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无法愈合的伤口并发症。本课题的目的是阐明我们最近发现的内源性促愈合类巨噬细胞类脂质(PLAM)在糖尿病创面愈合损伤中的作用机制。这些绒毛包括神经保护素(NPD)和松脂，绒毛是作用于其生物合成部位的金刚藤类化合物。巨噬细胞(Mf)在创面愈合过程中发挥着重要作用。在我们的初步研究中，我们发现，健康的小鼠MF和创伤后的皮肤会产生NPD1、MAREST-1和其他新的脂质介质。相比之下，在类似的MF和糖尿病小鼠的伤口中，这些PLAM的水平降低。伤口PLAM水平与氧化应激标志物F4-神经前列腺素水平呈负相关，F4-神经前列腺素是DHA的自氧化衍生物。此外，我们还发现，高血糖可减少糖尿病小鼠MF所致的PLAM形成。我们还表明，用这些PLAM治疗糖尿病小鼠的MF，可以挽救糖尿病小鼠伤口的MF促进愈合功能。这些新的发现引出了我们的假设：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可以挽救糖尿病受损的巨噬细胞对伤口和血管生成的促进愈合功能。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、体内和体外的血管形成评估以及包括傅立叶变换红外光谱在内的串联质谱仪(MS)分析来验证我们的假设。具体目标为：目的1.检验创伤愈合过程中PLAM形成的预测及其与MF、糖尿病高血糖和氧化应激的关系。我们将使用基于我们的靶向LC-UV-MS/MS的介体-脂组分析来研究WENTS和MF，以检验预测：1。代表糖尿病氧化应激的F2-异前列腺素和F4-神经前列腺素水平与糖尿病小鼠伤口中PLAM的形成减少呈时间和负相关。1B.模拟高血糖可减少MF所致的PLAM形成。1C。去除MF可显著减少非糖尿病小鼠伤口中的PLAM形成。目的2.测试补充神经保护素、松脂或新型PLAMS可以挽救糖尿病受损的MF功能，促进伤口愈合。我们将用这些药片处理糖尿病小鼠产生的MF，并将其应用于糖尿病小鼠的伤口，以确定其作用。目的3.测试补充PLAMS可以挽救糖尿病受损的MF功能，促进高血糖时伤口愈合所需的血管生成。我们将在高糖条件下用PLAMS治疗糖尿病受损的MF，以确定PLAMS是否能在促进伤口愈合的关键血管生成过程中拯救MF。这项拟议研究的完成将有助于揭示新型内源性类脂类化合物拯救糖尿病受损的巨噬细胞对伤口愈合至关重要的分子和细胞机制，并为开发治疗糖尿病患者伤口的新疗法提供新的见解。 与公共健康相关：拟议的研究将确定补充乳脂素、神经保护素以及新型促愈合类脂类和副类固醇的候选物质拯救糖尿病受损巨噬细胞的机制，从而促进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无法愈合的潜在靶点。

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.