| 1. |
- Bazov, Igor, 1973-, et al.
(author)
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Neuronal Expression of Opioid Gene is Controlled by Dual Epigenetic and Transcriptional Mechanism in Human Brain
- 2018
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In: Cerebral Cortex. - : Oxford University Press (OUP). - 1047-3211 .- 1460-2199. ; 28:9, s. 3129-3142
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Journal article (peer-reviewed)abstract
- Molecular mechanisms that define patterns of neuropeptide expression are essential for the formation and rewiring of neural circuits. The prodynorphin gene (PDYN) gives rise to dynorphin opioid peptides mediating depression and substance dependence. We here demonstrated that PDYN is expressed in neurons in human dorsolateral prefrontal cortex (dlPFC), and identified neuronal differentially methylated region in PDYN locus framed by CCCTC-binding factor binding sites. A short, nucleosome size human-specific promoter CpG island (CGI), a core of this region may serve as a regulatory module, which is hypomethylated in neurons, enriched in 5-hydroxymethylcytosine, and targeted by USF2, a methylation-sensitive E-box transcription factor (TF). USF2 activates PDYN transcription in model systems, and binds to nonmethylated CGI in dlPFC. USF2 and PDYN expression is correlated, and USF2 and PDYN proteins are co-localized in dlPFC. Segregation of activatory TF and repressive CGI methylation may ensure contrasting PDYN expression in neurons and glia in human brain.
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| 2. |
- Henriksson, Richard, et al.
(author)
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Elevated synaptophysin I in the prefrontal cortex of human chronic alcoholics
- 2008
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In: Synapse. - : Wiley. - 0887-4476 .- 1098-2396. ; 62:11, s. 829-33
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Journal article (peer-reviewed)abstract
- Convergent lines of evidence suggest potentiation of glutamatergic synapses after chronic ethanol exposure, and indicate that the presynaptic effect hereof is on modulators of synaptic strength rather than on executors of glutamate release. To address this hypothesis in the context of ethanol dependence in humans, we used semiquantitative immunoblotting to compare the immunoreactivities of synaptophysin I, syntaxin 1A, synaptosome-associated protein 25, and vesicle-associated membrane protein in the prefrontal and motor cortices between chronic alcoholics and control subjects. We found a region-specific elevation in synaptophysin I immunoreactivity in the prefrontal cortex of alcoholics, but detected no significant differences between the groups in the immunoreactivities of the other three proteins. Our findings are consistent with an effect of repeated ethanol exposure on modulators of synaptic strength but not on executors of glutamate release, and suggest a role for synaptophysin I in the enduring neuroplasticity in the prefrontal cortical glutamate circuitry that is associated with ethanol dependence.
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| 3. |
- Johansson, Sofia, et al.
(author)
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Dysregulation of cell death machinery in the prefrontal cortex of human alcoholics
- 2009
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In: International Journal of Neuropsychopharmacology. - 1461-1457 .- 1469-5111. ; 12:1, s. 109-115
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Journal article (peer-reviewed)abstract
- In human alcoholics, the cell density is decreased in the prefrontal cortex (PFC) and other brain areas. This may be due to persistent activation of cell death pathways. To address this hypothesis, we examined the status of cell death machinery in the dorsolateral PFC in alcoholics. Protein and mRNA expression levels of several key pro- and anti-apoptotic genes were compared in post-mortem samples of 14 male human alcoholics and 14 male controls. The findings do not support the hypothesis. On the contrary, they show that several components of intrinsic apoptotic pathway are decreased in alcoholics. No differences were evident in the motor cortex, which is less damaged in alcoholics and was analysed for comparison. Thus, cell death mechanisms may be dysregulated by inhibition of intrinsic apoptotic pathway in the PFC in human alcoholics. This inhibition may reflect molecular adaptations that counteract alcohol neurotoxicity in cells that survive after many years of alcohol exposure and withdrawal.
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| 4. |
- Johansson, Sofia, et al.
(author)
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Validation of endogenous controls for quantitative gene expression analysis : Application on brain cortices of human chronic alcoholics
- 2007
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In: Brain Research. - : Elsevier BV. - 0006-8993 .- 1872-6240. ; 1132:1, s. 20-8
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Journal article (peer-reviewed)abstract
- Real-time PCR is frequently used for gene expression quantification due to its methodological sensitivity and reproducibility. The gene expression is quantified by normalization to one or more reference genes, usually beta-actin (ACTB), glyceraldehyde-3-phosphate dehydrogenase (GAPD) or to ribosomal RNA (18S). However, different environmental or pathological conditions might also influence the expression of normalizing genes, which could severely skew the interpretation of quantitative results. This study evaluates whether 16 genes frequently used as endogenous controls in expression studies, can serve as such for comparison of human brain tissues of chronic alcoholics and control subjects. The prefrontal and motor cortices that are affected differently by chronic alcohol consumption were analyzed. The reference genes that have no or small differences in expression in alcoholics and control subjects, were found to be specific for each region: beta-actin (ACTB) and ribosomal large P0 (RPLP0) for the prefrontal cortex while importin 8 (IPO8) and RNA polymerase II (POLR2A) for the motor cortex. Four out of sixteen analyzed genes demonstrated significant differences in expression between alcoholics and controls: phosphoglycerate kinase (PGK1), hypoxanthine phosphoribosyl transferase (HPRT1) and peptidylprolyl isomerase A (PPIA) in the motor cortex and beta-2-microglobulin (B2M) in the prefrontal cortex. Our study demonstrates the importance of validation of endogenous control genes prior to real-time PCR analysis of human brain tissues. Prescribed and non-prescribed drugs, pathological or environmental conditions along with alcohol abuse may differentially influence expression of reference genes.
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| 5. |
- Kuzmin, Alexander, et al.
(author)
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Expression of pronociceptin and its receptor is downregulated in the brain of human alcoholics
- 2009
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In: Brain Research. - : Elsevier BV. - 0006-8993 .- 1872-6240. ; 1305:Suppl. 1, s. S80-85
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Journal article (peer-reviewed)abstract
- Animal studies demonstrated a role of neuropeptide nociceptin (NC) and its receptor (opiate receptor like-1, OPRL1) in ethanol-induced reward; activation of the OPRL1 by natural or synthetic ligands reduced ethanol self-administration and prevented relapse to ethanol drinking. The endogenous NC may function in neuronal circuits involved in reinforcing or conditioning effects of ethanol as a "brake" to limit ethanol intake (Roberto, M., Siggins, G.R. 2006. Nociceptin/orphanin FQ presynaptically decreases GABAergic transmission and blocks the ethanol-induced increase of GABA release in central amygdala. Proc. Natl. Acad. Sci. USA 103. 9715-9720), whereas repeated ethanol intake may downregulate the endogenous NC/OPRL1 system resulting in activation of ethanol consumption. To address this hypothesis, we evaluated whether expression of the pronociceptin (PNOC) and OPRL1 genes is altered in human alcoholics. mRNAs transcribed from these genes were analyzed by quantitative RT-PCR in the prefrontal and orbitofrontal cortices, central amygdala and hippocampal dentate gyrus, structures controlling alcohol consumption. Reduction in PNOC mRNA (1.7-fold) was found in the hippocampus of alcoholics, whereas OPRL1 mRNA levels were decreased (1.4-fold) in the central amygdala. No changes in expression of these genes in other brain areas analyzed were evident. We hypothesise that chronic ethanol intake downregulates PNOC and OPRL1 gene expression in the hippocampus and amygdala, respectively. The findings may be also interpreted as inherited molecular differences between alcoholics and controls. The PNOC/OPRL1 downregulation may underlie impairment of cognitive control over alcohol seeking in alcoholics. Stimulation of the OPRL1 receptors with synthetic agonists may increase threshold for activation of ethanol-related behaviour by environmental cues, and thus may reduce cue- or stress-primed relapse to ethanol consumption.
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| 6. |
- Taqi, Malik Mumtaz, et al.
(author)
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Prodynorphin CpG-SNPs associated with alcohol dependence : elevated methylation in the brain of human alcoholics
- 2011
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In: Addiction Biology. - : Wiley. - 1355-6215 .- 1369-1600. ; 16:3, s. 499-509
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Journal article (peer-reviewed)abstract
- The genetic, epigenetic and environmental factors may influence the risk for neuropsychiatric disease through their effects on gene transcription. Mechanistically, these effects may be integrated through regulation of methylation of CpG dinucleotides overlapping with single-nucleotide polymorphisms (SNPs) associated with a disorder. We addressed this hypothesis by analyzing methylation of prodynorphin (PDYN) CpG-SNPs associated with alcohol dependence, in human alcoholics. Postmortem specimens of the dorsolateral prefrontal cortex (dl-PFC) involved in cognitive control of addictive behavior were obtained from 14 alcohol-dependent and 14 control subjects. Methylation was measured by pyrosequencing after bisulfite treatment of DNA. DNA binding proteins were analyzed by electromobility shift assay. Three PDYN CpG-SNPs associated with alcoholism were found to be differently methylated in the human brain. In the dl-PFC of alcoholics, methylation levels of the C, non-risk variant of 3'-untranslated region (3'-UTR) SNP (rs2235749; C > T) were increased, and positively correlated with dynorphins. A DNA-binding factor that differentially targeted the T, risk allele and methylated and unmethylated C allele of this SNP was identified in the brain. The findings suggest a causal link between alcoholism-associated PDYN 3'-UTR CpG-SNP methylation, activation of PDYN transcription and vulnerability of individuals with the C, non-risk allele(s) to develop alcohol dependence.
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| 7. |
- Watanabe, Hiroyuki, et al.
(author)
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FOSB proteins in the orbitofrontal and dorsolateral prefrontal cortices of human alcoholics
- 2009
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In: Addiction Biology. - : Wiley. - 1355-6215 .- 1369-1600. ; 14:3, s. 294-297
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Journal article (peer-reviewed)abstract
- The transcription factor DeltaFosB is accumulated in the addiction circuitry, including the orbitofrontal and medial prefrontal cortices of rodents chronically exposed to ethanol or other drugs of abuse, and has been suggested to play a direct role in addiction maintenance. To address this hypothesis in the context of substance dependence in humans, we compared the immunoreactivities of FOSB proteins in the orbitofrontal and dorsolateral prefrontal cortices (OFC and DLPFC respectively) between controls and alcoholics using semiquantitative immunoblotting. In both structures, we detected three forms of FOSB, one of which was DeltaFOSB, but in neither case did their immunoreactivities differ between the groups. Our results indicate that the DeltaFOSB immunoreactivity in the human brain is very low, and that it is not accumulated in the OFC and DLPFC of human alcoholics, suggesting that it may not be directly involved in addiction maintenance, at least not in ethanol dependence.
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| 8. |
- Ökvist, Anna, et al.
(author)
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Neuroadaptations in human chronic alcoholics : dysregulation of the NF-κB system
- 2007
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In: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 2:9, s. e930-
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Journal article (peer-reviewed)abstract
- BACKGROUND: Alcohol dependence and associated cognitive impairments apparently result from neuroadaptations to chronic alcohol consumption involving changes in expression of multiple genes. Here we investigated whether transcription factors of Nuclear Factor-kappaB (NF-kappaB) family, controlling neuronal plasticity and neurodegeneration, are involved in these adaptations in human chronic alcoholics. METHODS AND FINDINGS: Analysis of DNA-binding of NF-kappaB (p65/p50 heterodimer) and the p50 homodimer as well as NF-kappaB proteins and mRNAs was performed in postmortem human brain samples from 15 chronic alcoholics and 15 control subjects. The prefrontal cortex involved in alcohol dependence and cognition was analyzed and the motor cortex was studied for comparison. The p50 homodimer was identified as dominant kappaB binding factor in analyzed tissues. NF-kappaB and p50 homodimer DNA-binding was downregulated, levels of p65 (RELA) mRNA were attenuated, and the stoichiometry of p65/p50 proteins and respective mRNAs was altered in the prefrontal cortex of alcoholics. Comparison of a number of p50 homodimer/NF-kappaB target DNA sites, kappaB elements in 479 genes, down- or upregulated in alcoholics demonstrated that genes with kappaB elements were generally upregulated in alcoholics. No significant differences between alcoholics and controls were observed in the motor cortex. CONCLUSIONS: We suggest that cycles of alcohol intoxication/withdrawal, which may initially activate NF-kappaB, when repeated over years downregulate RELA expression and NF-kappaB and p50 homodimer DNA-binding. Downregulation of the dominant p50 homodimer, a potent inhibitor of gene transcription apparently resulted in derepression of kappaB regulated genes. Alterations in expression of p50 homodimer/NF-kappaB regulated genes may contribute to neuroplastic adaptation underlying alcoholism.
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