Mitochondrial complex I defect induces ROS release and degeneration in trabecular meshwork cells of POAG patients: Protection by antioxidants
Mitochondrial complex I defect induces ROS release and degeneration in trabecular meshwork cells of POAG patients: Protection by antioxidants
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DOI：
10.1167/iovs.07-1361
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发表时间：
2008-04-01
影响因子：
4.4
通讯作者：
Ge, Jian
中科院分区：
文献类型：
--
作者：
He, Yuan;Leung, Kar Wah;Ge, Jian
目的.越来越多的证据表明，氧化应激有助于原发性开角型青光眼（POAG）的进展，这是全球不可逆失明的主要原因。作者提供的证据表明，线粒体功能障碍是POAG患者小梁网（TM）细胞丢失的可能机制。通过标准手术小梁切除术获得POAG患者（GTM）和年龄匹配的无疾病受试者（NTM）的TM。用以下线粒体呼吸链抑制剂之一处理原代TM培养物：鱼藤酮（ROT，复合物I抑制剂）、噻吩甲酰三氟丙酮（TTFA，复合物II抑制剂）、粘噻唑或抗霉素A（MYX，AM-复合物III抑制剂）;线粒体通透性转换（MPT）抑制剂环孢菌素A（CsA）;和抗氧化剂维生素E（Vit E）或N-乙酰半胱氨酸（NAC）。分别用荧光探针5，5 '，6，6'-四氯-1，1 '3，3'-四乙基苯甲亚胺偶氮羰花青碘化物（JC-1）和基于ATP酶的ATP测定法通过线粒体膜电位（Δ Psim）和三磷酸腺苷（ATP）产生的变化来测定线粒体功能。通过H-2-DCF-DA测定活性氧（ROS）水平，通过乳酸脱氢酶活性和Annexin V-FITC标记测定细胞死亡。GTM细胞具有较高的内源性ROS水平、较低的ATP水平和降低的Δ Psim，并且它们比它们的正常对应物对线粒体复合物I抑制更敏感。ROT诱导GTM细胞中ROS产生的进一步增加、细胞色素c的释放以及ATP水平和Δ Psim的降低，最终导致细胞凋亡。复合物II和III抑制对细胞几乎没有影响。抗氧化剂通过抑制ROS的产生和细胞色素c的释放来防止ROT诱导的死亡。作者提出，线粒体复合物I缺陷与POAG患者的TM细胞变性有关，抗氧化剂和MPT抑制剂可以减缓这种疾病的进展。
PURPOSE. There is growing evidence that oxidative stress contributes to the progression of primary open-angle glaucoma (POAG), a leading cause of irreversible blindness worldwide. The authors provide evidence that mitochondrial dysfunction is a possible mechanism for the loss of trabecular meshwork (TM) cells in persons with POAG.METHODS. TM from patients with POAG (GTM) and age-matched subjects without disease (NTM) were obtained by standard surgical trabeculectomy. Primary TM cultures were treated with one of the following mitochondrial respiratory chain inhibitors: rotenone (ROT, complex I inhibitor), thenoyltrifluoroacetone (TTFA, complex II inhibitor), myxothiazol or antimycin A (MYX, AM-complex III inhibitors); mitochondrial permeability transition (MPT) inhibitor cyclosporine A (CsA); and antioxidants vitamin E (Vit E) or N-acetylcysteine (NAC). Mitochondrial function was determined by changes in mitochondrial membrane potential (Delta Psi m) and adenosine triphosphate (ATP) production with the fluorescent probes 5,5', 6,6'-tetrachloro-1,1'3,3'-tetraethylbenzimid azolocarbocyanine iodide (JC-1) and a luciferin/luciferase-based ATP assay, respectively. Reactive oxygen species (ROS) level, determined by H-2-DCF-DA, and cell death, measured by lactate dehydrogenase activity and Annexin V-FITC labeling, were also examined.RESULTS. GTM cells have higher endogenous ROS levels, lower ATP levels, and decreased Delta Psi m and they are more sensitive to mitochondrial complex I inhibition than their normal counterparts. ROT induces a further increase in ROS production, the release of cytochrome c, and decreases in ATP level and Delta Psi m in GTM cells, eventually leading to apoptosis. Complex II and III inhibition had little effect on the cells. Antioxidants protect against ROT-induced death by inhibiting ROS generation and cytochrome c release.CONCLUSIONS. The authors propose that a mitochondrial complex I defect is associated with the degeneration of TM cells in patients with POAG, and antioxidants and MPT inhibitors can reduce the progression of this condition.