Protein carbonylation and aggregation precede neuronal apoptosis induced by partial glutathione depletion.
Protein carbonylation and aggregation precede neuronal apoptosis induced by partial glutathione depletion.
复制标题
蛋白羰基和聚集在部分谷胱甘肽消耗引起的神经元细胞凋亡之前。
DOI：
10.1042/an20110064
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发表时间：
2012-04-10
期刊：
影响因子：
4.7
通讯作者：
Bizzozero OA
中科院分区：
文献类型：
--
作者：
Dasgupta A;Zheng J;Bizzozero OA
虽然细胞内氧化蛋白质的积累被认为是有毒的，但目前没有证据表明蛋白质羰基化和细胞死亡有关。在本研究中，我们发现，与50 μM DEM（马来酸二乙酯）孵育的nPC 12（神经元样PC 12）细胞导致谷胱甘肽（GSH）的部分和短暂的耗尽。伴随着GSH的消失，PCO（蛋白质羰基）的积累和细胞死亡（坏死和凋亡）增加。免疫细胞化学研究还揭示了羰基化和细胞凋亡之间的时间/空间关系。此外，如果氧化蛋白不被蛋白酶体降解去除，PCO积累、蛋白质聚集和细胞死亡这三者的程度都会增加。此外，羰基清除剂肼屈嗪、组氨酸酰肼和甲氧基胺在防止细胞死亡方面的有效性将PC 0鉴定为有毒物质。使用充分表征的细胞凋亡抑制剂的实验将蛋白质羰基化置于线粒体过渡孔开口的下游和半胱天冬酶激活的上游。虽然这项研究主要集中在nPC 12细胞上，但在原代神经元培养物中的实验也得到了相同的结果。这些发现也不局限于DEM诱导的细胞死亡，因为在星形孢菌素和丁硫堇亚砜胺处理的nPC 12细胞中发现了羰基化和细胞凋亡之间类似的关系。总之，上述结果首次显示了羰基化、蛋白质聚集和经历氧化损伤的神经元凋亡之间的因果关系。据我们所知，这是第一个研究直接（氧化）蛋白质羰基化的凋亡途径。
While the build-up of oxidized proteins within cells is believed to be toxic, there is currently no evidence linking protein carbonylation and cell death. In the present study, we show that incubation of nPC12 (neuron-like PC12) cells with 50 μM DEM (diethyl maleate) leads to a partial and transient depletion of glutathione (GSH). Concomitant with GSH disappearance there is increased accumulation of PCOs (protein carbonyls) and cell death (both by necrosis and apoptosis). Immunocytochemical studies also revealed a temporal/spatial relationship between carbonylation and cellular apoptosis. In addition, the extent of all three, PCO accumulation, protein aggregation and cell death, augments if oxidized proteins are not removed by proteasomal degradation. Furthermore, the effectiveness of the carbonyl scavengers hydralazine, histidine hydrazide and methoxylamine at preventing cell death identifies PCOs as the toxic species. Experiments using well-characterized apoptosis inhibitors place protein carbonylation downstream of the mitochondrial transition pore opening and upstream of caspase activation. While the study focused mostly on nPC12 cells, experiments in primary neuronal cultures yielded the same results. The findings are also not restricted to DEM-induced cell death, since a similar relationship between carbonylation and apoptosis was found in staurosporine- and buthionine sulfoximine-treated nPC12 cells. In sum, the above results show for the first time a causal relationship between carbonylation, protein aggregation and apoptosis of neurons undergoing oxidative damage. To the best of our knowledge, this is the first study to place direct (oxidative) protein carbonylation within the apoptotic pathway.