The role of SerpinB2 in insulin resistance and inflammation - 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.
