lung function and hemodynamic stability. . This prospective nonblinded single institutional randomized study involved 51 patients who were randomly assigned to receive or not receive ECVVH, an ECVVH group ( = 24) and a non-ECVVH group ( = 27). Besides standard therapies, patients in ECVVH group underwent CVVH for 72 h. . At 0 and 24 h after initiation of treatment, arterial partial pressure of oxygen/fraction of inspired oxygen (PaO/FiO) ratio, extravascular lung water index (EVLWI), and E-selectin level were not significantly different between groups (all > 0.05). Compared to non-ECVVH group, PaO/FiO is significantly higher and EVLWI and E-selectin level are significantly lower in ECVVH group (all < 0.05) at 48 h and 72 h after initiation of treatment. The lengths of mechanical ventilation and stay in intensive care unit (ICU) were shorter in ECVVH group (all < 0.05), but there was no difference in 28-day mortality between two groups. . In patients with septic-shock-induced ARDS (not AKI), treatment with ECVVH in addition to standard therapies improves endothelial function, lung function, and hemodynamic stability and reduces the lengths of mechanical ventilation and stay in ICU. |
| 2016 | Cold-inducible RNA-binding protein causes endothelial dysfunction via activation of Nlrp3 inflammasome. | Sci Rep | Cold-inducible RNA-binding protein (CIRP) is a damage-associated molecular pattern (DAMP) molecule which stimulates proinflammatory cytokine release in hemorrhage and sepsis. Under these medical conditions, disruption of endothelial homeostasis and barrier integrity, typically induced by proinflammatory cytokines, is an important factor contributing to morbidity and mortality. However, the role of CIRP in causing endothelial dysfunction has not been investigated. In this study, we show that intravenous injection of recombinant murine CIRP (rmCIRP) in C57BL/6 mice causes lung injury, evidenced by vascular leakage, edema, increased leukocyte infiltration and cytokine production in the lung tissue. The CIRP-induced lung damage is accompanied with endothelial cell (EC) activation marked by upregulation of cell-surface adhesion molecules E-selectin and ICAM-1. Using in vitro primary mouse lung vascular ECs (MLVECs), we demonstrate that rmCIRP treatment directly increases the ICAM-1 protein expression and activates NAD(P)H oxidase in MLVECs. Importantly, CIRP stimulates the assembly and activation of Nlrp3 inflammasome in MLVECs accompanied with caspase-1 activation, IL-1β release and induction of proinflammatory cell death pyroptosis. Finally, our study demonstrates CIRP-induced EC pyroptosis in the lungs of C57BL/6 mice for the first time. Taken together, the released CIRP in shock can directly activate ECs and induce EC pyroptosis to cause lung injury. |
| 2016 | Endothelial Activation and Blood-Brain Barrier Injury as Risk Factors for Delirium in Critically Ill Patients. | Crit Care Med | During critical illness, impaired endothelial vascular reactivity predicts prolonged acute brain dysfunction, but relationships between endothelial activation, blood-brain barrier/neurological injury, and acute brain dysfunction, including delirium, remain unexamined. We tested the hypothesis that elevated plasma markers of endothelial activation and blood-brain barrier/neurological injury are associated with delirium duration during critical illness.Prospective cohort study.Medical and surgical ICUs in an academic medical center.Adults in acute respiratory failure and/or shock.None.We enrolled subjects within 72 hours of organ failure diagnosis in the ICU. We measured plasma concentrations of plasminogen activator inhibitor-1, E-selectin, and angiopoietin-2 as markers of endothelial activation and S100B as a marker of blood-brain barrier/neurological injury in blood collected at enrollment. We assessed patients for delirium and coma twice daily after enrollment using the Confusion Assessment Method for the ICU and the Richmond Agitation-Sedation Scale. Among 134 patients with a median (interquartile) age of 57 years (46-66 yr) and Acute Physiology and Chronic Health Evaluation II of 26 (19-31), delirium occurred in 94 patients (70%) with a median duration of 2 days (0-4 d). Higher plasminogen activator inhibitor-1 (p = 0.002), E-selectin (p = 0.02), and S100B (p < 0.001) concentrations were associated with fewer delirium/coma-free days after adjusting for age, Charlson comorbidity index, modified Sequential Organ Failure Assessment score, and severe sepsis. Similarly, higher plasminogen activator inhibitor-1 (p = 0.007) and S100B (p = 0.01) concentrations were associated with longer delirium duration in survivors. Adjusting for S100B did not alter plasminogen activator inhibitor-1 and E-selectin associations with delirium, suggesting that these associations were not mediated by blood-brain barrier/neurological injury.Elevated plasma markers of endothelial activation and blood-brain barrier/neurological injury during critical illness are associated with prolonged delirium after biomarker measurement. Future research is needed to determine whether these processes have pathophysiologic roles in delirium and whether therapies targeted at the endothelium or blood-brain barrier can prevent and/or treat delirium during critical illness. |
| 2016 | The value of serial serum cell adhesion molecules in predicting acute kidney injury after severe sepsis in adults. | Clin Chim Acta | Septic acute kidney injury (AKI) is a common complication of severe sepsis. We tested the hypothesis that serum cell adhesion molecule levels are substantially increased in early septic AKI and decreased after antimicrobial therapy and their level can predict prognosis.Seventy-two nontraumatic, nonsurgical adult patients with severe sepsis admitted to the emergency department were evaluated. Serum adhesion molecules were collected and assessed. We evaluated their relationship with early septic AKI compared with other clinical predictors and biomarkers.Forty-five patients (62.5%) experienced early septic AKI. Patients with septic AKI also were more likely to experience septic shock and respiratory failure and had higher in-hospital mortality. Stepwise logistic regression model revealed that E-selectin level, septic shock, and respiratory failure were independently associated with septic AKI and each 1ng/ml increase in serum E-selectin level increased the risk of septic AKI by 1%. Furthermore, the E-selectin levels in the septic AKI group were significantly higher than those in the non-AKI group at two different times (days 1 and 4).WE show that early septic AKI implies a higher mortality in severe sepsis patients and that E-selectin level at presentation is a powerful predictor of early septic AKI. |
| 2015 | Characterization of Adsorbents for Cytokine Removal from Blood in an In Vitro Model. | J Immunol Res | Cytokines are basic targets that have to be removed effectively in order to improve the patient's health status in treating severe inflammation, sepsis, and septic shock. Although there are different adsorbents commercially available, the success of their clinical use is limited. Here, we tested different adsorbents for their effective removal of cytokines from plasma and the resulting effect on endothelial cell activation.The three polystyrene divinylbenzene (PS-DVB) based adsorbents Amberchrom CG161c and CG300m and a clinically approved haemoperfusion adsorbent (HAC) were studied with regard to cytokine removal in human blood. To induce cytokine release from leucocytes, human blood cells were stimulated with 1 ng/ml LPS for 4 hours. Plasma was separated and adsorption experiments in a dynamic model were performed. The effect of cytokine removal on endothelial cell activation was evaluated using a HUVEC-based cell culture model. The beneficial outcome was assessed by measuring ICAM-1, E-selectin, and secreted cytokines IL-8 and IL-6. Additionally the threshold concentration for HUVEC activation by TNF-α and IL-1β was determined using this cell culture model.CG161c showed promising results in removing the investigated cytokines. Due to its pore size the adsorbent efficiently removed the key factor TNF-α, outperforming the commercially available adsorbents. The CG161c treatment reduced cytokine secretion and expression of cell adhesion molecules by HUVEC which underlines the importance of effective removal of TNF-α in inflammatory diseases.These results confirm the hypothesis that cytokine removal from the blood should approach physiological levels in order to reduce endothelial cell activation. |
| 2015 | Plasma fractalkine is a sustained marker of disease severity and outcome in sepsis patients. | Crit Care | Fractalkine is a chemokine implicated as a mediator in a variety of inflammatory conditions. Knowledge of fractalkine release in patients presenting with infection to the Intensive Care Unit (ICU) is highly limited. The primary objective of this study was to establish whether plasma fractalkine levels are elevated in sepsis and associate with outcome. The secondary objective was to determine whether fractalkine can assist in the diagnosis of infection upon ICU admission.Fractalkine was measured in 1103 consecutive sepsis patients (including 271 patients with community-acquired pneumonia (CAP)) upon ICU admission and at days 2 and 4 thereafter; in 73 ICU patients treated for suspected CAP in whom this diagnosis was refuted in retrospect; and in 5 healthy humans intravenously injected with endotoxin.Compared to healthy volunteers, sepsis patients had strongly elevated fractalkine levels. Fractalkine levels increased with the number of organs failing, were higher in patients presenting with shock, but did not vary by site of infection. Non-survivors had sustained elevated fractalkine levels when compared to survivors. Fractalkine was equally elevated in CAP patients and patients treated for CAP but in whom the diagnosis was retrospectively refuted. Fractalkine release induced by intravenous endotoxin followed highly similar kinetics as the endothelial cell marker E-selectin.Plasma fractalkine is an endothelial cell derived biomarker that, while not specific for infection, correlates with disease severity in sepsis patients admitted to the ICU. |
| 2015 | [Impacts of high-volume hemofiltration on hemodynamics and vascular endothelial permeability in children with septic shock]. | Zhongguo Dang Dai Er Ke Za Zhi | To investigate the effects of high-volume hemofiltration (HVHF) on hemodynamics, vasoactive factors, and vascular endothelial permeability in children with septic shock by a comparative analysis.Thirty-six children who were diagnosed with septic shock between January 2013 and September 2014 were randomly divided into control and observation groups (n=18 each). Children in the control group were treated with the standard-volume hemofiltration (SVHF), while children in the observation group were treated with HVHF. The hemodynamic indices and levels of vasoactive factors including 6-keto-prostaglandin F1α (6-keto-PGF1α), thromboxane B2 (TXB2), soluble E-selectin (sE-selectin), and endothelium-derived relaxing factor (EDRF) were determined before and after treatment. In addition, the effects of ultrafiltrate on endothelial cell permeability were assessed.Compared with the control group, the observation group had significantly higher mean arterial pressure, significantly higher blood oxygen saturation, and a significantly lower heart rate after treatment (P<0.05). The levels of TXB2 and sE-selectin were significantly lower in the observation group than in the control group (P<0.05), while the levels of 6-keto-PGF1α and EDRF were significantly higher in the observation group than in the control group (P<0.05). Compared with the control group, the ultrafiltrate significantly attenuated the transepithelial electrical resistance in the observation group (P<0.05).Compared with SVHF, HVHF is a more effective approach for improving the hemodynamics and levels of vasoactive factors and reducing the vascular endothelial permeability in children with septic shock. |
| 2015 | Histone Deacetylase Inhibition and IκB Kinase/Nuclear Factor-κB Blockade Ameliorate Microvascular Proinflammatory Responses Associated With Hemorrhagic Shock/Resuscitation in Mice. | Crit Care Med | To investigate the consequences of histone deacetylase inhibition by histone deacetylase inhibitor valproic acid and IκB kinase/nuclear factor-κB signaling blockade by IκB kinase inhibitor BAY11-7082 on (microvascular) endothelial cell behavior in vitro as well as in mice subjected to hemorrhagic shock/resuscitation in vivo.Prospective, randomized laboratory investigation using an established mouse model of hemorrhagic shock.Research laboratory at university teaching hospital.Endothelial cells and C57BL/6 male mice.Endothelial cells were incubated with tumor necrosis factor-α in the absence or presence of valproic acid or BAY11-7082 in vitro. Mice were subjected to hemorrhagic shock by blood withdrawn until the mean arterial pressure of 30 mm Hg and maintained at this pressure for 90 minutes. At 90 minutes, subgroups of mice were resuscitated with 4% human albumin in the absence or presence of vehicle, valproic acid (300 μg/g body weight) or BAY11-7082 (400 μg per mouse). Mice were killed 1 hour and 4 hours after resuscitation.Valproic acid and BAY11-7082 selectively diminished tumor necrosis factor-α-induced endothelial proinflammatory activation in vitro. In vivo, both systemic and local inflammatory responses were significantly induced by hemorrhagic shock/resuscitation. The decreased histone acetylation in kidneys after hemorrhagic shock/resuscitation was restored by valproic acid treatment. In glomerular endothelial cells, the nuclear translocation of nuclear factor-κB, which was induced by hemorrhagic shock/resuscitation, was eliminated by BAY11-7082 treatment while enhanced in the presence of valproic acid. Both valproic acid and BAY11-7082 significantly attenuated the hemorrhagic shock/resuscitation-induced protein expression of endothelial cell adhesion molecules E-selectin and vascular cell adhesion molecule-1 in the microvasculature of kidneys and liver, although messenger RNA expression levels of these molecules analyzed in whole-organ lysates of kidneys, lungs, and liver were not extensively affected. The reduced protein expression of adhesion molecules was paralleled by diminishing the adhesion/transmigration of neutrophils in kidneys and liver after hemorrhagic shock/resuscitation.Suppression of histone deacetylase activity and blockade of IκB kinase/nuclear factor-κB signaling during resuscitation ameliorate microvascular endothelial proinflammatory responses in organs in mice after hemorrhagic shock. |
| 2015 | Attenuating systemic inflammatory markers in simulated high-altitude exposure by heat shock protein 70-mediated hypobaric hypoxia preconditioning in rats. | J Formos Med Assoc | The primary goal of this study was to test whether high-altitude exposure (HAE: 0.9% O(2) at 0.47 ATA for 24 hours) was capable of increasing the systemic inflammatory markers as well as the toxic organ injury indicators in rats, with a secondary goal to test whether preinduction of heat shock protein (HSP) 70 by hypobaric hypoxia preconditioning (HHP: 18.3% O(2) at 0.66 ATA for 5 h/day on 5 days consecutively for 2 weeks) attenuated the proposed increased serum levels of both the systemic inflammatory markers and the toxic organ injury indicators.Rats were assigned to: (1) non-HHP (21% O(2) at 1.0 ATA)+non-HAE (21% O(2) at 1.0 ATA) group; (2) non-HHP+HAE group; (3) HHP+non-HAE group; (4) HHP+HAE group; and (5) HHP+HSP70 antibodies (Ab)+HAE group. For the HSP70Ab group, a neutralizing HSP70Ab was injected intravenously at 24 hours prior to HAE. All the physiological and biochemical parameters were obtained at the end of HAE or the equivalent time period of non-HAE. Blood samples were obtained for determination of both the systemic inflammatory markers (e.g., serum tumor necrosis factor-α, interleukin-1β, E-selectin, intercellular adhesion molecule-1, and liver myeloperoxidase activity) and the toxic organ injury indicators (e.g., nitric oxide metabolites, 2,3-dihydroxybenzoic acid, and lactate dehydrogenase).HHP, in addition to inducing overexpression of tissue HSP70, significantly attenuated the HAE-induced hypotension, bradycardia, hypoxia, acidosis, and increased tissue levels of both the systemic inflammatory markers and the toxic organ injury indicators. The beneficial effects of HHP in inducing tissue overexpression of HSP70 as well as in preventing the HAE-induced increased levels of the systemic inflammatory markers and the toxic organ injury indicators could be significantly reduced by HSP70Ab preconditioning.These results suggest that HHP may downgrade both the systemic inflammatory markers and the toxic organ injury indicators in HAE by upregulating tissue HSP70. |
| 2014 | Abrupt reflow enhances cytokine-induced proinflammatory activation of endothelial cells during simulated shock and resuscitation. | Shock | Circulatory shock and resuscitation are associated with systemic hemodynamic changes, which may contribute to the development of MODS (multiple organ dysfunction syndrome). In this study, we used an in vitro flow system to simulate the consecutive changes in blood flow as occurring during hemorrhagic shock and resuscitation in vivo. We examined the kinetic responses of different endothelial genes in human umbilical vein endothelial cells preconditioned to 20 dyne/cm unidirectional laminar shear stress for 48 h to flow cessation and abrupt reflow, respectively, as well as the effect of flow cessation and reflow on tumor necrosis factor-α (TNF-α)-induced endothelial proinflammatory activation. Endothelial CD31 and VE-cadherin were not affected by the changes in flow in the absence or presence of TNF-α. The messenger RNA levels of proinflammatory molecules E-selectin, VCAM-1 (vascular cell adhesion molecule 1), and IL-8 (interleukin 8) were significantly induced by flow cessation respectively acute reflow, whereas ICAM-1 (intercellular adhesion molecule 1) was downregulated on flow cessation and induced by subsequent acute reflow. Flow cessation also affected the Ang/Tie2 (Angiopoietin/Tie2 receptor tyrosine kinase) system by downregulating Tie2 and inducing its endothelial ligand Ang2, an effect that was further extended on acute reflow. Furthermore, the induction of proinflammatory adhesion molecules by TNF-α under flow cessation was significantly enhanced on subsequent acute reflow. This study demonstrated that flow alterations per se during shock and resuscitation contribute to endothelial activation and that these alterations interact with proinflammatory factors coexisting in vivo such as TNF-α. The abrupt reflow-related enhancement of cytokine-induced endothelial proinflammatory activation supports the concept that sudden regain of flow during resuscitation has an aggravating effect on endothelial activation, which may play a significant role in vascular dysfunction and consequent organ injury. This study implies that the improvement of resuscitation strategies and the pharmacological interference with proinflammatory signaling cascades at the right time of resuscitation of shock patients may be beneficial to regain and/or maintain organ function in patients after circulatory shock. |
| 2014 | Normal saline influences coagulation and endothelial function after traumatic brain injury and hemorrhagic shock in pigs. | Surgery | Traumatic brain injury (TBI) and hemorrhagic shock (HS) are the leading causes of trauma-related deaths. These insults disrupt coagulation and endothelial systems. This study investigated whether previously reported differences in lesion size and brain swelling during normal saline (NS), colloids (Hextend [HEX]), and fresh frozen plasma (FFP) resuscitation are associated with differential effects on coagulation and endothelial systems.We subjected 15 Yorkshire swine to TBI and HS (40% blood volume), and kept in HS for 2 hours before resuscitation with NS, HEX, or FFP. Markers of endothelial activation (E-selectin, Intercellular adhesion molecule [ICAM]-1), coagulation activation (prothrombin fragment 1 + 2), and natural anticoagulation (activated protein C [aPC]) were determined in serum and brain whole cell lysates.Serum levels of aPC were greater in the NS group (203 ± 30 pg/mL) compared with HEX (77 ± 28 pg/mL; P = .02) and FFP (110 ± 28 pg/mL; P = .09), as was PF 1 + 2 in the brain when compared with FFP (PF 1 + 2, 89 ± 46 vs 37 ± 14 ng/mL; P = .035). Brain E-selectin was greater in the NS group compared with FFP (3.36 ± 0.02 vs 3.31 ± 0.01 ng/mL; P = .029). Circulating ICAM-1 levels were increased in the NS group (151 ± 9 ng/mL) compared with the HEX (100 ± 9 ng/mL; P < .01) and FFP (108 ± 9 ng/mL; P = .01).In this clinically realistic large animal model of TBI + HS, NS resuscitation was associated with an early activation of coagulation, natural anticoagulation, and endothelial systems, compared with HEX and FFP. |
| 2014 | Methylene blue modulates adhesion molecule expression on microvascular endothelial cells. | Inflamm Res | As methylene blue (MB) has been recently proposed to preserve blood pressure in case of vasoplegic syndrome and shock, an entity directly related to systemic inflammation, we aimed to elucidate the effect of MB on the expression of adhesion-molecules in endothelial-cells.Human microvascular endothelial-cells (HuMEC-1) were treated with 10, 30 or 60 µM MB for 30 min and 2 h each. Additionally, the treated HuMEC-1 were co-cultured with either human peripheral blood mononuclear cells (PBMCs) or Jurkat cells (human T-lymphocytes) for 2 h.HuMEC-1 were analyzed after MB treatment and after co-culture experiments for expression of different adhesion-molecules (ICAM-1, VCAM-1, L-selectin, E-selectin) via FACS measurement and western blot analysis. The supernatants of the experiments were analyzed with regard to the soluble forms of the adhesion molecules.We found that MB is able to modulate the expression of adhesion-molecules on EC. Administration of MB increases the expression of E-selectin and VCAM-1 depending on the dosage and time of exposure. ICAM-1 measurements provide evidence that different circulating blood cells can differently alter the adhesion-molecule expression on EC after MB exposure.Our results provide evidence regarding the immunomodulatory effect of MB upon endothelial-cells after inflammation. |
| 2014 | Human endothelial cell-based assay for endotoxin as sensitive as the conventional Limulus Amebocyte Lysate assay. | Biomaterials | Endotoxin, also known as lipopolysaccharide (LPS) produced by bacteria can be present in any liquid or on any biomaterial even if the material is sterile. Endotoxin in mammals can cause fever, inflammation, cell and tissue damage and irreversible septic shock and death. In the body, endothelial cells making up the blood vasculature and endothelial cells in vitro rapidly react to minute amounts of endotoxin resulting in a rapid induction of the cell adhesion molecule E-selectin. In this study we have used immunofluorescent staining to evaluate the expression of E-selectin on human microvascular endothelial cells from the skin (HDMEC) and human umbilical vein endothelial cells (HUVEC) exposed to various concentrations of LPS. In addition, the sensitivity of detection was compared with the most widely used assay for the presence of endotoxin, the Limulus Amebocyte Lysate assay (LAL). The detection of E-selectin on endothelial cells in the presence of LPS for 4 h was found to be at least as sensitive in detecting the same concentration using the LAL assay. A cell adhesion molecule-enzyme immunosorbent assay was also developed and used to quantify LPS using the endothelial cell model. A comparison of LAL and the immunofluorescent staining method was carried out with solutions, nanoparticles, biomaterial extracts and endothelial cells grown directly on biomaterials. Under all conditions, the endothelial/E-selectin model system was positive for the test samples that were positive by LAL. Thus, we propose the use of this highly sensitive, rapid, reproducible assay for the routine testing of endotoxin in all steps in the manufacturing process of materials destined for use in humans. This can give a rapid feedback and localization of bacterial contamination sources with the LAL being reserved for the testing of the final product. |
| 2013 | GroEL1, a heat shock protein 60 of Chlamydia pneumoniae, impairs neovascularization by decreasing endothelial progenitor cell function. | PLoS One | The number and function of endothelial progenitor cells (EPCs) are sensitive to hyperglycemia, hypertension, and smoking in humans, which are also associated with the development of atherosclerosis. GroEL1 from Chlamydia pneumoniae has been found in atherosclerotic lesions and is related to atherosclerotic pathogenesis. However, the actual effects of GroEL1 on EPC function are unclear. In this study, we investigate the EPC function in GroEL1-administered hind limb-ischemic C57BL/B6 and C57BL/10ScNJ (a toll-like receptor 4 (TLR4) mutation) mice and human EPCs. In mice, laser Doppler imaging, flow cytometry, and immunohistochemistry were used to evaluate the degree of neo-vasculogenesis, circulating level of EPCs, and expression of CD34, vWF, and endothelial nitric oxide synthase (eNOS) in vessels. Blood flow in the ischemic limb was significantly impaired in C57BL/B6 but not C57BL/10ScNJ mice treated with GroEL1. Circulating EPCs were also decreased after GroEL1 administration in C57BL/B6 mice. Additionally, GroEL1 inhibited the expression of CD34 and eNOS in C57BL/B6 ischemic muscle. In vitro, GroEL1 impaired the capacity of differentiation, mobilization, tube formation, and migration of EPCs. GroEL1 increased senescence, which was mediated by caspases, p38 MAPK, and ERK1/2 signaling in EPCs. Furthermore, GroEL1 decreased integrin and E-selectin expression and induced inflammatory responses in EPCs. In conclusion, these findings suggest that TLR4 and impaired NO-related mechanisms could contribute to the reduced number and functional activity of EPCs in the presence of GroEL1 from C. pneumoniae. |
| 2013 | αB-crystallin/HspB5 regulates endothelial-leukocyte interactions by enhancing NF-κB-induced up-regulation of adhesion molecules ICAM-1, VCAM-1 and E-selectin. | Angiogenesis | αB-crystallin is a small heat shock protein, which has pro-angiogenic properties by increasing survival of endothelial cells and secretion of vascular endothelial growth factor A. Here we demonstrate an additional role of αB-crystallin in regulating vascular function, through enhancing tumor necrosis factor α (TNF-α) induced expression of endothelial adhesion molecules involved in leukocyte recruitment. Ectopic expression of αB-crystallin in endothelial cells increases the level of E-selectin expression in response to TNF-α, and enhances leukocyte-endothelial interaction in vitro. Conversely, TNF-α-induced expression of intercellular adhesion molecule 1, vascular cell adhesion molecule 1 and E-selectin is markedly inhibited in endothelial cells isolated from αB-crystallin-deficient mice. This is associated with elevated levels of IκB in αB-crystallin deficient cells and incomplete degradation upon TNF-α stimulation. Consistent with this, endothelial adhesion molecule expression is reduced in inflamed vessels of αB-crystallin deficient mice, and leukocyte rolling velocity is increased. Our data identify αB-crystallin as a new regulator of leukocyte recruitment, by enhancing pro-inflammatory nuclear factor κ B-signaling and endothelial adhesion molecule expression during endothelial activation. |
| 2013 | Clinical review: Statins and trauma--a systematic review. | Crit Care | Statins, in addition to their lipid-lowering properties, have anti-inflammatory actions. The aim of this review is to evaluate the effect of pre-injury statin use, and statin treatment following injury. MEDLINE, EMBASE, and CENTRAL databases were searched to January 2012 for randomised and observational studies of statins in trauma patients in general, and in patients who have suffered traumatic brain injury, burns, and fractures. Of 985 identified citations, 7 (4 observational studies and 3 randomised controlled trials (RCTs)) met the inclusion criteria. Two studies (both observational) were concerned with trauma patients in general, two with patients who had suffered traumatic brain injury (one observational, one RCT), two with burns patients (one observational, one RCT), and one with fracture healing (RCT). Two of the RCTs relied on surrogate outcome measures. The observational studies were deemed to be at high risk of confounding, and the RCTs at high risk of bias. Three of the observational studies suggested improvements in a number of clinical outcomes in patients taking statins prior to injury (mortality, infection, and septic shock in burns patients; mortality in trauma patients in general; mortality in brain injured patients) whereas one, also of trauma patients in general, showed no difference in mortality or infection, and an increased risk of multi-organ failure. Two of three RCTs on statin treatment in burns patients and brain injured patients showed improvements in E-selectin levels and cognitive function. The third, of patients with radial fractures, showed no acceleration in fracture union. In conclusion, there is some evidence that pre-injury statin use and post-injury statin treatment may have a beneficial effect in patients who have suffered general trauma, traumatic brain injury, and burns. However, these studies are at high risk of confounding and bias, and should be regarded as 'hypothesisgenerating'. A well-designed RCT is required to determine the therapeutic efficacy in improving outcomes in this patient population. |
| 2013 | Biomarkers of endothelial activation/dysfunction in infectious diseases. | Virulence | Endothelial dysfunction contributes to the pathogenesis of a variety of potentially serious infectious diseases and syndromes, including sepsis and septic shock, hemolytic-uremic syndrome, severe malaria, and dengue hemorrhagic fever. Because endothelial activation often precedes overt endothelial dysfunction, biomarkers of the activated endothelium in serum and/or plasma may be detectable before classically recognized markers of disease, and therefore, may be clinically useful as biomarkers of disease severity or prognosis in systemic infectious diseases. In this review, the current status of mediators of endothelial cell function (angiopoietins-1 and -2), components of the coagulation pathway (von Willebrand Factor, ADAMTS13, and thrombomodulin), soluble cell-surface adhesion molecules (soluble E-selectin, sICAM-1, and sVCAM-1), and regulators of vascular tone and permeability (VEGF and sFlt-1) as biomarkers in severe infectious diseases is discussed in the context of sepsis, E. coli O157:H7 infection, malaria, and dengue virus infection. |
| 2013 | Biomarkers of endothelial cell activation in early sepsis. | Shock | The objective of this study was to investigate the association of endothelial-related markers with organ dysfunction and in-hospital mortality to validate our earlier findings in a multicenter study. We hypothesize that (i) endothelial biomarkers will be associated with organ dysfunction and mortality in sepsis and that (ii) soluble fms-like tyrosine kinase 1 (sFlt-1) holds promise as a novel prognostic marker in sepsis.This was a prospective, multicenter, observational study of a convenience sample of emergency department (ED) patients with a suspected infection presenting to one of four urban, academic medical center EDs between January 2009 and January 2010. We collected plasma while the patients were in the ED and subsequently assayed endothelial-related biomarkers, namely, sFlt-1, soluble E-selectin, soluble intercellular adhesion molecule 1, soluble vascular cell adhesion molecule 1, and plasminogen activator inhibitor 1 (PAI-1). Outcomes were organ dysfunction and in-hospital mortality.We enrolled a total of 166 patients: 63 with sepsis (38%), 61 with severe sepsis (37%), and 42 with septic shock (25%). All endothelial biomarkers were significantly associated with sepsis severity, P < 0.002. We found a significant intercorrelation between all biomarkers, strongest between sFlt-1 and PAI-1 (r = 0.61, P < 0.001) and PAI-1 and soluble E-selectin and soluble intercellular adhesion molecule 1 (r = 0.49, P < 0.001). Among the endothelial biomarkers, sFlt-1 had the strongest association with Sequential Organ Failure Assessment score (r = 0.58, P < 0.001). Soluble fms-like tyrosine kinase 1 and PAI-1 had the highest area under the operating receiver characteristic curve for mortality of 0.87.This multicenter validation study confirms that markers of endothelial activation are associated with sepsis severity, organ dysfunction, and mortality in sepsis. This supports the hypothesis that the endothelium plays a central role in the pathophysiology of sepsis and may serve as a more accurate prediction tool and a target for therapies aimed at ameliorating endothelial cell dysfunction. In addition, sFLT-1 holds promise as a novel sepsis severity biomarker. |
| 2013 | Milan PM1 induces adverse effects on mice lungs and cardiovascular system. | Biomed Res Int | Recent studies have suggested a link between inhaled particulate matter (PM) exposure and increased mortality and morbidity associated with cardiorespiratory diseases. Since the response to PM1 has not yet been deeply investigated, its impact on mice lungs and cardiovascular system is here examined. A repeated exposure to Milan PM1 was performed on BALB/c mice. The bronchoalveolar lavage fluid (BALf) and the lung parenchyma were screened for markers of inflammation (cell counts, tumor necrosis factor-α (TNF-α); macrophage inflammatory protein-2 (MIP-2); heme oxygenase-1 (HO-1); nuclear factor kappa-light-chain-enhancer of activated B cells p50 subunit (NFκB-p50); inducible nitric oxide synthetase (iNOS); endothelial-selectin (E-selectin)), cytotoxicity (lactate dehydrogenase (LDH); alkaline phosphatase (ALP); heat shock protein 70 (Hsp70); caspase-8-p18), and a putative pro-carcinogenic marker (cytochrome 1B1 (Cyp1B1)). Heart tissue was tested for HO-1, caspase-8-p18, NFκB-p50, iNOS, E-selectin, and myeloperoxidase (MPO); plasma was screened for markers of platelet activation and clot formation (soluble platelet-selectin (sP-selectin); fibrinogen; plasminogen activator inhibitor 1 (PAI-1)). PM1 triggers inflammation and cytotoxicity in lungs. A similar cytotoxic effect was observed on heart tissues, while plasma analyses suggest blood-endothelium interface activation. These data highlight the importance of lung inflammation in mediating adverse cardiovascular events following increase in ambient PM1 levels, providing evidences of a positive correlation between PM1 exposure and cardiovascular morbidity. |
| 2013 | Ex vivo lung evaluation of prearrest heparinization in donation after cardiac death. | Ann Surg | To evaluate the effects of prearrest heparin administration on lung quality in a model of donation after cardiac death (DCD), and to assess the potential application of ex vivo lung perfusion (EVLP) in the identification of better grafts from the DCD donor pool.Cardiac death was induced by electric shock in 10 pigs. One group received a prearrest heparin dose of 300 units/kg (H group, n = 5) and the other did not (NH group, n = 5). Animals remained at room temperature for 1 hour without ventilation, defining the warm ischemic time. After harvest, the lungs underwent 6 hours of cold ischemia before being evaluated with EVLP for 4 hours.Static compliance 28 ± 3 versus 29 ± 2 (Cstat-cm H2O), pulmonary vascular resistance (PVR) 593 ± 127 versus 495 ± 70 (PVR-dyn·s/cm), and oxygenation 327 ± 32 versus 330 ± 28 (ΔPO2-mm Hg) remained stable from the beginning until the end of EVLP in the H group. In the NH group, Cstat started to decline after the first hour (25 ± 2 vs 21 ± 2), ΔPO2 after hour 2 (265 ± 44 vs 207 ± 44), and PVR started to increase after hour 3 (765 ± 132 vs 916 ± 168). Significant differences between the groups were observed at the end of EVLP (P < 0.001). Parameters of lung quality after EVLP also showed significant differences between the groups: wet weight-to-dry weight ratio (P < 0.001), protein in the bronchial lavage (P < 0.01), Na + K-ATPase activity (P < 0.001), and E-selectin (P < 0.001) in the perfusate.Prearrest heparin administration improved organ function by preserving endothelial homeostasis. EVLP proved to be a useful platform for assessing DCD lungs, providing reliable means of discriminating injured grafts. |
| 2012 | Resuscitation of traumatic hemorrhagic shock patients with hypertonic saline-without dextran-inhibits neutrophil and endothelial cell activation. | Shock | Posttraumatic inflammation and excessive neutrophil activation cause multiple organ dysfunction syndrome (MODS), a major cause of death among hemorrhagic shock patients. Traditional resuscitation strategies may exacerbate inflammation; thus, novel fluid treatments are needed to reduce such posttraumatic complications. Hypertonic resuscitation fluids inhibit inflammation and reduce MODS in animal models. Here we studied the anti-inflammatory efficacy of hypertonic fluids in a controlled clinical trial. Trauma patients in hypovolemic shock were resuscitated in a prehospital setting with 250 mL of either 7.5% hypertonic saline (HS; n = 9), 7.5% hypertonic saline + 6% dextran 70 (HSD; n = 8), or 0.9% normal saline (NS; n = 17). Blood samples were collected on hospital admission and 12 and 24 h after resuscitation. Multicolor flow cytometry was used to quantify neutrophil expression of cell-surface activation/adhesion (CD11b, CD62L, CD64) and degranulation (CD63, CD66b, CD35) markers as well as oxidative burst activity. Circulating concentrations of soluble intercellular adhesion molecule-1 (sICAM-1), vascular cell adhesion molecule-1 (sVACM-1), P- and E-selectins, myeloperoxidase (MPO), and matrix metalloproteinase 9 (MMP-9) were assessed by immunoassay. Multiple organ dysfunction syndrome, leukocytosis, and mortality were lower in the HS and HSD groups than in the NS group. However, these differences were not statistically significant. Hypertonic saline prevented priming and activation and neutrophil oxidative burst and CD11b and CD66b expression. Hypertonic saline also reduced circulating markers of neutrophil degranulation (MPO and MMP-9) and endothelial cell activation (sICAM-1, sVCAM-1, soluble E-selectin, and soluble P-selectin). Hypertonic saline + 6% dextran 70 was less capable than HS of suppressing the upregulation of most of these activation markers. This study demonstrates that initial resuscitation with HS, but neither NS nor HSD, can attenuate posttraumatic neutrophil and endothelial cell activation in hemorrhagic shock patients. These data suggest that hypertonic resuscitation without dextran may inhibit posttraumatic inflammation. However, despite this effect, neither HS nor HSD reduced MODS in trauma patients with hemorrhagic shock. |
| 2012 | MicroRNA-126 contributes to renal microvascular heterogeneity of VCAM-1 protein expression in acute inflammation. | Am J Physiol Renal Physiol | Endothelial cells in different microvascular segments of the kidney have diverse functions and exhibit differential responsiveness to disease stimuli. The responsible molecular mechanisms are largely unknown. We previously showed that during hemorrhagic shock, VCAM-1 protein was expressed primarily in extraglomerular compartments of the kidney, while E-selectin protein was highly induced in glomeruli only (van Meurs M, Wulfert FM, Knol AJ, de Haes A, Houwertjes M, Aarts LPHJ, Molema G. Shock 29: 291-299, 2008). Here, we investigated the molecular control of expression of these endothelial cell adhesion molecules in mouse models of renal inflammation. Microvascular segment-specific responses to the induction of anti-glomerular basement membrane (anti-GBM), glomerulonephritis and systemic TNF-α treatment showed that E-selectin expression was transcriptionally regulated, with high E-selectin mRNA and protein levels preferentially expressed in the glomerular compartment. In contrast, VCAM-1 mRNA expression was increased in both arterioles and glomeruli, while VCAM-1 protein expression was limited in the glomeruli. These high VCAM-1 mRNA/low VCAM-1 protein levels were accompanied by high local microRNA (miR)-126 and Egfl7 levels, as well as higher Ets1 levels compared with arteriolar expression levels. Using miR-reporter constructs, the functional activity of miR-126 in glomerular endothelial cells could be demonstrated. Moreover, in vivo knockdown of miR-126 function unleashed VCAM-1 protein expression in the glomeruli upon inflammatory challenge. These data imply that miR-126 has a major role in the segmental, heterogenic response of renal microvascular endothelial cells to systemic inflammatory stimuli. |
| 2012 | Fusobacterium nucleatum GroEL induces risk factors of atherosclerosis in human microvascular endothelial cells and ApoE(-/-) mice. | Mol Oral Microbiol | Infection and inflammation are risk factors in the initiation and progression of atherosclerosis. Periodontitis is one of the most prevalent chronic inflammations of the oral cavity, and has been reported to be associated with systemic disease. In this study, we evaluated whether the heat-shock protein GroEL of Fusobacterium nucleatum, one of the most prevalent bacteria in periodontitis, induces factors that predispose to atherosclerosis in human microvascular endothelial cells (HMEC-1) and apolipoprotein E-deficient (ApoE(-/-)) mice. GroEL induced the expression of chemokines such as monocyte chemoattractant protein-1 and interleukin-8 as well as cell adhesion molecules, such as intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and E-selectin. GroEL induced the activity of tissue factor and reduced the activity of the tissue factor pathway inhibitor. Foam cell formation was induced by GroEL. GroEL-injected ApoE(-/-) mice showed significant atherosclerotic lesion progression compared with control mice. Serum levels of risk factors for atherosclerosis such as interleukin-6, C-reactive protein, and low-density lipoprotein were increased in GroEL-injected ApoE(-/-) mice compared with control mice, whereas serum levels of high-density lipoprotein were decreased. We could detect significantly higher levels of anti-F. nucleatum GroEL antibody in serum and F. nucleatum DNA in gingival crevicular fluid from patients with periodontitis than in that from healthy subjects. Our results indicate that the host response to the GroEL of periodontal pathogens like F. nucleatum may be a mechanism involved in atherosclerosis, supporting the association of periodontitis and systemic infection. |
| 2011 | The effect of resuscitation fluids on neutrophil-endothelial cell interactions in septic shock. | Shock | Fluid resuscitation is essential in the treatment of septic shock. This study examined the effect of resuscitative fluids (RFs) on sepsis-induced neutrophil-endothelial cell interactions. The RFs studied were 0.9% saline (NS), Ringer's lactate (RL), 7.5% saline and dextran-70 (DHS), 5% albumin (AL), and 6% hydroxyethyl starch (HS). Platelets and neutrophils were obtained from normal volunteers, and plasma was obtained from patients with septic shock. Microslides coated with human umbilical endothelial vein cell (HUVEC) and platelet-neutrophil solutions were primed with septic plasma with/without the RF. Neutrophil rolling velocity, leukoaggregation, and neutrophil adherence were determined. Separately, platelet-neutrophil solutions and endothelial cells were exposed to septic plasma with/without RFs, and cellular activation, neutrophil superoxide production, and endothelial cell E-selectin expression were assessed. Ringer's lactate decreased neutrophil rolling velocity and increased aggregation and adherence. Normal saline had no effect on these parameters. Hydroxyethyl starch and AL increased neutrophil rolling velocity and decreased adherence and aggregation when HUVECs were preincubated with the RF. Dextran-70 and 7.5% saline decreased neutrophil-endothelial cell interactions in both HUVECs and platelet/neutrophil preincubated experiments. Ringer's lactate increased activation of neutrophils and platelets, whereas AL decreased their activation. Other than NS, all the RFs increased neutrophil superoxide production. Ringer's lactate increased endothelial cell E-selectin release, whereas AL and HS both decreased its release. These data suggest that fluids used in the resuscitation of septic shock vary in their effects on sepsis-induced neutrophil-endothelial cell interactions. Ringer's lactate amplifies the effects of sepsis, while NS appears to have minimal impact. Dextran-70 and 7.5% saline, AL, and HS in varying degrees decrease sepsis-related neutrophil-endothelial cell interactions and activation. |
| 2011 | Coenzyme Q10 levels are low and may be associated with the inflammatory cascade in septic shock. | Crit Care | Mitochondrial dysfunction is associated with increased mortality in septic shock. Coenzyme Q10 (CoQ10) is a key cofactor in the mitochondrial respiratory chain, but whether CoQ10 is depleted in septic shock remains unknown. Moreover, statin therapy may decrease CoQ10 levels, but whether this occurs acutely remains unknown. We measured CoQ10 levels in septic shock patients enrolled in a randomized trial of simvastatin versus placebo.We conducted a post hoc analysis of a prospective, randomized trial of simvastatin versus placebo in patients with septic shock (ClinicalTrials.gov ID: NCT00676897). Adult patients with suspected or confirmed infection and the need for vasopressor support were included in the initial trial. For the current analysis, blood specimens were analyzed for plasma CoQ10 and low-density lipoprotein (LDL) levels. The relationship between CoQ10 levels and inflammatory and vascular endothelial biomarkers was assessed using either the Pearson or Spearman correlation coefficient.We analyzed 28 samples from 14 patients. CoQ10 levels were low, with a median of 0.49 (interquartile range 0.26 to 0.62) compared to levels in healthy control patients (CoQ10 = 0.95 μmol/L ± 0.29; P < 0.0001). Statin therapy had no effect on plasma CoQ10 levels over time (P = 0.13). There was a statistically significant relationship between plasma CoQ10 levels and levels of vascular cell adhesion molecule (VCAM) (r2 = 0.2; P = 0.008), TNF-α (r2 = 0.28; P = 0.004), IL-8 (r2 = 0.21; P = 0.015), IL-10 (r2 = 0.18; P = 0.025), E-selectin (r2 = 0.17; P = -0.03), IL-1ra (r2 = 0.21; P = 0.014), IL-6 (r2 = 0.17; P = 0.029) and IL-2 (r2 = 0.23; P = 0.009). After adjusting for LDL levels, there was a statistically significant inverse relationship between plasma CoQ10 levels and levels of VCAM (r2 = 0.24; P = 0.01) (Figure 3) and IL-10 (r2 = 0.24; P = 0.02).CoQ10 levels are significantly lower in septic shock patients than in healthy controls. CoQ10 is negatively associated with vascular endothelial markers and inflammatory molecules, though this association diminishes after adjusting for LDL levels. |
| 2011 | Hemorrhagic shock-induced endothelial cell activation in a spontaneous breathing and a mechanical ventilation hemorrhagic shock model is induced by a proinflammatory response and not by hypoxia. | Anesthesiology | The interaction between neutrophils and activated endothelium is essential for the development of multiple organ dysfunction in patients with hemorrhagic shock (HS). Mechanical ventilation frequently is used in patients with HS. The authors sought to investigate the consequences of mechanical ventilation of mice subjected to HS on microvascular endothelial activation in the lung and kidney.Anesthetized wild type C57BL/6 male mice were subjected to controlled hemorrhage; subgroups of mice were mechanically ventilated during the HS insult. To study the effect of acute hypoxia on the mice, the animals were housed in hypoxic cages. Gene expression levels was assessed by quantitative real-time polymerase chain reaction. Protein expression was assessed by immunohistochemistry and enzyme-linked immunosorbent assay.Ninety minutes after the shock induction, a vascular bed-specific, heterogeneous proinflammatory endothelial activation represented by E-selectin, vascular cell adhesion molecule 1, and intercellular adhesion molecule 1 expression was seen in kidney and lung. No differences in adhesion molecules between the spontaneously breathing and mechanically ventilated mice were found. Concentrations of the proinflammatory cytokines chemokine (C-X-C motif) ligand 1 (11.0-fold) and interleukin-6 (21.7-fold) were increased after 90 min of HS. Two hours of 6% oxygen did not induce the expression of E-selectin, vascular cell adhesion molecule 1, and intercellular adhesion molecule 1 in the kidneys and the lung.Hemorrhagic shock leads to an early and reversible proinflammatory endothelial activation in kidney and lung. HS-induced endothelial activation is not changed by mechanical ventilation during the shock phase. Hypoxia alone does not lead to endothelial activation. The observed proinflammatory endothelial activation is mostly ischemia- or reperfusion-dependent and not related to hypoxia. |
| 2011 | Endothelial cell activation in emergency department patients with sepsis-related and non-sepsis-related hypotension. | Shock | Previous studies found increased circulating levels of biomarkers related to endothelial cell activation in patients with sepsis, particularly in the most severe sepsis stages of sepsis shock. It remains unclear, however, whether this activation is mainly driven by sepsis-specific mechanisms or occurs as a generalized inflammatory response. The objective of this analysis was to compare patterns of biomarkers of endothelial cell activation in patients with hypotension due to sepsis and nonsepsis etiologies. This is a secondary analysis of a prospective, observational cohort study including emergency department patients older than17 years with an episode of hypotension defined as any systolic blood pressure measurement less than 100 mmHg. Etiology of hypotension episodes was classified as sepsis or nonsepsis (eg, cardiac or hemorrhagic). Endothelial activation biomarkers of cell adhesion (E-selectin, vascular cell adhesion molecule 1 [VCAM-1], and intercellular adhesion molecule 1 [ICAM-1]), coagulation (plasminogen activator inhibitor 1 [PAI-1]), and vascular endothelial growth factor (VEGF) signaling (VEGF, soluble fms-like tyrosine kinase 1 [sFLT-1]) were assayed. A total of 161 patients were analyzed. Hypotension was classified as sepsis (n = 69), nonsepsis (cardiac [n = 35], hemorrhagic [n = 12]), or indeterminate (n = 45). With the exception of PAI-1, median plasma levels of all endothelial markers were significantly higher in patients with sepsis compared with nonsepsis etiology (P < 0.05 for all comparisons). Logistic regression analysis, adjusted for age, sex, mean blood pressure level, and mortality, confirmed a significant association of E-selectin (odds ratio [OR], 3.7; 95% confidence interval [CI], 1.7-7.8, P < 0.001) and sFLT-1 (OR, 2.0; CI, 1.1-3.8; P < 0.03) with sepsis etiology. Biomarkers VCAM-1 (OR, 2.0; CI, 0.88-4.4; P = 0.1), VEGF (OR, 1.5; CI, 0.98-2.2; P = 0.06), ICAM-1 (OR, 1.5; CI, 0.9-2.6; P = 0.2), and PAI-1 (OR, 1.4; CI, 0.8-2.3; P = 0.2) did not reach statistical significance. This study found a sepsis-specific activation of endothelium activation markers, particularly E-selectin and sFLT-1, in emergency department patients with hypotension. |
| 2011 | Therapeutic effects of melatonin on heatstroke-induced multiple organ dysfunction syndrome in rats. | J Pineal Res | Melatonin reportedly exerts beneficial effects to attenuate multiple organ dysfunction syndrome (MODS) in septic shock. Heatstroke resembles septic shock in many aspects. Thus, this study was performed on the anesthetized rats by using heat exposure to induce heatstroke-associated MODS. We evaluated the effect of melatonin, a versatile molecule synthesized in the pineal gland and in many organs, in heatstroke rats and showed that melatonin (0.2-5.0 mg/kg of body weight, i.v., immediately after the start of heat stress) significantly (i) attenuated hyperthermia, hypotension and hypothalamic ischemia and hypoxia, (ii) reduced plasma index of the toxic oxidizing radicals like nitric oxide metabolites and hydroxyl radicals, (iii) diminished plasma index of hepatic and renal dysfunction like creatinine, blood urea nitrogen, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and lactate dehydrogenase, (iv) attenuated plasma systemic inflammation response molecules like soluble intercellular and lesion molecule-1, E-selectin, tumor necrosis factor-alpha, interleukin (IL)-1β, and IL-6, (v) promoted plasma levels of an anti-inflammatory cytokine IL-10, (vi) reduced an index of infiltration of polymorphonuclear neutrophils in the lung like myeloperoxidase activity, and (vii) promoted the survival time to fourfold compared with the heatstroke alone group. Thus, melatonin could be a novel agent for the treatment of heatstroke animals or patients in the early stage. |
| 2011 | Influence of diabetes on endothelial cell response during sepsis. | Diabetologia | Several endothelial pathways of cell adhesion, coagulation and vascular endothelial growth factor (VEGF) signalling are activated during sepsis. The objective of this analysis was to investigate the influence of diabetes on biomarkers of endothelial cell activation in sepsis.This was a prospective observational cohort study of a convenience sample of adult patients (age ≥ 18 years) for whom infection was clinically suspected and who presented to an urban tertiary care emergency department between February 2005 and November 2008. We investigated the association of diabetes and sepsis with various endothelial activation biomarkers of cell adhesion (E-selectin, vascular cell adhesion molecule 1 [VCAM-1] and intercellular adhesion molecule 1 [ICAM-1]), coagulation (plasminogen activator inhibitor 1 [PAI-1]) and VEGF signalling (soluble fms-like tyrosine kinase-1 [sFLT-1]).A total of 207 patients (34% with sepsis, 32% with severe sepsis and 34% with septic shock) were studied, including 63 (30%) with diabetes. Compared with patients without diabetes, patients with diabetes had significantly increased E-selectin and sFLT-1 levels overall; this was most pronounced during septic shock in the stratified analysis. Multivariate models including age, sex, sepsis severity and other variables as potential covariates confirmed the association of diabetes with elevated circulating plasma levels of E-selectin (standardised β 0.24, p < 0.001) and sFLT-1 (standardised β 0.19, p < 0.01), but there was no significant association with VCAM-1, ICAM-1 or PAI-1.During septic shock, patients with diabetes had higher levels of circulating biomarkers of endothelial cell adhesion (E-selectin) and VEGF signalling (sFLT-1). Future studies should address whether enhanced activation of the endothelium places patients with diabetes at increased risk for the development of sepsis and worsening morbidity and mortality. |
| 2010 | GroEL and lipopolysaccharide from Francisella tularensis live vaccine strain synergistically activate human macrophages. | Infect Immun | Francisella tularensis, the causative agent of tularemia, interacts with host cells of innate immunity in an atypical manner. For most Gram-negative bacteria, the release of lipopolysaccharide (LPS) from their outer membranes stimulates an inflammatory response. When LPS from the attenuated live vaccine strain (LVS) or the highly virulent Schu S4 strain of F. tularensis was incubated with human umbilical vein endothelial cells, neither species of LPS induced expression of the adhesion molecule E-selectin or secretion of the chemokine CCL2. Moreover, a high concentration (10 microg/ml) of LVS or Schu S4 LPS was required to stimulate production of CCL2 by human monocyte-derived macrophages (huMDM). A screen for alternative proinflammatory factors of F. tularensis LVS identified the heat shock protein GroEL as a potential candidate. Recombinant LVS GroEL at a concentration of 10 microg/ml elicited secretion of CXCL8 and CCL2 by huMDM through a TLR4-dependent mechanism. When 1 microg of LVS GroEL/ml was added to an equivalent amount of LVS LPS, the two components synergistically activated the huMDM to produce CXCL8. Schu S4 GroEL was less stimulatory than LVS GroEL and showed a lesser degree of synergy when combined with Schu S4 LPS. These findings suggest that the intrinsically low proinflammatory activity of F. tularensis LPS may be increased in the infected human host through interactions with other components of the bacterium. |
| 2008 | The altered expression of inflammation-related molecules and secretion of IL-6 and IL-8 by HUVEC from newborns with maternal inactive systemic lupus erythematosus is modified by estrogens. | Lupus | Systemic lupus erythematosus (SLE) predominantly affects women, especially those in reproductive age. Genetic contributions to disease susceptibility as well as immune dysregulation, particularly persistent inflammatory responses, are considered essential features. Our aim was to determine whether human umbilical vein endothelial cells (HUVEC) isolated from healthy newborns to women with inactive SLE show inflammation-related abnormalities that might lead to an early development of SLE in the offsprings. HUVEC isolated from six women with inactive SLE were stimulated with 2.5 ng/mL of TNF-alpha and/or physiological and pharmacological doses of 17-I(2) estradiol (E2). Then the expression of VCAM-1, ICAM-1, E-selectin, toll-like receptor-9 (TLR-9), heat shock protein 70 (HSP70) and HSP90 were measured. The concentrations of IL-6, IL-8, and IL-10 were also determined in maternal serum and in TNF-alpha stimulated and non-stimulated HUVEC culture supernatant. HUVEC from children with no family history of autoimmune disease served as controls. Our results showed that in HUVEC from SLE+ mothers, a constitutively low expression of adhesion molecules was enhanced by TNF-alpha treatment. The E2 (1 ng/mL) increased the expression of adhesion molecules but had no effect upon TNF-alpha-treated cells. IL-6 was constitutively higher in SLE+ HUVEC, whereas IL-8 was lower; E2 treatment diminished the latter. The E2 had no effect upon IL-6 and IL-8 secretions in TNF-alpha-treated cells. SLE+ HUVEC showed a disordered cytoskeleton and overexpressed HSP70, HSP90, and TLR-9. Our results indicate that endothelial cells of newborns to SLE+ mothers are in a proinflammatory condition which can be upregulated by estrogens. |
| 2009 | The importance of the effect of shear stress on endothelial cells in determining the performance of hemoglobin based oxygen carriers. | Biomaterials | The lack of blood donations and the threat of infections from blood and blood products have led to extensive research into the development of blood substitutes. The latest generation of hemoglobin based oxygen carriers (HBOC) has been shown to induce side effects like hypertension, vasoconstriction, inflammation and oxidative stress. HBOC are able to restore volemia and transport oxygen after a hemorrhagic shock, the reperfusion leading to the restoration of the blood flow in vessels. We propose an innovative approach, more closely emulating clinical situations, to assess the impact of HBOC perfusion on endothelial cells (EC) in vitro. Through this approach we quantified levels of oxidative stress, vasoactive factors and inflammation. EC were cultivated under a laminar flow to reproduce the return of shear stress (SS) during the reperfusion. We showed that heme oxygenase I transcription correlated with changes in oxidatively modified heme and methemoglobin; all were lower under SS. SS induced increased nitric oxide production, which may have implications for the mechanism of in vivo vasoconstriction and hypertension. E-selectin changes under SS were greater than those of ICAM-1. Our results demonstrate how it is essential to include SS in assays attempting to understand the potential vascular side effects of HBOC perfusion. |
| 2008 | Differential dose-dependent effects of prednisolone on shedding of endothelial adhesion molecules during human endotoxemia. | Immunol Lett | Low dose prednisolone was shown to be beneficial in the treatment of the Acute respiratory distress syndrome (ARDS) and septic shock. One corticosteroid-induced effect, postulated to mediate corticosteroid-induced anti-inflammatory effects, is decreased expression of adhesion molecules on endothelial cells, thereby preventing leukocyte recruitment at inflammatory sites. The current study aimed to investigate the effect of increasing doses of prednisolone on the release of soluble adhesion molecules in healthy volunteers challenged with endotoxin. Therefore, 32 healthy, male volunteers received prednisolone orally at doses of 0mg, 3mg, 10mg or 30 mg at 2h before injection of endotoxin prepared from Escherichia coli (4 ng/kg) and levels of soluble E-selectin (sE-selectin), soluble VCAM-1 (sVCAM-1) and soluble ICAM-1 (sICAM-1) were measured. Levels of all markers were increased after induction of endotoxemia. Levels of sE-selectin were inhibited by a dose of 3mg prednisolone and levels of sVCAM-1 were decreased after a dose of 10mg. Maximal inhibition of both sE-selectin and sVCAM-1 levels was achieved by the highest dose of prednisolone 30 mg. Remarkably, prednisolone 3mg potentiated endotoxin-induced sVCAM-1 release. Levels of sICAM-1 were not affected by prednisolone. Together, the data suggest that prednisolone differentially and dose-dependently influences the release of soluble endothelial adhesion molecules during human endotoxemia. |
| 2008 | Soluble intercellular adhesion molecule-1 (sICAM-1) and aortic valve calcification in the multi-ethnic study of atherosclerosis (MESA). | J Heart Valve Dis | Previous studies have reported associations between individual inflammatory biomarkers and aortic valve disease, but none has examined associations with, baseline prevalence or severity of aortic valve calcium (AVC), as measured with cardiac computed tomography (CT). The study aim was to determine whether specific inflammatory markers were associated with AVC in the Multi-Ethnic Study of Atherosclerosis (MESA) cohort.The associations of inflammatory biomarkers, including C-reactive protein (CRP), interleukin-6 (IL-6), fibrinogen, D-dimer, soluble intercellular adhesion molecule-1 (sICAM-1), heat shock protein 60 (Hsp60), soluble tumor necrosis factor receptor-1 (sTNF-R1), soluble tissue factor (sTF), soluble E-selectin and matrix metalloproteinases-3 and -9 (MMP-3 and -9) with baseline AVC prevalence and severity were examined.After adjusting for age, gender, race and cardiovascular risk factors (smoking, hypertension, diabetes, total cholesterol, HDL, serum creatinine, body mass index and lipid-lowering therapy), the point prevalence ratios (95% confidence interval) for prevalent AVC were 1.20 (1.04,1.39) for sICAM-1,1.13 (1.03, 1.24) for IL-6, and 1.11 (1.02, 1.21) for D-dimer. No other associations were statistically significant. When CRP, sICAM-1, IL-6 and D-dimer were modeled together, only sICAM-1 remained associated with increased AVC prevalence (1.18 (1.02, 1.38)). Only sICAM-1 was associated with increased AVC severity (relative difference (95% CI): 1.18 (1.00, 1.39)).In this large, multi-ethnic, asymptomatic cohort, sICAM-1- a marker of endothelial perturbation - was the only biomarker associated with both an increased prevalence and severity of AVC. |
| 2009 | Synthetic oligopeptides related to the [beta]-subunit of human chorionic gonadotropin attenuate inflammation and liver damage after (trauma) hemorrhagic shock and resuscitation. | Shock | Severe hemorrhagic shock (HS) followed by resuscitation induces a massive inflammatory response, which may culminate into systemic inflammatory response syndrome, multiple organ dysfunction syndrome, and, finally, death. Treatments that effectively prevent this inflammation are limited so far. In a previous study, we demonstrated that synthetic oligopeptides related to the primary structure of human chorionic gonadotropin (HCG) can inhibit the inflammatory response and mortality that follow high-dose LPS-induced inflammation. Considering this powerful anti-inflammatory effect, we investigated whether administration of similar synthetic HCG-related oligopeptides (LQGV, AQGV, LAGV) during HS were able to attenuate the inflammatory response associated with this condition. Hemorrhagic shock was induced in rats for 60 min by blood withdrawal until a MAP of 40 mmHg was reached. Rats received a single injection with one of the hCG-related oligopeptides (LQGV, AQGV or LAGV) or 0.9% NaCl solution as control 30 min after induction of HS. Treatment with LQGV, AQGV, or LAGV prevented systemic release of TNF-[alpha] and IL-6 and was associated with reduced TNF-[alpha], IL-6, and E-selectin mRNA transcript levels in the liver. LQGV treatment prevented neutrophil infiltration into the liver and was associated with reduced liver damage. Our data suggest that HCG-related oligopeptides, in particular LQGV, have therapeutic potential by attenuating the life-threatening inflammation and organ damage that is associated with (trauma) HS and resuscitation. |
| 2008 | The anti-endotoxic effects of the KSL-W decapeptide on Escherichia coli O55:B5 and various oral lipopolysaccharides. | J Periodontal Res | Host responses following the recognition of bacterial lipopolysaccharide can range from acute inflammation to septic shock. The aim of this study was to evaluate the ability of the KSL-W decapeptide to bind to and block the endotoxic effects of lipopolysaccharide.An enzyme-linked immunosorbent assay-based binding assay using fluorescently labeled KSL-W to detect adsorbed Escherichia coli O55:B5 lipopolysaccharide was employed. A commercially available recombinant Factor C lipopolysaccharide detection assay, hemagglutination of rabbit erythrocytes as well as E-selectin expression in human umbilical vein endothelial cells were used to assess the anti-endotoxic effects after KSL-W exposure to E. coli lipopolysaccharide as well as to oral lipopolysaccharide samples.Lipopolysaccharide-binding assays using E. coli O55:B5 lipopolysaccharide revealed both a higher maximal binding range (532-713 microM) and a half-maximum binding concentration (70-185 microM) for the KSL-W peptide when compared with its analog control. Significant inhibition of E-selectin expression in human umbilical vein endothelial cells (p < 0.0001) as well as hemagglutination of rabbit erythrocytes occurred after the interaction of KSL-W with E. coli lipopolysaccharide. Recombinant Factor C enzyme detection inhibition revealed dose-dependent inhibition values ranging from 1.0-51.8 microM. which were dependent upon the type of lipopolysaccharide sample tested.These results demonstrate that for the concentrations tested, the KSL-W decapeptide was nontoxic to mammalian cells and could bind to and block the host recognition and response towards enteric, as well as oral, lipopolysaccharide samples. |
| 2008 | Pathophysiological changes of gram-negative bacterial infection can be reproduced by a synthetic peptide mimicking loop L7 sequence of Haemophilus influenzae porin. | Microbes Infect | Several in vivo models have been used to dissect the molecular mechanisms that contribute to activate the coagulation and fibrinolytic systems by bacteria and bacterial products but many aspects remain poorly understood. In this study we examined the in vivo effect of the synthetic peptide corresponding to loop L7 from Haemophilus influenzae type b (Hib) porin to evaluate its role on the coagulative/fibrinolytic cascade and the circulating markers of endothelial injury. Plasma was obtained from rats injected intravenously with loop L7, Hib porin or a scrambled peptide and tested for fragment 1+2 (F1+2), tissue-type plasminogen activator (tPA), plasminogen activator inhibitor type I (PAI-1) antigen, von Willebrand factor (vWF) and soluble E-selectin (sE-selectin). The coagulative/fibrinolytic cascade was impaired as shown by PAI-1 level increased. Concomitantly, E-selectin, a marker of endothelial injury, was also significantly elevated. In addition either loop L7 or Hib porin injection induced hyperglycaemia and inflammatory cytokine production. The data were correlated with hemodynamic functions. The results indicate that loop L7 plays an essential role in the pathophysiologic events observed during gram-negative infection. These findings may have implications for the development of alternative therapies to counteract excessive inflammatory responses during septic shock. |
| 2008 | [Regulation of inflammatory responses by endothelial cells--understanding the molecular mechanism(s) and its therapeutic application to sepsis]. | Masui | Endothelial cells are activated by shear-stress and inflammatory mediators that are capable of activating sensory neurons. Activated endothelial cells increase the production of nitric oxide and prostaglandins, thereby regulating inflammatory responses induced by various insults. Dysfunction of sensory neurons and excess inflammatory mediators released from activated neutrophils damage endothelial cells, thereby increasing inflammatory responses such as an increase in tumor necrosis factor production. Pulmonary endothelial dysfunction plays a critical role in the development of acute lung injury and shock, leading to multi-organ failure. Determination of soluble E-selectin in serum samples of patients with sepsis predicts the future development of acute lung injury. Therapeutic agents that are capable of stimulating sensory neurons or inhibiting leukocyte activation might be useful in the treatment of severe sepsis especially when these agents are administered in the early stage of severe sepsis. |
| 2008 | Early organ-specific endothelial activation during hemorrhagic shock and resuscitation. | Shock | Multiple organ dysfunction syndrome (MODS) is a complication of hemorrhagic shock (HS) and related to high morbidity and mortality. Interaction of activated neutrophils and endothelial cells is considered to play a prominent role in the pathophysiology of MODS. Insight in the nature and molecular basis of endothelial cell activation during HS can assist in identifying new rational targets for early therapeutic intervention. In this study, we examined the kinetics and organ specificity of endothelial cell activation in a mouse model of HS. Anesthetized male mice were subjected to controlled hemorrhage to a MAP of 30 mmHg. Mice were killed after 15, 30, 60, or 90 min of HS. After 90 min of hemorrhagic shock, a group of mice was resuscitated with 6% hydroxyethyl starch 130/0.4. Untreated mice and sham shock mice that underwent instrumentation and 90 min of anesthesia without shock served as controls. Gene expression levels of inflammatory endothelial cell activation (P-selectin, E-selectin, vascular cell adhesion molecule 1, and intercellular adhesion molecule 1) and hypoxia-responsive genes (vascular endothelial growth factor and hypoxia-inducible factor 1alpha) were quantified in kidney, liver, lung, brain, and heart tissue by quantitative reverse-transcription-polymerase chain reaction. Furthermore, we examined a selection of these genes with regard to protein expression and localization using immunohistochemical analysis. Induction of inflammatory genes occurred early during HS and already before resuscitation. Expression of adhesion molecules was significantly induced in all organs, albeit to a different extent depending on the organ. Endothelial genes CD31 and VE-cadherin, which function in endothelial cell homeostasis and integrity, were not affected during the shock phase except for VE-cadherin in the liver, which showed increased mRNA levels. The rapid inflammatory activation was not paralleled by induction of hypoxia-responsive genes. This study demonstrated the occurrence of early and organ-specific endothelial cell activation during hemorrhagic shock, as presented by induced expression of inflammatory genes. This implies that early therapeutic intervention at the microvascular level may be a rational strategy to attenuate MODS. |
| Effects of melatonin on Candida sepsis in an experimental rat model. | Adv Ther | As an immunomodulator, melatonin reportedly exhibits protective effects in severe sepsis/shock induced by bacterial lipopolysaccharides in animal models. The present study was conducted to evaluate the possible protective effects of melatonin against experimental Candida sepsis in rats. A total of 40 adult male Wistar rats were randomly assigned to 4 groups: control, melatonin-treated control, septic, and melatonin-treated septic. Melatonin (200 microg/kg/d, intraperitoneally) injections were begun a week prior to sepsis induction and were continued daily for 3 wk until the end of the study. Cyclophosphamide was administered to animals in all groups as an immunosuppressive agent as a single dose 4 d prior to yeast inoculation. To cause sepsis, the Candida albicans (ATCC 10259) strain was administered intravenously. Amphotericin B was given as an antimycotic therapeutic agent as a single dose to septic rats. Plasma levels of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), vascular cell adhesion molecule-1, and E-selectin were measured on the first and 15th days of sepsis. IL-6, TNF-alpha, vascular cell adhesion molecule-1, and E-selectin levels of septic rats were higher than those of controls. Melatonin reduced IL-6 levels and shortened time to improvement in animals with Candida sepsis. Levels of TNF-alpha and adhesion molecules in melatonin-treated septic rats were decreased compared with those in septic rats, but this difference was not statistically significant. In light of the current results, investigators conclude that melatonin may have therapeutic benefits in Candida sepsis and in classic antimycotic treatment because of its immune regulatory effects. |
| 2007 | Protective role of PI3-kinase-Akt-eNOS signalling pathway in intestinal injury associated with splanchnic artery occlusion shock. | Br J Pharmacol | Endothelial NO synthase (eNOS) is a dynamic enzyme tightly controlled by co- and post-translational lipid modifications, phosphorylation and regulated by protein-protein interactions. Here we have pharmacologically modulated the activation of eNOS, at different post-translational levels, to assess the role of eNOS-derived NO and of these regulatory mechanisms in intestinal injury associated with splanchnic artery occlusion (SAO) shock.SAO shock was induced by clamping both the superior mesenteric artery and the celiac trunk for 45 min followed by 30 min of reperfusion. During ischemia, 15 min prior to reperfusion, mice were given geldanamycin, an inhibitor of hsp90 recruitment to eNOS, or LY-294002 an inhibitor of phosphatidylinositol 3-kinase (PI3K), an enzyme that initiates Akt-catalysed phosphorylation of eNOS on Ser1179. After 30 min of reperfusion, samples of ileum were taken for histological examination or for biochemical studies.Either LY-294002 or geldanamycin reversed the increased activation of eNOS and Akt observed following SAO shock. These molecular effects were mirrored in vivo by an exacerbation of the intestinal damage. Histological damage also correlated with neutrophil infiltration, assessed as myeloperoxidase activity, and with an increased expression of the adhesion proteins: ICAM-I, VCAM, P-selectin and E-selectin.Overall these results suggest that activation of the Akt pathway in ischemic regions of reperfused ileum is a protective event, triggered in order to protect the intestinal tissue from damage induced by ischaemia/reperfusion through a fine tuning of the endothelial NO pathway. |
| 2007 | Preoperative unstable angina causes venous adaptation to surgical graft injury. | Basic Res Cardiol | Ischemic preconditioning may provide a systemic organ protection, evident as the phenomenon known as remote preconditioning. Unstable angina may be a clinical analogue to ischemic preconditioning. Vein graft harvesting induces inflammation of the graft wall. We hypothesized that preoperative unstable angina preconditions vein grafts and reduces the inflammatory response to graft harvesting. Consecutive patients with stable or unstable angina undergoing open heart surgery (n = 12 in each group) were studied. Saphenous vein biopsies were collected at the start of graft harvesting, and when the last proximal anastomosis to the aorta was finished (average 112 minutes later). Gene expression of inflammatory mediators (tumor necrosis factor alpha, interleukin-1beta (IL-1beta), E-selectin (CD62E), intercellular leukocyte adhesion molecule 1, inducible nitric oxide synthase, endothelin-1) increased after surgical handling (semiquantitative RT-PCR). In vein grafts from unstable patients the increase was attenuated for Il-1beta (p < 0.004) and CD62E (p < 0.001). In stable patients the protein expression of IkappaBalpha and heat shock protein72 was reduced by surgical handling (p < 0.04), but was not influenced in unstable patients (immunoblotting). In vitro relaxation to acetylcholine was enhanced, and contractions to phenylephrine and endothelin-1 were attenuated in veins rings from unstable patients (p < 0.003). In conclusion, surgical handling of vein grafts induces inflammation of the vessel wall. This response was reduced in grafts from patients with unstable angina, indicating a possible systemic preconditioning-like effect of acute coronary syndromes. |
| 2006 | Elevated soluble thrombomodulin in the febrile stage related to patients at risk for dengue shock syndrome. | Pediatr Infect Dis J | Children with dengue hemorrhagic fever (DHF) are at risk to develop dengue shock syndrome (DSS) for which neither marker has been demonstrated.The study was designed to investigate the markers of vascular endothelial cell injuries and dysfunction that might be used as early predictors of the subsequent manifestation of DSS.The blood samples from 111 patients with dengue fever, DHF and other febrile illness (OFI) were collected daily from the day of admission until discharge and at convalescent stage. The sample from the day of defervescence was defined as day 0, 1 day before defervescence was defined as day -1 and so on. Also, 1 day after defervescence was defined as day +1 and so on.Increased soluble thrombomodulin (sTM) was demonstrated in dengue-infected patients via an enzyme-linked immunosorbent assay. Patients with DSS (DHF grades III and IV) had higher concentrations of sTM than those with dengue fever, DHF grade I, II and OFIs from day -3 until day +2. Increased circulating endothelial cells were detected from day 0 until day +2 in DSS patients as compared with other groups. In addition, increased soluble vascular cell adhesion molecule-1, soluble intercellular adhesion molecule-1 and soluble E-selectin were also found in dengue virus-infected patients as compared with OFIs.Blood sTM may be useful as an early predictor of DSS in dengue infected patients in the febrile stage. However, a further evaluation in a large prospective series is needed. |
| 2006 | Proinflammatory reaction and cytoskeletal alterations in endothelial cells after shock wave exposure. | J Investig Med | Although the effects on human organs by shock waves (SWs) induced by medical treatments or high-energy trauma are well recognized, little is known about the effects on the cellular level. Since blood vessel injury is a common finding after SW exposure, we assessed the in vitro effects of SWs on human umbilical vein endothelial cells (HUVECs).An in vitro trauma model was used to expose HUVEC monolayers to focused SWs or to shock waves plus cavitation (SWC), a subsequent phenomenon that is often considered the main cause of SW vascular injury.SWs alone did not cause any changes in the studied variables. In contrast, HUVEC monolayers exposed to SWC exhibited discrete central lesions with extensive cell death. Cells peripheral to the main lesion area displayed disassembly of dense peripheral bands and formation of actin stress fibers, indicating increased intercellular gaps. Expression of P-selectin was enhanced 11-fold compared with controls, whereas expression of E-selectin and intercellular adhesion molecule 1 was enhanced 8-fold (p < .05) and 1.5-fold (p < .01), respectively. The latter responses were preceded by nuclear translocation of nuclear factor kappaB subunit p65 by 16% (p < .01). When compared with mechanically produced lesions used as controls, SWC lesions exhibited an impaired regeneration rate of the endothelial cell layer (p < .001). Redistribution of centrosomes toward the lesion borders was less effective in the SWC samples compared with the mechanically produced lesions (p < .01).SWC lesions were associated with a switch to an endothelial proinflammatory phenotype, with an impaired regeneration rate and changes in cytoskeletal functions. |
| 2005 | Ehrlichia under our noses and no one notices. | Arch Virol Suppl | Ehrlichia chaffeensis, an obligately intracellular bacterium, resides within a cytoplasmic vacuole in macrophages, establishes persistent infection in natural hosts such as white-tailed deer and canids, and is transmitted transstadially and during feeding by ticks, particularly Amblyomma americanum. Ehrlichial cell walls contain glycoproteins and a family of divergent 28 kDa proteins, but no peptidoglycan or lipopolysaccharide. The dense-cored ultrastructural form preferentially expresses certain glycoproteins, including a multiple repeat unit-containing adhesin. Ehrlichiae attach to L-selectin and E-selectin, inhibit phagolysosomal fusion, apoptosis, and JAK/STAT activation, and downregulate IL-12, IL-15, IL-18, TLR2 and 3, and CD14. Mouse models implicate overproduction of TNF-alpha by antigen-specific CD8 T lymphocytes in pathogenesis and strong type 1 CD4 and CD8 T lymphocyte responses, synergistic activities of IFN-gamma and TNF-alpha, and IgG2a antibodies in immunity. Human monocytotropic ehrlichiosis (HME) manifests as a flu-like illness that progresses in severity to resemble toxic shock-like syndrome, with meningoencephalitis or adult respiratory distress syndrome in some patients, and requires hospitalization in half. In immunocompromised patients, HME acts as an overwhelming opportunistic infection. In one family physician's practice, active surveillance for three years revealed an incidence of 1000 cases per million population. Diagnosis employs serology or polymerase chain reaction, which are not utilized sufficiently to establish the true impact of this emerging virus-like illness. |
| 2005 | Pretreatment with hyperbaric oxygen and its effect on neuropsychometric dysfunction and systemic inflammatory response after cardiopulmonary bypass: a prospective randomized double-blind trial. | J Thorac Cardiovasc Surg | Animal studies have shown that pretreatment with hyperbaric oxygen can induce central nervous system ischemic tolerance and also modulate the inflammatory response. We evaluated this therapy in patients undergoing cardiopulmonary bypass.Sixty-four patients were prospectively randomized to group A (n = 31; atmospheric air, 1.5 atmospheres absolute) or group B (n = 33; hyperbaric oxygen, 2.4 atmospheres absolute) before on-pump coronary artery bypass grafting. Age, sex, body mass index, diabetes, hypertension, smoking, coronary disease severity, left ventricular function, Parsonnet score, Euroscore, bypass time, myocardial ischemia time, and number of grafts were comparable in both groups. Canadian Cardiovascular Society angina, New York Heart Association dyspnea, and previous myocardial infarction were significantly higher in group B. Inflammatory markers were analyzed before surgery and 2 and 24 hours after bypass. Neuropsychometric testing was performed 48 hours before surgery and 4 months after surgery and included trail making A and B, the Rey auditory verbal learning test, grooved peg board, information processing table A, and digit span forward and backward. Neuropsychometric dysfunction was defined as more than 1 SD deterioration in more than 2 neuropsychometric tests. Chi-square tests, Fisher tests, t tests, and analysis of variance were used as appropriate for statistical analysis.Group A had a significant postoperative increase in the inflammatory markers soluble E-selectin, CD18, and heat shock protein 70. This was not observed in group B. Neuropsychometric dysfunction was also significantly higher in group A compared with group B. There was no difference in any other early postoperative clinical outcome.Our results seem to indicate that pretreatment with hyperbaric oxygen can reduce neuropsychometric dysfunction and also modulate the inflammatory response after cardiopulmonary bypass. However, further multicenter randomized trials are needed to clinically evaluate this form of therapy. |
| 2005 | C1 inhibitor: molecular and clinical aspects. | Springer Semin Immunopathol | C1 inhibitor (C1-INH) is a serine protease inhibitor (serpins) that inactivates several different proteases in the complement, contact, coagulation, and fibrinolytic systems. By its C-terminal part (serpin domain), characterized by three beta-sheets and an exposed mobile reactive loop, C1-INH binds, and blocks the activity of its target proteases. The N-terminal end (nonserpin domain) confers to C1-INH the capacity to bind lipopolysaccharides and E-selectin. Owing to this moiety, C1-INH intervenes in regulation of the inflammatory reaction. The heterozygous deficiency of C1-INH results in hereditary angioedema (HAE). The clinical picture of HAE is characterized by bouts of local increase in vascular permeability. Depending on the affected site, patients suffer from disfiguring subcutaneous edema, abdominal pain, vomiting and/or diarrhoea for edema of the gastrointestinal mucosa, dysphagia, and dysphonia up to asphyxia for edema of the pharynx and larynx. Apart from its genetic deficiency, there are several pathological conditions such as ischemia-reperfusion, septic shock, capillary leak syndrome, and pancreatitis, in which C1-INH has been reported to either play a pathogenic role or be a potential therapeutic tool. These potential applications were identified long ago, but controlled studies have not been performed to confirm pilot experiences. Recombinant C1-INH, produced in transgenic animals, has recently been produced for treatment of HAE, and clinical trials are in progress. We can expect that the introduction of this new product, along with the existing plasma derivative, will renew interest in exploiting C1-INH as a therapeutic agent. |
| 2005 | Hydroxyethyl starch inhibits neutrophil adhesion and transendothelial migration. | Shock | A resuscitation strategy that significantly alters the state of neutrophil (PMN) activation may impact organ function and survivability after shock. Various resuscitative fluids have been shown to elicit a severe immune activation and an upregulation of cellular injury markers, whereas other fluids have been shown to be protective. Recent studies have demonstrated that hydroxyethyl starch (HES), an artificial colloid, may exert significant anti-inflammatory effects, whereas conflicting studies with the same substance have shown an increase in PMN activation. Successful manipulation of the early immune events associated with hemorrhage and resuscitation will require a better understanding of the possible pro- or anti-inflammatory effects of resuscitation fluids. Our study investigated the effect of HES directly on PMN and cultured vascular endothelial cells in vitro. The effect of HES on PMN surface expression of CD11b and L-selectin was measured by flow cytometry. PMN activation response to HES was measured using a shape-change assay in response to formyl-methionyl-leucyl-phenylalanine (f-MLP). The effect of HES on endothelial cell surface expression of E-selectin, P-selectin, vascular cell adhesion molecule-1(VCAM-1), and intracellular adhesion molecule-1 (ICAM-1) was evaluated by enzyme-linked immunoabsorbant assay. PMN rolling, adhesion, and migration events were measured using direct microscopy under conditions simulating microvascular flow. PMN surface expression of CD11b and L-selectin in whole blood samples and isolated PMNs were unaffected by exposure to HES. HES had no effect on the normal f-MLP dose-dependent increase in PMN activation. In the absence of IL-1 stimulation, there was a small but statistically significant (P < 0.05) increase in ICAM-1 after exposure to HES. After stimulation with IL-1 (10 U/mL), HES had no effect on the expression of P-selectin, E-selectin, ICAM-1, or VCAM-1. Under simulated microvascular flow conditions in vitro, HES significantly diminished the PMN tethering rate (P < 0.05) and the transendothelial migration rate (P < 0.05) in a dose-dependent manner. HES significantly alters the function of the PMN at the interface of the PMN responding to activated endothelium. The effect occurs, surprisingly, without a coincident effect on the state of PMN activation or a significant change in the surface expression of the adhesion molecules responsible for PMN-endothelial interaction. |
| 2005 | 15-Deoxy-delta(12,14)-prostaglandin J(2) (15D-PGJ(2)), a peroxisome proliferator activated receptor gamma ligand, reduces tissue leukosequestration and mortality in endotoxic shock. | Shock | Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a nuclear receptor that requires ligand activation for transcription. Experimental studies have shown that 15-deoxy-Delta-PGJ2 (15d-PGJ2) is a natural PPARgamma ligand which has potent anti-inflammatory properties. This study was designed to examine the effect and the molecular mechanisms of 15d-PGJ2 on tissue neutrophil infiltration and survival in endotoxic shock. Male Swiss albino mice were subjected to intraperitoneal injection of Escherichia coli lipopolysaccharide (LPS, 25 mg/kg). Three hours after LPS mice received vehicle or 15d-PGJ2 (1 mg/kg) and continued treatment every 12 hours. Survival was monitored for 72 hours. In a separate experiment, mice were sacrificed 6 hours after LPS and tissue examined. In vehicle-treated mice, LPS injection resulted in a survival rate of 9%. Marked lung injury was characterized by hemorrhage, infiltration of inflammatory cells and reduction of alveolar space. Elevated levels of myeloperoxidase activity in lung and small intestine were indicative of infiltration of neutrophils. Increased expression of intercellular adhesion molecule-1, vascular cellular adhesion molecule-1 and E-selectin were observed in the lung and small intestine. These inflammatory events were associated with reduced expression of PPARgamma and with activation of nuclear factor-kappaB (NF-kappaB) in the lung. Treatment with 15d-PGJ2 improved survival rate to 55%, downregulated expression of adhesion molecules and reduced neutrophil infiltration in tissues. These beneficial effects were associated with reduced activation of NF-kappaB DNA binding, whereas expression and DNA binding of PPARgamma and expression of the cytoprotective heat shock protein (HSP) 70 were increased in the lung. Our data demonstrate that 15d-PGJ2 ameliorates endotoxic shock most likely through repressing the proinflammatory pathway of NF-kappaB and enhancement of the cytoprotective heat shock response. |
| 2005 | Serial changes in neutrophil-endothelial activation markers during the course of sepsis associated with disseminated intravascular coagulation. | Thromb Res | For systematic elucidation of serial changes in neutrophil-endothelial activation markers as well as to investigate the correlationship among the inflammation markers, disseminated intravascular coagulation (DIC), and multiple organ dysfunction syndrome (MODS) in patients with sepsis, we made this prospective study.Forty-five patients with sepsis, severe sepsis, and septic shock were subdivided into two groups, 27 with DIC and 18 without DIC. Eight normal healthy volunteers served as control subjects. Serial levels of soluble L-, P-, and E-selectins, intercellular adhesion molecule-1 (sICAM-1), vascular cell adhesion molecule-1 (sVCAM-1), thrombomodulin (sTM), and neutrophil elastase were measured within 12 h after the diagnosis of sepsis (day 0) and on days 1-4 after the diagnosis. The numbers of systemic inflammatory response syndrome (SIRS) criteria that patients met and the DIC score were determined simultaneously.Acute Physiology and Chronic Health Evaluation (APACHE) II score was identical between the two groups. In the DIC patients, higher DIC scores, lower platelet counts, and more maximum numbers of SIRS criteria being met were observed compared with the non-DIC patients. The incidence of MODS and the number of the dysfunctioning organs were higher in the patients with DIC than those without DIC, and the DIC patients had poor outcome. Soluble L-selectin (sL-selectin) levels in both groups tended to be lower than those in the control subjects. All other parameters both in the two groups were continuously higher than those in the control subjects during study period. The levels of soluble E-selectin (sE-selectin), sICAM-1, sVCAM-1, neutrophil elastase, and sTM were more elevated in the DIC patients than those in the non-DIC patients. There were no differences in the sP-selectin levels between the two groups; however, more increased sP-selectin levels per platelet were found in the DIC patients compared with the non-DIC patients. Maximum DIC scores in the DIC group positively correlated with the peak levels of neutrophil elastase and sTM and the number of the dysfunctioning organs.We found close relations among the neutrophil-endothelial cell interactions, DIC, and MODS in patients with sepsis, severe sepsis, and septic shock. The results indirectly confirm the concept that DIC can produce organ dysfunction and that DIC reflects an inflammatory disorder of the microvasculature. |
| 2005 | Natriuretic peptides and E-selectin as predictors of acute deterioration in patients with inotrope-dependent heart failure. | Eur J Cardiothorac Surg | In patients with inotrope-dependent end-stage heart failure the timely application of the most suitable treatment, i.e. heart transplantation, implantation of a ventricular assist device or conservative treatment, is a key issue for therapeutic success.Seventy-six inotrope-dependent patients with end-stage heart failure were enrolled. Measurements of hemodynamics, routine laboratory parameters, and clinical examination were performed daily. Additionally, natriuretic peptides (BNP and NT-proBNP) and E-selectin were measured at the end of the study. The patients were retrospectively divided into groups with regard to the following end-points: Group I-deterioration into cardiogenic shock after an initially stable clinical course (n=26); Group II-stable clinical course without deterioration into cardiogenic (n=41); Group III-weaning from inotropic support (n=9).One day before cardiogenic shock occurred, BNP, NT-proBNP and E-selectin were significantly elevated in group I compared with group II. A logistic regression model showed that only BNP and E-selectin were independent predictors of clinical deterioration on the following day. The odds ratio (OR) for E-selectin using a cut-off point of 65ng/ml was 8.7 and for BNP using a cut-off of 500pg/ml it was 4.8. In combination, the OR increased to 11.1. Continuous decrease of NT-proBNP predicted patients in whom weaning from inotropes was possible.While routine parameters did not predict the clinical course, elevated BNP and E-selectin independently predicted cardiogenic shock on admission and 1 day before its occurrence. The combination showed increased predictive value. |
| Effects of brain death on stress and inflammatory response in the human donor kidney. | Transplant Proc | After kidney transplantation, a decreased graft survival is seen in grafts from brain dead donors compared to living donors, possibly related to a progressive inflammatory reaction in the graft. In this study, we focused on the effects of brain death on the inflammatory response (adhesion molecules, leukocyte infiltration, and gene expression) and stress-related heat shock proteins in the human kidney. Research outcomes and clinical donor parameters were linked to outcome data after transplantation.Human kidney biopsy specimens were obtained during organ retrieval from brain dead and living organ donor controls. On these specimens, immunohistochemistry and semiquantitative RT-PCR were performed. Regression analyses were performed connecting results to outcome data of kidney recipients.In brain death, immunohistochemistry showed an increase of E-selectin and interstitial leukocyte invasion versus controls; RT-PCR showed an increase of gene expression of HO-1 and Hsp70. One and 3 years after transplantation, high ICAM and VCAM expression proved to have a negative effect on kidney function in brain dead and living kidneys, while HO-1 proved to have a strongly positive effect, but only in kidneys from living donors.E-selectin expression and interstitial leukocyte accumulation in brain dead donor kidneys indicate an early phase inflammatory state prior to organ retrieval. Also, brain death causes a stress-related response resulting in upregulation of potentially protective heat shock proteins. The upregulation of HO-1 is beneficial in living donor kidneys, but might be inadequate in brain death. |
| 2005 | Protective role of angiopoietin-1 in endotoxic shock. | Circulation | Angiopoietin-1 (Ang1) plays an essential role in embryonic vasculature development, protects the adult peripheral vasculature from leakage, and has antiinflammatory properties. Because endotoxin-induced shock is a condition with microvascular leakage resulting from inflammation, we examined the potential therapeutic benefit of Ang1 in a murine model of lipopolysaccharide (LPS)-induced endotoxic shock.To induce endotoxic shock, LPS was injected intraperitoneally into C57BL/6 mice. Half of the mice received an intravenous application of 1.0x10(9) plaque-forming units of an adenoviral construct expressing human Ang1 (AdhAng1); in the other half an identical vector expressing green fluorescent protein (AdGFP) was injected as a control. In the AdhAng1-treated mice, hepatic transfection and high expression of circulating Ang1 protein were observed. Whereas in LPS-treated control mice, hemodynamic function was severely depressed 12 hours after LPS injection (decrease of blood pressure from 91+/-3 to 49+/-7 mm Hg, dP/dt(max) from 7284+/-550 to 2699+/-233 mm Hg/s, cardiac output from 11.3+/-1.2 to 2.8+/-0.8 mL/min; P<0.0005), in LPS-treated AdhAng1 mice blood pressure fell only to 76+/-3 mm Hg, dP/dt(max) to 5091+/-489 mm Hg/s, and cardiac output to 6.7+/-1.4 mL/min (P<0.05). This resistance to LPS-induced hemodynamic changes was reflected by an improved Kaplan-Meier survival rate of the AdhAng1 mice. Histological analysis revealed that lung injury after LPS injection was markedly attenuated in AdhAng1 mice. In addition, LPS-induced increase in lung water content and pulmonary myeloperoxidase activity was significantly reduced. Furthermore, LPS-induced increases in the expression level of vascular cell adhesion molecule-1, intracellular adhesion molecule-1, and E-selectin protein in the lungs were markedly lower in AdhAng1 mice than in control mice. Finally, in the mice overexpressing Ang1, pulmonary endothelial NO synthase (eNOS) expression and activity remained preserved after LPS challenge, providing evidence that the beneficial effect of Ang1 in endotoxic shock is mediated by eNOS-derived NO.Our study demonstrates an improved mortality rate in mice with endotoxic shock pretreated with an adenoviral construct encoding Ang1. The enhanced survival rate induced by Ang1 was accompanied by an improvement in hemodynamic function, reduced lung injury, a lower expression of inflammatory adhesion molecules, and preserved eNOS activity in the lung tissue. Ang1 may therefore have utility as an adjunctive agent for the treatment of septic shock condition. |
| 2004 | Effect of brain death on gene expression and tissue activation in human donor kidneys. | Transplantation | After kidney transplantation, decreased graft survival is seen in grafts from brain dead (BD) donors compared with living donors. This might result partly from a progressive nonspecific inflammation in the graft. In this study, we focused on the effects of BD on inflammatory response (adhesion molecules, leukocyte invasion, gene expression) and stress-related heat shock proteins in the human kidney. Research outcomes and clinical donor parameters were then linked to outcome data after transplantation.Kidney biopsy specimens and serum were obtained during organ retrieval from BD and living organ donor controls. Immunohistochemistry and semiquantitative reverse transcriptase-polymerase chain reaction were performed on the biopsy specimens. Clinical and laboratory parameters from BD donors were recorded and connected to outcome data of the recipients of the kidneys studied.After brain death, immunohistochemistry showed an increase of E-selectin (P<0.01) and interstitial leukocyte invasion (P<0.05) compared with controls. Also, reverse transcriptase-polymerase chain reaction showed a threefold increased heme oxygenase-1 (P<0.05) and Hsp70 (P<0.01) gene expression after BD. Levels of monocyte chemotactic protein-1 and transforming growth factor-beta were twice as high after brain death but did not reach significance. Transplantation outcome was influenced by several donor variables: positively most notably by donor treatment with desmopressin and negatively by high serum urea levels during brain death and by high intercellular adhesion molecule and vascular cell adhesion molecule expression in the kidney. Heme oxygenase-1 proved to have a protective function, but only in kidneys from living donors.The presence of interstitial leukocytes and the early adhesion molecule E-selectin in BD donor kidneys indicates an early-phase inflammatory process during organ retrieval. Elevated levels of monocyte chemotactic protein-1 and transforming growth factor-beta suggest a role for monocytes/macrophages in this phase. We suggest that BD causes a stress-related response against which protective heat shock proteins are formed in the future graft. This stress response may be too severe to be fully counteracted by elevated heat shock proteins. Which systemic and/or local factors trigger brain death-related graft injury is currently under investigation. |
| 2004 | Andrographolide attenuates inflammation by inhibition of NF-kappa B activation through covalent modification of reduced cysteine 62 of p50. | J Immunol | NF-kappaB is a central transcriptional factor and a pleiotropic regulator of many genes involved in immunological responses. During the screening of a plant extract library of traditional Chinese herbal medicines, we found that NF-kappaB activity was potently inhibited by andrographolide (Andro), an abundant component of the plant Andrographis that has been commonly used as a folk remedy for alleviation of inflammatory disorders in Asia for millennia. Mechanistically, it formed a covalent adduct with reduced cysteine (62) of p50, thus blocking the binding of NF-kappaB oligonucleotide to nuclear proteins. Andro suppressed the activation of NF-kappaB in stimulated endothelial cells, which reduced the expression of cell adhesion molecule E-selectin and prevented E-selectin-mediated leukocyte adhesion under flow. It also abrogated the cytokine- and endotoxin-induced peritoneal deposition of neutrophils, attenuated septic shock, and prevented allergic lung inflammation in vivo. Notably, it had no suppressive effect on IkappaBalpha degradation, p50 and p65 nuclear translocation, or cell growth rates. Our results thus reveal a unique pharmacological mechanism of Andro's protective anti-inflammatory actions. |
| 2004 | Glucocorticoid-induced TNF receptor family gene (GITR) knockout mice exhibit a resistance to splanchnic artery occlusion (SAO) shock. | J Leukoc Biol | In the present study, we used glucocorticoid-induced tumor necrosis factor (TNF) receptor family gene knockout (GITR-KO) mice to evaluate a possible role of GITR on the pathogenesis of splanchnic artery occlusion (SAO) shock, which was induced in mice by clamping the superior mesenteric artery and the celiac artery for 30 min, followed thereafter by release of the clamp (reperfusion). At 60 min after reperfusion, animals were killed for histological examination and biochemical studies. There was a marked increase in the lipid peroxidation in the ileum of the SAO-shocked, GITR wild-type (WT) mice after reperfusion. The absence of GITR significantly reduced the lipid peroxidation in the intestine. SAO-shocked WT mice developed a significant increase of ileum tissue, TNF-alpha, and myeloperoxidase activity and marked histological injury. SAO shock was also associated with a significant mortality (5% survival at 24 h after reperfusion). Reperfused ileum tissue sections from SAO-shocked WT mice showed positive staining for P-selectin, intercellular adhesion molecule 1 (ICAM-1), and E-selectin. The intensity and degree of P-selectin, E-selectin, and ICAM-1 were markedly reduced in tissue section from SAO-shocked, GITR-KO mice. SAO-shocked, GITR-KO mice also showed a significant reduction of the TNF-alpha production and neutrophil infiltration into the reperfused intestine, an improved histological status of the reperfused tissues, and an improved survival. Taken together, our results clearly demonstrate that GITR plays an important role in the ischemia and reperfusion injury and put forward the hypothesis that modulation of GITR expression may represent a novel and possible strategy. |
| 2004 | Vbeta-restricted T cell adherence to endothelial cells: a mechanism for superantigen-dependent vascular injury. | Arthritis Rheum | To investigate the potential for endothelial cells to operate as superantigen-presenting cells for T cells and the potential for such an interaction to cause endothelial cell activation and injury.Class II major histocompatibility complex (MHC)-positive human umbilical vein endothelial cells (HUVECs) were cocultured for 4 hours with purified T cells and the superantigens staphylococcal enterotoxin B (SEB) or toxic shock syndrome toxin 1 (TSST-1). After staining with fluorescence-conjugated monoclonal antibodies, flow cytometric analysis was performed on the HUVECs and T cells to examine V(beta)-restricted T cell adherence to the endothelial cell monolayer, V(beta)-restricted T cell activation (CD69 up-regulation), surface expression of endothelial cell activation markers, and generation of endothelial microparticles (EMPs).Coculture of purified T cells with class II MHC-positive HUVECs and either TSST-1 or SEB resulted in V(beta)-restricted CD69 up-regulation by CD4 and CD8 cells (V(beta)2 activation for TSST-1; V(beta)3, V(beta)5.1, and V(beta)12 activation for SEB). Additionally, there was CD4 and CD8 T cell V(beta)-restricted adherence to the HUVEC monolayer at 4 hours. Expression of intercellular adhesion molecule 1, E-selectin, and vascular cell adhesion molecule 1 was up-regulated on the class II MHC-positive HUVECs following exposure to superantigen in the presence of T cells, and there was increased EMP release from activated HUVECs, which occurred earlier and was of greater magnitude than that observed in response to tumor necrosis factor alpha.Class II MHC-positive endothelial cells operate as competent superantigen-presenting cells for CD4 and CD8 lymphocytes in vitro. Dual signaling between endothelial cells and T cells results in V(beta)-restricted activation and adherence to endothelial monolayers and endothelial cell activation and release of EMPs expressing inducible cell adhesion molecules. It is proposed that this mechanism could account in part for the vascular injury associated with superantigen-mediated diseases including Kawasaki disease. |
| 2004 | Effects of anti-endothelial cell antibodies in leprosy and malaria. | Infect Immun | As a result of damaging endothelial cells (ECs), Mycobacterium leprae triggers the production of antibodies (Abs). These anti-EC Abs (AECAs) can be divided into two types. The first type nonspecifically reacts with components of the cytosol (CY) and can be detected by enzyme-linked immunosorbent assay (ELISA). The second specifically reacts with the EC membrane (MB) and requires fluorescence-activated cell sorter (FACS) analysis to be detected. The presence of both types of AECAs was determined in 68 leprosy patients. The ELISA was positive for 35 of them but also for 30 of 34 malaria patients and 17 of 50 healthy African controls. However, whereas FACS analysis showed MB reactivity in only three malaria patients and four controls, this reactivity was found in 27 leprosy patients, more of those having the lepromatous than the tuberculoid form. Specificity for MB, which we failed to absorb by incubation with CY lysates, predominated over that for CY in leprosy, unlike malaria, where the EC reactivity was restricted to the CY. Western blot analysis and two-dimensional electrophoresis revealed that calreticulin, vimentin, tubulin, and heat shock protein 70 were targeted by AECAs from leprosy patients, but other proteins remained unidentified. These auto-Abs, but not those from malaria patients, did activate ECs, as indicated by the E-selectin and intercellular adhesion molecule 1 upregulation, and/or induced them into apoptosis, as documented by four different methods. Our findings suggest that, in some but not all leprosy patients, AECAs may play a role in pathogenesis. |
| 2003 | Porphyromonas gingivalis lipopolysaccharide antagonizes Escherichia coli lipopolysaccharide at toll-like receptor 4 in human endothelial cells. | Infect Immun | E. coli lipopolysaccharide (LPS) induces cytokine and adhesion molecule expression via the toll-like receptor 4 (TLR4) signaling complex in human endothelial cells. In the present study, we investigated the mechanism by which Porphyromonas gingivalis LPS antagonizes E. coli LPS-dependent activation of human endothelial cells. P. gingivalis LPS at 1 micro g/ml inhibited both E. coli LPS (10 ng/ml) and Mycobacterium tuberculosis heat shock protein (HSP) 60.1 (10 micro g/ml) stimulation of E-selectin mRNA expression in human umbilical vein endothelial cells (HUVEC) without inhibiting interleukin-1 beta (IL-1beta) stimulation. P. gingivalis LPS (1 micro g/ml) also blocked both E. coli LPS-dependent and M. tuberculosis HSP60.1-dependent but not IL-1beta-dependent activation of NF-kappaB in human microvascular endothelial (HMEC-1) cells, consistent with antagonism occurring upstream from the TLR/IL-1 receptor adaptor protein, MyD88. Surprisingly, P. gingivalis LPS weakly but significantly activated NF-kappaB in HMEC-1 cells in the absence of E. coli LPS, and the P. gingivalis LPS-dependent agonism was blocked by transient expression of a dominant negative murine TLR4. Pretreatment of HUVECs with P. gingivalis LPS did not influence the ability of E. coli LPS to stimulate E-selectin mRNA expression. Taken together, these data provide the first evidence that P. gingivalis LPS-dependent antagonism of E. coli LPS in human endothelial cells likely involves the ability of P. gingivalis LPS to directly compete with E. coli LPS at the TLR4 signaling complex. |
| 2003 | High mobility group 1 B-box mediates activation of human endothelium. | J Intern Med | Severe sepsis and septic shock is a consequence of a generalized inflammatory systemic response because of an invasive infection that may result in acute organ dysfunction. Mortality is high despite access to modern intensive care units. The nuclear DNA binding protein high mobility group 1 (HMGB1) protein has recently been suggested to act as a late mediator of septic shock via its function as a macrophage-derived pro-inflammatory cytokine (J Exp Med 2000; 192: 565, Science1999; 285: 248). We investigated the pro-inflammatory activities of the A-box and the B-box of HMGB1 on human umbilical venular endothelial cells (HUVEC).The HUVEC obtained from healthy donors were used for experiments. Recombinant human full-length HMGB1, A-box and B-box were cloned by polymerase chain reaction (PCR) amplification from a human brain quick-clone cDNA. The activation of HUVEC was studied regarding (i) upregulation of adhesion molecules, (ii) the release of cytokines and chemokines, (iii) the adhesion of neutrophils to HUVEC, (iv) the activation of signalling transduction pathways and (v) the involvement of the receptor for advanced glycation end-products (RAGE).The full-length protein and the B-box of HMGB1 dose-dependently activate HUVEC to upregulate adhesion molecules such as ICAM-1, VCAM-1 and E-selectin and to release IL-8 and G-CSF. The activation of HUVEC could be inhibited to 50% by antibodies directed towards the RAGE. HMGB1-mediated HUVEC stimulation resulted in phosphorylation of the ELK-1 signal transduction protein and a nuclear translocation of p65 plus c-Rel, suggesting that HMGB1 signalling is regulated in endothelial cells through NF-kappaB.The HMGB1 acts as a potent pro-inflammatory cytokine on HUVEC and the activity is mainly mediated through the B-box of the protein. HMGB1 may be a key factor mediating part of the pro-inflammatory response occurring in septic shock and severe inflammation. |
| 2003 | 5-lipoxygenase knockout mice exhibit a resistance to splanchnic artery occlusion shock. | Shock | In the present study, we used 5-lipoxygenase (5-LO) knockout (KO) mice to evaluate the possible role of 5-LO on the pathogenesis of splanchnic artery occlusion (SAO) shock. SAO shock was induced in mice by clamping both the superior mesenteric artery and the celiac artery for 30 min, followed thereafter by release of the clamp (reperfusion). At 120 min after reperfusion, animals were sacrificed for histological examination and biochemical studies. There was a marked increase in the lipid peroxidation in the ileum as well as in the lung of the SAO-shocked 5-LO wild-type (WT) mice after reperfusion. The absence of 5-LO did not reduce the lipid peroxidation in the intestine or the lung. SAO-shocked WT mice developed a significant increase of tissue (ileum and lung) myeloperoxidase activity and marked histological injury. SAO shock was also associated with a significant mortality (50% survival at 5 h after reperfusion). Reperfused ileum and lung tissue sections from SAO-shocked WT mice showed positive staining for P-selectin, ICAM-1, and E-selectin that was mainly localized in the vascular endothelial cells. The intensity and degree of P-selectin, E-selectin, and ICAM-1 were markedly reduced in tissue section from SAO-shocked 5-LOKO mice. SAO-shocked 5-LOKO mice showed also a significant reduction of the neutrophils infiltration into the reperfused intestine as well as in the lung as evidenced by reduced myeloperoxidase activity, an improved histological status of the reperfused tissues, and an improved survival. Taken together, our results clearly demonstrate that 5-LO plays an important role in ischemia and reperfusion injury and put forward the hypothesis that inhibition of 5-LO may represent a novel and possible strategy in the treatment of ischemia and reperfusion injury. Part of this effect may be due to inhibition of the expression of adhesion molecules and subsequent reduction of neutrophil-mediated cellular injury. |
| 2003 | The effects of fresh frozen plasma on neutrophil-endothelial interactions. | Anesth Analg | Leukocyte adhesion to endothelial cells contributes to microcirculatory disturbances during severe shock syndromes. Whereas certain plasma expanders inhibit leukocyte adhesion, contaminants of plasma protein solutions upregulate endothelial cell adhesion molecules in certain cases. We performed this study to determine whether fresh frozen plasma (FFP) affects neutrophil-endothelial interactions in cocultures of neutrophils and human umbilical vein endothelial cells (HUVEC) in vitro. HUVEC (n = 9) were incubated with either 20% FFP or 20% serum in medium for 6 h. Expression of E-selectin, intercellular adhesion molecule 1, and vascular cell adhesion molecule-1 was induced by tumor necrosis factor alpha (0.5 ng/mL for 4 h) and measured by flow cytometry. Neutrophil adhesion was examined in a parallel plate flow chamber in which isolated neutrophils were perfused over pretreated HUVEC under postcapillary flow conditions. Incubation with FFP decreased E-selectin and intercellular adhesion molecule 1 on activated HUVEC by 28% and 22%, respectively (P < or = 0.01; analysis of covariance). Consequently, neutrophil adhesion decreased by 20%-41% in FFP-treated cocultures (n = 4; P < or = 0.01; paired Student's t-test). We conclude that FFP attenuates the inflammatory response of endothelial cells with regard to neutrophil-endothelial interactions. Because the composition of patients' plasma is affected not only by transfusion, but more frequently by shock treatment with IV fluids, plasma dilution in critically ill patients could be important.During shock, fluid administration leads to a massive dilution of plasma. Apart from maintaining hemodynamics, this might affect tissue damage by influencing leukocyte accumulation in the microvasculature. Using endothelial cells, isolated neutrophils, and a parallel plate flow chamber, we studied the effects of fresh frozen plasma on neutrophil-endothelial interactions. |
| Cytokine and adhesion molecule expression in primary human endothelial cells stimulated with fever-range hyperthermia. | Int J Hyperthermia | Migration of blood-borne lymphocytes into lymphoid tissues and sites of inflammation is initiated by vascular adhesion molecules and proinflammatory cytokines. Previous in vivo studies have shown that febrile temperatures dynamically stimulate adhesion in differentiated high endothelial venules (HEV), which are portals for lymphocyte extravasation. This report examines the direct effect of fever-range hyperthermia on the expression of adhesion molecules and cytokines by primary cultured endothelial cells. In both macrovascular (HUVEC) and microvascular (HMVEC) endothelial cells, fever-range hyperthermia (40 degrees C for 6-12 h) did not affect expression of adhesion molecules (ICAM-1, E-selectin, VCAM-1, P-selectin, PECAM-1, PNAd, MAdCAM-1), cytokine release (IL-1beta, TNF-alpha, IFN-gamma, IL-6, IL-11, IL-12, IL-13), or chemokine secretion (IL-8, RANTES, MCP-1, MIP-1beta, MIG). This is in contrast to the stimulatory effects of TNF-alpha or 43 degrees C heat shock. However, a novel role for fever-range hyperthermia was identified in augmenting actin polymerization in cultured endothelial cells and enhancing the ability of endothelial-derived factors to transactivate the alpha4beta7 integrin lymphocyte homing receptor. These findings provide insight into the tightly regulated effects of fever-range hyperthermia that exclude induction of adhesion in non-activated endothelium of normal blood vessels. Through these mechanisms, it is proposed that febrile temperatures associated with infection or clinical hyperthermia avoid the unproductive exodus of lymphocytes to non-involved extralymphoid tissues while simultaneously promoting lymphocyte delivery to sites of immune activation. |
| 2003 | Immunologic and hemodynamic effects of "low-dose" hydrocortisone in septic shock: a double-blind, randomized, placebo-controlled, crossover study. | Am J Respir Crit Care Med | Within the last few years, increasing evidence of relative adrenal insufficiency in septic shock evoked a reassessment of hydrocortisone therapy. To evaluate the effects of hydrocortisone on the balance between proinflammatory and antiinflammation, 40 patients with septic shock were randomized in a double-blind crossover study to receive either the first 100 mg of hydrocortisone as a loading dose and 10 mg per hour until Day 3 (n = 20) or placebo (n = 20), followed by the opposite medication until Day 6. Hydrocortisone infusion induced an increase of mean arterial pressure, systemic vascular resistance, and a decline of heart rate, cardiac index, and norepinephrine requirement. A reduction of plasma nitrite/nitrate indicated inhibition of nitric oxide formation and correlated with a reduction of vasopressor support. The inflammatory response (interleukin-6 and interleukin-8), endothelial (soluble E-selectin) and neutrophil activation (expression of CD11b, CD64), and antiinflammatory response (soluble tumor necrosis factor receptors I and II and interleukin-10) were attenuated. In peripheral blood monocytes, human leukocyte antigen-DR expression was only slightly depressed, whereas in vitro phagocytosis and the monocyte-activating cytokine interleukin-12 increased. Hydrocortisone withdrawal induced hemodynamic and immunologic rebound effects. In conclusion, hydrocortisone therapy restored hemodynamic stability and differentially modulated the immunologic response to stress in a way of antiinflammation rather than immunosuppression. |
| 2003 | Combined anticoagulant and antiselectin treatments prevent lethal intravascular coagulation. | Blood | Widespread microvascular injury followed by vessel obstruction may lead to disseminated intravascular coagulation (DIC). We describe a murine model wherein leukocytes interacting with inflamed microvessels in vivo are activated by antibodies. Treatment of tumor necrosis factor alpha (TNF-alpha)-primed mice with anti-Ly-6G antibodies reproduced many of the features of septic or traumatic shock including microvessel obstruction and coagulation, severe vasculitis, respiratory difficulties, and vascular leakage. Mice lacking either E-selectin or P-selectin were protected from this reaction as were animals treated with a combination of either selectin-blocking antibodies and heparin or a selectin antagonist plus heparin. Combined blockade of leukocyte/platelet adhesion and coagulation may provide convincing protection in DIC. |
| [Inflammatory mechanisms, arteriosclerosis and ischemic stroke: clinical data and perspectives]. | Rev Neurol | The atherosclerosis is the most common cause of death and disability in developed countries by causing ischemic cardiopathic and stroke. The ischemic atherotrombotic stroke is the most frequent form of the last one. In this sense we review herein the mechanisms underlying the artherosclerotic process.It is understood as an inflammatory disease, by taking into account the widely accepted hypothesis by Ross: it was firstly stated in structural terms, as macrophages and T/B linfocities were present in the arterial wall from the first stages of the disease (fatty streak) to the last and complicated ones. The starting point is a functional endothelial damage, secondary to mechanical or vascular risk factors and called response to injury hypothesis . The next step is an inflammatory cascade that involves humoral (citokines, growth factors) and cellular (increased quimiotaxis, adherece and infiltration of inflamatory cells) mechanisms. They interact among them, outbalanced and in a progresssive way that leads to the final fibroproliferative response. Every stage has his own inflammatory components and interactive pathways. The following elements are outstanding in this process: 1) Adhesion molecules, including E selectin, ICAM 1 and VCAM 1, that are increased locally in the plaques and as circulating elements; plaquetary receptors of the type IIb/IIIa are integrins wich belong to the same family; 2) Citokines with either proinflammatory activity like IL 1, the TNF a and linfocitary ligands like the CD 40, or with antiinflammatory activity like the gamma interpheron; 3) Growth factors, with plaquetary (PDGF) and fibroblastic (FGF) variants as the cornerstone; 4) Markers of systemic inflammation, overall plasma C reactive protein and fibrinogen, that predict the risk of stroke and cardiovascular death; IL 6, complement, thrombin and heat shock proteins (HSP) would act in a similar but less conclusive way.The evidences of the pivotal role of the inflammation in the stroke allow to develop therapeutical strategies to prevent the disease: fostering natural antiinflamatory mechanisms, or inhibiting inflammatory elements by selective (monoclonal antibodies) or non selective (IIb/IIIa receptors, antiinflammatory drugs) pathways are distinctily glimpsed, ongoing or fully developed. |
| Endothelial accumulation of hydroxyethyl starch and functional consequences on leukocyte-endothelial interactions. | Eur Surg Res | To date, accumulation of hydroxyethyl starch (HES) has been studied mainly in skin specimens, but there are no detailed reports available regarding starch accumulation in the endothelium. Because endothelial cells play an essential role during shock, we studied the accumulation of HES in human umbilical venous endothelial cells (HUVEC). HUVEC (n = 9) were incubated with a fluorescein-conjugated HES 200/0.5 (FITC-HES) at 0.5-20 mg/ml for 1-72 h. FITC-HES was internalized dose- and time-dependently by pinocytosis into secondary lysosomes. Asymptotic elimination curves showed that 50% of the formerly ingested molecules could not be eliminated. Despite accumulation, starch molecules did not attenuate the expression of E-selectin, ICAM-1 or VCAM-1 on TNF-alpha-activated HUVEC. However, apart from adhesion molecule expression, perfusion studies showed that HES reduced neutrophil adhesion by direct inhibition of integrin-mediated interactions. |
| 2002 | A fish oil emulsion used for parenteral nutrition attenuates monocyte-endothelial interactions under flow. | Shock | Monocyte adhesion contributes to perfusion abnormalities, tissue damage, and activation of the coagulation system seen during trauma, shock, or overwhelming inflammation. This study was performed to determine whether an intravenous fish oil emulsion used for parenteral nutrition attenuates monocyte-endothelial interactions under flow and reduces procoagulant activity, measured as tissue factor (TF) expression on adherent monocytes in vitro. Endothelial cell monolayers were incubated with either an intravenous fish oil emulsion or a conventional omega-6 lipid emulsion at 0.05 to 1 mg/ml for 24 h. Six hours following activation with TNFalpha (25 ng/ml), expression of endothelial cell adhesion molecules was measured by flow cytometry. Adhesion of isolated monocytes to pretreated endothelium was examined in a parallel plate flow chamber at a shear stress of 1.5 dynes/cm2. Following perfusion, the cells were cocultured for an additional 4 h and TF expression on monocytes was determined by flow cytometry. In contrast to omega-6 lipids, fish oil down-regulated E-selectin, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 in a dose-dependent manner. P-selectin, however, remained unchanged. In addition, firm adhesion was reduced to 54%, whereas rolling interactions remained unchanged. Fish oil exhibited no effect on the TF expression on cocultured monocytes. We conclude that intravenous fish oil emulsions reduce both endothelial cell adhesion molecule expression and monocyte adhesion. However, under postcapillary flow conditions, rolling interactions via P-selectin remain unaltered. The functional importance of this effect is illustrated by the corresponding upregulation of TF in response to residual monocyte-endothelial interactions. |
| 2002 | Heterogeneous expression of cell adhesion molecules by endothelial cells in ARDS. | J Pathol | ARDS (acute respiratory distress syndrome) can be associated with septic shock and multiple organ failure caused by an uncontrolled systemic inflammatory response to Gram-negative bacterial infection. While in animal models the key role of the endothelial adhesion molecules ICAM-1, E-selectin, and VCAM in ARDS has been extensively studied, there are scarcely any corresponding pathomorphological studies of human lung tissue. Hence, little is known about whether there is a comparable, or even heterogeneous, expression pattern of these molecules in the human pulmonary vasculature. This study was therefore undertaken to investigate the immunohistochemical expression of the constitutively expressed PECAM (CD31) and the inducible molecules ICAM-1, E-selectin, and VCAM in ARDS lungs from patients who had died in septic shock induced by Gram-negative bacteria. While in all specimens (ARDS and normal lungs) there was homogeneous strong expression of PECAM in all vessels, ICAM-1 was clearly up-regulated in ARDS lungs. E-selectin and VCAM were not expressed by endothelial cells (ECs) in normal lungs, but in ARDS lungs there was strong expression of both molecules in larger vessels, while in the capillaries there was only mosaic-like weak expression of a few ECs. This immunohistochemical investigation demonstrates the induction and up-regulation of adhesion molecules in human ARDS lungs, comparable to that described in animal models. There is also markedly heterogeneous expression of E-selectin and VCAM, indicating toporegional differences in the function of pulmonary ECs. |
| 2002 | Trauma/hemorrhagic shock mesenteric lymph upregulates adhesion molecule expression and IL-6 production in human umbilical vein endothelial cells. | Shock | Trauma/hemorrhagic shock (T/HS) is associated with significant lung injury, which is mainly due to an inflammatory process, resulting from the local activation and subsequent interaction of endothelial cells and leukocytes. Adhesion molecules expressed by both cell types play a crucial role in the process of neutrophil-mediated endothelial cell injury. We have previously shown that mesenteric lymph duct ligation prevents T/HS-induced lung leukocyte infiltration and endothelial injury, suggesting that inflammatory factors originating from the gut and carried in the lymph are responsible for the lung injury observed following T/HS. Based on these observations, we hypothesized that inflammatory substances in T/HS lymph trigger lung injury by a mechanism involving the upregulation of adhesion molecules. To test this hypothesis, we examined whether T/HS mesenteric lymph induces the expression of E-selectin, P-selectin, and intracellular adhesion molecule-1 (ICAM-1) in human umbilical vein endothelial cells (HUVECs). Furthermore, because the cytokine IL-6 is an important component of the endothelial inflammatory process, we investigated how T/HS lymph affects the production of IL-6 by HUVECs. Mesenteric lymph from T/HS rats increased both E- and P-selectin, as well as ICAM-1 expression on HUVECS, as compared to trauma/sham shock (T/SS) lymph or medium only groups. However, T/HS lymph failed to induce the shedding of E-selectin. In HUVECs treated with T/HS lymph, IL-6 concentrations were higher than HUVECs treated with T/SS lymph. These findings suggest that mesenteric lymph produced after hemorrhagic shock potentiates lung injury by the upregulation of endothelial cell adhesion molecule expression and IL-6 production. |
| 2002 | Systemic endothelial activation is greater in septic than in traumatic-hemorrhagic shock but does not correlate with endothelial activation in skin biopsies. | Crit Care Med | Sepsis and severe trauma result in endothelial activation and damage. The activated endothelium expresses adhesion receptors that control leukocyte trafficking. After activation, some adhesion molecules are also released into plasma as soluble forms. The present study was designed to compare the expression of soluble cell adhesion molecules (sCAMs) in three groups of patients: those with septic shock, severe sepsis, and traumatic-hemorrhagic shock. In addition, the endothelial expression of these adhesive molecules was examined in skin biopsies.Prospective observational studyIntensive care unit at a university hospitalThe study included 15 patients with septic shock (by Bone's definition), 11 patients with severe sepsis (by Bone's definition), and 13 patients with traumatic-hemorrhagic shock. Fifteen healthy blood donors served as controls.Measurements of sCAMs were performed on days 1, 2, and 3 of the disease. On day 1, when compared with controls, sE-selectin, sP-selectin, soluble vascular cell adhesion molecule (sVCAM)-1, and soluble intercellular adhesion molecule (sICAM)-1 were markedly elevated in septic shock patients, whereas these sCAMs, except for sP-selectin, were within normal ranges in traumatic-hemorrhagic shock patients. In patients with severe sepsis, an earlier stage than septic shock in the sepsis continuum, intermediate values of sCAMs were found. In skin biopsies of septic shock patients, the endothelial cells expressed a bright staining of constitutive endothelial molecules (CD146, CD144, CD131). Inducible molecules (ICAM-1, VCAM-1, and E-selectin) were positively expressed with bright staining. The biopsies from traumatic-hemorrhagic shock patients showed a similar positive expression of endothelial molecules.The patterns of sCAMs indicate that the systemic activation of the endothelium is different in the three clinical entities, maximum in septic shock, intermediate in severe sepsis, and not different from controls in traumatic-hemorrhagic shock. Comparable endothelial activation as evidenced by skin biopsies suggests that caution is required in the interpretation of CAMs in plasma, which does not necessarily reflect the in situ activation state of endothelium. |
| 1999 | [The effect of early escheractomy on expression of ICAM-1 and E-selectin mRNA of endothelial cells]. | Zhonghua Wai Ke Za Zhi | To examine the effect of burned serum on change in endothelial cell ICAM-1 and E-selectin mRNA expression and whether these alterations can be corrected by the means of escharectomy during burn shock period.One hundred and seventy-six Wistar rats with 30% III degree TBSA on back were studied. Endothelial cell ICAM-1 and E-selectin mRNA were tested with the method of RT-PCR.ICAM-1 and E-selectin expression showed an increase after stimulation by serum of 4 hours post burn, reaching peak levels 168 hours after injury. ICAM-1 and E-selectin expression levels could decrease to baseline within 168 hours in the rats which underwent escharectomy during shock stage, but kept higher than normal in rats without escharectomy or underwent 96 hours post burn.These findings suggest that eschar can induce endothelial cells to express adhesion molecule. Early escharectomy is important to prevent adhesion molecule expression and SIRS. |
| 2000 | [The role of bacterial endotoxins, receptors and cytokines in the pathogenesis of septic (endotoxin) shock]. | Vutr Boles | Sepsis, resistant to therapy, results in the development of septic (endotoxin) shock. The latter is caused by the endotoxins of different Gram-negative bacteria. Endotoxin (bacterial lipopdisacharide--LPS) interacts with cells through specific membrane or plasma soluble endotoxin receptors (sCD14, mlD14, LBP, CD13/CD14, CD16, CD116/CD18, L-selectin, etc.). Endotoxin interaction with the mCD14 receptor of the monocytes, macrophages and the neutrophils results in the production of a number of proinflammatory cytokines--tumor necrosis factor alpha (TNF alpha), interleukines 1 and 6 (IL-1 and IL-6, etc), antiinflammatory cytokines--interleukines 10 and 12 (IL-10 and IL-12), cell adhesion molecules (P-selectin, E-selectin, ICAM-1, VCAM-1, etc.) and inducible enzymes: inducible NO synthase (iNOS), inducible phospholipase A2 (cPL-A2), inducible cyclooxygenase (COX-2). All pathologic processes in the structure and function of human body during endotoxin shock are a result of the disbalance of a number of mediators with a proinflammatory and antiinflammatory effects. |
| 2001 | Characterization of equine E-selectin. | Immunology | Expression of E-selectin on activated endothelium is a critical initial step that leads to extravasation of leucocytes during inflammation, yet E-selectin is largely uncharacterized in several animal species including the horse. We have sequenced and compared E-selectin genes derived from activated cultures of purified equine (horse), cervid (black-tailed deer) and ovine (sheep) pulmonary artery endothelial cells (ECs). Phylogenetic and amino acid sequence comparisons indicate that bovine, cervid and ovine E-selectin are similar, whereas human and equine E-selectin are more closely related to each other than to the ruminant molecules. Human E- and P-selectin-specific monoclonal antibodies that also recognize equine E-selectin were identified and used to characterize its expression. Expression of E-selectin was more readily induced by lipopolysaccharide treatment in equine ECs than in human ECs and supported adhesion and activation of neutrophils, consistent with the extreme sensitivity of horses to endotoxaemia and septic shock. |
| 2001 | The value of immune modulating parameters in predicting the progression from peritonitis to septic shock. | Shock | Intra-abdominal infection is one of the major causes of septic shock and multiple organ failure. To date, what causes the disease's progression remains unclear and therefore the relevance of immune modulating therapies remains speculative. The primary outcome measure of this study was to investigate immune modulating mediators at the onset of peritonitis before the development of subsequent septic shock. The secondary outcome measure was to investigate the usefulness of these immune parameters in predicting progression from peritonitis to septic shock. Fifty-eight peritonitis patients were included in this study: 14 patients subsequently developed septic shock. All patients were examined on "diagnosis of peritonitis" (<4 h within establishment of diagnosis), during "early septic shock" (<12 h following the onset of septic shock), and once again during "late septic shock" (within 72-98 h following the onset of septic shock). The immune modulating parameters tumor necrosis factor-alpha (TNF-alpha), the soluble TNF-alpha receptors I and II (sTNF-alpha RI and sTNF-alpha RII), interleukines (IL) -1beta, -6, -8, and -10, and the adhesions molecules endothelial-leukocyte-adhesion-molecule (E-Selectin), intercellular-adhesion-molecule-1 (ICAM-1), and vascular-adhesion-molecule-1 (VCAM-1), in addition to nitrate and nitrite, were determined. In the peritonitis group with subsequent septic shock, TNF-alpha, sTNF-alpha RI + RII IL-1beta, IL-8, IL-10, and nitrate were significantly increased before the onset of septic shock. TNF-alpha had an area under the receiver operating characteristics curve (AUC) of 0.84 and was reliable in predicting the progression from peritonitis to septic shock. The AUC of the other immune modulating parameters, despite being significantly elevated, ranged from 0.71 to 0.76. The AUC of the conventional laboratory markers such as leukocytes and C-reactive protein ranged from 0.64 to 0.68. In peritonitis that progressed to septic shock, an early immune response had already occurred before the onset of septic shock. The progression was best predicted by TNF-alpha. Therefore, mediator therapy might be considered in high-risk peritonitis patients who show an exaggerated immune response before the progression to septic shock. |
| 2000 | Human lactoferrin interacts with soluble CD14 and inhibits expression of endothelial adhesion molecules, E-selectin and ICAM-1, induced by the CD14-lipopolysaccharide complex. | Infect Immun | Lipopolysaccharides (LPS), either in the free form or complexed to CD14, a LPS receptor, are elicitors of the immune system. Lactoferrin (Lf), a LPS-chelating glycoprotein, protects animals against septic shock. Since optimal protection requires administration of Lf prior to lethal doses of LPS, we hypothesized that interactions between Lf and soluble CD14 (sCD14) exist. In a first step, human sCD14 and human Lf (hLf) were used to determine the kinetic binding parameters of hLf to free sCD14 in an optical biosensor. The results demonstrated that hLf bound specifically and with a high affinity (K(d) = 16+/-7 nM) to sCD14. Affinity chromatography studies showed that hLf interacted not only with free sCD14 but also, though with different binding properties, with sCD14 complexed to LPS or lipid A-2-keto-3-deoxyoctonic acid-heptose. In a second step, we have investigated whether the capacity of hLf to interact with sCD14 could modulate the expression of endothelial-leukocyte adhesion molecule 1 (E-selectin) or intercellular adhesion molecule 1 (ICAM-1) induced by the sCD14-LPS complex on human umbilical vein endothelial cells (HUVEC). Our experiments show that hLf significantly inhibited both E-selectin and ICAM-1 expressions at the surface of HUVEC. In conclusion, these observations suggest that the anti-inflammatory effects of hLf are due not only to the ability of the molecule to chelate LPS but also to its ability to interact with sCD14 and with the sCD14 complexed to LPS, thus modifying the activation of endothelial cells. |
| 2000 | Endotoxin-induced lung inflammation is independent of the complement membrane attack complex. | Infect Immun | Several products of the activated complement system are known to modulate endothelial cell function in vitro. It has been shown that the membrane attack complex (MAC) (C5b-C9) can enhance tumor necrosis factor alpha (TNF-alpha)-induced expression of P- and E-selectin and intercellular adhesion molecule type 1 in cell cultures of human umbilical vein endothelial cells. In the present study the potential role of this synegism for lung injury during endotoxin-mediated septic shock in vivo was examined using a model of C6-deficient PVG (C-) (RT1(C)) rats and the congenic PVG (C+) (RT1(C)) strain. Following administration of a high (5 mg/kg) or low (0.5 mg/kg) dose of lipopolysaccharide (LPS) (Escherichia coli O55:B5), we determined the expression of cytokines, chemokines, and adhesion molecules as well as the recruitment of leukocytes in the lung. Challenge with intraperitoneal i.p. injections of LPS resulted in a strong induction of TNF-alpha, interleukin-1alpha/beta, cytokine-induced neutrophil chemoattractant, interferon-inducible protein 10, macrophage inflammatory proteins 1alpha and 2, macrophage chemotactic protein 1, and P-selectin. However, there were no significant differences between PVG (C-) and PVG (C+) rats. Immunoperoxidase staining showed a similar increase of lung infiltration by CD11b/c(+) leukocytes in both rat strains. We therefore conclude that the described synergism between TNF-alpha and the MAC of the complement system on the induction of endothelial adhesion molecules is dispensable for inflammatory processes during endotoxin-mediated septic shock in vivo. |
| 1999 | Circulating intercellular adhesion molecule-1 as an early predictor of hepatic failure in patients with septic shock. | Crit Care Med | To investigate whether endotoxin, interleukin-6, and circulating adhesion molecules, measured sequentially in blood, can predict mortality and organ dysfunction in sepsis.Inception cohort study with follow-up for 28 days.Surgical intensive care unit at a university hospital.A total of 14 consecutive patients were enrolled in the study within the first 24 hrs after onset of septic shock. Seven healthy subjects were studied as controls.Patients were analyzed for mortality and development of organ dysfunction.At the end of the 28-day follow-up period, seven of the patients were still alive (survivors) but the other seven (nonsurvivors) had died. At the time of enrollment in the study (day 0), the Acute Physiology and Chronic Health Evaluation II score was 28.4 in survivors (n = 7) and 28.7 in nonsurvivors (n = 7). In contrast, circulating intercellular adhesion molecule-1 (ICAM-1) was significantly higher in nonsurvivors than in survivors. Circulating ICAM-1 predicted mortality in patients with septic shock with a sensitivity and a specificity of 71.4% each. Endotoxin, interleukin-6, circulating L-selectin, P-selectin, E-selectin, and platelet endothelial cell adhesion molecule-1, however, did not distinguish between survivors and nonsurvivors. In addition, circulating ICAM-1 at day 0 showed a significant correlation with the highest serum bilirubin observed during the entire study period (r2 = 0.963).Because only circulating ICAM-1 was higher in nonsurvivors than in survivors at day 0, circulating ICAM-1 may serve as an early prognostic marker for outcome in septic shock. In addition, measurement of circulating ICAM-1 facilitates identification of those patients with the highest risk of developing liver dysfunction. |
| 1999 | Markers of systemic inflammation predicting organ failure in community-acquired septic shock. | Clin Sci (Lond) | To obtain predictors of organ failure (OF), we studied markers of systemic inflammation [circulating levels of interleukin-6 (IL-6), IL-8, soluble IL-2 receptor (sIL-2R), soluble E-selectin and C-reactive protein, and neutrophil and monocyte CD11b expression] and routine blood cell counts in 20 patients with systemic inflammatory response syndrome and positive blood culture. Eight patients with shock due to community-acquired infection developed OF, whereas 11 normotensive patients and one patient with shock did not (NOF group). The first blood sample was collected within 48 h after taking the blood culture (T1). OF patients, as compared with NOF patients, had at T1 a lower monocyte count, a lower platelet count, higher levels of CD11b expression on both neutrophils and monocytes, and higher concentrations of IL-6, IL-8 and sIL-2R. C-reactive protein and soluble E-selectin concentrations did not differ between groups. No parameter alone identified all patients that subsequently developed OF. However, a sepsis-related inflammation severity score (SISS), developed on the basis of the presence or absence of shock and on the levels of markers at T1, identified each patient that developed OF. The maximum SISS value was 7. The range of SISS values in OF patients was 2-5, and that in NOF patients was 0-1. In conclusion, high levels of CD11b expression, depressed platelet and monocyte counts, and high concentrations of IL-6, IL-8 and sIL-2R predict OF in patients with community-acquired septic shock, and the combination of these markers may provide the means to identify sepsis patients who will develop OF. |
| 1999 | E5531, a synthetic non-toxic lipid A derivative blocks the immunobiological activities of lipopolysaccharide. | Br J Pharmacol | 1. The major pathological responses to Gram-negative bacterial sepsis are triggered by endotoxin or lipopolysaccharide. As endotoxin is shed from the bacterial outer membrane, it induces immunological responses that lead to release of a variety of cytokines and other cellular mediators. As part of a program aimed at developing a therapeutic agent for septic shock, we have developed E5531, a novel synthetic lipopolysaccharide antagonist. 2. As measured by release by tumour necrosis factor-alpha, human monocytes or whole blood can be activated by lipopolysaccharide, lipid A, and lipoteichoic acid (from Gram-positive bacteria). E5531 potently antagonizes activation by all these agents while itself being devoid of agonistic activity. 3. The inhibitory activity of E5531 was dependent on time of addition. When 10 nM E5531 was added simultaneously with lipopolysaccharide or 1 - 3 h before addition of lipopolysaccharide, production of tumour necrosis factor-alpha was inhibited by more than 98%. The addition of E5531 1 h after lipopolysaccharide reduced the efficacy of E5531 by 47%. 4. Antagonistic activity of E5531 was specific for lipopolysaccharide as it was ineffective at inhibiting interferon-gamma mediated NO release of RAW 264.7 cells, phorbor 12-myristate 13-acetate stimulated superoxide anion production in human neutrophils, concanavalin A stimulated mitogenic activity in murine thymocytes and tumor necrosis factor-alpha induced E-selectin expression in human umbilical vein endothelial cells. 5. E5531 as well as MY4, an anti-CD14 antibody, inhibited radiolabelled lipopolysaccharide binding in human monocytes. 6. These results support our contention that E5531 is a potent antagonist of lipopolysaccharide-induced release of tumour necrosis factor-alpha and other cellular mediators and may be an effective therapeutic agent for human septic shock due to Gram-negative bacteria. |
| 1998 | [Soluble cell adhesion molecules in chronic renal graft rejection]. | Pol Arch Med Wewn | Cell-bound adhesion molecules are involved in immune and inflammatory responses. Soluble forms of adhesion molecules (s.a.m.) can be detected in the blood. The elevated blood levels of s.a.m. were found as a response to variety disease processes (e.g. septic shock, acute graft rejection, atherosclerosis). The objective of the present study was to measure the serum levels of s.a.m. in patients with chronic renal allograft rejection and in recipients with a stable graft function. Evaluated was also the effect of activity of graft rejection (ch. g. r.) and risk factors of graft lesion on the levels of the investigated s.a.m. 34 patients with ch.g.r. were examined (Group I), 50 patients with a stable allograft function (Group II), and 25 healthy subjects (control). Group I patients were 76 +/- 34 months and Group II patients were 59 +/- 36 months after transplantation. Both groups of patients were treated with immunosuppressive drugs (CsA, azathioprine and prednisone) Group I patients had a higher plasma levels of creatinine and uric acid, increased arterial blood pressure and triglycerides concentrations, and lower plasma levels of HDL cholesterol, as compared to Group II patients. In all the examined subjects, serum concentrations of s.a.m. from the immunoglobulin and selectin families (s.ICAM-1, s.VCAM-1, s.E-selectin) were measured by the immunoenzymatic method. The investigations of s.a.m. in ch.g.r. patients revealed a statistically significant increase the serum levels of s.ICAM-1, s.VCAM-1 and s.E-selectin. Some disorders of the release of s.a.m. into blood were also found in patients without graft disfunction. In this patients were observed: increased levels of s.VCAM-1 and s.E-selectin. S.ICAM-1, s.VCAM-1 and s.E-selectin serum levels showed a correlation with plasma uric acid concentration and arterial pressure, whereas the other two molecules with the plasma level of triglycerides. Each of the three molecules had a negative correlation with the HDL cholesterol level. The regression analysis revealed a correlation of s.ICAM-1 and s.VCAM-1 with IL-6. The correlation of the molecules with chemokines (s.VCAM-1 and s. E-selectin with IL-8, and s. E-selectin with MCP-1) may results from their release in the course of the inflammatory process. The increased levels of circulating s.VCAM-1 and s.E-selectin found in renal allograft patients suggest a chronic stimulation and activation of the endothelium. Non-immunological mechanisms (such as arterial hypertension or metabolic disorders) contributed to the generation of the s.a.m. in patients with ch.g.r. and in those with stable graft function. The negative correlation of HDL with s.a.m. (s.ICAM-1, s.VCAM-1) suggests a protective role of HDL on the vascular endothelium by inhibiting the generation of these mediators. |
| 1999 | Regulation of adhesion molecules during human endotoxemia. No acute effects of aspirin. | Am J Respir Crit Care Med | Gram-negative septic shock is mediated in part by endotoxin (lipopolysaccharide; LPS), and animal models have shown that blockade of even single adhesion molecules considerably improves survival. Thus interference with the adhesion cascade may provide a useful therapeutic approach in human sepsis. Young healthy men (n = 30) each received a bolus of 4 ng/kg LPS intravenously to study the effects of endotoxemia on adhesion processes in humans and to identify potential targets for pharmacologic intervention. One third of subjects received pretreatment with 1,000 mg aspirin and 1,000 mg paracetamol to study potential antiinflammatory effects of aspirin or effects of antipyresis. Circulating neutrophils dropped by -80% at 67 min after LPS, monocytes by -96% at 90 min, and lymphocytes by -85% at 240 min. L-selectin expression decreased, particularly on monocytes. Circulating (c)E-selectin levels increased by 820%, von Willebrand factor-Ag (vWF), soluble thrombomodulin, circulating (c)P-selectin, circulating intercellular adhesion molecule-1 (cICAM-1), and circulating vascular cell adhesion molecule-1 (cVCAM-1) by a mean of 65 to 98% (p < 0.001 for all), but cL-selectin by only 15%. Urinary excretion of soluble adhesion molecules was negligible. Aspirin had no influence on the LPS-induced changes of adhesion parameters, but paracetamol blunted the relative increase in vWF while having no effects on the other parameters measured. The consistent, profound, and early upregulation of cE-selectin during endotoxemia indicates that cE-selectin may be a better surrogate marker to monitor the activation status of endothelial cells in systemic inflammation than the other markers measured. Although aspirin did not have any antiinflammatory effects in this model, paracetamol lowered the relative increase in vWF. |
| 1999 | Role of tumor necrosis factor and interferon gamma in endotoxin-induced E-selectin expression. | Shock | Tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma), potent inflammatory cytokines, are released by macrophages during endotoxin shock. However, the contribution of these cytokines to endotoxin-induced inflammation has not been defined. The expression of E-selectin, measured using the dual radiolabeled monoclonal antibody (mAb) technique, was monitored in different tissues of endotoxin-challenged wild-type and IFN-gamma-deficient mice receiving a mAb to TNF-alpha (TN3). A significant elevation in E-selectin expression occurred in all tissues of wild-type mice challenged with endotoxin. Injection of TN3 in wild-type mice significantly attenuated the endotoxin-induced up-regulation of E-selectin in all tissues (p < .05) except the pancreas. The level of reduction in endotoxin-induced E-selectin expression ranged between 30% in the stomach to 60% in the small intestine. E-selectin expression in endotoxin-challenged, IFN-gamma-deficient mice was significantly reduced in the small and large intestines, when compared with endotoxin-challenged wild-type mice. Although IFN-gamma deficiency had no effect on E-selectin expression in other tissues, administration of TN3 to endotoxin-challenged, IFN-gamma-deficient mice significantly reduced E-selectin expression to levels observed in endotoxin-challenged, wild-type mice that received TN3. These findings indicate that TNF-alpha is essential for achievement of maximal E-selectin expression in most vascular beds during endotoxemia, whereas the contribution of IFN-gamma is largely confined to the small intestine. |
| 1999 | Chlamydial and human heat shock protein 60s activate human vascular endothelium, smooth muscle cells, and macrophages. | J Clin Invest | Both chlamydial and human heat shock protein 60s (HSP 60), which colocalize in human atheroma, may contribute to inflammation during atherogenesis. We tested the hypothesis that chlamydial or human HSP 60 activates human endothelial cells (ECs), smooth muscle cells (SMCs), and monocyte-derived macrophages. We examined the expression of adhesion molecules such as endothelial-leukocyte adhesion molecule-1 (E-selectin), intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1), and the production of the proinflammatory cytokine interleukin-6 (IL-6). We also tested whether either HSP 60 induces nuclear factor-kappaB (NF-kappaB), which contributes to the gene expression of these molecules. Either chlamydial or human HSP 60 induced E-selectin, ICAM-1, and VCAM-1 expression on ECs similar to levels induced by Escherichia coli lipopolysaccharide (LPS). Each HSP 60 also significantly induced IL-6 production by ECs, SMCs, and macrophages to an extent similar to that induced by E. coli LPS, as assessed by enzyme-linked immunosorbent assay (ELISA). In ECs, either HSP 60 triggered activation of NF-kappaB complexes containing p65 and p50 Rel proteins. Heat treatment abolished all these effects, but did not alter the ability of E. coli LPS to induce these functions. Chlamydial and human HSP 60s therefore activate human vascular cell functions relevant to atherogenesis and lesional complications. These findings help to elucidate the mechanisms by which a chronic asymptomatic chlamydial infection might contribute to the pathophysiology of atheroma. |
| 1998 | Prediction of clinical severity and outcome of ventilator-associated pneumonia. Comparison of simplified acute physiology score with systemic inflammatory mediators. | Am J Respir Crit Care Med | Systemic kinetics of three inflammatory mediators (bactericidal/permeability-increasing protein [BPI], soluble intercellular adhesion molecule [sICAM], and soluble E-selectin [sE-selectin]) were studied during the development of ventilator-associated pneumonia (VAP) (n = 42), diagnosed on quantitative cultures of bronchoscopic samples. From a pool of collected samples, nested samples were used to measure mediators on Days -4, -2, 0, and +2, relative to diagnosis. Correlations between systemic levels of mediators and clinical severity of infection (VAP with or without severe sepsis or septic shock) and patient outcome (mortality at Day 10 after diagnosis) were studied. Predictive values of inflammatory mediators were compared with daily Simplified Acute Physiology Score II (SAPS II) values and the logarithmic number of bacteria in bronchoscopic samples. During the development of VAP, increasing SAPS II scores and rising systemic mediator levels were only found in patients in whom VAP was accompanied with severe sepsis or septic shock. Values of SAPS II and plasma levels of BPI and sE-selectin, but not sICAM, increased from the day of diagnosis on in patients who died within 10 d of diagnosis. Systemic levels of inflammatory mediators did not better predict clinical severity or patient outcome than daily SAPS II scores. The logarithmic number of bacteria in bronchoscopic samples poorly correlated with circulating levels of inflammatory mediators, severity of infection, and patient outcome. Our findings show that a clinical scoring system (SAPS II score) is at least as good as a predictor for the clinical severity of infection and patient outcome, and provide new information on the kinetics of inflammatory mediators during the development of VAP. |
| 1998 | Effects of explosive brain death on cytokine activation of peripheral organs in the rat. | Transplantation | The success rate of transplanted organs from brain-dead cadaver donors is consistently inferior to that of living sources. As cadaver and living unrelated donors are equally genetically disparate with a given recipient, the difference must lie within the donor himself and/or the effects of organ preservation and storage. We have hypothesized that irreversible central nervous system injury may up-regulate proinflammatory mediators and cell surface molecules in peripheral organs to be engrafted, making them more prone to host inflammatory and immunological responses.Rats undergoing surgically induced acutely increased intracranial pressure (explosive brain death) were followed for 6 hr. Their peripheral tissues were examined by reverse transcriptase polymerase chain reaction and immunohistology, serum factors were assessed by enzyme-linked immunosorbent assay, and the influence of inflammatory molecules in the blood stream was determined by cross-circulation experiments with normal animals.mRNA expression of both lymphocyte- and macrophage-associated products increased dramatically in all tissues. Similar factors in serum were coincidentally increased; these were shown to be active in vivo by cross-circulation with normal animals. The organs of all control groups, including animals with important ischemic injury and with hemorrhagic shock, were negative. Up-regulation of MHC class I and II antigens and the co-stimulatory molecule B7 suggests increased immunogenicity of the peripheral organs. These changes could be inhibited by: (i) administration of a recombinant soluble P-selectin glycoprotein ligand-Ig, a P- and E-selectin antagonist; and (ii) a fusion protein, cytotoxic T lymphocyte antigen 4-Ig, which blocks B7-mediated T-cell co-stimulation.Activation of peripheral organs following explosive brain death may be caused by various interrelated events, including the effects of massive acute central injury, hypotension, and circulating factors. Almost complete suppression of these changes could be produced by biological agents. Such interventions, if reproducible in humans, could improve the quality of organs from "marginal" donors, broadening the criteria for donor acceptance. |
| 1998 | Cytokines and adhesion molecules in elective and accidental trauma-related ischemia/reperfusion. | J Trauma | The major pathophysiologic role of cytokines such as tumor necrosis factor (TNF)-alpha, interleukin (IL)-1, and IL-6, as well as of the (soluble) adhesion molecules ICAM-1 and E-selectin, has been identified using different experimental models of ischemia/reperfusion injury. Moreover, in intensive care management, evaluation of these agents as diagnostic or prognostic tools is of great interest in ischemia/reperfusion injury caused by surgical or accidental trauma. For this reason, inflammatory mediators including those mentioned above were investigated in three different groups of surgical patients.The first group (A, n = 13) comprised patients undergoing elective limb surgery without a tourniquet. The second group (B, n = 36) included patients subjected to limb surgery with a tourniquet. The third group (C, n = 30) was composed of accidental trauma patients who were retrospectively divided into those with and without multiple organ dysfunction (+MOD and -MOD, respectively) as defined by the Denver Score. Serial blood samples were taken during a 5-day (elective surgery) or 14-day (accidental trauma) observation period for monitoring of cytokines and soluble adhesion molecules. The clinical course and the degree of MOD were recorded daily.Only when a tourniquet was applied for a mean time of 105 minutes did elective limb surgery result in significantly increased serum levels of IL-6, IL,-1ra, and IL-10 but not TNF-RII. Yet, the increase in cytokine levels was not sufficient to cause an enhanced shedding of adhesion molecules, and both soluble ICAM-1 and soluble E-selectin remained unchanged in groups A and B throughout the 5-day observation period. In patients with multiple injuries (group C), all parameters increased early after trauma up to 10- to 20-fold in comparison with the elective limb surgery patients in groups A and B. When the accidental trauma patients were divided according to the Denver Score for +MOD (n = 8, mean Injury Severity Score = 33.8) and -MOD (n = 22, mean Injury Severity Score = 31.2), a clear difference became evident in serum IL-6 and IL-1ra levels within the first 4 days and in serum IL-10 levels for the first 2 days after trauma, with cytokine levels being significantly higher in the +MOD patients 3 to 4 days before the onset of MOD. Although highly elevated, TNF-RII levels did not differentiate between +MOD and -MOD at any time. The increase in serum cytokine levels was associated with a remarkable expression and shedding of ICAM-1 and E-selectin made obvious by significantly increased soluble serum ICAM-1 levels in +MOD patients compared with the -MOD group between days 3 and 5 after trauma and increased soluble serum E-selectin levels between days 2 and 4 after trauma.The release of cytokines and soluble adhesion molecules into the circulation correlates well with the degree of trauma (elective surgery vs. accidental multiple trauma), depending on the extent of the associated ischemia/reperfusion injury. Both groups of mediators are also clearly related to the development of MOD in patients with multiple injuries with generalized ischemia/reperfusion injury caused by hemorrhagic shock. They may be predictive of patients at risk for MOD when measured early in the posttraumatic period. |
| 1998 | Elevated circulating E-selectin, intercellular adhesion molecule 1, and von Willebrand factor in patients with severe infection. | Am J Respir Crit Care Med | To investigate interactions between the endothelium and leukocytes in patients with sepsis, we measured soluble adhesion molecules (sE-selectin and sICAM-1), von Willebrand factor antigen (vWf:Ag), myeloperoxidase (MPO), and lactoferrin (Lacto-f) as plasma markers of endothelial and neutrophil activation. We tested whether the five proteins were predictors of clinical severity, which was evaluated by simplified acute physiological score (SAPS), number of organ failures (MOF), acute lung injury (ALI), and subsequent final outcome. Levels of the five plasma markers were higher in patients with severe infection (n = 25) than in patients without sepsis (n = 7) and healthy volunteers (n = 9). In the study population, levels of sE-selectin, sICAM-1, and vWf:Ag were higher for nonsurvivors as well as for patients with septic shock or with bacteremia, and they were correlated with SAPS and MOF. Survival outcome was predicted with high sensitivity and specificity by initial plasma levels of sICAM-1 and vWf:Ag. The initial sICAM-1 level appeared to be an independent prognostic variable, based on a logistic regression analysis. Unlike sE-selectin, sICAM-1 remained at high levels indefinitely in nonsurvivors. We conclude that, unlike neutrophil activation markers, levels of endothelium-derived soluble adhesion molecules and vWf:Ag in severe sepsis syndrome are correlated with the severity of illness and may be considered as predictors of survival outcome. |
| 1997 | Superantigens but not mitogens are capable of inducing upregulation of E-selectin ligands on human T lymphocytes. | Exp Dermatol | Bacterial infections can exacerbate immune mediated dermatoses, possibly via superantigens produced by these bacteria. Therefore, we asked whether superantigens induce the expression of adhesion molecules which may then facilitate invasion of highly activated T cells into different organs. The influence of exfoliative toxin (ET) and toxic shock syndrome toxin-1 (TSST-1) stimulation on the expression of a broad panel of adhesion and costimulatory molecules was investigated by flow cytometry. We found that only the E-selectin ligands cutaneous lymphocyte-associated antigen (CLA) and sialylated Lewis(x) (CD15s) are significantly upregulated by these superantigens but not by mitogen stimulation. In contrast, the mucosal lymphocyte-associated antigen (MLA) recognized by the monoclonal antibody Ber-Act8 was not differentially induced by mitogen or superantigen stimulation. Therefore, T lymphocyte stimulation by bacterial superantigens might directly influence their skin homing capacity. Furthermore, the superantigen-driven induction of CD15s-an adhesion molecule which is absent or only weakly expressed by resting or mitogen stimulated T cells-may indicate a role of this antigen for T cell skin homing. An additional adhesion pathway via E-selectin may thus be available to lymphocytes, comparable to granulocytes which constitutively express CD15s. |
| 1997 | Induction of acute inflammatory lung injury by staphylococcal enterotoxin B. | J Immunol | Superantigens stimulate T lymphocytes at high frequency by interacting with appropriate Vbeta segments of the TCR. Challenge of mice with bacterial superantigens such as staphylococcal enterotoxin B (SEB) induces the systemic release of cytokines resulting in septic shock and death of sensitized animals. Analysis of putative pathogenic mechanisms of T cell-dependent septic shock revealed that administration of SEB results in acute inflammatory lung injury characterized by a profound increase in vascular permeability. Injury was associated with marked leukocyte infiltration of the lung and induction of cell adhesion molecules including VCAM-1, ICAM-1, and P-selectin, but not E-selectin. Lung infiltrating leukocytes consisted of granulocytes, mononuclear phagocytes and NK cells with granulocytes representing the major fraction. Consistent with a role of neutrophils as cellular mediators of inflammatory organ injury, we demonstrate activation of circulating granulocytes in mice treated with SEB. When compared with granulocytes of control mice, peripheral blood granulocytes of SEB-treated mice were found to express increased levels of cell surface Mac-1, to down-regulate expression of L-selectin, and to respond with an increased production of toxic oxygen metabolites upon exposure to FMLP. Interestingly, TNF-alpha further enhanced FMLP-induced oxidant production by granulocytes from SEB-treated but not control mice, suggesting that the systemic response to SEB increases granulocyte sensitivity to TNF-alpha-mediated signals. Together, these results suggest that acute inflammatory lung injury may contribute to the pathogenesis of T cell-dependent lethal shock in mice challenged with bacterial superantigens and indicate common pathogenic mechanisms of lung injury induced by a large number of distinct inflammatory stimuli. |
| 1997 | Cytokine and adhesion molecule expression in human monocytes and endothelial cells stimulated with bacterial heat shock proteins. | Infect Immun | Bacterial heat shock proteins (HSPs) from Escherichia coli (GroES, GroEL, and DNAk) were tested for their ability to induce by themselves the expression and release of interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-alpha), and granulocyte-monocyte colony-stimulating factor (GM-CSF) by human monocytes and GM-CSF, IL-6, E-selectin, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) by human umbilical vein endothelial cells (HUVEC). Our study demonstrated that treatment of monocytes with DNAk increased IL-6, TNF-alpha, and GM-CSF release in a dose-dependent manner. The same effect was elicited by GroEL but at a lower rate. Treatment of HUVEC cultures with DNAk and GroEL also increased GM-CSF, IL-6, E-selectin, ICAM-1, and VCAM-1 release in a dose-dependent fashion. In any case, the greatest release was obtained by using DNAk and GroEL at a concentration of 1 microg/ml. DNAk and GroEL were also able to up-regulate the surface expression of E-selectin, ICAM-1, and VCAM-1. As detected by reverse transcription-PCR analysis, DNAk and GroEL also increased the steady-state levels of cytokines and adhesion molecules in human monocytes and endothelial cells. In our study GroES showed a significant activity only on the release, surface expression, and mRNA transcription of E-selectin. Adhesion molecule expression seems to be a direct effect of HSPs and not via cytokines. Furthermore, these effects are due to HSPs properties because they are inhibited by specific monoclonal antibodies. These findings support the potential role of HSPs in modulating cell interactions during immunological and inflammatory responses. |
| 1996 | Comparison of endothelial activation during endotoxic and posttraumatic conditions by serum analysis of soluble E-selectin in nonhuman primates. | J Lab Clin Med | Because the presence of E-selectin, which is one of the adhesion molecules on the endothelium, cannot be determined directly in vivo except by invasive biopsy techniques, the only available kinetic information concerning its activity is the serum level of the soluble form. Therefore we tried to measure soluble E-selectin levels and compare the degree of endothelial activation in a trauma and endotoxic situation, where endothelial activation is supposed to occur. To perform these studies, an enzyme-linked immunosorbent assay with two monoclonal antibodies was set up and used in (1) a model of endotoxic shock in baboons (n = 8) (1.5 mg/kg Escherichia coli endotoxin as a 10-minute intravenous infusion), (2) a hemorrhagic traumatic shock model in baboons (n = 6), where trauma was simulated by infusion of zymosan-activated serum and hemorrhage. Soluble E-selectin was released in vivo after the application of endotoxin, and it reached a peak level after 24 hours (13.82 +/- 5.38 ng/ml). In baboons with hemorrhagic shock, much lower levels (5.03 +/- 1.98 ng/ml) of soluble E-selectin were found. A lower soluble E-selectin level indicates a lower level of endothelial activation after experimental hemorrhagic traumatic shock (with endotoxin translocation from the gut) as compared with endotoxic shock probably caused by much lower endotoxin levels in traumatic shock. |
| 1996 | Elevated selectin levels after severe trauma: a marker for sepsis and organ failure and a potential target for immunomodulatory therapy. | J Trauma | Severe injury is frequently complicated by sepsis and organ failure. Activated neutrophils adherent to inflamed endothelia have been implicated in the pathogenesis of these complications. Identification of high-risk patients to target immunomodulatory therapy, however, remains an elusive goal. We postulated that (1) patients at risk for sepsis and organ failure could be identified by measuring shed selectin adhesions molecules as a marker of endothelial activation after injury and reperfusion, and (2) these elevated selectin levels would correlate with injury severity, shock, major complications, and mortality.Blood samples were drawn from 50 patients with multiple trauma every 2 hours after admission for the first 24 hours, and every 6 hours for the subsequent 24 hours, and assayed for levels of shed E- and P-selectin. Patients were then stratified according to Injury Severity Score (ISS), presence or absence of shock, presence or absence of organ failure and/or infectious complications, and finally, death versus survival.Trauma patients who had ISS < 30, who did not develop shock, sepsis, or organ dysfunction, had minimal increase in circulating E- and P-selectin over admission levels. In patients who subsequently developed infectious complications, organ dysfunction, or both, or subsequently went on to die, elevated levels of E-selectin levels were evident by 36 hours, and in some cases, earlier. Differences between nonsurvivors and survivors was statistically significant. There was also a trend toward increased levels of P-selectin in the same group of patients, although these differences were not significant. There was no differentiation in either of the two selections when patients were stratified according to ISS or presence of shock.A subset of major trauma patients manifest increased levels of circulating E-selectin adhesion molecules after resuscitation. These patients seem to be at increased risk of death and possibly at risk for infections complications and organ failure. Selectin blockade is a potential new immunomodulatory strategy in this subgroup of patients. |
| 1996 | Protection against septic shock in mice with SJC13, an azaindolidine derivative that is a cell adhesion molecule inhibitor. | Inflamm Res | An azaindolidine derivative, SJC13, selectively inhibits expression and mRNA synthesis of E-selectin and vascular cell adhesion molecule-1 (VCAM-1) in human umbilical vein endothelial cells (HUVEC) stimulated with lipopolysaccharide (LPS). The present experiments were performed to determine the in vivo effects of SJC13 against the lethality of LPS. In a mouse model of septic shock, intravenous administration of SJC13 5 min prior to LPS injection prevented significantly the lethality at doses of 3 mg/kg and 10 mg/kg. The prophylactic effect was dose-dependent. When injected up to 1 h after LPS injection, SJC13 inhibited significantly the lethality. Neutrophil emigration into lung tissues during sepsis induced with LPS, as assessed by lung myeloperoxidase (MPO) content and histological examination, was significantly prevented by SJC13 administration. These data demonstrate that SJC13 has therapeutic anti-inflammation potential in vivo. |
| 1996 | Effects of E-selectin and P-selectin blockade on neutrophil sequestration in tissues and neutrophil oxidative burst in burned rats. | Crit Care Med | Neutrophil deposition in tissues (leukosequestration) after shock may produce local tissue injury from proteases and high-energy oxygen species released from sequestered neutrophils. The initial step in the binding of neutrophils to capillary endothelium is the interaction of adhesion molecule (selectin) receptors between neutrophils and endothelial cells. We quantified leukosequestration in the tissues of burned rats using two methods of analysis: a) measurement of lung myeloperoxidase; and b) measurement of radiolabeled neutrophils and erythrocytes deposited in multiple tissues. We then determined the ability of a selectin receptor blocking agent to affect neutrophil deposition in tissues after burn injury.Prospective, controlled, laboratory study.University research laboratory.Male Wistar rats (200 to 300 g).After tracheostomy and venous cannulation, rats received 17% total body surface area full-thickness contact burns and were resuscitated with saline (20 mL i.p.). Experimental animals received 2 mg/kg body weight i.v. administration of a P- and E-selectin blocking monoclonal antibody, CY-1747, immediately after burn. Lung tissue neutrophils were estimated by measuring myeloperoxidase in lung tissue. Neutrophil retention in lung, liver, spleen, gut, skin, muscle, kidney, and brain tissues was determined by removing (preburn) and differentially radiolabeling neutrophils (111In) and erythrocytes (51Cr), reinfusing cells 4.5 hrs after burn, and measuring tissue radioactivity 30 mins later. Edema was estimated by measuring extravasated 125 I-labeled albumin in the various tissues. Peripheral blood neutrophils were analyzed for intracellular hydrogen peroxide content, utilizing a fluorescent dye that reacts with hydrogen peroxide, coupled with analysis of cell fluorescence by flow cytometry.Myeloperoxidase concentration was increased in lungs 5 hrs after burn (p < .05), indicating neutrophil deposition. Radioisotope studies demonstrated significant (p < .05) leukosequestration into the lung, gut, kidney, skin, and brain tissues at 5 hrs after burn. Flow cytometry showed increased intracellular hydrogen peroxide content in peripheral blood neutrophils 5 hrs after burn. Tissue edema, manifested by radiolabeled albumin retention, was not seen in any tissues. Postburn neutrophil deposition in lungs and liver was blocked (p < .05) by administration of CY-1747 after burn, but maximal neutrophil hydrogen peroxide content was unaffected.Burn injury in rats results in accumulation of neutrophils in multiple tissues. Neutrophil deposition in the lungs and liver is blocked by administration of the E/P-selectin blocking antibody, CY-1747. Since sequestration of metabolically active neutrophils may induce tissue injury, therapies that block postburn leukosequestration may improve clinical outcomes by limiting remote tissue injury. |
| 1996 | Heat shock protein 65 induces CD62e, CD106, and CD54 on cultured human endothelial cells and increases their adhesiveness for monocytes and granulocytes. | J Immunol | Expression of members of the heat shock protein 60 (hsp60) family in tissues has been reported to coincide with leukocyte infiltration, but it is not known whether these proteins are directly involved in the extravasation of leukocytes. Extravasation of leukocytes requires their adhesion to endothelial cells (EC) via an interaction between adhesion molecules expressed on both cell types. The aim of the present study was to investigate the effect of recombinant mycobacterial hsp65 on the adhesive characteristics of EC for monocytes and granulocytes. Incubation of EC with hsp65 induces a concentration- and time-dependent increase in adhesiveness of these EC for monocytes and granulocytes. The effect was maximal after incubation of EC with hsp65 for 4 to 6 h. In addition, incubation of EC with hsp65 induced the expression of endothelial CD62E (E-selectin), CD106 (vascular cell adhesion molecule-1) and CD54 (intercellular adhesion molecule-1). The increased adhesion of granulocytes to hsp65-stimulated EC was inhibited completely by blocking Ab against CD62E. mAb against endothelial CD62E, CD106, or CD54 or against the monocyte adhesion molecules CD14 or CD49d (very late Ag-4) did not inhibit the increased adhesion of monocytes to hsp65-stimulated EC; however, mAb against the monocyte adhesion molecule CD18 (beta2-integrin) inhibited monocyte adhesion to hsp65-stimulated EC to the same extent as monocyte adhesion to nonstimulated EC. Hsp65 did not exert its effect in an autocrine or paracrine fashion via the endogenous production of IL-1, TNF-alpha, or other factors or via contaminating LPS. Together these results indicate that hsp65 can play an important role in the adhesion of monocytes and granulocytes to EC at sites of inflammation via modulation of the adhesive characteristics of EC and thus may facilitate extravasation of these phagocytes. |
| 1996 | Increased concentrations of cytokines and adhesion molecules in patients after repair of abdominal aortic aneurysm. | Eur J Surg | To evaluate the association between inflammatory mediators and clinical outcome in patients after repair of abdominal aortic aneurysms.Prospective study.University hospital, The Netherlands.30 Consecutive patients who had undergone elective or acute repair of abdominal aortic aneurysms.Plasma concentrations of the cytokines tumour necrosis factor (TNF), interleukin-6 (IL-6) and interleukin-8 (IL-8) as well as soluble TNF receptors and the soluble (s) adhesion molecules E-selectin and intercellular adhesion molecule 1 (ICAM-1) were measured and correlated with the degree of systemic hypotension (shock: hypotension more than 15 minutes) and clinical outcome.Peak plasma concentrations of TNF and IL-6 were significantly higher in shocked patients (p < 0.005 and p < 0.0005, respectively) and those who died (both p < 0.01), whereas concentrations of IL-8 increased only when shock complicated rupture of the aneurysm (p < 0.01). Increases in the concentrations of TNF receptors reflected impaired renal function. In contrast to sE-selectin concentrations, peak sICAM-1 concentrations were significantly higher in shocked patients (p < 0.01) and those that died (p < 0.01).These results strongly suggest that increased concentrations of sICAM-1 and IL-6 reflect the inflammatory response induced by ischaemia after repair of an abdominal aortic aneurysm, and indicate that the postoperative course is likely to be complicated. |
| 1996 | Administration of an antibody to E-selectin in patients with septic shock. | Crit Care Med | To determine the safety and pharmacokinetics of a murine monoclonal antibody to E-selectin in patients with newly developed septic shock.Open-label, prospective, phase II pilot study with escalating doses of the antibody.Intensive care unit of a 900-bed university hospital.Nine patients who survived the first 24 hrs of septic shock.In addition to standard therapy, an intravenous bolus of a murine monoclonal antibody to E-selectin, CY1787, was given at doses of 0.1 mg/kg (n = 3), 0.33 mg/kg (n = 3), and 1.0 mg/kg (n = 3).CY1787 was well tolerated in all patients. Signs of shock resolved in all patients, and organ failure entirely reversed in eight patients. All patients survived the 28-day follow-up. Administration of CY1787 was associated with an early and brisk increase in PaO2/FIO2 ratio (p < .001), from 146 +/- 38 mm Hg (19.5 +/- 5.1 kPa) to 205 +/- 45 mm Hg (27.3 +/- 6.0 kPa) after 2 hrs, and 250 +/- 58 mm Hg (33.3 +/- 7.7 kPa) after 12 hrs. A dose-related effect of CY1787 was suggested by an earlier weaning from catecholamine therapy and a faster resolution of organ failure in the high-dose group. Development of antimouse antibodies was documented in eight patients.This pilot study indicates that this antibody to E-selectin appears to be safe and may represent a promising form of therapy in septic shock. |
| 1995 | Tumor necrosis factor induces E-selectin production in splanchnic artery occlusion shock. | Am J Physiol | Splanchnic arteries were clamped for 45 min to induce splanchnic artery occlusion (SAO) shock in anesthetized rats. Sham-operated animals were used as controls. Survival time, serum tumor necrosis factor-alpha (TNF-alpha), white blood cell (WBC) count, mean arterial blood pressure, myeloperoxidase (MPO) activity, and serum levels of soluble E-selectin (sE-selectin) were investigated. SAO-shocked rats exhibited decreased survival time (95 +/- 11 min, whereas sham-shocked rats survived for > 5 h), reduced mean arterial blood pressure, increased serum levels of TNF-alpha (185 +/- 8 U/ml) and MPO activity in the ileum (0.11 +/- 0.03 U x 10(-3)/g tissue) and lung (1.5 +/- 0.4 U x 10(-3)/g tissue), leukopenia, and enhanced serum levels of sE-selectin. Furthermore SAO-shocked rats showed histological alterations in the ileum and lung. Administration of cloricromene (2 mg/kg i.v.), an inhibitor of TNF-alpha, significantly increased survival time (225 +/- 10 min), decreased serum levels of TNF-alpha and sE-selectin, reduced leukopenia and MPO activity in the ileum (0.035 +/- 0.003 U x 10(-3)/g tissue) and lung (0.3 +/- 0.005 U x 10(-3)/g tissue), improved the cardiovascular changes, and reduced the histological changes in the ileum and lung. Finally, an anti-E-selectin antibody protected rats against SAO shock. Our findings are consistent with an involvement of E-selectin, "in vivo," in the pathogenesis of SAO shock. |
| 1995 | E-selectin involvement in the pathogenesis of splanchnic artery occlusion shock. | Eur J Pharmacol | We investigated the involvement of E-selectin in the pathogenesis of splanchnic artery occlusion shock. Splanchnic artery occlusion shock was induced in anaesthetized rats by clamping splanchnic arteries for 45 min. Sham-operated animals were used as controls. Survival time, serum tumor necrosis factor-alpha, while blood cell count, mean arterial blood pressure and myeloperoxidase activity were determined. Splanchnic artery occlusion-shocked rats had a decreased survival time (85 +/- 8 min, while sham-shocked rats survived more than 4 h), reduced mean arterial blood pressure, increased serum levels of tumor necrosis factor-alpha (186 +/- 9 U/ml) and myeloperoxidase activity in the ileum (0.10 +/- 0.04 U x 10(-3)/g tissue) and in the lung (1.5 +/- 0.06 U x 10(-3)/g tissue). Shocked rats showed histological alterations in the ileum and in the lung. Administration of a hyperimmune serum containing specific antibodies raised against E-selectin significantly increased survival time (225 +/- 10 min), reduced leukopenia and myeloperoxidase activity both in the ileum (0.035 +/- 0.001 U x 10(-3)/g tissue) and in the lung (0.3 +/- 0.005 U x 10(-3)/g tissue), improved the cardiovascular changes and reduced the histological alterations in the ileum and lung. Our data are consistent with an involvement of E-selectin in the pathogenesis of splanchnic artery occlusion shock. |
| 1993 | Expression of tissue factor, thrombomodulin, and E-selectin in baboons with lethal Escherichia coli sepsis. | Am J Pathol | Disseminated intravascular thrombosis is a frequent complication of endotoxic shock, and modulation of endothelial cell hemostatic properties has been proposed to play a role in its pathogenesis based on studies of endothelial cells in culture. This study examined the in vivo expression of tissue factor (TF) and thrombomodulin (TM) in a baboon model of lethal Escherichia coli sepsis using immunohistochemistry with monospecific antibodies. Expression of E-selectin (E-sel) was also determined as a marker of endothelial cell activation. Correlation of immunoreactivity with procoagulant activity in lipopolysaccharide-stimulated cultured human endothelial cells showed that immunohistochemistry was sufficiently sensitive to detect as little as 5% of the maximum in vitro endothelial cell TF response. Vascular endothelium of control animals expressed TM but had no detectable TF or E-sel. Following E. coli infusion, widespread E-sel expression and microvascular fibrin deposition was evident within 6 hours. However, expression of TF by endothelial cells became detectable only in the splenic microvasculature, where endothelial specificity of TF expression was confirmed by dual immunofluorescence of TF with von Willebrand's factor and with TM. In the spleen, there was a dissociation of expression of TF and E-sel, with marginal zone vessels being TF-positive and E-sel-negative, whereas sinusoidal endothelium was E-sel-positive but TF-negative. TM expression was unchanged from controls. Additionally, expression of TF by lung alveolar epithelial cells, splenic macrophages, and epithelial cells of the renal glomeruli was observed to be enhanced in septic animals. This study documents endothelial cell expression of TF in vivo in a relevant pathological setting. At the same time, compared with endothelial cells in culture, there is in vivo both significantly greater control of TF expression than expected, given the strong positive stimuli present in lethal E. coli septic shock and an unpredicted heterogeneity of activation responses. |
| 1993 | Soluble E-selectin is found in supernatants of activated endothelial cells and is elevated in the serum of patients with septic shock. | J Immunol | A quantitative sandwich ELISA for E-selectin in the fluid phase (soluble E-selectin, sEs) has been developed that is sensitive to 100 pg/ml. The assay shows no reactivity with either L- or P-selectins. We have used this to determine the fate of E-selectin after cell-surface expression and to test whether levels measured in vivo may represent the state of endothelial activation. E-selectin was first detectable in supernatants of IL-1-stimulated endothelial cells at 24 h, and increased slowly up until 72 h. However, over this time period the total E-selectin detectable in the system (cells plus supernatants) declined dramatically. 125I-surface-labeled endothelial cells cultured for 24 h show an E-selectin of reduced m.w. in the supernatant, indicating that the molecule is shed from the surface. The shed form also appears to be slightly smaller than the intact membrane form as determined from immunoprecipitation and molecular sieving studies. In addition, the cytoplasmic domain of the molecule found in supernatants of activated endothelial cells and in serum is not intact as determined by loss of reactivity with an antipeptide antibody specific for the cytoplasmic domain. We have examined the sera of 71 normal individuals. Without exception, sEs was found in serum in the range of 0.13 to 2.8 ng/ml, suggesting that even in the absence of overt inflammatory processes E-selectin is being synthesized and released into the bloodstream. In addition, bacteremic patients with hypotension, but not those without, showed markedly elevated sEs values. As determined by cell-binding studies, the blood-derived form of E-selectin is biologically active. |