| 2018 | Intergenerational Transmission of DNA Methylation Signatures Associated with Early Life Stress. | Curr Genomics | Early life stress in humans (i.e. maltreatment, violence exposure, loss of a loved one) and in rodents (i.e. disrupted attachment or nesting, electric shock, restraint, predator odor) occurs during critical steps of neural circuit formation. ELS in humans is associated with increased risk for developmental psychopathology, including anxious and depressive phenotypes. The biological mechanisms underlying these potentially persistent maladaptive changes involve long-term epigenetic modifications, which have been suggested to be potentially transmissible to subsequent generations. DNA methylation is an epigenetic mechanism that modifies gene expression patterns in response to environmental challenges and influences mutation rates. It remains to be seen whether a functionally relevant fraction of DNA methylation marks can escape genome-wide erasures that occur in primordial germ cells and after fertilization within the zygote. Early life-stress-triggered changes in epigenetic mediated transmission of acquired behavioral traits among humans have been assessed mainly by targeting genes involved in the hypothalamic-pituitary-adrenal (HPA) axis, such as NR3C1 and FKBP5. Recently, researchers examining epigenetic transmission have begun to apply genome-wide approaches. In humans, reduced representation bisulfite sequencing (RRBS) was performed on peripheral samples that were obtained from individuals who were prenatally exposed to the "Dutch Hunger Winter", resulting in two Differentially Methylated Regions (DMRs) in INSR and CPTIA genes that were functionally, biologically and technically validated, and significantly associated with birth weights and LDL cholesterol levels in offspring. In rodents, non-genomic intergenerational transmission of anxiety which was associated with differentially methylated enhancers that were putatively involved in lipid signaling and synaptic/neurotransmission in hippocampal granule cells, was discovered also using RRBS. Finally, transgenerational transmission of altered behaviors was associated with sperm-derived microRNAs produced by ELS male mice. The field of epigenetic transmission is just beginning to enter the epigenomic era by using genome-wide analyses. Such approaches remain of strong interest to human studies, first in order to help to assess the relevance of the previous targeted studies, and second to discover new important epigenetic modifications of potential clinical importance. New discoveries may help to assess how transmittable the negative impact of stress may be to offspring. The latter may open doors for future treatments and resilience-promoting interventions, as well as new approaches to treat the effects of childhood trauma before the onset of psychiatric disorder. |
| 2015 | Expression profile of six stress-related genes and productive performances of fast and slow growing broiler strains reared under heat stress conditions. | Meta Gene | High temperature is one of the prominent environmental factors causing economic losses to the poultry industry as it negatively affects growth and production performance in broiler chickens. We used One Step TaqMan real time RT-PCR (reverse transcription polymerase chain reaction) technology to study the effects of chronic heat stress on the expression of genes codifying for the antioxidative enzymes superoxide dismutase (SOD), and catalase (CAT), as well as for heat shock protein (HSP) 70, HSP90, glucocorticoid receptor (NR3C1), and caspase 6 (CASP6) in the liver of two different broiler genetic strains: Red JA Cou Nu Hubbard (CN) and Ross 508 Aviagen (RO). CN is a naked neck slow growing broiler intended for the free range and/or organic markets, whereas RO is selected for fast growing. We also analysed the effect of chronic heat stress on productive performances, and plasma corticosterone levels as well as the association between transcriptomic response and specific SNPs (single nucleotide polymorphisms) in each genetic strain of broiler chickens. RO and CN broilers, 4 weeks of age, were maintained for 4 weeks at either 34 °C or 22 °C. The results demonstrated that there was a genotype and a temperature main effect on the broilers' growth from the 4th to the 8th week of age, but the interaction effect between genotype and temperature resulted not statistically significant. By considering the genotype effect, fast growing broilers (RO) grew more than the slow growing ones (CN), whereas by considering the temperature effect, broilers in unheated conditions grew more than the heat stressed ones. Corticosterone levels increased significantly in the blood of heat stressed broilers, due to the activation of the HPA (hypothalamic-pituitary-adrenocortical axis). Carcass yield at slaughter was of similar values in the 4 cohorts (genotype/temperature combinations or treatment groups), ranging from 86.5 to 88.6%, whereas carcass weight was negatively influenced by heat stress in both broiler strains. Heat stress affected gene expression by downregulating CASP6 and upregulating CAT transcript levels. HSPs, SOD and NR3C1 mRNA levels remained unaffected by heat stress. The differences found in the mRNA copies of CASP6 gene could be partly explained by SNPs. |
| 2014 | Inhibitory avoidance learning in zebrafish (Danio rerio): effects of shock intensity and unraveling differences in task performance. | Zebrafish | The zebrafish (Danio rerio) is increasingly used as a model in neurobehavioral and neuroendocrine studies. The inhibitory avoidance paradigm has been proposed as tool to study mechanisms underlying learning and memory in zebrafish. In this paradigm subjects receive a shock after entering the black compartment of a black-white box. On the next day, latency to enter the black compartment is assessed; higher latencies are indicative of increased avoidance learning. Here, we aimed to understand the effects of different shock intensities (0, 1, 3, and 9 V) and to unravel variation in inhibitory avoidance learning in an in-house reared Tuebingen Long-Fin zebrafish (D. rerio) strain. While median latencies had increased in the 1, 3, and 9 V groups, no increase in median latency was found in the 0 V group. In addition, higher shock intensities resulted in a higher number of avoiders (latency ≥180 s) over nonavoiders (latency <60 s). Both changes are indicative of increased avoidance learning. We assessed whole-body cortisol content and the expression levels of genes relevant to stress, anxiety, fear, and learning 2 h after testing. Shock intensity was associated with whole-body cortisol content and the expression of glucocorticoid receptor alpha [nr3c1(alpha)], cocaine- and amphetamine-regulated transcript (cart4), and mineralocorticoid receptor (nr3c2), while avoidance behavior was associated with whole-body cortisol content only. The inhibitory avoidance paradigm in combination with measuring whole-body cortisol content and gene expression is suitable to unravel (genetic) mechanisms of fear avoidance learning. Our data further show differences in brain-behavior relationships underlying fear avoidance learning and memory in zebrafish. These findings serve as starting point for further unraveling differences in brain-behavior relationships underlying (fear avoidance) learning and memory in zebrafish. |
| 2011 | Acetyl-L-Carnitine Modulates TP53 and IL10 Gene Expression Induced by 3-NPA Evoked Toxicity in PC12 Cells. | Curr Neuropharmacol | The neurotoxicity induced by the mitochondrial inhibitor 3-nitropropionic acid (3-NPA) is associated with a decrease of ATP synthesis and an increase of free radical production which can lead to apoptosis or necrosis. We have used the PC12, neuron-like rat pheochromocytoma cell line, to study further the mechanism of 3-NPA-evoked neurotoxicity and the effects of acetyl-L-carnitine (ALC) which has neuroprotective actions against various types of mitochondrial inhibitors.Cultured PC 12 cells were exposed to a low dose of 3-NPA 50 (microM) in the presence or absence of 5 mM ALC. The dose of 3-NPA was sub toxic and no changes in pro-apoptotic Bax or anti-apoptotic Bcl-2 gene expression were observed. We followed specific genetic markers to look for changes evoked by 3-NPA toxicity and also changes associated with neuroprotection exerted by the ALC treatment, using RT-PCR arrays (delta-delta method). 3-NPA exposure evoked a decrease in expression of the Tp53 gene. This down regulation was prevented by pretreatment of the cells with ALC. The Tp53 gene responds to cellular stresses and the effects seen here are possibly associated with the 3-NPA evoked changes in mitochondrial metabolism. Other genes associated with stress and apoptosis, Parp-1, Bcl-2, and Bax were not affected by 3-NPA or ALC. The decrease of inflammatory response Il-10 gene expression due to 3-NPA was further lowered by presence of ALC. Other inflammation related genes, Il1rn, Nr3c1 and Cxcr4 were not affected. Interestingly, the glutamate transporter slc17a7, carnitine-acylcarnitine translocase Slc25a20 and heat shock proteins genes, Hsp27, Hmox1 (Hsp32, HO1) as well as Hspa 1a (Hsp 70) increased only when both ALC and small dose of 3-NPA were present. The alterations in gene expression detected in this study suggest role of several intracellular pathways in the neurotoxicity of 3-NPA and the neuroprotection against 3-NPA-induced neurotoxicity by ALC. |
| 2010 | Biological and diurnal variation in glucocorticoid sensitivity detected with a sensitive in vitro dexamethasone suppression of cytokine production assay. | J Clin Endocrinol Metab | Glucocorticoid resistance due to mutations of the glucocorticoid receptor (NR3C1) are rare, but reduced glucocorticoid sensitivity may play a role in steroid-resistant asthma, inflammatory bowel disease, and septic shock. A rapid and sensitive functional assay to detect glucocorticoid resistance will be advantageous.We describe a rapid in vitro monocyte dexamethasone suppression of cytokine production (DSCP) assay with a 3-h incubation. The DSCP assay was compared with the reference stimulated lymphocyte proliferation method. We characterized the effect of delayed processing, time of sampling, and biological variation on the DSCP assay.The DSCP assay clearly distinguished subjects with a known heterozygous mutation of the NR3C1 gene from control subjects, whereas the reference method failed. Decreased glucocorticoid sensitivity was demonstrated in samples collected in the afternoon. Intra-individual variation from samples collected in the morning was 13.0 and 12.7% for the IL-6 and TNF-alpha responses with the respective inter-individual variations of 9.7 and 7.9%.The DSCP assay was superior to the reference method and was sufficiently sensitive to detect diurnal variation in control subjects. The biological variation data supported the use of a population-based reference interval. The assay is suitable for screening of glucocorticoid resistance phenotypes and may provide insight into cortisol metabolism in critically ill patients, asthmatics, and patients with inflammatory bowel disease provided that the variation due to delayed processing and time of collection are considered. |