The Role of Macrophage Migration Inhibitory Factor (MIF) Family Proteins in Wound Repair and Obesity

肥胖是一种与慢性脂肪组织炎症(CAI)相关的疾病，是包括2型糖尿病(T2 DM)在内的多种疾病的危险因素。重要的是，肥胖患者经常表现出伤口修复延迟和伤口愈合障碍风险增加，这仍然是一个悬而未决的问题，特别是在整形和重建手术中。大量证据表明，趋化素样炎性细胞因子巨噬细胞移动抑制因子(MIF)促进肥胖、T2 DM、CAI，同时抑制创面修复。相比之下，人们对最近在这方面发现的MIF同系物D-DOPAChrome互换擦除酶(DDT)知之甚少。申请者的初步工作提供了证据，证明对急性伤口愈合障碍患者的脂肪组织中的MIF和DDT进行了相互调节。在小鼠注射脂多糖的模型中，观察到炎症脂肪组织中MIF和DDT的相互调节，脂肪细胞被认为是MIF和DDT差异表达的细胞来源。有趣的是，MIF和DDT的上调和下调分别一致，被认为对体外伤口修复有不利影响。此外，只有MIF通过其受体CXCR2和CXCR4调节单核细胞迁移到脂多糖注射的脂肪组织中，而DDT似乎没有显示出促进迁移的作用。本研究旨在通过应用小鼠体内模型、患者组织标本和力学实验，综合研究MIF和DDT在肥胖条件下的CAI、胰岛素抵抗和创面修复中的作用。首先，野生型小鼠和全球DDT基因敲除小鼠将接受高脂饮食(HFD)，以调查迄今未知的DDT对肥胖、胰岛素抵抗和脑梗塞的影响。接下来，喂食HFD的具有脂肪组织特异性MIF和DDT(或对照中的野生型)基因敲除的小鼠将接受切开和切除伤，以表征肥胖条件下脂肪组织来源的MIF和DDT在伤口修复中的确切作用。我们预测，在我们的初步实验中观察到的MIF和DDT的相反作用也将适用于我们的体内模型。为了建立潜在的治疗策略，还将在体内模型中研究给予重组MIF/DDT蛋白和/或MIF/DDT中和抗体的可行性。在第三部分，体内实验研究将通过评估肥胖者和瘦肉者的原始人类脂肪组织样本来补充。为此，将使用脂肪组织样本和人成纤维细胞/角质形成细胞的体外共培养试验来研究依赖MIF和DDT的伤口愈合过程。最后，为了确定参与MIF和DDT相互作用的受体，将对CD74、CD44、CXCR2和/或CXCR4的作用以及相关的信号通路进行机制研究。

Obesity is a condition associated with chronic adipose tissue inflammation (CAI) and represents a risk factor for several diseases including type 2 diabetes mellitus (T2DM). Importantly, obese patients often show delayed wound repair and an increased risk of wound healing disorders which remains an unsolved issue, particularly in Plastic and Reconstructive Surgery. A strong body of evidence suggests that the chemokine-like inflammatory cytokine macrophage migration inhibitory factor (MIF) promotes obesity, T2DM, CAI, while inhibiting wound repair. In contrast, little is known about MIFs recently discovered homolog D-dopachrome tautomerase (DDT) in this context. Preliminary work by the applicants leading up to this proposal has provided evidence for a reciprocal regulation of MIF and DDT in adipose tissue of patients with acute wound healing disorders. Reciprocal MIF and DDT regulation in inflamed adipose tissue was observed in a murine LPS injection model and adipocytes were identified as the cellular source for differential MIF and DDT expression. Interestingly, the coincident up-regulation versus down-regulation of MIF and DDT, respectively, was seen to have a detrimental effect on wound repair in vitro. Furthermore, Only MIF regulated monocyte migration into LPS-injected adipose tissue via its receptors CXCR2 and CXCR4 while DDT did not appear to exhibit pro-migratory effects.Here we aim to comprehensively study the role of MIF and DDT in the context of CAI, insulin resistance and wound repair under obesity, applying murine in vivo models, patient tissue specimens, and mechanistic experiments. First, wildtype mice and mice with a global Ddt knockout will undergo a high fat diet (HFD) to investigate the hitherto unknown effect of DDT on obesity, insulin resistance and CAI. Next, mice with an adipose tissue-specific knockout of Mif and Ddt (or wildtype in the control) that are fed a HFD will undergo incisional and excisional wounding to characterize the precise role of adipose tissue-derived MIF and DDT on wound repair under obese conditions. We predict that the inverse role of MIF and DDT that was observed in our preliminary experiments also will apply to our in vivo models. To establish potential therapeutic strategies, the feasibility of administering recombinant MIF/DDT protein and/or MIF/DDT neutralizing antibodies will additionally be studied in the in vivo models. In a third part, the experimental in vivo studies will be complemented by evaluation of primary human adipose tissue samples of obese and lean individuals. To this end, MIF- and DDT-dependent wound healing processes will be examined using in vitro co-culture assays of the adipose tissue samples and human fibroblasts/keratinocytes. Lastly, to identify the receptors contributing to MIF and DDT interactions, mechanistic studies will be performed to address the role of CD74, CD44, CXCR2, and/or CXCR4 as well as the relevant signaling pathways.