Munemasa Yasunari, Kim Seok Hwan, Ahn Jae Hong, Kwong Jacky M K, Caprioli Joseph, Piri Natik
Jules Stein Eye Institute, University of California at Los Angeles, Los Angeles, California 90095, USA.
Invest Ophthalmol Vis Sci. 2008 Aug;49(8):3535-43. doi: 10.1167/iovs.08-1716. Epub 2008 Apr 25.
Oxidative stress has been implicated in retinal ganglion cell (RGC) death pathways after optic nerve transection (ONT) and during glaucomatous neuropathy. The authors investigated the expression and cell-protective roles of thioredoxins (cytosolic Trx1 and mitochondrial Trx2), important regulators of the cellular redox state, on RGCs after ONT and pharmacologic oxidative stress induction.
ONT was performed on adult Wistar rats. Trx1 and Trx2 quantitative and spatial expression were examined with Western blot and immunohistochemistry, respectively. Electroporation and calcium phosphate-mediated procedures were used to deliver Trx1 and Trx2 expression constructs to RGCs in vivo and to cultured RGC-5 cells, respectively. Cell-protective effects of Trx1 and Trx2 overexpression on RGCs after ONT and on RGC-5 cells treated with glutamate/buthionine sulfoximine (BSO) were determined by RGC density analysis and cell viability assay, respectively.
Upregulation of Trx1 and Trx2 was observed in RGCs at different times after ONT and in RGC-5 cells after glutamate/BSO treatment. Trx1 and Trx2 overexpression in RGC-5 cells increased their survival rate by approximately twofold and threefold 24 and 48 hours after glutamate/BSO treatment, respectively. A neuroprotective effect of Trx1 and Trx2 overexpression on RGCs was also observed in vivo; the survival rate of RGCs was increased by 35% and 135%, respectively, 1 and 2 weeks after ONT.
These findings provide evidence for in vitro and in vivo cell-protective effects of Trx1 and Trx2 on RGCs against oxidative stress-induced neurodegeneration.
氧化应激与视神经横断（ONT）后及青光眼性神经病变期间的视网膜神经节细胞（RGC）死亡途径有关。作者研究了细胞氧化还原状态的重要调节因子硫氧还蛋白（胞质Trx1和线粒体Trx2）在ONT后及药理学诱导氧化应激时对RGC的表达及细胞保护作用。
对成年Wistar大鼠进行ONT。分别用蛋白质印迹法和免疫组织化学法检测Trx1和Trx2的定量及空间表达。采用电穿孔法和磷酸钙介导法分别将Trx1和Trx2表达构建体导入体内的RGC及培养的RGC-5细胞。分别通过RGC密度分析和细胞活力测定，确定Trx1和Trx2过表达对ONT后RGC及用谷氨酸/丁硫氨酸亚砜胺（BSO）处理的RGC-5细胞的细胞保护作用。
ONT后不同时间的RGC及谷氨酸/BSO处理后的RGC-5细胞中观察到Trx1和Trx2上调。谷氨酸/BSO处理后24小时和48小时，RGC-5细胞中Trx1和Trx2过表达分别使其存活率提高了约两倍和三倍。在体内也观察到Trx1和Trx2过表达对RGC的神经保护作用；ONT后1周和2周，RGC的存活率分别提高了35%和135%。
这些发现为Trx1和Trx2在体外和体内对RGC抵抗氧化应激诱导的神经退行性变的细胞保护作用提供了证据。
