Peroxynitrite-induced apoptosis in photoreceptor cells.
Peroxynitrite-induced apoptosis in photoreceptor cells.
复制标题
过氧亚硝酸盐诱导感光细胞凋亡。
DOI：
10.1076/ceyr.28.1.17.23488
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Rao,NarsingA
中科院分区：
文献类型：
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作者：
Ito,Shinji;Wu,Guey-Shuang;Kimoto,Takashi;Hisatomi,Toshio;Ishibashi,Tatsuro;Rao,NarsingA
目的研究过氧亚硝基阴离子对视网膜光感受器细胞的损伤机制。方法取20日龄刘易斯大鼠视网膜，用不同浓度的SIN-1处理不同时间。采用原位末端标记（TUNEL）法和DNA片段化法检测光感受器细胞凋亡。处理所选视网膜样品进行超微结构分析以确认细胞凋亡。通过免疫组织化学和Western印迹分析检测暴露于SIN-1的视网膜中caspase-3的表达。视网膜也进行了评估的存在下的半胱天冬酶抑制剂苄氧基羰基-Val-Ala-Asp-氟甲基酮（Z-VAD-faturalketone）的凋亡的预防。结果暴露于SIN-1的视网膜外植体显示TUNEL阳性感光细胞的存在下显着增加。类似地，在SIN-1浓度增加的情况下，观察到TUNEL阳性细胞的线性增加。SIN-1作用后可见DNA梯状条带形成。在超微结构上，SIN-1暴露的视网膜揭示了感光细胞核中典型的凋亡变化。与单独暴露于SIN-1的视网膜相比，用尿酸盐预孵育6小时并暴露于SIN-1 16小时的视网膜显示出显著更少的TUNEL阳性细胞（p < 0.05）。此外，SIN-1暴露的视网膜显示caspase- 3的表达。这种表达，以及凋亡的光感受器的数量，显着降低Z-VAD-fasten.ConclusionsThese结果表明，过氧亚硝酸盐诱导光感受器细胞凋亡，这种视网膜损伤似乎是介导的caspase-3的存在。过亚硝酸盐清除剂尿酸盐以及半胱天冬酶抑制剂Z-VAD-favorite可以使凋亡过程最小化。
PurposeTo study the mechanisms of peroxynitrite-induced photoreceptor cell damage, using retinal cultures and a peroxynitrite donor, 3-morpholinosydnonimine (SIN-1).MethodsRetinal explants obtained from 20-day-old Lewis rat pups, were exposed to SIN-1 for varying lengths of time at varying concentrations. Apoptosis in the photoreceptor cells was detected using the TdT-mediated dUTP-biotin nick end labeling (TUNEL) method and a DNA fragmentation assay. Selected retinal samples were processed for an ultra-structural analysis to confirm apoptosis. The retinas exposed to SIN-1 were tested for the expression of caspase-3 by immunohistochemistry and a Western blot analysis. The retinas were also evaluated for the prevention of apoptosis in the presence of the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (Z-VAD-fmk).ResultsThe retinal explants exposed to SIN-1 showed a significant increase in the presence of TUNEL-positive photoreceptor cells. Similarly lineal increases in TUNELpositive cells were seen in the presence of increasing concentrations of SIN-1. DNA ladder formation was seen with the exposure of SIN-1. Ultrastructurally, SIN-1 exposed retinas revealed typical apoptotic changes in the photoreceptor cell nuclei. The retinas preincubated with urate for 6 hours and exposed to SIN-1 for 16 hours showed significantly fewer TUNEL-positive cells compared to the retinas exposed to SIN-1 alone (p < 0.05). Moreover, the retinas exposed to SIN-1 showed the expression of caspase- 3. This expression, as well as the number of apoptotic photoreceptors, significantly decreased in the presence of Z-VAD-fmk.ConclusionsThese results show that peroxynitrite induces apoptosis in photoreceptor cells and that such retinal damage appears to be mediated by caspase-3. The apoptotic process can be minimized by peroxynitrite scavenger urate, as well as by the caspase inhibitor Z-VAD-fmk.