Tissue transglutaminase expression and activity in normal and glaucomatous human trabecular meshwork cells and tissues
Tissue transglutaminase expression and activity in normal and glaucomatous human trabecular meshwork cells and tissues
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DOI：
10.1167/iovs.07-0835
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发表时间：
2008-02-01
影响因子：
4.4
通讯作者：
Wordinger, Robert J.
中科院分区：
文献类型：
--
作者：
Tovar-Vidales, Tara;Roque, Rouel;Wordinger, Robert J.
目的.青光眼是世界范围内不可逆视力损害和失明的主要原因。青光眼的一个主要危险因素是眼内压升高，这是由于通过小梁网（TM）的房水流出阻力增加所致。在昏迷性TM中，由于ECM沉积和降解之间的正常平衡被破坏，细胞外基质（ECM）材料的积累增加。组织转氨酶（TGM 2）属于钙依赖性酶家族，其通过交联蛋白催化ECM的翻译后修饰，从而使这些蛋白质抵抗酶促和物理降解。可能的是，ECM蛋白在肿瘤TM中的增加是由于TGM 2的交联活性增加。本研究的目的是确定是否有正常和青光眼TM细胞和组织中的TGM 2的表达和活性的差异。使正常（n = 3 NTM）和肉瘤（n = 3 GTM）人TM细胞系生长直至汇合。使用细胞裂解物的Western免疫印迹分析来比较NTM和GTM细胞中的TGM 2蛋白水平。通过使用生物素尸胺测定法研究NTM和GTM细胞之间的TGM 2酶活性。此外，三个正常和三个肿瘤TM组织的免疫组化被用来评估在体内表达的TGM 2，纤连蛋白（FN）和γ-（γ-谷氨酰）赖氨酸（GGEL）蛋白。蛋白质印迹分析和免疫组织化学证明NTM和GTM细胞中都存在TGM 2蛋白。与NTM细胞相比，GTM细胞中的TGM 2蛋白增加，并且与NTM细胞相比，GTM细胞中的TGM 2酶活性也增加。免疫组化结果显示TGM 2和FN在GTM组织中表达增加。FN和GGEL蛋白在GTM组织中共定位，表明FN与TGM 2有明显的交联作用。这项研究表明，NTM和GTM细胞都表达TGM 2。此外，TGM 2蛋白水平和酶活性在GTM细胞中升高。GTM组织中FN和GGEL蛋白的共定位也增加。这些结果表明，TGM 2可能通过交联ECM蛋白如FN，从而使ECM更耐降解，在青光眼的发病机制中发挥重要作用。
PURPOSE. Glaucoma is a leading cause of irreversible visual impairment and blindness in the world. A major risk factor for glaucoma is elevated intraocular pressure due to increased resistance of aqueous humor outflow through the trabecular meshwork (TM). In the glaucomatous TM, there is increased accumulation of extracellular matrix (ECM) material due to a disruption of the normal balance between ECM deposition and degradation. Tissue transglutaminase (TGM2) belongs to a family of calcium-dependent enzymes that catalyze the posttranslational modification of the ECM by cross-linking proteins, thus making these proteins resistant to enzymatic and physical degradation. It is possible that the increase in ECM proteins in the glaucomatous TM is due to increased cross-linking activity of TGM2. The purpose of this study was to determine whether there are differences in expression and activity of TGM2 between normal and glaucoma TM cells and tissues.METHODS. Normal (n = 3 NTM) and glaucomatous (n = 3 GTM) human TM cell lines were grown until confluent. Western immunoblot analysis of cell lysates was used to compare TGM2 protein levels in NTM and GTM cells. TGM2 enzyme activity between NTM and GTM cells was studied by using a biotin cadaverine assay. In addition, immunohistochemistry of three normal and three glaucomatous TM tissues was used to evaluate the in vivo expression of TGM2, fibronectin (FN) and epsilon-(gamma-glutamyl) lysine (GGEL) proteins.RESULTS. Western blot analysis and immunohistochemistry demonstrated the presence of TGM2 protein in both NTM and GTM cells. There was an increase in TGM2 protein in GTM cells compared with NTM cells, and GTM cells also had increased in TGM2 enzyme activity compared with NTM cells. Immunohistochemical results demonstrated increased expression of TGM2 and FN in GTM tissues. FN and GGEL proteins were colocalized in GTM tissues, indicating significant cross-linking of FN by TGM2.CONCLUSIONS. This study demonstrated that both NTM and GTM cells express TGM2. In addition, TGM2 protein levels and enzyme activities were elevated in GTM cells. There was also an increase in colocalization of FN and GGEL protein in GTM tissues. These results indicate that TGM2 may play an important role in the pathogenesis of glaucoma by cross-linking ECM proteins such as FN and thus making the ECM more resistant to degradation.