Role of Matrix Metalloproteinases in Subcapsular Cataract Formation

基质金属蛋白酶在囊下白内障形成中的作用

• 批准号: 7386538
• 负责人: Judith A West-Mays
• 金额: $ 20.55万
• 依托单位: MCMASTER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7386538
• 关键词: Anterior; Cataract; Conditioned Culture Media; Crystalline Lens; Data; Development; Disease; E-Cadherin; Epithelial; Epithelial Cells; Exhibits; Family member; Fibrosis; Figs - dietary; Gelatinase A; Gelatinase B; Genes; Genetic; Goals; In Vitro; Knockout Mice; MAP Kinase Gene; MAP Kinase Signaling Pathways; MAPK14 gene; MMPI; Malignant Neoplasms; Matrix Metalloproteinases; Mediating; Mesenchymal; Mitogen-Activated Protein Kinases; Modeling; Mus; Mutant Strains Mice; Myofibroblast; Pathology; Pathway interactions; Phenotype; Phosphorylation; Play; Rattus; Research Personnel; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Smooth Muscle Actin Staining Method; System; Testing; Time; Western Blotting; Wound Healing; cytokine; in vitro Model; in vivo; kinase inhibitor; laser capture microdissection; lens; morphogens; mouse model; prevent; programs; repaired

The cytokine TGFP is a pleotropic morphogen that modulates the tissue repair phenotype and also plays an important role in the development of fibrotic repair pathologies. In the lens of the eye, fibrotic pathologies mediated by TGFP include anterior subcapsular cataracts (ASC) and posterior capsular opacification (PCO). The cellular changes that precede fibrosis in ASC and PCO include an increased proliferation of lens epithelial cells (LECs), which under go an epithelial-mesenchymal transformation (EMT) into myofibroblasts, involving loss of E-cadherin and induced a-smooth muscle actin (aSMA) expression. The long-term goal of this project is to determine the TGFp-mediated signals, which alter the genetic makeup and phenotype of lens epithelial cells during ASC. Using a previously developed rat lens culture model in which exogenous TGFP induces ASC we have shown that treatment with agents that inhibit enzymatic activity of Matrix Metalloproteinase (MMP) family members (MMPIs) suppresses the TGFp-induced cataractous changes. Further preliminary findings suggest the testable hypothesis that MMPIs act to inhibit TGFP- induced EMT and ASC in the lens by suppressing MMP-mediated E-cadherin degradation by shedding. The RSmad, SmadS, is a common effector of TGFP signaling, which has been identified in the lens. However, we have found using two different mouse models that in the absence of SmadS (in Smad3 KO mice) TGFpl can induce EMT in the lens, demonstrating the involvement of SmadS-independent signaling in ASC formation. Additional findings suggest that the hypothesis that TGFp-induced ASC involves signaling MAP kinase pathways, specifically p38,which act independently of Smad3. To test these hypotheses we will utilize the in vitro TGFp-induced rat lens model of ASC, as well as, genetically modified mice, including MMP-2, MMP-9 and SmadS KO mice. The effect of MMPIs on E-cadherin expression and shedding will be further examined using RT-QPCR in combination with laser capture microdissection, western blotting and immunolocalization. The requirement for activated pSSMAPKin ASC formation will be investigated in both the in vivo and in vitro models using specific Pp38 kinase inhibitors. These data will aid in defining the TGFp-mediated pathways controlling EMT and fibrosis in ASC. Furthermore, since the genes and signaling mechanisms in ASC formation are similar to those, which occur, in other fibrotic diseases and cancer, the information gained from these studies in the lens, may also have relevance to these disease entities.

细胞因子TGFP是一种多形形态，可调节组织修复表型，并且还播放 在纤维化修复病理发展中的重要作用。在眼睛的镜头中，纤维化病理 由TGFP介导的包括前下囊型白内障（ASC）和后囊膜不透明（PCO）。 ASC和PCO纤维化之前的细胞变化包括晶状体增殖的增加 上皮细胞（LEC），在上皮 - 间质转化（EMT）下 肌纤维细胞涉及损失E-钙粘蛋白并诱导A-Smooth肌肉肌动蛋白（ASMA）表达。这 该项目的长期目标是确定TGFP介导的信号，该信号改变了遗传构成和 ASC期间晶状体上皮细胞的表型。使用先前开发的老鼠透镜培养模型，其中 外源性TGFP诱导ASC我们表明，用抑制抑制酶活性的药物的治疗 基质金属蛋白酶（MMP）家族成员（MMPI）抑制TGFP诱导的白内障 更改。进一步的初步发现表明，MMPI作用以抑制TGFP-的可检验假设 通过通过抑制MMP介导的E-钙粘着蛋白降解来诱导镜头中的EMT和ASC 脱落。 RSMAD SMADS是TGFP信号传导的常见效应器，已在 镜片。但是，我们发现使用两种不同的鼠标模型在没有SMAD的情况下（在SMAD3 KO中 小鼠）TGFPL可以在镜头中诱导EMT，证明与SMADS无关的信号参与 ASC形成。其他发现表明，TGFP诱导的ASC的假设涉及信号 MAP激酶途径，特别是p38，独立于SMAD3。为了检验这些假设我们 将利用ASC的体外TGFP诱导的大鼠透镜模型以及转基因的小鼠，包括 MMP-2，MMP-9和SMADS KO小鼠。 MMPI对电子钙粘蛋白表达和脱落的影响将是 使用RT-QPCR与激光捕获微分解，蛋白质印迹和 免疫定位。激活的PSSMAPKIM ASC形成的要求将在两者中研究 使用特定PP38激酶抑制剂的体内和体外模型。这些数据将有助于定义 TGFP介导的途径控制ASC中的EMT和纤维化。此外，由于基因和信号传导 ASC形成的机制与其他纤维性疾病和癌症中发生的机制相似， 从镜头中的这些研究中获得的信息也可能与这些疾病实体有关。