Gene Expression Profiling on Insulin Resistance and Obes

为了鉴定可能导致胰岛素抵抗的新基因，我们使用1）含有6800个基因和35000个表达序列标签（EST）的人寡核苷酸微阵列和2）差异显示PCR检测了非糖尿病纯种皮马印第安人的骨骼肌基因表达谱。我们还与Ulf Smith博士（瑞典Sahlovenska大学医院）和Samuel Cushman博士（NIDDK）合作，在选定的胰岛素抵抗和胰岛素敏感瑞典受试者的骨骼肌和脂肪细胞样本中进行了基因表达谱分析。将通过以下方式评价差异表达基因/EST在胰岛素抵抗中的潜在作用：检查1）它们在相关代谢途径中的功能，另外对于Pima样本：2）在Pima人群中进行全基因组扫描时，它们相对于与胰岛素作用和糖尿病相关的峰值的染色体位置。使用实时PCR进一步研究所选基因的骨骼肌表达水平。该项目的手稿已经提交。先前的研究表明原代培养的骨骼肌细胞的生化特性与供体的临床参数之间存在很强的相关性。为了确定胰岛素抵抗与胰岛素敏感受试者的肌肉组织中哪些差异表达的基因可能是原发性的，而不是继发性的异常，我们还建议比较同源培养的成肌细胞和肌管的基因谱。我们已经开始进行类似的实验，以确定肥胖易感基因使用新鲜分离的脂肪细胞和培养的前脂肪细胞从瘦和肥胖的皮马印第安人。使用寡核苷酸微阵列，我们比较了脂肪细胞的表达谱从10瘦与10肥胖皮马印第安人。我们正在分析现有的数据，并为这个项目收集更多的样本。

To identify novel genes that may contribute to insulin resistance, we examined skeletal muscle gene expression profiles in non-diabetic full-blooded Pima Indians using 1) human oligonucleotide microarrays containing 6800 genes and 35000 expressed sequence tags (ESTs), and 2) differential display PCR. We have also performed gene expression profiling in skeletal muscle and adipocyte samples from selected insulin resistant and insulin sensitive Swedish subjects in collaboration with Dr. Ulf Smith (Sahlgrenska University Hospital, Sweden)and Dr. Samuel Cushman (NIDDK). The potential roles of the differentially expressed genes/ESTs in insulin resistance will be evaluated by examining 1) their functions in relevant metabolic pathways, and additionally for Pima samples: 2) their chromosomal locations relative to peaks of linkage to insulin action and diabetes in a genome-wide scan in the Pima population. Skeletal muscle expression levels of selected genes were further investigated using real-time PCR. The manuscript on this project has been submitted. Previous studies have indicated strong correlations between biochemical characteristics of primary cultured skeletal muscle cells and clinical parameters of the donors. To determine which of the differentially expressed genes in muscle tissue of insulin resistant vs. insulin sensitive subjects may be primary, rather than secondary abnormalities, we propose also to compare the gene profiles of the cognate cultured myoblasts and myotubes. We have started to perform similar experiments to identify obesity susceptibility genes using freshly isolated adipocytes and cultured pre-adipocytes from lean and obese Pima Indians. Using oligonucleotide microarrays, we compared the expression profiles of adipocytes from 10 lean vs. 10 obese Pima Indians. We are analyzing the current data and collecting more samples for this project.