Pathology of Renal Matrix Metabolism

描述(申请人提供)：间质纤维化和肾小管萎缩是所有形式进行性肾纤维化的特征。在过去的十年里，这个实验室一直专注于特定的(MMP2和MT1-MMPs)在这一过程中的作用。基质金属蛋白酶-2与MT1-基质金属蛋白酶结合，足以诱导极化的上皮细胞向成纤维细胞表型的转化，这一过程被称为上皮间充质转化(EMT)。从数量上讲，EMT与间质成纤维细胞群的激活和扩张一起，是肾小管萎缩和进行性肾纤维化的主要因素。在体外，EMT由多种细胞因子和环境因素驱动；然而，我们假设肾脏EMT是由三个主要转录网络驱动的亚稳态：转化生长因子-β/Smad；E-钙粘蛋白/β-连环蛋白/Wnt/Lef/Tcf；以及MAPK/ERK信号转导通路。我们已经确定了MAPK/ERK信号级联的转录靶点是MAPK-2和MT1-MMPs，并确定了一个特定的AP-1复杂成分Fra-2足以在体外驱动EMT的过程。这一建议的主要假设是持续的MAPK/ERK信号，伴随着Fra-2的增强生成，驱动肾脏EMT所需的一组确定的基因的转录。解决这一问题的方法包括三个特定的目标，即利用微阵列分析来描述一系列表现出一系列上皮到间充质特征的上皮细胞克隆群体中Fra-2调控的基因。已确定的基因的功能意义将通过体外和体内方法进行验证，包括通过在肾近端小管中转基因表达活性基质金属蛋白酶-2而产生的独特的肾脏EMT模型。最后，Fra-2单独在体内诱导EMT的能力将通过该转录因子的转基因近端小管表达来测试。这些研究旨在确定肾脏EMT所需的基因集，从而有望为肾脏疾病的治疗提供新的治疗靶点。

DESCRIPTION (provided by applicant): Interstitial fibrosis and tubular atrophy are the hallmarks of all forms of progressive renal fibrosis. This laboratory has focused over the past decade on the role of specific (MMP-2 and MT1-MMP) in this process. MMP-2, in conjunction with MT1-MMP, is sufficient to induce the conversion of the polarized epithelial cell to a fibroblastic phenotype, a process termed epithelial mesenchymal transition (EMT). Quantitatively, EMT, in conjunction with activation and expansion of the interstitial fibroblast population, is a major contributor to tubular atrophy and progressive renal fibrosis. In vitro, EMT is driven by a diverse number of cytokines and environmental factors; however, we have postulated that renal EMT represents a metastable state driven by three dominant transcriptional networks: TGF-beta/Smad; E- cadherin/beta-catenin/Wnt/LEF/TCF; and MAPK/ERK signaling cascades. We have identified both MMP-2 and MT1-MMP as transcriptional targets of the MAPK/ERK signaling cascades and determined that a specific AP-1 complex component, Fra-2, is sufficient to drive the process of EMT in vitro. The primary hypothesis of this proposal is that sustained MAPK/ERK signaling, with enhanced generation of Fra-2, drives the transcription of a defined cohort of genes required for renal EMT. The approaches to this problem include three Specific Aims proposing to characterize, using microarray analysis, Fra-2-regulated genes in a series of clonal populations of epithelial cells displaying a range of epithelial to mesenchymal features. The functional significance of identified genes will be validated using in vitro and in vivo approaches, including a unique model of renal EMT generated by the transgenic expression of active MMP-2 in the renal proximal tubule. Finally, the ability of Fra-2, alone, to induce EMT in vivo will be tested by the transgenic proximal tubule expression of this transcription factor. These studies are designed to identify those gene sets required for renal EMT and thereby hopefully provide new therapeutic targets for the treatment of renal disease.