ENDOPLASMIC RETICULUM STRESS RESPONSE IN HUMAN TYPE 2 DIABETES

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。2型糖尿病（T2DM）是一种常见的遗传性疾病，似乎是由大量常见DNA序列变异引起的。尽管进行了广泛的调查，但人类的早期病理事件仍然不确定。最近的动物数据表明，所有细胞和所有生物体都有一个共同的途径，即内质网应激反应（ERSR）在肥胖的脂肪细胞和肝细胞中被激活，也可能在胰腺细胞中被激活，但没有人类数据可用。我们将结合人体生理学和分子研究与基因组学方法来鉴定广泛转录级联中的序列变异，该序列变异是由未折叠蛋白质的反应启动的，并可能导致蛋白质降解，转录改变和细胞凋亡。我们假设1)与非肥胖、健康、糖耐量个体相比，糖耐量受损的肥胖个体和糖耐量受损的肥胖个体的皮下脂肪组织中ERSR被激活，这些途径被脂肪细胞中的噻唑烷二酮下调，常见的顺式作用DNA序列变异改变了关键ERSR基因的表达，增加了对T2DM的易感性。为了验证这些假设，我们提出了四个具体目标，其中只有第一个目标目前被提议供IRB和GCRC批准。我们将在非洲裔美国人和白种人这两个种族的3组（血糖正常、瘦对照、肥胖、血糖正常和糖耐量受损）中每组测试40人。我们将使用静脉葡萄糖耐量试验和最小模型来表征胰岛素敏感性，并通过穿刺活检获得脂肪和肌肉。我们将通过测量关键ERSR基因的转录和磷酸化来确定ERSR。在接下来的目标中，我们将使用本研究和其他正在进行的研究中收集的组织进行研究，我们将测量吡格列酮治疗前后脂肪和肌肉中最重要基因的转录变化，我们将使用一种新的方法，通过检测脂肪、肌肉和爱stein -bar病毒转化的淋巴细胞来识别ERSR基因的顺式作用序列变异，以寻找mRNA水平失衡的证据。最后，我们将通过寻找额外的编码或调节序列变异来检查每个显示转录或RNA稳定性改变证据的基因，建立这些变异在人群中（单倍型结构）之间的关联，并测试那些识别这些单倍型与T2DM、胰岛素抵抗和胰岛素分泌紊乱相关的变异。目前只提出了Aim 1的研究，并将寻求收集试点数据，这些数据可能用于准备向NIH申请，以提供足够的资金来执行拟议的研究。这项工作将最终确定潜在的遗传变异，这些变异可能提供药物靶点或一种方法，以确定将从旨在改变ERSR基因转录的药物中受益的个体。因此，这项工作有可能对T2DM和代谢综合征的预防和护理产生直接影响。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Type 2 diabetes (T2DM) is a common, inherited disease that appears to result from a large number of common DNA sequence variants. Despite extensive investigation, the early pathological events in humans remain uncertain. Recent animal data suggest that a pathway common to all cells and all organisms, the endoplasmic reticulum stress response (ERSR) is activated in adipocytes and hepatocytes with obesity, and likely in pancreatic $-cells, but no human data are available. We will combine human physiological and molecular studies with a genomics approach to identify sequence variants in the extensive transcription cascade that is initiated with the response to unfolded proteins and may lead to protein degradation, altered transcription, and cellular apoptosis. We hypothesize that 1) ERSR is activated in subcutaneous adipose tissue from both glucose tolerant obese individuals and individuals with impaired glucose tolerance relative to nonobese, healthy, glucose tolerant individuals, that these pathways are downregulated by thiazolidinediones in adipocytes, and that common cis-acting DNA sequence variants alter the expression of key ERSR genes and increase susceptibility to T2DM. To test these hypotheses we propose four Specific Objectives, of which only the first is proposed at present for IRB and GCRC approval. We will test 40 individuals in each of 3 groups (normoglycemic, lean controls; obese, normoglycemic individuals, and individuals with impaired glucose tolerance) in two ethnic groups, African American and Caucasian. We will characterize insulin sensitivity using the intravenous glucose tolerance test and Minimal Model, and will obtain adipose and muscle by needle biopsy. W e will determine ERSR by measuring transcription and phosphorylation of key ERSR genes. In subsequent aims, to be performed using tissues gathered in this an other ongoing studies, we will measure the change in transcription of the most significant genes in adipose and muscle before and after treatment with pioglitazone, and we will use a novel approach to identify cis-acting sequence variants in ERSR genes by testing adipose, muscle, and epstein-bar virus transformed lymphocytes for evidence of an imbalance in mRNA levels. Finally, we will examine each gene which shows evidence for altered transcription or RNA stability by searching for additional coding or regulatory sequence variants, establishing the associations between these variants in a population (haplotype structure), and testing those variants that identify these haplotypes for an association with T2DM, insulin resistance, and disordered insulin secretion. Only studies of Aim 1 are proposed at the present time, and will seek to collect pilot data that may be used to prepare an application to NIH to provide adequate funding to perform the proposed studies. This work will eventually identify potential genetic variants that might provide drug targets or a means to identify individuals who would benefit from drugs aimed at altering ERSR gene transcription. This work thus has the potential to have a direct impact on the prevention and care of T2DM and metabolic syndrome.