Epigenetic regulation of keratinocyte function in normal and pathologic skin repair

项目概要/摘要 2型糖尿病（T2 D）患者的不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T2 D患者伤口愈合失败是最常见的原因 在美国，截肢手术的5年死亡率接近50%。因此，迫切需要了解 T2 D的伤口愈合缺陷，以开发靶向治疗。我们将利用遗传（db/db）和 T2 D的饮食（饮食诱导的肥胖）鼠模型以及收集的人伤口组织和血液样品 从2型糖尿病患者中探索受损伤口愈合的机制。我们目前的数据使用人类单细胞 RNA测序和小鼠糖尿病伤口模型，抑制性组蛋白甲基转移酶，SETDB 2， 在糖尿病伤口角质形成细胞中减少，导致炎性基因的产生增加，即 TNFα。此外，我们已经确定，干扰素-β（IFN γ），通过janus激酶（JAK 1）/信号转导， 转录激活因子（STAT）机制诱导伤口角质形成细胞中的SETDB 2。我们的初步数据还 发现IL-17 A在糖尿病伤口中增加，并且除了糖尿病伤口中IFN γ水平降低外， 伤口，抑制糖尿病伤口角质形成细胞中的SETDB 2。这些结果导致我们的假设， 在伤口角质形成细胞中诱导SETDB 2抑制NF κ B介导的基因的表达（即，TNFα）， 维持伤口炎症并直接损害角质形成细胞迁移，从而促进组织修复。我们 假设在糖尿病伤口中，未能诱导SETDB 2并抑制NFκ B介导的炎症反应， (Tnfα）角质形成细胞中阻止炎症消退并损害角质形成细胞迁移的基因， 导致伤口愈合不良。我们将通过三个具体的目标来测试我们的假设：目标1： 正常和糖尿病伤口角质形成细胞中SETDB 2对NFκ B介导的基因表达的体内调节。目的 2：确定调节角质形成细胞特异性SETDB 2表达的JAK/STAT介导的机制 在正常和糖尿病伤口组织中。目的3：检查SETDB 2调节的治疗功效和时机。 TNFα对角质形成细胞迁移和炎性伤口Mφ表型的抑制。在这种平移方法中， 我们的数据将为开发有前景的治疗药物铺平道路， 介导糖尿病伤口角质形成细胞功能从而促进糖尿病伤口的表观遗传途径 修复.

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 will utilize 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. We present data using human single cell RNA sequencing and murine diabetic wound models, that the repressive histone methyltransferase, SETDB2, is decreased in diabetic wound keratinocytes resulting in increased production of inflammatory genes, namely TNFα. Further, we have identified that interferon-beta (IFNꞵ), via a janus kinase (JAK1) / signal transducer and activator of transcription (STAT) mechanism induces SETDB2 in wound keratinocytes. Our preliminary data also identified that IL-17A is increased in diabetic wounds and may, in addition to reduced levels of IFNꞵ in diabetic wounds, suppress SETDB2 in diabetic wound keratinocytes. These results have led to our hypothesis that induction of SETDB2 in wound keratinocytes represses expression of NFkB-mediated genes (i.e., TNFα) that maintain wound inflammation and directly impair keratinocyte migration, thereby promoting tissue repair. We postulate that in diabetic wounds, it is the failure to induce SETDB2 and repress NFκB-mediated inflammatory (Tnfα) genes in keratinocytes that prevents resolution of inflammation and impairs keratinocyte migration and results in poor wound healing. We will test our hypotheses through three specific aims: Aim 1: To define the in vivo regulation of NFκB-mediated gene expression by SETDB2 in normal and diabetic wound keratinocytes. Aim 2: To determine the JAK/STAT-mediated mechanism(s) that regulate keratinocyte-specific SETDB2 expression in normal and diabetic wound tissue. Aim 3: To examine the therapeutic efficacy and timing of SETDB2-regulated TNFα inhibition on keratinocyte migration and inflammatory wound Mφ phenotype. In this translational approach, our data will pave the way for the development of promising therapeutic agents aimed at the targeting of epigenetic pathways that mediate diabetic wound keratinocyte function and thereby promote diabetic wound repair.