GSH Levels Serve As a Biological Redox Switch Regulating Sulforaphane-Induced Cell Fate in Human Lens Cells.
GSH Levels Serve As a Biological Redox Switch Regulating Sulforaphane-Induced Cell Fate in Human Lens Cells.
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GSH水平是调节人晶状体细胞中硫烷诱导的细胞命运的生物氧化还原转换。
DOI：
10.1167/iovs.62.15.2
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发表时间：
2021-12-01
影响因子：
4.4
通讯作者：
Wormstone IM
中科院分区：
文献类型：
--
作者：
Huynh TPN;Bowater RP;Bernuzzi F;Saha S;Wormstone IM
萝卜硫素（SFN）是一种治疗许多健康状况的植物化学剂。sfn诱导的细胞毒性在预防后囊膜混浊（PCO）方面具有前景。在当前的研究中，我们旨在阐明SFN治疗与晶状体细胞死亡之间的关键过程和机制。以人晶状体上皮细胞系FHL124和中央前上皮为实验模型。通过显微镜观察和细胞损伤/活力测定来评估细胞死亡。采用TaqMan RT-PCR或免疫印迹法检测基因或蛋白水平。免疫荧光法检测线粒体网络和DNA损伤。采用不同的光报告法测定线粒体膜电位、激活转录因子6 （ATF6）活性、还原性谷胱甘肽（GSH）与氧化性谷胱甘肽（GSSG）的比值和谷胱甘肽还原酶（GR）活性。采用LC-MS/MS分析SFN代谢物。n -乙酰半胱氨酸（NAC）是一种活性氧清除剂，在两种模型中均可预防sfn诱导的细胞死亡。NAC还能显著保护FHL124细胞免受sfn诱导的线粒体功能障碍、内质网应激（ERS）、DNA损伤和自噬的影响。SFN显著减少了眼内主要抗氧化剂GSH，并降低了GR活性，尽管其蛋白质水平增加了一倍。应用SFN后，晶状体细胞中检测到的最丰富的SFN共轭物是SFN - gsh。GSH的加入保护晶状体细胞免受sfn诱导的所有细胞事件的影响。SFN通过结合和抑制GR活性来消耗晶状体细胞中的GSH水平。这导致活性氧和氧化应激增加，从而引发线粒体功能障碍、ERS、自噬和DNA损伤，导致细胞死亡。总之，本研究为支持SFN治疗PCO和其他疾病提供了机制理解。
Sulforaphane (SFN) is a therapeutic phytochemical agent for many health conditions. SFN-induced cytotoxicity is shown to have promise in preventing posterior capsule opacification (PCO). In the current study, we aimed to elucidate key processes and mechanisms linking SFN treatment to lens cell death. The human lens epithelial cell line FHL124 and central anterior epithelium were used as experimental models. Cell death was assessed by microscopic observation and cell damage/viability assays. Gene or protein levels were assessed by TaqMan RT-PCR or immunoblotting. Mitochondrial networks and DNA damage were assessed by immunofluorescence. Mitochondrial membrane potential, activating transcription factor 6 (ATF6) activity, ratio of reduced glutathione (GSH) to oxidized glutathione (GSSG), and glutathione reductase (GR) activity were measured using different light reporter assays. SFN metabolites were analyzed by LC-MS/MS. Treatment with N-acetylcysteine (NAC), a reactive oxygen species scavenger, prevented SFN-induced cell death in both models. NAC also significantly protected FHL124 cells from SFN-induced mitochondrial dysfunctions, endoplasmic reticulum stress (ERS), DNA damage and autophagy. SFN significantly depleted GSH, the major antioxidant in the eye, and reduced GR activity, despite doubling its protein levels. The most abundant SFN conjugate detected in lens cells following SFN application was SFN–GSH. The addition of GSH protected lens cells from all SFN-induced cellular events. SFN depletes GSH levels in lens cells through conjugation and inhibition of GR activity. This leads to increased reactive oxygen species and oxidative stress that trigger mitochondrial dysfunction, ERS, autophagy, and DNA damage, leading to cell death. In summary, the work presented provides a mechanistic understanding to support the therapeutic application of SFN for PCO and other disorders.