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: 7589653
• 负责人: Judith A West-Mays
• 金额: $ 20.97万
• 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/7589653
• 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是一种多效性形态因子，它调节组织修复表型，也扮演着 在纤维化修复病理的发展中起着重要作用。在眼睛的晶状体中，纤维性病变 TGFP介导的后囊混浊包括前囊下白内障(ASC)和后囊混浊(PCO)。 ASC和PCO纤维化前的细胞变化包括晶状体的增殖增加。 上皮细胞(LECs)，在上皮-间充质转化(EMT)下转化为 肌成纤维细胞，涉及E-钙粘附素的丢失和诱导α-平滑肌肌动蛋白(ASMA)的表达。这个 这个项目的长期目标是确定TGFp介导的信号，这些信号改变了基因的组成和 晶状体上皮细胞在ASC过程中的表型。使用先前开发的大鼠晶状体培养模型 外源性TGFP诱导ASC我们已经证明，用抑制TGFP酶活性的药物治疗 基质金属蛋白酶家族成员抑制TGFp诱导的白内障 改变。进一步的初步发现表明，MMPIs抑制TGFP的作用是可检验的假说。 通过抑制基质金属蛋白酶介导的E-钙粘附素降解诱导晶状体内EMT和ASC 正在脱落。RSmad，SMADS，是TGFP信号的常见效应器，已在 镜头。然而，我们发现使用两种不同的鼠标模型，在没有SMAD3 KO的情况下 小鼠)TGFp1可诱导晶状体上皮细胞发生EMT，表明Smads非依赖性信号转导途径参与了 ASC队形。其他发现表明，TGFP诱导的ASC涉及信号转导的假设 MAP激酶通路，特别是p38，独立于Smad3发挥作用。为了检验这些假设，我们 将利用转化生长因子诱导的体外ASC大鼠晶状体模型，以及转基因小鼠，包括 MMP2、MMP9和Smads KO小鼠。MMPIs对E-钙粘素表达和脱落的影响将是 进一步应用RT-QPCR结合激光捕获显微切割、Western blotting和 免疫定位。在ASC形成中激活pSSMAPK的需求将在两个项目中进行研究 使用特异性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ASC EMT和纤维化的调控途径。此外，由于基因和信号 ASC的形成机制类似于其他纤维性疾病和癌症中发生的机制 从这些研究中获得的信息，也可能与这些疾病实体相关。