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: 7186673
• 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/7186673
• 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

DESCRIPTION: The cytokine TGF¿ 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 TGF¿ 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 TGF¿-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 TGF¿ induces ASC we have shown that treatment with agents that inhibit enzymatic activity of Matrix Metalloproteinase (MMP) family members (MMPIs) suppresses the TGF¿-induced cataractous changes. Further preliminary findings suggest the testable hypothesis that MMPIs act to inhibit TGF¿-induced EMT and ASC in the lens by suppressing MMP-mediated E-cadherin degradation by shedding. The RSmad, Smad3, is a common effector of TGF¿ 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) TGF¿1 can induce EMT in the lens, demonstrating the involvement of Smad3-independent signaling in ASC formation. Additional findings suggest that the hypothesis that TGF¿-induced ASC involves signaling MAP kinase pathways, specifically p38, which act independently of Smad3. To test these hypotheses we will utilize the in vitro TGF¿-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 p38MAPK in 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 TGF¿-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.

描述：细胞因子TGF¿是一种多效性形态因子，调节组织修复表型，在纤维化修复病理的发展中起重要作用。在眼晶状体中，TGF¿介导的纤维化病理包括前囊下白内障（ASC）和后囊膜混浊（PCO）。ASC和PCO纤维化前的细胞变化包括晶状体上皮细胞（LECs）增殖增加，晶状体上皮细胞在上皮-间充质转化（EMT）为肌成纤维细胞，包括e -钙粘蛋白的缺失和诱导的a-平滑肌肌动蛋白（aSMA）表达。该项目的长期目标是确定TGF -介导的信号，这些信号在ASC期间改变晶状体上皮细胞的基因组成和表型。利用先前开发的外源性TGF诱导ASC的大鼠晶状体培养模型，我们发现用抑制基质金属蛋白酶（MMP）家族成员（MMPIs）酶活性的药物治疗可以抑制TGF诱导的白内障变化。进一步的初步研究结果支持可验证的假设，即MMPIs通过抑制mmp介导的e -钙粘蛋白脱落降解来抑制TGF¿-诱导的晶状体EMT和ASC。RSmad，即Smad3，是TGF¿信号的常见效应体，已在晶状体中发现。然而，我们通过使用两种不同的小鼠模型发现，在缺乏smad的情况下（在Smad3 KO小鼠中），TGF¿1可以诱导晶状体中的EMT，这表明Smad3独立信号参与ASC的形成。其他研究结果表明，TGF¿-诱导的ASC涉及MAP激酶信号通路，特别是p38，它独立于Smad3起作用。为了验证这些假设，我们将利用TGF -诱导的体外大鼠ASC晶状体模型，以及转基因小鼠，包括MMP-2、MMP-9和SmadS KO小鼠。MMPIs对E-cadherin表达和脱落的影响将采用RT-QPCR结合激光捕获显微解剖、western blotting和免疫定位进一步研究。使用特定的Pp38激酶抑制剂，将在体内和体外模型中研究ASC形成对活化p38MAPK的需求。这些数据将有助于确定TGF介导的ASC中控制EMT和纤维化的途径。此外，由于ASC形成中的基因和信号机制与其他纤维化疾病和癌症中的基因和信号机制相似，因此从晶状体研究中获得的信息也可能与这些疾病实体相关。