Epigenetic Regulation of Prostaglandin E2 (PGE2) Synthesis Alters Macrophage Function to Promote Inflammation and Impair Diabetic Wound Healing

项目摘要/摘要 2型糖尿病(T2D)患者的不可愈合伤口是发病率和死亡率的主要原因 正以惊人的速度增长。T2D患者的伤口愈合失败是最常见的 截肢原因在美国，5年死亡率接近50%。因此，迫切需要 了解T2D的伤口愈合缺陷，以便开发有针对性的治疗方法。我们已经利用了这两个 遗传性(db/db)和饮食(饮食诱导的肥胖)小鼠T2D模型以及人类伤口组织和 采集T2D患者的血液样本，以探讨创面愈合受损的机制。我们出版的 初步数据表明，巨噬细胞(M-φ)在协调适当的创伤中起着关键作用 愈合并证明创伤MφS在糖尿病小鼠和T2D患者中的特征是 持续的炎症状态、吞噬/杀伤功能受损和免疫调节剂的过度产生 脂类，前列腺素E2(PGE2)。我们的数据表明，关键基因的表观遗传调控对 产生PGE2，即胞浆磷脂酶A2(CPLA2)和环氧合酶-2(COX-2)，并显示 前列腺素E_2在M-φS细胞中的过表达导致白细胞介素2等炎症介质的产生增加 1白介素1(β，IL1β)和宿主对经常定植在伤口床上的细菌病原体的防御能力减弱。我们的 初步数据首次证实糖尿病创面MφS和前列腺素E_2水平升高。 这一途径可能受多种表观遗传机制的调节，包括DNA甲基化和组蛋白 在饮食诱导和遗传糖尿病模型中都存在甲基化。这些结果支持我们的假设 抑制环氧合酶-2/前列腺素E_2通路在M-φS消退炎症和正确的宿主防御中起关键作用 这是有效修复伤口所必需的。这些结果导致了我们的假设COX-2/PGE2 糖尿病创面M-φ通路受表观遗传调控而增加S 炎症、宿主防御功能受损和伤口修复缺陷。我们的数据表明，WIND Mφ函数 可通过FDA批准的COX抑制剂(S)、转化生长因子β信号受体的M-TRAN靶向治疗而恢复 和/或第一个开发的EP2特异性拮抗剂。为了检验我们的假设，我们将：目标1：确定 调控cPLA2在糖尿病创面Mφ释放AA和促进COX-2/PGE_2的产生。目的2： 确定转化生长因子β诱导的miR-29b是否导致COX-2基因低甲基化以增加COX-2 和前列腺素E_2在糖尿病创面M-φ中的表达及评价靶向COX-2的M-φ的治疗效果 转化生长因子β受体拮抗剂。目的3：确定M-φ特异性前列腺素E_2介导的机制(S) 调节正常和糖尿病伤口的炎症、宿主防御功能和成纤维细胞的串扰 组织。

PROJECT SUMMARY/ABSTRACT Non-healing wounds in patients with Type 2 Diabetes (T2D) are a major cause of morbidity and mortality and are increasing at an alarming rate. Failure of wound healing in T2D patients represents the most common cause of amputation in the US with a 5-year mortality rate of nearly 50%. Thus, a critical need exists for understanding the wound healing defects in T2D in order to develop targeted therapies. We have utilized both genetic (db/db) and dietary (diet-induced obese) murine models of T2D as well as human wound tissue and blood samples collected from T2D patients to explore mechanisms of impaired wound healing. Our published and preliminary data point to a pivotal role for macrophage (Mφ) function in orchestrating appropriate wound healing and demonstrate that wound Mφs in diabetic mice and patients with T2D are characterized by a persistent inflammatory state, impaired phagocytosis/killing and the over-production of the immunomodulatory lipid, prostaglandin E2 (PGE2). Our data demonstrate epigenetic regulation of key genes important for the production of PGE2, namely cytosolic phospholipase A2 (cPLA2) and cyclooxygenase-2 (COX-2), and show overexpression of PGE2 in Mφs results in increased production of inflammatory mediators such as interleukin 1β (IL1β) and impaired host defense against bacterial pathogens that often colonize the wound bed. Our preliminary data are the first to identify that cPLA2/COX-2 and PGE2 are increased in diabetic wound Mφs and that this pathway may be regulated by multiple epigenetic mechanisms, including DNA methylation and histone methylation, in both diet-induced and genetic models of diabetes. These results support our hypothesis that inhibition of the COX-2/PGE2 pathway in Mφs is critical for resolution of inflammation and proper host defense that is required for effective wound repair. These results have led to our hypothesis that the COX-2/PGE2 pathway is epigenetically regulated and increased in diabetic wound Mφs and this results in increased inflammation, impaired host defense and defective wound repair. Our data suggest that wound Mφ function may be restored via Mφ-targeted treatment of FDA-approved COX inhibitor(s), TGFβ signaling receptors and/or the first-ever developed EP2-specific antagonist. To test our hypotheses, we will: Aim 1: Determine the regulation of cPLA2 to release AA and promote COX-2/PGE2 production in diabetic wound Mφs. Aim 2: Determine whether TGFβ-induced miR-29b causes hypomethylation of the COX-2 gene to increase COX-2 and PGE2 production in diabetic wound Mφ and evaluate the therapeutic efficacy of Mφ targeted COX-2 inhibition and TGFβ receptor antagonists. Aim 3: Determine the Mφ-specific PGE2-mediated mechanism(s) that modulate inflammation, host-defense functions and fibroblast crosstalk in normal and diabetic wound tissue.