The role of SerpinB2 in insulin resistance and inflammation

项目摘要/摘要： 纤溶酶原激活物抑制物-1(PAI-1)和纤溶酶原激活物抑制物-2(PAI-2)是组织中的丝氨酸蛋白酶抑制物 纤溶酶原激活剂和尿激酶。由SerpinB2编码的PAI-2已被证明是关键的 不同疾病的发病机制，包括癌症和线虫感染。然而，它在肥胖中的作用- 相关的胰岛素抵抗尚不清楚。我们的初步实验显示，糖尿病患者 与非糖尿病患者相比，大网膜脂肪组织中SerpinB2+细胞的数量显著减少 个人。此外，我们观察到SerpinB2+细胞的频率与身体呈负相关 患者的体重指数，提示SerpinB2在糖尿病中具有保护作用。始终如一地，SerpinB2缺乏 小鼠表现出糖耐量受损。在内脏脂肪组织中的所有造血细胞中，只有 常驻巨噬细胞表达可检测到的高数量的SerpinB2。这个巨噬细胞亚群 与瘦身对照相比，肥胖人类和小鼠的SerpinB2表达显著降低。此外， 肥胖的人和小鼠的VAT中驻留巨噬细胞的数量减少是由于它们的高密度 细胞凋亡，这与众所周知的SerpinB2的抗凋亡作用是一致的。另外，我们发现， SerpinB2对于增值税居民产生抗炎细胞因子，如IL-4和IL-13是必不可少的 巨噬细胞。这些细胞因子对维持胰岛素敏感性至关重要。基于这些观察，我们 假设肥胖患者SerpinB2表达减少会触发VAT驻留的巨噬细胞凋亡， 增加炎症，促进胰岛素抵抗。我们将在两个具体目标上检验这一假设。瞄准。 1.为了确定SerpinB2在胰岛素抵抗中的作用，我们将使用三种方法：a)SerpinB2-/-小鼠，b) 巨噬细胞特异性SerpinB2缺陷小鼠(LysMcre/+SerpinB2fl/fl)和c)SerpinB2在VAT中的沉默 利用脂类纳米粒中配制的siRNA在野生型小鼠体内的巨噬细胞。此外，我们还将 确定肥胖患者T细胞来源的干扰素-γ是否会降低SerpinB2的表达。我们还将测试SerpinB2是否- 介导的抗炎细胞因子的产生依赖于Kruppel样因子4(KLF4)和有丝分裂原 活化蛋白激酶(MAPK)激活剂ERK1/2。2.从分子水平上探讨黄曲霉毒素的分子机制 SerpinB2对细胞凋亡的抑制作用。我们的初步数据显示SerpinB2缺乏 巨噬细胞含有转谷氨酰胺酶2(TG2)，这是一种已知的半胱氨酸天冬氨酸氨基转移酶(Caspase)调节剂。 此外，我们观察到SerpinB2直接与TG2结合。确定SerpinB2是否具有抗细胞凋亡作用 如果是TG2介导的，我们将在TG2缺陷的巨噬细胞中过表达SerpinB2，并使用特异性TG2抑制剂。

Project Summary/ Abstract: Plasminogen activator inhibitor type 1 (PAI-1) and type 2 (PAI-2) are serine protease inhibitors of tissue plasminogen activator and urokinase. PAI-2, which is encoded by SerpinB2, has been shown to be critical to the pathogenesis of different diseases including cancer and nematode infection. However, its role in obesity- associated insulin resistance is not known. Our preliminary experiments revealed that diabetic patients harbored significantly reduced number of SerpinB2+ cells in omental adipose tissue compared to non-diabetic individuals. Moreover, we observed an inverse correlation between the frequency of SerpinB2+ cells and body mass index in patients, suggesting a protective role of SerpinB2 in diabetes. Consistently, SerpinB2-deficient mice exhibited impaired glucose tolerance. Among all hematopoietic cells in visceral adipose tissue (VAT), only resident macrophages expressed detectable and high amount of SerpinB2. This macrophage subset had significantly reduced SerpinB2 expression in obese humans and mice compared to lean control. Furthermore, VAT of obese humans and mice contained diminished number of resident macrophages due to their high apoptosis, which is in line with the well-known anti-apoptotic role of SerpinB2. Additionally, we found that SerpinB2 is essential for the production of anti-inflammatory cytokines, such as IL-4 and IL-13, by VAT resident macrophages. These cytokines are crucial for maintaining insulin sensitivity. Based on these observations, we hypothesize that reduced SerpinB2 expression in obesity triggers VAT resident macrophage apoptosis, increases inflammation and promotes insulin resistance. We will test this hypothesis in two specific aims. Aim. 1. To determine the role of SerpinB2 in insulin resistance, we will use 3-fold approaches: a) SerpinB2-/- mice, b) macrophage-specific SerpinB2-deficient mice (LysMcre/+ SerpinB2fl/fl) and c) SerpinB2 silencing in VAT macrophages in vivo in wild type mice using siRNA formulated in lipidoid nanoparticles. Furthermore, we will determine if T cell-derived IFN-γ in obesity decreases SerpinB2 expression. We will also test if SerpinB2- mediated production of anti-inflammatory cytokines depends on Kruppel-like factor 4 (Klf4) and mitogen activated protein kinase (MAPK) activators ERK1/2. Aim. 2. We will investigate the molecular mechanisms of the prevention of apoptosis by SerpinB2. Our preliminary data demonstrated that SerpinB2-deficient macrophages contained diminished levels of transglutaminase 2 (TG2), a known modulator of caspase. Additionally, we observed that SerpinB2 directly binds to TG2. To determine if anti-apoptotic effect of SerpinB2 is TG2-mediated, we will overexpress SerpinB2 in TG2-deficient macrophages and use specific TG2 inhibitors.