Signal Transduction of Human Keratinocyte Migration

人类角质形成细胞(HK)的运动在人类皮肤创伤的再上皮化过程中起着重要作用。细胞外基质(ECM)和血清生长因子(GFS)是控制HK迁移的两个主要刺激因素。ECM和GFS与它们的同源受体结合，激活不同但重叠的信号网络。ECM和GFS在控制香港移民方面的具体作用仍不清楚，我们的初步研究表明，启动移民的是ECM，而不是GFS。然而，ECM诱导的迁移是部分和非定向的，而GFS的作用似乎是指导和优化迁移。我们的假设是，HK在结缔组织上的迁移是由ECM发起的，ECM与GFS协同工作，以优化流动性并提供方向性。在这里，我们建议研究细胞外基质启动的随机运动和生长因子优化的定向运动的信号机制。这些研究的重点将集中在没有GFS(随机迁移)的胶原基质和添加GFS(定向和最佳迁移)的胶原基质的信号转导。具体地说，我们将：1)在存在或不存在GFS的情况下，通过胶原基质研究Rho家族GTP酶在香港迁移中的作用。不同的Rho家族GTPase成员在肌动蛋白细胞骨架和细胞迁移的调节中具有不同的功能。它们在香港运动中的作用尚不清楚。将使用药理学和遗传学的方法来研究HKS中的这些GTP酶；2)研究p38-MAPK和PKC-Delta如何调控随机迁移和定向迁移。我们最近的发现表明，p38-MAPK和PKC-Delta是HK在胶原基质上运动所独立需要的(在印刷中)。在这里，我们将进一步研究这两条通路在HKS整合素和生长因子受体信号转导中的具体功能；3)通过一种新的“转化生长因子-β阻断”方法识别和表征HK迁移相关基因。我们将利用转化生长因子-β阻止HKS增殖而不是迁移的事实。来自经转化生长因子-β处理的1)非迁移性HKS、2)随机迁移性HKS和3)定向迁移性HKS的CRNAs将接受DNA微阵列分析。微阵列确定的基因将进一步接受“途径筛选”方法，以缩小rac1-p38-MAPK途径和PKC-Delta途径诱导的基因范围。这些研究将为伤口再上皮化的分子机制提供新的线索。

Human keratinocyte (HK) motility plays an important role in the re-epithelialization of human skin wounds. Extracellular matrices (ECMs) and serum growth factors (GFs) are the two main stimuli that control HK migration. ECMs and GFs bind to their cognate receptors and activate distinct, yet overlapping, signaling networks. The SPECIFIC function of ECMs versus GFs in the control of HK migration remains unclear, Our preliminary studies indicate that ECMs, but not GFs, initiate migration. The ECM-induced migration, however, is partial and non-directional, while GFs' role appears to direct and optimize migration. Our hypothesis is that HK migration on connective tissue is initiated by ECMs, which work in concert with GFs to optimize the molility and provide directionality. Here, we propose to study the signaling mechanisms of ECM-initiated random motility and growth factor-optimized directional motility. The focus of these studies will be on the signal transduction by a collagen matrix without GFs (random migration) and by a collagen matrix plus GFs (directional and optimal migration). Specifically, we will: 1) study the role of the Rho family GTPases in HK migration by a collagen matrix in the presence or absence of GFs. Different Rho family GTPase members have distinct functions in the regulation of the actin cytoskeleton and cell migration. Their functions in HK motility are not clear. Both pharmacological and genetic approaches will be used to study these GTPases in HKs; 2) study how p38-MAPK and PKC-delta regulate random versus directional migration. Our recent findings show that p38-MAPK and PKC-delta are independently required for HK motility on a collagen matrix (in press). Here, we will further investigate the specific function of these two pathways in the integrin and the growth factor receptor signaling in HKs; 3) identify and characterize HK migration-linked genes by a novel "TGF-beta block" approach. We will take advantage of the fact that TGF-beta blocks proliferation but not migration in HKs. cRNAs from TGF-beta- treated 1) non-migrating HKs, 2) randomly migrating HKs, and 3) directionally migrating HKs will be subjected to DNA microarray analysis. The microarray-identified genes will be further subjected to a "pathway-screening" approach to narrow down the Rac1-p38-MAPK pathway- and the PKC-delta pathway-induced genes. These studies collectively will shed new light on the molecular mechanisms of wound re-epithelialization.