Regulation of wound healing pathways by NLRP10 in cutaneous Leishmaniasis

项目总结 皮肤利什曼病会导致皮肤溃疡，需要几个月到几年的时间才能消除，从而导致 严重的发病率。帮助这些损伤愈合的药物干预将减少发病率，但 调节利什曼病损修复的机制还不是很清楚。此外，还有 关于天然免疫核苷酸结合区和富含亮氨酸重复序列的作用研究很少 受体(NLR)家族成员在调节伤口愈合中的作用。使用皮肤利什曼病的小鼠模型， 我们发现了NLR家族成员NLRP10在感染伤口愈合中的新角色，就像缺乏 与野生型小鼠相比，NLRP10会发展成更大的无法愈合的损伤。虽然大多数对NLR家族的研究 在这个模型中，模式识别受体已经确定了这些受体在造血细胞中的作用。 NLRP10调节非造血细胞群体的伤口愈合。重要的是，虽然损伤在 NLPRP10缺陷小鼠比野生型小鼠大，这不是由于未能控制 寄生虫，因为两组之间的寄生虫负担没有差异。相比之下，无菌伤口的愈合 在NLRP10缺陷小鼠中是完整的，这表明对NLRP10的需求是特定于 伤口里有致病微生物。在这项提案中，我们的目标是确定NLRP10 调节皮肤利什曼病的伤口修复。我们发现NLRP10缺陷的成纤维细胞有 钝化的扩散。在评估失败的原因时，我们发现这些成纤维细胞阻止了它们的 在细胞周期的G0/G1期增殖。我们还发现NLRP10缺陷的成纤维细胞 衰老标志物增加，β-半乳糖苷酶染色增加，p16和p21增加 表情。NLRP10缺失的成纤维细胞也表现出衰老的形态特征， 促炎症细胞因子和趋化因子的分泌，与这些细胞衰老相一致- 相关分泌表型(SASP)。我们认为NLRP10在皮肤成纤维细胞中起作用。 利什曼病损害以减轻衰老，并在这样做的帮助伤口修复反应。利用 P16-3MR小鼠模型，这将使我们能够识别并选择性地杀死衰老细胞，我们将能够 明确衰老在皮肤利什曼病伤口修复中的作用，并确定NLRP10如何 调节这一过程。我们将在L.主要病变中确定推动成纤维细胞衰老的因素 NLRP10缺失。我们将进一步定义NLRP10对SASP的影响并确定影响因素 用于修复皮肤利什曼病伤口。我们的研究将确定NLRP10在调节 衰老途径和寻找新的治疗靶点以促进多种皮肤创面愈合 病理性的东西。

PROJECT SUMMARY Cutaneous leishmaniasis results in skin ulcerations that can take months to years to resolve resulting in significant morbidity. Pharmacologic interventions to aid in healing of these lesions would lessen morbidity, but the mechanisms that regulate the repair of Leishmania lesions are not well understood. In addition, there has been little investigation into the role of innate immune Nucleotide-Binding Domain and Leucine Rich Repeat Receptor (NLR) family members in regulating wound healing. Using a mouse model of cutaneous leishmaniasis, we have found a novel role for the NLR family member NLRP10 in healing of infected wounds, as mice that lack NLRP10 develop larger lesions that fail to heal compared to wild-type mice. While most studies of the NLR family of pattern recognitions receptors have identified roles for these receptors in hematopoietic cells, in this model NLRP10 regulates wound healing from a non-hematopoietic cell population. Importantly, while the lesions in NLPRP10-deficient mice were larger than those of wild-type mice, this was not due to a failure to control the parasite, as there was no difference in parasite burden between the groups. In contrast, healing of sterile wounds was intact in NLRP10-deficient mice, suggesting the requirement for NLRP10 is specific to the presence of pathogenic microorganisms in the wound. In this proposal, we aim to identify the mechanism by which NLRP10 regulates wound repair in cutaneous leishmaniasis. We have found that NLRP10-deficient fibroblasts have blunted proliferation. While evaluating the reason for this failure, we identified that these fibroblasts arrested their proliferation at the G0/G1 phase of the cell cycle. We also found NLRP10-deficient fibroblasts displayed increased markers of senescence, with elevated β-galactosidase staining and increased p16 and p21 expression. NLRP10-deficient fibroblasts also showed morphologic features of senescence and had increased secretion of pro-inflammatory cytokines and chemokines, consistent with these cells having a senescence- associated secretory phenotype (SASP). We propose that NLRP10 acts within fibroblasts in cutaneous leishmaniasis lesions to mitigate senescence and in doing so aids in the wound repair response. Utilizing the p16-3MR mouse model, which will allow us to both identify and selectively kill senescent cells, we will be able to define the contribution of senescence to wound repair in cutaneous leishmaniasis and determine how NLRP10 regulates this process. We will determine the factors in L. major lesions that drive fibroblast senescence in the absence of NLRP10. We will further define the influence of NLRP10 on the SASP and determine the factor required for repair of cutaneous leishmaniasis wounds. Our studies will define the role of NLRP10 in regulating senescence pathways and identify novel therapeutic targets to accelerate cutaneous wound healing in a variety of pathologies.