Keratinocyte Integrin Crosstalk During Wound Healing.

项目概要 表皮角质形成细胞通过恢复表皮屏障和分泌细胞因子，对正常伤口愈合至关重要。 旁分泌因子控制着不同的过程，包括伤口血管生成和肌成纤维细胞功能。在 在致病环境下，表皮功能受损会导致长期功能不足（例如糖尿病性溃疡）或过度 旺盛的愈合（例如，肥厚性疤痕）。我们的长期目标是通过以下方式开发治疗范例 可以操纵哪些整合素来调节致病性角质形成细胞的功能。虽然已经很完善 整合素调节增殖、迁移和生长因子信号传导，以及它们在协调伤口中的作用 角质形成细胞的功能仍然是个谜。此外，虽然正常和伤口角质形成细胞表达整合素 91，在体内，根据解释，整合素 91 丢失，混淆了先前研究中的观察结果， 体外。使用基因定义的、病毒转导的角质形成细胞，表达整合素 α3β1 和/或 α9β1 不同的组合，我们在上一个项目期间发现91对 伤口细胞功能和基因表达受 31 控制，包括促进 内皮细胞功能和调节基底膜组装的自分泌信号。利用基因 定义了我们衍生的在表皮中表达 α3β1 和/或 α9β1 不同组合的小鼠，我们 还发现表皮中α9β1的缺失促进了伤口血管生成并增强了层粘连蛋白α2 损伤后在再生表皮基底膜中的处理。根据我们最近发布的 研究和新的基础数据，我们现在假设 91 交叉抑制 31 依赖性 角质形成细胞通过抑制新型 31-FAK-YAP/TAZ 信号轴发挥作用。我们进一步假设 该信号轴控制促进角质形成细胞伤口功能的基因表达程序， 包括内皮细胞和成纤维细胞的旁分泌刺激以及基底细胞的自分泌调节 膜组件。该假设将使用共培养模型的组合在三个目标中进行测试， qPCR 阵列、蛋白质组学、PAC-seq mRNA 分析、细胞生物学和定义的遗传小鼠模型。在 在本项目期结束时，我们将在第一个项目期开发的基础上阐明 角质形成细胞中整合素信号下游的复杂信号网络，控制旁分泌和 正常伤口中的自分泌信号。我们还将确定这些整合素信号传导途径是如何进行的 在血管生成和其他伤口过程持续存在的表皮肿瘤中发生改变。在此过程中，我们将 开发了新型整合素靶向疗法来调节角质形成细胞功能和伤口的基础 结果。

PROJECT SUMMARY Epidermal keratinocytes are vital to normal wound healing by restoring the epidermal barrier and secreting paracrine factors that govern diverse processes including wound angiogenesis and myofibroblast function. In pathogenic settings, impaired epidermal function results in chronically insufficient (e.g., diabetic ulcers) or over- exuberant healing (e.g., hypertrophic scars). Our long-term goal is to develop therapeutic paradigms through which integrins can be manipulated to modulate pathogenic keratinocyte function. While it is well established that integrins regulate proliferation, migration and growth factor signaling, their roles in orchestrating wound keratinocyte functions remain enigmatic. Moreover, while normal and wound keratinocytes express integrin 91, in vivo, upon explanation integrin 91 is lost, confounding observations made in previous studies, in vitro. Using genetically defined, virally transduced keratinocytes that express integrins 31 and/or 91 in different combinations, we discovered in the last project period that 91 exerts a cross-suppressive effect on wound cell function and gene expression that is governed by 31, including paracrine signals that promote endothelial cell function and autocrine signals that regulate basement membrane assembly. Using genetically defined mice that we have derived expressing different combinations of 31 and/or 91 in the epidermis, we also found that deletion of 91 from epidermis promoted wound angiogenesis and enhanced laminin 2 processing in the regenerating, epidermal basement membrane after injury. Based on our recently published studies and new foundation data, we now hypothesize that 91 cross-suppresses 31-dependent keratinocyte functions through inhibition of a novel 31-FAK-YAP/TAZ signaling axis. We further hypothesize that this signaling axis controls a gene expression program that promotes keratinocyte wound functions, including paracrine stimulation of endothelial cells and fibroblasts and autocrine regulation of basement membrane assembly. This hypothesis will be tested in three Aims using a combination of co-culture models, qPCR arrays, proteomics, PAC-seq mRNA analysis, cell biology, and defined genetic mouse models. At the end of this project period, we will have built on the foundation developed in the first project period to elucidate the complex signaling network downstream of integrin signaling in keratinocytes that governs paracrine and autocrine signaling in normal wounds. We will also have determined how these integrin signaling pathways are altered in epidermal tumors in which angiogenesis and other wound processes persist. In doing so, we will have developed the basis for novel integrin targeting therapeutics to modulate keratinocyte function and wound outcome.