Postmortem Brain Tissue Examination in Neuropsychiatric Disorders

在过去的一年中，我们专注于精神分裂症和情感障碍的几个候选基因，并研究了这些基因的多个转录本的表达及其与精神分裂症风险相关基因型的关联。例如，对于DARPP-32基因，我们研究了两个主要DARPP-32转录本，全长转录本和全长转录本的表达之间的关联。（FL-DARPP-32）和截短（t-DARPP-32），DARPP-32的遗传变体在接受多巴胺能输入的三个大脑区域中，并与精神分裂症有关（背外侧前额叶皮层DLPFC，海马和尾状核）在一组更大的死后样本中，来自精神分裂症，双相情感障碍，抑郁症和正常对照组（>700名受试者）。我们发现t-DARPP-32在精神分裂症和双相情感障碍患者的DLPFC中的表达增加，并且与SNP（rs 879606、rs 90974和rs3764352）的基因型以及先前鉴定的与认知功能相关的7-SNP单倍型强烈相关。预测认知能力较差的遗传变异与较高的t-DARPP-32表达相关。我们的研究结果表明，PPP 1 R1 B的变化影响剪接变体t-DARPP-32 mRNA的丰度，并可能反映精神分裂症和情感障碍的潜在分子机制。 我们还研究了与各种神经发育行为障碍相关的拷贝数变异（CNVs）。 我们分析了1 M SNP基因型阵列（Illumina BeadArrays），以获得先前报道的复发性CNV的证据，并在600个大脑中的DNA中富集了全基因组CNV负荷，其中包括441名患有各种精神病诊断的个体。我们使用Illumina BeadArrays（总共568名受试者）和66-92名受试者使用定量实时PCR研究了选定的CNV病例和其他受试者的背外侧前额叶皮层中的基因表达。 在4/193例诊断为精神分裂症的患者（1q21.1，11 q25，15q11.2，22 q11），4/238例情感障碍患者（11 q25，15q11.2，22 q11）和1/10例自闭症患者（2p16.3）中鉴定出先前报道的基因组区域中的CNV。 在精神分裂症或情绪障碍病例中没有观察到全基因组CNV负荷增加的证据，尽管该研究在观察罕见事件方面的效力不足。 mRNA表达模式提示观察到的CNVs的不完全分子遗传学，特别是在复制中。 我们的数据证实，在脑DNA中，在一小部分精神病诊断患者中存在某些复发性CNVs。 最后，我们探索了人类前额叶皮层（PFC）发育过程中的表观遗传变化，PFC是大脑的主谋，是最后成熟的大脑区域之一。这也是一个涉及精神分裂症和其他主要精神疾病的区域。为了研究表观遗传学在PFC发生中的作用，我们检测了108名受试者从胎儿到老年的27，000个CpG位点的14，500个基因的DNA甲基化，这些基因集中在5个启动子区域。PFC中的DNA甲基化在整个生命中显示出独特的时间模式。最快的变化发生在产前时期，出生后明显放缓，并随着年龄的增长继续进一步放缓。在基因组水平上，从胎儿到出生后的转变是以方向的逆转为代表的，从产前的去甲基化到出生后的甲基化增加。DNA甲基化与基因型变异密切相关，并与一组基因的表达相关，包括参与大脑发育和从头DNA甲基化的基因。我们的研究结果表明，在人类PFC启动子DNA甲基化是一个高度动态的过程中修改的遗传变异和调节基因转录。我们通过使用我们团队创建的独立应用程序BrainCloudMeth，使科学家的其他发现成为可能。 我们已经进行了几次调查，使用死后人脑标本主要集中在了解除了其他复杂的神经精神疾病的SZ的病理生理。除了我们自己的研究之外，该部门还继续向NIH内外的研究人员和实验室提供死后人脑组织。

Over the past year, we focused on several candidate genes for schizophrenia and affective disorders, and studied expression of multiple transcripts of these genes and their associations with schizophrenia risk-associated genotypes. For instance, for a DARPP-32 gene, we examined the association of expression of two major DARPP-32 transcripts, full-length (FL-DARPP-32) and truncated (t-DARPP-32), with genetic variants of DARPP-32 in three brain regions receiving dopaminergic input and implicated in schizophrenia (the dorsolateral prefrontal cortex DLPFC, hippocampus, and caudate) in a much larger set of postmortem samples from patients with schizophrenia, bipolar disorder, major depression and normal controls (>700 subjects). We found that the expression of t-DARPP-32 was increased in the DLPFC of patients with schizophrenia and bipolar disorder and was strongly associated with genotypes at SNPs (rs879606, rs90974 and rs3764352), as well as the previously identified 7-SNP haplotype related to cognitive functioning. The genetic variants that predicted worse cognitive performance were associated with higher t-DARPP-32 expression. Our results suggest that variation in PPP1R1B affects the abundance of the splice variant t-DARPP-32 mRNA and may reflect potential molecular mechanisms implicated in schizophrenia and affective disorders. We also examined copy number variations (CNVs) associated with diverse neurodevelopmental behavioral disorders. We analyzed 1M SNP genotype arrays (Illumina BeadArrays) for evidence of previously reported recurrent CNVs and enriched genome wide CNV burden in DNA from 600 brains, including 441 individuals with various psychiatric diagnoses. We explored gene expression in the dorsolateral prefrontal cortex in selected cases with CNVs and in other subjects using Illumina BeadArrays (568 subjects in total), and additionally in 66-92 subjects using quantitative real-time PCR. CNVs in previously reported genomic regions were identified in 4/193 patients with the diagnosis of schizophrenia (1q21.1, 11q25, 15q11.2, 22q11), 4/238 patients with mood disorders (11q25, 15q11.2, 22q11), and 1/10 patients with autism (2p16.3). No evidence of increased genome wide CNV burden was observed in cases with schizophrenia or mood disorders although the study is underpowered to observe rare events. mRNA expression patterns suggested incomplete molecular penetrance of observed CNVs, particularly in the duplications. Our data confirm in brain DNA the presence of certain recurrent CNVs in a small percentage of patients with psychiatric diagnoses. Finally, we explored epigenetic changes during development of the human prefrontal cortex (PFC), a mastermind of the brain, which is one of the last brain regions to mature. It is also a region implicated in schizophrenia and other major mental disorders. To investigate the role of epigenetics in the development of PFC we examined DNA methylation in 14,500 genes at 27,000 CpG loci focused on 5 promoter regions in 108 subjects ranging from fetal to old age. DNA methylation in the PFC shows unique temporal patterns across life. The fastest changes occur during the prenatal period, slow down markedly after birth and continue to slow further with aging. At the genome level, the transition from fetal to postnatal life is typified by a reversal of direction, from demethylation prenatally to increased methylation postnatally. DNA methylation is strongly associated with genotypic variants and correlates with expression of a subset of genes, including genes involved in brain development and in de novo DNA methylation. Our results indicate that promoter DNA methylation in the human PFC is a highly dynamic process modified by genetic variance and regulating gene transcription. We have made additional discovery by the scientists possible by using a stand-alone application BrainCloudMeth created by our team. We have conducted several investigations using postmortem human brain specimens focused primarily on understanding the pathophysiology of SZ in addition to other complex neuropsychiatric disorders. In addition to our own studies, the Section continues to provide postmortem human brain tissues to researchers and labs within and outside NIH.