Estrogen Regulation of Fetal Microvessel Development During Primate Pregnancy: Impact on Insulin Sensitivity in Offspring

我们最近发现，雌激素（E2）抑制狒狒的后代表现出胰岛素抵抗，葡萄糖耐受不良，第一阶段胰岛素分泌不足，进展为2型糖尿病（T2 DM）的步骤。然而，这种E2调节事件的基础机制尚不清楚。微血管（MV）单元（即小动脉和相关毛细血管[cap]）通过使胰岛素和葡萄糖能够递送至靶组织（特别是骨骼肌（SM）），在胰岛素作用中具有根本性的重要作用。在胎儿发育过程中，胰岛素靶组织内形成了广泛的MV网络，然而， 对胎儿这一至关重要的发育过程的调节是已知的。血管生成是 血管内皮生长因子是胎儿发育期间帽状网络扩张的基础，并且血管内皮生长因子-A（VEGF）是SM中血管生成的主要调节剂。因此，这项“健康和疾病的发育起源”研究的过度高度新颖的概念是子宫内的E2促进胎儿中SM MV的发育，并因此形成广泛的MV网络，该网络对于将胰岛素和葡萄糖递送至后代中SM中的胰岛素作用和葡萄糖稳态至关重要。在目标1中，我们将测试的假设，E2促进SM MV的形态和功能的发展，狒狒胎儿的一个重要步骤，导致后代的胰岛素敏感性。将在胎儿中期（第100天）和晚期（第100天）对SM VEGF表达、帽密度和MV成熟和形态进行定量。（第165-175天;足月= 184天）妊娠和2岁时的后代，3岁和4岁的狒狒未接受治疗，或其中E2的产生/水平已被母体给予芳香酶抑制剂来曲唑抑制，并通过来曲唑加E2恢复。在将后代递送至E2缺乏/补充狒狒的血管激发之前/期间，通过肱动脉血流介导的扩张和帽流（通过对比增强超声/微泡技术定量）评估SM血管功能。目的2将确定E2促进胎儿SM血管生成的机制，如目的1所述。我们将检验E2快速刺激SM VEGF表达、帽状内皮细胞（EC）紧密连接（TJ）/粘附连接（AJ）破坏和帽状EC粘附连接（AJ）破坏的假设。 在妊娠第165天静脉推注E2给来曲唑处理的狒狒胎儿后0-24 h，作为血管生成早期步骤的增殖。拟定研究具有临床意义，因为早产、芳香酶突变、类固醇硫酸酯酶缺乏、雌激素受体无效突变和母体/胎儿暴露于内分泌干扰物（减少胎儿暴露于E2的正常升高或作用）与人类后代胰岛素抵抗/T2 DM的发生率增加相关。确定E2对胎儿MV发育和灵长类后代胰岛素敏感性发作的重要性为人类的治疗应用提供了基础。

We recently showed that offspring delivered to estrogen (E2)-suppressed baboons exhibited insulin resistance, glucose intolerance, and a deficit in first phase insulin secretion, steps that progress to type 2 diabetes mellitus (T2DM). However, the mechanism(s) underpinning this E2 regulated event are unknown. The microvessel (MV) unit (i.e. arterioles and associated capillaries [cap]) has a fundamentally important role in insulin action by enabling insulin and glucose delivery to target tissue, notably skeletal muscle (SM). An extensive MV network forms within insulin target tissues during fetal development, however, little is known about the regulation of this critically important developmental process in the fetus. Angiogenesis is foundational for expansion of the cap network during fetal development and vascular endothelial growth factor-A (VEGF) is a predominant regulator of angiogenesis in SM. Therefore, the over-arching highly novel concept of this “developmental origin of health and disease” study is that E2 in utero promotes SM MV development in the fetus and consequently formation of an extensive MV network critical for the delivery of insulin and glucose to and thus insulin action and glucose homeostasis within SM in the offspring. In Aim 1, we will test the hypothesis that E2 promotes SM MV morphological and functional development in the baboon fetus as an essential step leading to insulin sensitivity in the offspring. SM VEGF expression, cap density and MV maturation and morphology will be quantified in the fetus at mid (day 100) and late (days 165-175; term = 184 days) gestation and in offspring at 2, 3 and 4 years of age delivered to baboons untreated or in which E2 production/levels have been suppressed by maternal administration of the aromatase inhibitor letrozole and restored by letrozole plus E2. SM vascular function will be assessed by brachial artery flow-mediated dilation and by cap flow, as quantified by contrast-enhanced ultrasound/microbubble technology, before/during vasochallenge of offspring delivered to E2-deprived/-replenished baboons. Aim 2 will determine the mechanisms by which E2 acts to promote SM angiogenesis in the fetus as established in Aim 1. We will test the hypothesis that E2 rapidly stimulates SM VEGF expression, cap endothelial cell (EC) tight junction (TJ)/adherens junction (AJ) breakdown, and cap EC proliferation as early steps in angiogenesis on day 165 of gestation 0-24 h after an iv bolus injection of E2 to fetuses of letrozole-treated baboons. The proposed study is clinically significant since preterm birth, aromatase mutation, steroid sulfatase deficiency, estrogen receptor null mutation, and maternal/fetal exposure to endocrine disruptors, which curtail exposure of the fetus to the normal elevation in or action of E2, are associated with increased incidence of insulin resistance/T2DM in human offspring. Establishing the importance of E2 to fetal MV development and onset of insulin sensitivity in primate offspring provides a basis for therapeutic application to the human.