Neuroprotectins and Maresins for Macrophages in Diabetic Wound Healing

描述（由申请人提供）：许多美国人患有与糖尿病相关的伤口不愈合并发症。本项目的目标是阐明我们最近发现的巨噬细胞内源性促愈合脂质autacoids（PLAM）在糖尿病伤口愈合障碍中的作用机制。这些PLAMs包括神经保护素（NPD）和maresins，PLAMs是作用于其生物合成位点的自泌素。巨噬细胞（Mf）在伤口愈合中起着关键作用。在我们的初步研究中，我们表明，健康的小鼠MF和皮肤受伤后产生NPD 1，maresin-1，和其他新的脂质介质。相比之下，这些PLAMs的水平在糖尿病小鼠的可比Mf和伤口中减少。伤口PLAM水平与氧化应激标志物F4-神经前列腺素（DHA的自氧化衍生物）的水平呈负相关。此外，我们表明，高血糖症减少PLAM形成的MF糖尿病小鼠。我们还表明，用这些PLAMs治疗糖尿病小鼠的Mf拯救了糖尿病小鼠伤口中的Mf促愈合功能。这些新的发现导致我们的以下假设：1）糖尿病并发症减少了正常伤口愈合所需的PLAM的形成; 2）补充这些PLAM挽救了糖尿病受损的巨噬细胞对伤口和血管生成的促愈合功能。 我们将使用小鼠基因敲除II型糖尿病的伤口愈合模型、Mf耗竭、体内和体外血管化评估以及通过串联质谱（MS）（包括傅立叶变换FTMS）分析来测试我们的假设。 具体目标是：目标1。检测创伤愈合过程中PLAM形成的预测及其与Mf、糖尿病、高血糖和氧化应激的关系。我们将使用我们的基于靶向LC-UV-MS/MS的介体脂质组学分析来研究伤口和Mf，以测试以下预测：代表糖尿病氧化应激的F2-异前列烷和F4-神经前列烷的水平与糖尿病小鼠伤口中减少的PLAM形成在时间上呈负相关。1B.模拟的高血糖减少了由Mf形成的PLAM。1C. MF耗竭显著减少非糖尿病小鼠伤口中的PLAM形成。目标二。测试补充神经保护素、maresins或新型PLAMs拯救糖尿病受损的促进伤口愈合的Mf功能。我们将这些PLAMs处理的糖尿病小鼠产生的Mf应用于糖尿病小鼠的伤口，以确定它们的作用。目标3。测试补充PLAMs拯救糖尿病受损的Mf功能，促进高血糖症中伤口愈合所需的血管生成。我们将在高糖条件下使用PLAMs治疗糖尿病受损的Mf，以确定PLAMs是否可以拯救Mf，促进伤口愈合的关键血管生成过程。 这项拟议研究的完成将有助于揭示拯救糖尿病受损的巨噬细胞功能对伤口愈合至关重要的新型内源性脂质autacoids的分子和细胞机制，并为开发治疗糖尿病患者伤口的新型治疗方式提供新的见解。 公共卫生相关性：拟议的研究将确定补充maresins，神经保护素以及新型促愈合脂质autacoids和paracoids的候选物拯救糖尿病受损巨噬细胞的机制，这些巨噬细胞可增强糖尿病受损伤口的愈合。这些机制可能是开发治疗策略以治愈糖尿病受损伤口不愈合的潜在靶点。

DESCRIPTION (provided by applicant): Many Americans suffer from the non-healing wound complications associated with diabetes. The goal of this project is to elucidate the mechanisms of action of endogenous pro-healing lipid autacoids of macrophages (PLAM), which we discovered recently, in diabetic impairments of wound healing. These PLAMs include neuroprotectins (NPDs) and maresins, PLAMs are autacoids acting on the site of their biosynthesis. Macrophages (Mf) play critical roles in wound healing. In our preliminary studies, we show that healthy murine Mf and skin after wounding generate NPD1, maresin-1, and other novel lipid mediators. In contrast, the levels of these PLAMs are diminished in comparable Mf and wounds of diabetic mice. Wound PLAM levels are inversely correlated with the levels of the oxidative stress marker F4-neuroprostane, an autooxidized derivative of DHA. Moreover, we show that hyperglycemia diminishes PLAM formation by Mf of diabetic mice. We also show that treating Mf of diabetic mice with these PLAMs rescue Mf pro-healing functions in wounds of diabetic mice. These novel findings led to our following hypotheses: 1) Diabetic complications diminish the formation of PLAMs required for normal wound healing; 2) Supplementation with these PLAMs rescues the pro-healing function of diabetes-impaired macrophages on wounds and angiogenesis. We will test our hypotheses using a wound healing model of murine gene-knockout type II diabetes, Mf depletion, vascularization assessments in vivo and in vitro, and analysis via tandem mass spectrometry (MS), including Fourier transform FTMS. The Specific Aims are: Aim 1. Test the predictions about PLAM formation and its relationship with Mf and diabetic hyperglycemia and oxidative stress during wound healing. We will study wounds and Mf using our targeted LC-UV-MS/MS based mediator-lipidomic analysis to test the prediction that: 1A. the levels of F2-isoprostanes and F4-neuroprostanes representing diabetic oxidative-stress are temporally and inversely correlated with the diminished PLAM formation in wounds of diabetic mice. 1B. simulated hyperglycemia diminishes PLAM formation by Mf. 1C. Mf depletion significantly reduces the PLAM formation in wounds of non-diabetic mice. Aim 2. Test that supplementation of neuroprotectins, maresins, or novel PLAMs rescues diabetes-impaired Mf functions that promote wound healing. We will apply diabetic mouse-produced Mf treated with these PLAMs to wounds of diabetic mice to establish their actions. Aim 3. Test that supplementation with PLAMs rescues diabetes-impaired Mf functions that promote wound-healing-required angiogenesis in hyperglycemia. We will treat diabetes-impaired Mf with PLAMs under high-glucose conditions to determine if PLAMs can rescue Mf in promoting crucial angiogenic processes of wound healing. Completion of this proposed research will contribute to revealing the molecular and cellular mechanisms for novel endogenous lipid autacoids necessary to rescue diabetes-impaired macrophage functions vital to wound healing, and provide new insight for developing novel therapeutic modalities for treating wounds in diabetics. PUBLIC HEALTH RELEVANCE: The proposed research will define the mechanisms that supplementation of maresins, neuroprotectins, and candidates of novel pro-healing lipid autacoids and paracoids rescues diabetes-impaired macrophages, which enhance the healing in diabetes-impaired wounds. These mechanisms may be potential targets for developing therapeutical strategies to cure the nonhealing of diabetes-impaired wounds.