Liraglutide alleviates H2O2-induced retinal ganglion cells injury by inhibiting autophagy through mitochondrial pathways
Liraglutide alleviates H2O2-induced retinal ganglion cells injury by inhibiting autophagy through mitochondrial pathways
复制标题
利拉鲁肽通过线粒体途径抑制自噬减轻 H2O2 诱导的视网膜神经节细胞损伤
DOI：
10.1016/j.peptides.2017.04.008
复制
发表时间：
2017-06-01
期刊：
影响因子：
3
通讯作者：
Kuang, Hongyu
中科院分区：
文献类型：
--
作者：
Ma, Xuefei;Lin, Wenjian;Kuang, Hongyu
视网膜神经节细胞(RGCs)存在于视网膜内部，是糖尿病视网膜病变(DR)早期损伤的视网膜神经元。利拉鲁肽是一种GLP-1类似物，通过与受体(GLP-1R)结合发挥生物学功能，我们的团队于2012年首次发现GLP-1R在RGC-5细胞中表达。据报道，利拉鲁肽可预防糖尿病患者的视网膜神经变性。然而，自噬和线粒体平衡等机制在利拉鲁肽诱导的视网膜保护中的作用尚不清楚。在此，我们旨在研究利拉鲁肽对视网膜RGC的保护作用，并探讨其可能的保护机制。用过氧化氢和/或利拉鲁肽处理RGC-5细胞。CCK-8试剂盒检测细胞存活率。免疫印迹分析检测轴突标记物GAP43、自噬和有丝分裂吞噬标志物LC3A/B、Beclin-1、p62、Parkin、BCL2/腺病毒E1B 19 kDa蛋白相互作用蛋白3样蛋白(BNIP3L)和线粒体生物发生关键调控因子PGC-1a。用ImageXpress Micro XLS系统和透射电子显微镜对自噬进行了评估。用ImageXpress Micro XLS系统检测线粒体膜电位、活性氧(ROS)和荧光染色。我们的结果表明，利拉鲁肽可显著阻止过氧化氢诱导的细胞活力下降、线粒体形态恶化和自噬的诱导，表现为Lc3 II/I和Beclin-1的表达增加以及p62的降解。此外，利拉鲁肽可抑制过氧化氢诱导的GAP43表达下降，从而保护细胞。然而，雷帕霉素诱导了自噬并阻断了保护过程。利拉鲁肽还提供了线粒体保护，并似乎减轻了H_2O_2诱导的ROS过量产生和线粒体膜电位下降，部分是通过促进线粒体的生成和减弱有丝分裂。总之，利拉鲁肽通过维持线粒体生物发生和有丝分裂吞噬之间的平衡来抑制自噬，从而减轻H_2O_2诱导的RGC-5细胞损伤。
Retinal ganglion cells (RGCs), which exist in the inner retina, are the retinal neurons which can be damaged in the early stage of diabetic retinopathy (DR). Liraglutide, a glucagon-lilce peptide-1 (GLP-1) analog, exerts biological functions by binding the receptor (GLP-1R), the expression of which in RGC-5 cells was first shown by our team in 2012. It was reported that liraglutide prevented retinal neurodegeneration in diabetic subjects. However, the involvement of mechanisms such as autophagy and mitochondrial balance in liraglutide-induced retinal protection is unknown. Here, we aimed to investigate the protective effects of liraglutide and explore the potential mechanisms of liraglutide-induced retinal RGC protection. RGC-5 cells were treated with H2O2 and/or liraglutide. Cell viability was detected with the CCK-8 kit. The axon marker GAP43, autophagy and mitophagy indicators LC3A/B, Beclin-1, p62, Parkin, BCL2/Adenovirus E1B 19 kDa protein-interacting protein 3-like (BNIP3L) and the key regulator of mitochondrial biogenesis PGC-la were examined via western blot analysis. Autophagy was also evaluated using the ImageXpress Micro XLS system and transmission electron microscopy (TEM). Reactive oxygen species (ROS), mitochondrial membrane potential and fluorescent staining for mitochondria were also measured using the ImageXpress Micro XLS system. Our results showed that pretreatment with liraglutide significantly prevented H2O2-induced cell viability decline, mitochondrial morphological deterioration and induction of autophagy, which appeared as increased expression of LC3 II/I and Beclin-1, along with p62 degradation. Moreover, liraglutide suppressed the H2O2-induced decline in GAP43 expression, thus protecting cells. However, rapamycin induced autophagy and blocked the protective process. Liraglutide also provided mitochondrial protection and appeared to alleviate H2O2-induced ROS overproduction and a decline in mitochondrial membrane potential, partially by promoting mitochondrial generation and attenuating mitophagy. In conclusion, liraglutide attenuates H2O2 induced RGC-5 cell injury by inhibiting autophagy through maintaining a balance between mitochondrial biogenesis and mitophagy.