4 research outputs found
Image_1_A pilot investigation of differential hydroxymethylation levels in patient-derived neural stem cells implicates altered cortical development in bipolar disorder.TIF
IntroductionBipolar disorder (BD) is a chronic mental illness characterized by recurrent episodes of mania and depression and associated with social and cognitive disturbances. Environmental factors, such as maternal smoking and childhood trauma, are believed to modulate risk genotypes and contribute to the pathogenesis of BD, suggesting a key role in epigenetic regulation during neurodevelopment. 5-hydroxymethylcytosine (5hmC) is an epigenetic variant of particular interest, as it is highly expressed in the brain and is implicated in neurodevelopment, and psychiatric and neurological disorders.MethodsInduced pluripotent stem cells (iPSCs) were generated from the white blood cells of two adolescent patients with bipolar disorder and their same-sex age-matched unaffected siblings (n =β4). Further, iPSCs were differentiated into neuronal stem cells (NSCs) and characterized for purity using immuno-fluorescence. We used reduced representation hydroxymethylation profiling (RRHP) to perform genome-wide 5hmC profiling of iPSCs and NSCs, to model 5hmC changes during neuronal differentiation and assess their impact on BD risk. Functional annotation and enrichment testing of genes harboring differentiated 5hmC loci were performed with the online tool DAVID.ResultsApproximately 2 million sites were mapped and quantified, with the majority (68.8%) located in genic regions, with elevated 5hmC levels per site observed for 3β UTRs, exons, and 2-kb shorelines of CpG islands. Paired t-tests of normalized 5hmC counts between iPSC and NSC cell lines revealed global hypo-hydroxymethylation in NSCs and enrichment of differentially hydroxymethylated sites within genes associated with plasma membrane (FDRβ=β9.1βΓβ10β12) and axon guidance (FDRβ=β2.1βΓβ10β6), among other neuronal processes. The most significant difference was observed for a transcription factor binding site for the KCNK9 gene (p =β8.8βΓβ10β6), encoding a potassium channel protein involved in neuronal activity and migration. Proteinβprotein-interaction (PPI) networking showed significant connectivity (p =β3.2βΓβ10β10) between proteins encoded by genes harboring highly differentiated 5hmC sites, with genes involved in axon guidance and ion transmembrane transport forming distinct sub-clusters. Comparison of NSCs of BD cases and unaffected siblings revealed additional patterns of differentiation in hydroxymethylation levels, including sites in genes with functions related to synapse formation and regulation, such as CUX2 (p =β2.4βΓβ10β5) and DOK-7 (p =β3.6βΓβ10β3), as well as an enrichment of genes involved in the extracellular matrix (FDRβ=β1.0βΓβ10β8).DiscussionTogether, these preliminary results lend evidence toward a potential role for 5hmC in both early neuronal differentiation and BD risk, with validation and more comprehensive characterization to be achieved through follow-up study.</p
Image_2_A pilot investigation of differential hydroxymethylation levels in patient-derived neural stem cells implicates altered cortical development in bipolar disorder.TIFF
IntroductionBipolar disorder (BD) is a chronic mental illness characterized by recurrent episodes of mania and depression and associated with social and cognitive disturbances. Environmental factors, such as maternal smoking and childhood trauma, are believed to modulate risk genotypes and contribute to the pathogenesis of BD, suggesting a key role in epigenetic regulation during neurodevelopment. 5-hydroxymethylcytosine (5hmC) is an epigenetic variant of particular interest, as it is highly expressed in the brain and is implicated in neurodevelopment, and psychiatric and neurological disorders.MethodsInduced pluripotent stem cells (iPSCs) were generated from the white blood cells of two adolescent patients with bipolar disorder and their same-sex age-matched unaffected siblings (n =β4). Further, iPSCs were differentiated into neuronal stem cells (NSCs) and characterized for purity using immuno-fluorescence. We used reduced representation hydroxymethylation profiling (RRHP) to perform genome-wide 5hmC profiling of iPSCs and NSCs, to model 5hmC changes during neuronal differentiation and assess their impact on BD risk. Functional annotation and enrichment testing of genes harboring differentiated 5hmC loci were performed with the online tool DAVID.ResultsApproximately 2 million sites were mapped and quantified, with the majority (68.8%) located in genic regions, with elevated 5hmC levels per site observed for 3β UTRs, exons, and 2-kb shorelines of CpG islands. Paired t-tests of normalized 5hmC counts between iPSC and NSC cell lines revealed global hypo-hydroxymethylation in NSCs and enrichment of differentially hydroxymethylated sites within genes associated with plasma membrane (FDRβ=β9.1βΓβ10β12) and axon guidance (FDRβ=β2.1βΓβ10β6), among other neuronal processes. The most significant difference was observed for a transcription factor binding site for the KCNK9 gene (p =β8.8βΓβ10β6), encoding a potassium channel protein involved in neuronal activity and migration. Proteinβprotein-interaction (PPI) networking showed significant connectivity (p =β3.2βΓβ10β10) between proteins encoded by genes harboring highly differentiated 5hmC sites, with genes involved in axon guidance and ion transmembrane transport forming distinct sub-clusters. Comparison of NSCs of BD cases and unaffected siblings revealed additional patterns of differentiation in hydroxymethylation levels, including sites in genes with functions related to synapse formation and regulation, such as CUX2 (p =β2.4βΓβ10β5) and DOK-7 (p =β3.6βΓβ10β3), as well as an enrichment of genes involved in the extracellular matrix (FDRβ=β1.0βΓβ10β8).DiscussionTogether, these preliminary results lend evidence toward a potential role for 5hmC in both early neuronal differentiation and BD risk, with validation and more comprehensive characterization to be achieved through follow-up study.</p
Table_1_A pilot investigation of differential hydroxymethylation levels in patient-derived neural stem cells implicates altered cortical development in bipolar disorder.xlsx
IntroductionBipolar disorder (BD) is a chronic mental illness characterized by recurrent episodes of mania and depression and associated with social and cognitive disturbances. Environmental factors, such as maternal smoking and childhood trauma, are believed to modulate risk genotypes and contribute to the pathogenesis of BD, suggesting a key role in epigenetic regulation during neurodevelopment. 5-hydroxymethylcytosine (5hmC) is an epigenetic variant of particular interest, as it is highly expressed in the brain and is implicated in neurodevelopment, and psychiatric and neurological disorders.MethodsInduced pluripotent stem cells (iPSCs) were generated from the white blood cells of two adolescent patients with bipolar disorder and their same-sex age-matched unaffected siblings (n =β4). Further, iPSCs were differentiated into neuronal stem cells (NSCs) and characterized for purity using immuno-fluorescence. We used reduced representation hydroxymethylation profiling (RRHP) to perform genome-wide 5hmC profiling of iPSCs and NSCs, to model 5hmC changes during neuronal differentiation and assess their impact on BD risk. Functional annotation and enrichment testing of genes harboring differentiated 5hmC loci were performed with the online tool DAVID.ResultsApproximately 2 million sites were mapped and quantified, with the majority (68.8%) located in genic regions, with elevated 5hmC levels per site observed for 3β UTRs, exons, and 2-kb shorelines of CpG islands. Paired t-tests of normalized 5hmC counts between iPSC and NSC cell lines revealed global hypo-hydroxymethylation in NSCs and enrichment of differentially hydroxymethylated sites within genes associated with plasma membrane (FDRβ=β9.1βΓβ10β12) and axon guidance (FDRβ=β2.1βΓβ10β6), among other neuronal processes. The most significant difference was observed for a transcription factor binding site for the KCNK9 gene (p =β8.8βΓβ10β6), encoding a potassium channel protein involved in neuronal activity and migration. Proteinβprotein-interaction (PPI) networking showed significant connectivity (p =β3.2βΓβ10β10) between proteins encoded by genes harboring highly differentiated 5hmC sites, with genes involved in axon guidance and ion transmembrane transport forming distinct sub-clusters. Comparison of NSCs of BD cases and unaffected siblings revealed additional patterns of differentiation in hydroxymethylation levels, including sites in genes with functions related to synapse formation and regulation, such as CUX2 (p =β2.4βΓβ10β5) and DOK-7 (p =β3.6βΓβ10β3), as well as an enrichment of genes involved in the extracellular matrix (FDRβ=β1.0βΓβ10β8).DiscussionTogether, these preliminary results lend evidence toward a potential role for 5hmC in both early neuronal differentiation and BD risk, with validation and more comprehensive characterization to be achieved through follow-up study.</p
Image_3_A pilot investigation of differential hydroxymethylation levels in patient-derived neural stem cells implicates altered cortical development in bipolar disorder.TIFF
IntroductionBipolar disorder (BD) is a chronic mental illness characterized by recurrent episodes of mania and depression and associated with social and cognitive disturbances. Environmental factors, such as maternal smoking and childhood trauma, are believed to modulate risk genotypes and contribute to the pathogenesis of BD, suggesting a key role in epigenetic regulation during neurodevelopment. 5-hydroxymethylcytosine (5hmC) is an epigenetic variant of particular interest, as it is highly expressed in the brain and is implicated in neurodevelopment, and psychiatric and neurological disorders.MethodsInduced pluripotent stem cells (iPSCs) were generated from the white blood cells of two adolescent patients with bipolar disorder and their same-sex age-matched unaffected siblings (n =β4). Further, iPSCs were differentiated into neuronal stem cells (NSCs) and characterized for purity using immuno-fluorescence. We used reduced representation hydroxymethylation profiling (RRHP) to perform genome-wide 5hmC profiling of iPSCs and NSCs, to model 5hmC changes during neuronal differentiation and assess their impact on BD risk. Functional annotation and enrichment testing of genes harboring differentiated 5hmC loci were performed with the online tool DAVID.ResultsApproximately 2 million sites were mapped and quantified, with the majority (68.8%) located in genic regions, with elevated 5hmC levels per site observed for 3β UTRs, exons, and 2-kb shorelines of CpG islands. Paired t-tests of normalized 5hmC counts between iPSC and NSC cell lines revealed global hypo-hydroxymethylation in NSCs and enrichment of differentially hydroxymethylated sites within genes associated with plasma membrane (FDRβ=β9.1βΓβ10β12) and axon guidance (FDRβ=β2.1βΓβ10β6), among other neuronal processes. The most significant difference was observed for a transcription factor binding site for the KCNK9 gene (p =β8.8βΓβ10β6), encoding a potassium channel protein involved in neuronal activity and migration. Proteinβprotein-interaction (PPI) networking showed significant connectivity (p =β3.2βΓβ10β10) between proteins encoded by genes harboring highly differentiated 5hmC sites, with genes involved in axon guidance and ion transmembrane transport forming distinct sub-clusters. Comparison of NSCs of BD cases and unaffected siblings revealed additional patterns of differentiation in hydroxymethylation levels, including sites in genes with functions related to synapse formation and regulation, such as CUX2 (p =β2.4βΓβ10β5) and DOK-7 (p =β3.6βΓβ10β3), as well as an enrichment of genes involved in the extracellular matrix (FDRβ=β1.0βΓβ10β8).DiscussionTogether, these preliminary results lend evidence toward a potential role for 5hmC in both early neuronal differentiation and BD risk, with validation and more comprehensive characterization to be achieved through follow-up study.</p