Saad M J, Maeda L, Brenelli S L, Carvalho C R, Paiva R S, Velloso L A
Department of Internal Medicine, FCM-UNICAMP, Campinas, Sao Paulo, Brazil.
Diabetologia. 1997 Feb;40(2):179-86. doi: 10.1007/s001250050660.
Pregnancy is known to induce resistance, but the exact molecular mechanism involved is unknown. In the present study, we have examined the levels and phosphorylation state of the insulin receptor and of insulin receptor substrate 1 (IRS-1), as well as the association between IRS-1 and phosphatidylinositol 3-kinase (PI 3-kinase) in the liver and muscle of pregnant rats (day 20 of gestation) by immunoprecipitation and immunoblotting with anti-insulin receptor, anti-IRS-1, anti-PI 3-kinase and anti-phosphotyrosine antibodies. There were no changes in the insulin receptor concentration in the liver and muscle of pregnant rats. However, insulin stimulation of receptor autophosphorylation, as determined by immunoblotting with antiphosphotyrosine antibody, was reduced by 30 +/- 6% (p < 0.02) in muscle and 36 +/- 5% (p < 0.01) in liver at day 20 of gestation. IRS-1 protein levels decreased by 45 +/- 6% (p < 0.002) in liver and by 56 +/- 9% (p < 0.002) in muscle of pregnant rats. In samples previously immunoprecipitated with anti-IRS-1 antibody and blotted with antiphosphotyrosine antibody, the insulin-stimulated IRS-1 phosphorylation levels in the muscle and liver of pregnant rats decreased by 70 +/- 9% (p < 0.01) and 75 +/- 8% (p < 0.01), respectively. The insulin-stimulated IRS-1 association with PI 3-kinase decreased by 81 +/- 6% in muscle (p < 0.01) and 79 +/- 11% (p < 0.01) in the liver during pregnancy. These data suggest that changes in the early steps of insulin signal transduction may have a role in the insulin resistance observed in pregnancy.
已知怀孕会诱导胰岛素抵抗，但其中确切的分子机制尚不清楚。在本研究中，我们通过用抗胰岛素受体、抗胰岛素受体底物1（IRS-1）、抗磷脂酰肌醇3激酶（PI 3激酶）和抗磷酸酪氨酸抗体进行免疫沉淀和免疫印迹，检测了妊娠大鼠（妊娠第20天）肝脏和肌肉中胰岛素受体及IRS-1的水平和磷酸化状态，以及IRS-1与PI 3激酶之间的关联。妊娠大鼠肝脏和肌肉中的胰岛素受体浓度没有变化。然而，用抗磷酸酪氨酸抗体免疫印迹测定，在妊娠第20天时，肌肉中胰岛素刺激的受体自身磷酸化降低了30±6%（p<0.02），肝脏中降低了36±5%（p<0.01）。妊娠大鼠肝脏中IRS-1蛋白水平降低了45±6%（p<0.002），肌肉中降低了56±9%（p<0.002）。在用抗IRS-1抗体预先免疫沉淀并用抗磷酸酪氨酸抗体印迹的样品中，妊娠大鼠肌肉和肝脏中胰岛素刺激的IRS-1磷酸化水平分别降低了70±9%（p<0.01）和75±8%（p<0.01）。怀孕期间，胰岛素刺激的IRS-1与PI 3激酶的结合在肌肉中降低了81±6%（p<0.01），在肝脏中降低了79±11%（p<0.01）。这些数据表明，胰岛素信号转导早期步骤的变化可能在妊娠期间观察到的胰岛素抵抗中起作用。
