Kindlin-2调节整合素αⅤβ3活性对皮肤创伤愈合的作用和机制研究

The cellular and molecular mechanisms underpinning tissue repair and its failure to heal are still poorly understood, and current therapies are limited. Poor wound healing after trauma, surgery, acute illness, or chronic disease conditions affects millions of people worldwide each year and is the consequence of poorly regulated elements of the healthy tissue repair response, including inflammation, angiogenesis, matrix deposition, and cell recruitment. The integrin family of cell adhesion receptors regulates a diverse array of cellular functions crucial to cutaneous wound healing. Kindlins are a group of proteins that have recently attracted attention for their ability to bind and activate integrins. Kindlin-2 and αVβ3 integrin have also been linked to angiogenesis and cell recruitment. Here, wound healing will be analyze in fermt2 (Kindlin-2)-deficient mouse models. To delineate mechanism by which αVβ3 integrin and kindlin-2 regulate angiogenesis and cell recruitment, we will characterize VEGF-mediated signaling, integrin activation, and integrin-dependent functions in endothelial cells isolated from aortas of WT and kindlin-2 +/- mice. We will assay proliferation, adhesion and migration of the vascular smooth muscle cells isolated from WT and kindlin-2 -/fl mice. We will assay functions of αVβ3 integrin and kindlin-2 on fibroblasts, include its role in proliferation, adhesion and migration. Findings from this research in skin have contributed to the unraveling of novel fundamental principles in regenerative biology, which have relevance to function of other epithelial-mesenchymal tissues, such as intestine, lung, and liver.

创面异常愈合给患者带来极大的痛苦，给国家和个人带来巨大的经济负担。整合素通过介导黏附作用参与创面愈合整个过程，Kindlin结合整合素使其由中间活化状态变为完全活化状态。其中，整合素αⅤβ3和Kindlin-2参与调节血管新生、细胞募集。本课题将采用fermt2 (Kindlin-2)基因敲除小鼠，建立整合素活化异常模型，采用小鼠皮肤创伤修复模型，观察创面愈合；检测VEGFR通路活性、整合素活化及表达调节的内皮细胞血管生成特性；采用条件性敲除血管平滑肌细胞fermt2基因小鼠，观察平滑肌细胞增殖、黏附和迁移作用；检测TGF-β、整合素αⅤβ3活化及表达对成纤维细胞增殖、黏附和迁移的作用，研究创伤愈合中血管新生、成纤维细胞募集机制。本课题对揭示整合素介导黏附作用参与调节创面愈合的机制，对选择性干预创面异常愈合具有重要性，也对揭示再生生物学基本规律，理解创伤修复、组织再生机制具有重要意义。

创面愈合是指机体遭受外力作用，组织出现离断或缺损后的愈复过程，为包括炎症反应、血管新生、细胞募集和细胞外基质沉积的复杂组合，表现出各种过程的协同作用。当组织修复过程中的某一或某些环节受阻或发生紊乱时，受损伤的皮肤会出现创伤异常愈合现象，表现为创面不愈或瘢痕过度增生。创面异常愈合给患者带来极大的痛苦，给国家和个人带来巨大的经济负担，这已成为创伤外科领域需要解决的重要问题。. 本课题进行了以下研究：（1）采用野生型小鼠皮内注射AAV Mig-2（Kindlin-2） RNAi和kindlin-2+/-基因敲除小鼠，结果显示减少Kindlin-2表达延缓创面愈合；（2）创面愈合过程中，Kindlin-2调节新生血管成熟度；Kindlin-2通过增加血管内皮细胞形成紧密连接促进新生血管成熟；Kindlin-2调节血管内皮细胞Wnt、Notch、ERK和TGF-β等通路活性；（3）创面愈合过程中，Kindlin-2通过调节血管平滑肌细胞增殖、迁移促进新生血管成熟；Kindlin-2调节血管平滑肌细胞c-myc、SMAD3、SMAD4及snail等表达；（4） kindlin-2基因敲除减少创面ECM沉积；巨噬细胞是创面愈合过程中激活成纤维细胞、促进纤维性修复的主要细胞；细菌感染促进巨噬细胞浸润介导的纤维组织增生；（5）脂肪组织对创伤修复具有重要作用；创面愈合过程中，下调Kindlin-2抑制创面脂肪生成以及脂肪前体细胞成脂分化能力；Kindlin-2通过PI3K/AKT/mTOR信号通路调控3T3-L1脂肪前体细胞成脂分化。揭示了创面血管新生、纤维性修复过程的基本规律。. 在项目执行的4年期间，本课题组在国内、外知名学术期刊发表研究论文，培养硕士、博士研究生。此外，项目负责人亓发芝教授获得了重要奖励。