• Energy Research

Abstract Background Alcohol use disorders (AUDs) lead to alterations in central nervous system (CNS) architecture along with impaired learning and memory. Previous work from our group and that of others suggests that one mechanism underlying these changes is alteration of cell proliferation, apoptosis, and DNA-repair in neural stem cells (NSCs) produced as a consequence of ethanol-induced effects on the expression of genes related to p53-signaling. This study tests the hypothesis that changes in the expression of p53-signaling genes represent biomarkers of ethanol abuse which can be identified in the peripheral blood of rat drinking models and human AUD subjects and posits that specific changes may be correlated with differences in neuropsychological measures and CNS structure. Results Remarkably, microarray analysis of 350 genes related to p53-signaling in peripheral blood leukocytes (PBLs) of binge-drinking rats revealed 190 genes that were significantly altered after correcting for multiple testing. Moreover, 40 of these genes overlapped with those that we had previously observed to be changed in ethanol-exposed mouse NSCs. Expression changes in nine of these genes were tested for independent confirmation by a custom QuantiGene Plex (QGP) assay for a subset of p53-signaling genes, where a consistent trend for decreased expression of mitosis-related genes was observed. One mitosis-related gene (Pttg1) was also changed in human lymphoblasts cultured with ethanol. In PBLs of human AUD subjects seven p53-signaling genes were changed compared with non-drinking controls. Correlation and principal components analysis were then used to identify significant relationships between the expression of these seven genes and a set of medical, demographic, neuropsychological and neuroimaging measures that distinguished AUD and control subjects. Two genes (Ercc1 and Mcm5) showed a highly significant correlation with AUD-induced decreases in the volume of the left parietal supramarginal gyrus and neuropsychological measures. Conclusions These results demonstrate that alcohol-induced changes in genes related to proliferation, apoptosis, and DNA-repair are observable in the peripheral blood and may serve as a useful biomarker for CNS structural damage and functional performance deficits in human AUD subjects.

Abstract Background Alcohol use disorders (AUDs) lead to alterations in central nervous system (CNS) architecture along with impaired learning and memory. Previous work from our group and that of others suggests that one mechanism underlying these changes is alteration of cell proliferation, apoptosis, and DNA-repair in neural stem cells (NSCs) produced as a consequence of ethanol-induced effects on the expression of genes related to p53-signaling. This study tests the hypothesis that changes in the expression of p53-signaling genes represent biomarkers of ethanol abuse which can be identified in the peripheral blood of rat drinking models and human AUD subjects and posits that specific changes may be correlated with differences in neuropsychological measures and CNS structure. Results Remarkably, microarray analysis of 350 genes related to p53-signaling in peripheral blood leukocytes (PBLs) of binge-drinking rats revealed 190 genes that were significantly altered after correcting for multiple testing. Moreover, 40 of these genes overlapped with those that we had previously observed to be changed in ethanol-exposed mouse NSCs. Expression changes in nine of these genes were tested for independent confirmation by a custom QuantiGene Plex (QGP) assay for a subset of p53-signaling genes, where a consistent trend for decreased expression of mitosis-related genes was observed. One mitosis-related gene (Pttg1) was also changed in human lymphoblasts cultured with ethanol. In PBLs of human AUD subjects seven p53-signaling genes were changed compared with non-drinking controls. Correlation and principal components analysis were then used to identify significant relationships between the expression of these seven genes and a set of medical, demographic, neuropsychological and neuroimaging measures that distinguished AUD and control subjects. Two genes (Ercc1 and Mcm5) showed a highly significant correlation with AUD-induced decreases in the volume of the left parietal supramarginal gyrus and neuropsychological measures. Conclusions These results demonstrate that alcohol-induced changes in genes related to proliferation, apoptosis, and DNA-repair are observable in the peripheral blood and may serve as a useful biomarker for CNS structural damage and functional performance deficits in human AUD subjects.

Prenatal ethanol exposure is associated with, and is a risk factor for, developmental disorders with abnormal social behaviors, including autism spectrum disorders. We hypothesize that the specific effects of ethanol on social behavior are defined by the timing of the exposure as well as subsequent changes in brain regions such as the amygdala and ventral striatum. We recently reported that in utero ethanol exposure on gestational day 12 alters social behaviors of weanling [postnatal day (P) 28], adolescent (P42), and young adult (P75) rats. Male, but not female, offspring of the ethanol-exposed dams showed significant decreases in social investigation (sniffing of a social partner), contact behavior (grooming or crawling over/under the partner), and play fighting (following, chasing, nape attacks, or pinning) at all ages tested with maximal effects at P28 and P42. Furthermore, ethanol-exposed males and females showed evidence of social avoidance at P42 and P75. The present study sought to test whether a form of social enrichment could normalize any of the social deficits and what the molecular mechanisms of such effects might be. We found that housing rats with nonmanipulated control rats normalized the social avoidance phenotype normally seen when they are housed with sex-matched prenatal ethanol-exposed littermates. There was no mitigation of the other ethanol-induced behavioral deficits. Conversely, male control-treated rats housed with nonlittermates showed deficits in play fighting, social investigation and contact behavior. Molecular analyses of the amygdala and ventral striatum of adolescent rats following fetal ethanol exposure indicated several specific neurotransmitter systems and pathways that might underlie the social avoidance phenotype as well as its reversal.

Prenatal ethanol exposure is associated with, and is a risk factor for, developmental disorders with abnormal social behaviors, including autism spectrum disorders. We hypothesize that the specific effects of ethanol on social behavior are defined by the timing of the exposure as well as subsequent changes in brain regions such as the amygdala and ventral striatum. We recently reported that in utero ethanol exposure on gestational day 12 alters social behaviors of weanling [postnatal day (P) 28], adolescent (P42), and young adult (P75) rats. Male, but not female, offspring of the ethanol-exposed dams showed significant decreases in social investigation (sniffing of a social partner), contact behavior (grooming or crawling over/under the partner), and play fighting (following, chasing, nape attacks, or pinning) at all ages tested with maximal effects at P28 and P42. Furthermore, ethanol-exposed males and females showed evidence of social avoidance at P42 and P75. The present study sought to test whether a form of social enrichment could normalize any of the social deficits and what the molecular mechanisms of such effects might be. We found that housing rats with nonmanipulated control rats normalized the social avoidance phenotype normally seen when they are housed with sex-matched prenatal ethanol-exposed littermates. There was no mitigation of the other ethanol-induced behavioral deficits. Conversely, male control-treated rats housed with nonlittermates showed deficits in play fighting, social investigation and contact behavior. Molecular analyses of the amygdala and ventral striatum of adolescent rats following fetal ethanol exposure indicated several specific neurotransmitter systems and pathways that might underlie the social avoidance phenotype as well as its reversal.

There is currently a lack of reliable, minimally invasive biomarkers that could predict the extent of alcoholism-induced CNS damage. Developing such biomarkers may prove useful in reducing the prevalence of alcohol use disorders (AUDs). Extracellular microRNAs (miRNAs) can be informative molecular indicators of changes in neuronal gene expression. In this study, we performed a global analysis of extracellular miRNAs to identify robust biomarkers of early CNS damage in humans diagnosed with DSM-IV AUDs. We recruited a relatively young set of 20 AUD subjects and 10 age-matched controls. They were subjected to comprehensive medical, neuropsychological and neuroimaging tests, followed by comparison of miRNA levels found in peripheral blood serum. Employing a conservative strategy to identify candidate biomarkers, miRNAs were quantified using two independent high-throughput methods: microarray and next-generation RNA-sequencing. This improved our capacity to discover and validate relevant miRNAs.Our results identified several miRNAs with significant and reproducible expression changes in AUD subjects versus controls. Moreover, several significant associations between candidate miRNA biomarkers and various medical, neuropsychological and neuroimaging parameters were identified using Pearson correlation and unbiased hierarchical clustering analyses. Some of the top candidate biomarkers identified, such as mir-92b and mir-96 have established roles in neural development. Cross-species validation of miRNA expression was performed using two different in vivo rat drinking models and two different in vitro mouse neural stem cell exposure models. A systems level analysis revealed a remarkable degree of convergence in the top changes seen in all of these data sets, specifically identifying cell death, cell proliferation and cell cycle processes as most consistently affected. Though not necessarily the same molecules, the affected miRNAs within these pathways clearly influence common genes, such as p53 and TNF, which stand out as potential keystone molecules. Lastly, we also examined the potential tissue origins of these biomarkers by quantifying their levels in 15 different tissue types and show that several are highly-enriched in the brain.Collectively, our results suggest that serum miRNA expression changes can directly relate to alterations in CNS structure and function, and may do so through effects on highly specific cellular pathways.

There is currently a lack of reliable, minimally invasive biomarkers that could predict the extent of alcoholism-induced CNS damage. Developing such biomarkers may prove useful in reducing the prevalence of alcohol use disorders (AUDs). Extracellular microRNAs (miRNAs) can be informative molecular indicators of changes in neuronal gene expression. In this study, we performed a global analysis of extracellular miRNAs to identify robust biomarkers of early CNS damage in humans diagnosed with DSM-IV AUDs. We recruited a relatively young set of 20 AUD subjects and 10 age-matched controls. They were subjected to comprehensive medical, neuropsychological and neuroimaging tests, followed by comparison of miRNA levels found in peripheral blood serum. Employing a conservative strategy to identify candidate biomarkers, miRNAs were quantified using two independent high-throughput methods: microarray and next-generation RNA-sequencing. This improved our capacity to discover and validate relevant miRNAs.Our results identified several miRNAs with significant and reproducible expression changes in AUD subjects versus controls. Moreover, several significant associations between candidate miRNA biomarkers and various medical, neuropsychological and neuroimaging parameters were identified using Pearson correlation and unbiased hierarchical clustering analyses. Some of the top candidate biomarkers identified, such as mir-92b and mir-96 have established roles in neural development. Cross-species validation of miRNA expression was performed using two different in vivo rat drinking models and two different in vitro mouse neural stem cell exposure models. A systems level analysis revealed a remarkable degree of convergence in the top changes seen in all of these data sets, specifically identifying cell death, cell proliferation and cell cycle processes as most consistently affected. Though not necessarily the same molecules, the affected miRNAs within these pathways clearly influence common genes, such as p53 and TNF, which stand out as potential keystone molecules. Lastly, we also examined the potential tissue origins of these biomarkers by quantifying their levels in 15 different tissue types and show that several are highly-enriched in the brain.Collectively, our results suggest that serum miRNA expression changes can directly relate to alterations in CNS structure and function, and may do so through effects on highly specific cellular pathways.

AbstractTransforming growth factor (TGF) β1 and ethanol retard the migration of young, post‐mitotic neurons to the developing cerebral cortex. The coordination of this migration depends upon cell adhesion proteins (CAPs). We examined the effects of TGFβ1 and ethanol on genes related to both TGF and CAPs. Rat B104 neuroblastoma cells were treated with TGFβ1 (0 or 10 ng/mL) and ethanol (0 or 400 mg/dL) for 6–48 h. Total RNA was purified from each sample and analyzed using the Rat U34A GeneChip (Affymetrix). Candidate genes were those up‐ or down‐regulated by either TGFβ1 or ethanol. Twenty transcripts of CAPs were identified as being expressed by B104 cells and as being affected by treatment with TGFβ1 or ethanol. The expression was verified for five representative genes (neural cell adhesion molecule, L1, and integrins α1, α7, and β1) using assays with real‐time reverse transcriptase–polymerase chain reactions. Each of these genes showed time‐dependent changes. The changes were reflected in increases in protein expression that appeared within 24 or 48 h. Thus, the effects of TGFβ1 and ethanol on CAPs parallel changes described in vivo and likely underlie changes associated with ethanol‐induced alterations in neuronal migration.