青葙苷Ⅰ通过调节ROS介导的JNK/c-Jun信号通路增强视神经损伤模型视网膜神经节细胞的线粒体自噬  

Celoside I enhances mitophagy of retinal ganglion cells in optic nerve in⁃jury model by regulating ROS-mediated JNK/c-Jun signaling pathway

作　　者：韩易言 郑曲 赵磊[4] 宁志豪 董宝强[1,3] 左韬 HAN Yiyan;ZHENG Qu;ZHAO Lei;Ning Zhihao;DONG Baoqiang;ZUO Tao(Liaoning University of Traditional Chinese Medicine,Shenyang 110847,China;Key Laboratory of Acupuncture and Moxibustion Biology,Education Department of Liaoning Province,Shenyang 110847,China;Key Laboratory of Acupuncture and Moxibustion Health and Rehabilitation,Shenyang 110847,China;The Second Affiliated Hospital of Liaoning University of Traditional Chinese Medicine,Liaoning Academy of Traditional Chinese Medicine,Shenyang 110034,China)

机构地区：[1]辽宁中医药大学,辽宁沈阳110847 [2]辽宁省教育厅针灸生物学重点实验室,辽宁沈阳110847 [3]辽宁省针灸养生康复重点实验室,辽宁沈阳110847 [4]辽宁中医药大学附属第二医院,辽宁沈阳110034

出　　处：《中国病理生理杂志》2024年第10期1898-1905,共8页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.82104714);国家资助博士后研究人员计划(No.GZC20231025);国家中医药管理局中医药循证能力建设项目(No.2019XZZX-YK008);辽宁省教育厅青年项目(No.JYTQN2023458);辽宁省辽派中医学术经验和技能活态传承项目;辽宁省针灸养生康复重点实验室(No.18-006-0-06)。

摘　　要：目的:探讨青葙苷Ⅰ通过调节活性氧(ROS)介导的c-Jun氨基末端激酶(JNK)/c-Jun信号通路增强视神经损伤模型视网膜神经节细胞(RGCs)线粒体自噬的作用机制。方法:选取24只SPF级新西兰大白兔,随机分为4组,每组6只,分别为假手术组、模型组、甲钴胺组、实验组。模型组、甲钴胺组、实验组复制视神经损伤模型,假手术组仅切开球结膜后缝合。模型复制成功后,甲钴胺组给与0.15 mg/kg甲钴胺水溶液灌胃,实验组30 mg/kg青葙苷Ⅰ溶液灌胃,假手术组与模型组给与等体积生理盐水灌胃,各组每日干预一次,连续干预28 d。干预结束后,采用内窥镜检测各组大白兔眼底;苏木精-伊红(HE)染色法检测各组视网膜形态学变化;TUNEL法检测各组RGCs凋亡;免疫荧光染色法检测各组视网膜ROS、parkin及P62表达,Western blot检测各组视网膜JNK、c-Jun、parkin、P62及微管相关蛋白1轻链3(LC3)蛋白表达。结果:内窥镜结果显示,假手术组大白兔眼底表现正常,模型组可见眼底血供减少,视野呈现灰白色。与模型组比较,甲钴胺组与实验组可见眼底血流增加。HE结果显示,假手术组RGCs排列整齐,细胞形态规则,模型组可见RGCs数量减少、排列紊乱,与模型组相比,甲钴胺组与实验组RGCs形态改善,甲钴胺组仍存在部分RGCs形态异常,实验组RGCs总体数量较甲钴胺组多,RGCs形态异常少。与模型组相比,甲钴胺组与实验组视网膜凋亡指数较低(P<0.05),与甲钴胺组相比,实验组视网膜凋亡指数较低(P<0.05);免疫荧光法结果显示,与模型组相比,甲钴胺组与实验组视网膜组织中ROS、P62表达较低,parkin表达升高(P<0.05),与甲钴胺组比较,实验组视网膜组织中ROS、P62表达较低,parkin表达较高(P<0.05);Western blot结果显示,与模型组比较,甲钴胺组与实验组JNK、c-Jun、P62及LC3-Ⅰ/LC3-Ⅱ蛋白表达降低,parkin蛋白表达升高(P<0.05),与甲钴胺组比较,实验组JNK、c-Jun、P62AIM:This study aimed to investigate the mechanism by which celoside I enhances mitophagy in a model of optic nerve injury through regulation of reactive oxygen species(ROS)-mediated c-Jun N-terminal kinase(JNK)c-Jun signaling pathway.METHODS:Twenty-four New Zealand white rabbits were randomly divided into four groups:sham surgery,model,mecobalamin,and experimental group.Optic nerve injury was induced in the model,mecobalamin,and experimental groups,while the sham surgery group underwent a sham procedure.The mecobalamin group received mecobalamin,the experimental group received celoside I,and the sham surgery and model groups received saline.Interventions were administered daily for 28 d.Various techniques including endoscopy,hematoxylin-eosin(HE)staining,TUNEL method,immunofluorescence staining and Western blot were used to assess fundus condition,retinal morphology,apoptosis,ROS expression,and protein levels in the retina.RESULTS:Fundus examination revealed improved blood flow in the mecobalamin and experimental groups compared to the model group.Retinal morphology showed enhanced retinal ganglion cells(RGCs)in the mecobalamin and experimental groups.Apoptosis index was lower in the mecobalamin group compared to the experimental group.Immunofluorescence staining indicated reduced ROS and P62 expression and increased parkin and microtubule-associated protein light chain 3(LC3)expression in the experimental group compared to the mecobalamin group.Protein analysis showed decreased JNK,c-Jun,and P62 levels,and increased parkin and LC3 levels in the mecobalamin and experimental groups compared to the model group.CONCLUSION:Celoside I reduces ROS expression,inhibits the JNK/c-Jun pathway,enhances mitophagy,reduces apoptosis,and protects RGCs in optic nerve injury models.

关 键 词：青葙苷Ⅰ 视神经损伤 视网膜神经节细胞 线粒体自噬 

分 类 号：R774[医药卫生—眼科] R361.2[医药卫生—临床医学] R285.5