INSULIN SIGNALING DEFECTS IN PCOS

多囊卵巢综合征中的胰岛素信号缺陷

• 批准号: 7255640
• 负责人: Ricardo Azziz
• 金额: $ 31.65万
• 依托单位: CEDARS-SINAI MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-05 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7255640
• 关键词: 1-Phosphatidylinositol 3-Kinase; 70-kDa Ribosomal Protein S6 Kinases; A kinase anchoring protein; Abdomen; Adipocytes; Adipose tissue; Adrenal Glands; Affect; Age; Androgens; Basic Science; Binding; Binding Proteins; Biological Markers; Biopsy; Classification; Clinical; Collaborations; Consult; Critical Pathways; Cyclic AMP-Dependent Protein Kinases; Data; Defect; Deoxyglucose; Disease; Economic Burden; Endocrinologist; Etiology; FOXO1A gene; FRAT1 gene; Funding; Future; Glycogen Synthase Kinase 3; Hormonal; Hyperandrogenism; Hyperinsulinism; In Vitro; Insulin; Insulin Receptor; Insulin Resistance; Invasive; Laboratories; Longevity; Longitudinal Studies; MAP Kinase Gene; MAPK14 gene; MAPK8 gene; Mediating; Metabolic; Molecular; Nature; Needles; Ovarian; Pathway interactions; Patients; Phenotype; Phosphoinositide-3-Kinase, Catalytic, Gamma Polypeptide; Phosphorylation; Physiology; Play; Polycystic Ovary Syndrome; Polymerase Chain Reaction; Population; Procedures; Protein Kinase C; Proteins; Proto-Oncogene Proteins c-akt; Race; Receptor Signaling; Recruitment Activity; Regulation; Relative (related person); Research; Research Infrastructure; Research Personnel; Reverse Transcription; Role; Sampling; Scientist; Serine; Signal Transduction; Staging; Steroid biosynthesis; Study Subject; Surgeon; Techniques; Testing; Time; Tissue Sample; Tissues; Transfection; Translations; Tyrosine; Up-Regulation; Western Blotting; Woman; adenoviral-mediated; citrate carrier; concept; experience; glucose transport; glucose uptake; human MAP2K1 protein; in vitro Model; in vivo; inhibitor/antagonist; insulin receptor substrate 1 protein; insulin signaling; intravenous glucose tolerance test; novel; programs; receptor binding; reproductive; research study; response; skills; stress-activated protein kinase 1; subcutaneous; uptake

DESCRIPTION (provided by applicant): The polycystic ovary syndrome (PCOS) affects ~7% of women and ~70% demonstrate insulin resistance, with the resulting hyperinsulinemia stimulating androgen excess. The economic burden of the disorder is estimated to exceed 4 billion dollars annually, in the U.S. and during the reproductive life span alone. The broad long-term objective of our studies is to establish the molecular etiology(s) of the PCOS-associated insulin resistance. Overall, little is know about the molecular aspects of insulin signaling in PCOS. Insulin-stimulated glucose uptake is deficient in PCOS, suggesting an alteration along the IRS/PI-3 kinase/Akt cascade, although the mitogenic activity and MAPK pathway appears unaffected. Insulin receptor (IR) tyrosine autophosphorylation also appears to be lower, and serine phosphorylation higher. In addition, we have obtained preliminary data indicating that PCOS adipocytes have deficient serine (inhibitory) and increased tyrosine (activating) glycogen synthase kinase-3 (GSK3) phosphorylation, consistent with enhanced GSK3 action. This data suggests that GSK3 dysregulation may represent a novel mechanism for insulin resistance in PCOS. We propose the following studies: Aim 1: To determine the role that defective regulation of GSK3 plays in mediating the abnormal IR signaling and glucose transport of PCOS; we will phenotype, including performing a frequently sampled intravenous glucose tolerance test, 70 PCOS patients and 70 matched controls; and in the adipocytes of these subjects determine the association of GSK3 activity with 2-deoxyglucose uptake; the content of regulators and substrates for GSK phosphorylation, determined by RT-PCR and/or Western blot (including total and phosphorylated IR substrate-1 and 2 [IRS-1/2], Akt, the PI-3 kinase subunits p110a and p110p, the 220-kDa A-kinase anchoring protein [AKAP220], protein kinase C [PKC], p70S6K, and the 2 GSK3-binding proteins known as FRAT1 and FRAT2); the impact of specific PKA, PKB (Akt), and PKC inhibition; in PCOS, the impact of specific GSK3 inhibition; and, in controls, the effect of GSKSbeta upregulation, using adenoviral-mediated transfection. Aim 2: To test whether abnormal signaling of the IRS/PI-3 kinase/Akt, but not the MAPK cascade, is present in PCOS; determining the degree of IR binding and 2-deoxyglucose uptake; and by RT-PCR and/or Western blot, the total content and phosphorylation in response to insulin of the IR, total and translocated GLUT-4, and of critical intermediate proteins (e.g. FKHR of the PI-3 kinase/Akt cascade: c-Raf, MEK-1, ERK1/2, pQORSK, and the translational regulator p70S6, of the ERK1/2 cascade: JNK of the SAPK/JNK cascade; and p38 MAPK of the P38MAPK cascade). Overall, these studies have the potential of elucidating the etioloaic mechanism(s) in PCOS, and guiding our search for therapies and molecular markers.

描述（由申请人提供）：多囊卵巢综合征（PCOS）影响约7%的女性，约70%的女性表现出胰岛素抵抗，导致高胰岛素血症刺激雄激素过多。据估计，在美国，仅在生殖寿命期间，这种疾病的经济负担每年就超过40亿美元。我们研究的广泛的长期目标是建立PCOS相关胰岛素抵抗的分子病因学。总体而言，对PCOS中胰岛素信号传导的分子方面知之甚少。PCOS患者胰岛素刺激的葡萄糖摄取不足，提示IRS/PI-3激酶/Akt级联反应沿着改变，尽管促有丝分裂活性和MAPK途径似乎未受影响。胰岛素受体（IR）酪氨酸自磷酸化也似乎较低，丝氨酸磷酸化较高。此外，我们已经获得了初步的数据表明，PCOS脂肪细胞有缺陷的丝氨酸（抑制）和增加的酪氨酸（激活）糖原合成酶激酶-3（GSK 3）磷酸化，与增强GSK 3的行动一致。这些数据表明，GSK 3失调可能是PCOS胰岛素抵抗的一种新机制。目的1：通过对70例PCOS患者和70例正常对照者进行静脉葡萄糖耐量试验，检测脂肪细胞中GSK 3活性与2-脱氧葡萄糖摄取的关系，探讨GSK 3在PCOS患者胰岛素抵抗和葡萄糖转运中的作用。通过RT-PCR和/或Western印迹测定GSK磷酸化的调节剂和底物的含量（包括总的和磷酸化的IR底物-1和2 [IRS-1/2]、Akt、PI-3激酶亚基p110 a和p110 p、220-kDa A-激酶锚定蛋白[AKAP 220]、蛋白激酶C [PKC]、p70 S6 K、和2种称为FRAT 1和FRAT 2的GSK 3结合蛋白）;特异性PKA、PKB（Akt）和PKC抑制的影响;在PCOS中，特异性GSK 3抑制的影响;以及在对照中，使用腺病毒介导的转染，GSKSbeta上调的影响。目标二：检测PCOS中是否存在IRS/PI-3激酶/Akt信号异常，而非MAPK级联;测定IR结合和2-脱氧葡萄糖摄取的程度;并通过RT-PCR和/或Western印迹，测定IR、总GLUT-4和易位GLUT-4以及关键中间蛋白的总含量和对胰岛素应答的磷酸化。（例如PI-3激酶/Akt级联的FKHR：c-Raf、MEK-1、ERK 1/2、pQORSK和ERK 1/2级联的翻译调节因子p70 S6：SAPK/JNK级联的JNK;和P38 MAPK级联的p38 MAPK）。总之，这些研究有可能阐明PCOS的病因机制，并指导我们寻找治疗方法和分子标记物。