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: 7024380
• 负责人: Judith A West-Mays
• 金额: $ 21.6万
• 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/7024380
• 关键词: cadherins; cataract; lens; model; role

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.

产品说明：细胞因子TGF？是调节组织修复表型的多效性形态原，并且在纤维化修复病理学的发展中也起重要作用。在眼睛的透镜中，由TGF β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中纤维化之前的细胞变化包括透镜上皮细胞（LEC）增殖增加，其经历上皮-间充质转化（EMT）成肌成纤维细胞，涉及E-钙粘蛋白的损失和诱导的α-平滑肌肌动蛋白（aSMA）表达。该项目的长期目标是确定TGF β介导的信号，其在ASC期间改变透镜上皮细胞的遗传组成和表型。使用先前开发的大鼠透镜培养模型，其中外源性TGF诱导ASC，我们已经表明，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抑制TGF诱导的白内障变化。进一步的初步研究结果提示了一个可验证的假设，即MMPIs通过抑制MMP介导的E-钙粘蛋白降解（脱落）来抑制透镜中TGF β诱导的EMT和ASC。RSmad（Smad 3）是TGF β信号传导的常见效应物，其已在透镜中鉴定。然而，我们发现使用两种不同的小鼠模型，在SmadS缺失的情况下（在Smad 3 KO小鼠中），TGF？1可以诱导透镜中的EMT，这表明了非Smad 3依赖性信号传导参与ASC形成。其他研究结果表明，TGF β诱导的ASC涉及信号传导MAP激酶途径，特别是p38，其作用独立于Smad 3。为了检验这些假设，我们将利用体外TGF β诱导的ASC大鼠透镜模型，以及遗传修饰的小鼠，包括MMP-2、MMP-9和SmadS KO小鼠。将使用RT-QPCR结合激光捕获显微切割、蛋白质印迹和免疫定位进一步检查MMPI对E-钙粘蛋白表达和脱落的影响。将在体内和体外模型中使用特异性Pp 38激酶抑制剂研究ASC形成中活化p38 MAPK的需要。这些数据将有助于确定TGF β介导的控制ASC中EMT和纤维化的途径。 此外，由于ASC形成中的基因和信号传导机制与其他纤维化疾病和癌症中发生的基因和信号传导机制相似，因此从这些透镜研究中获得的信息也可能与这些疾病实体相关。