红景天苷通过NF-κB信号通路对高糖诱导HT22海马区神经元凋亡的保护作用  

Protective Role and Mechanism of Salidroside Against High Glucose-induced Apoptosis via NF-κB Pathway in HT22 Hippocampal Neurons

作　　者：丁佳媛 于洪丹 郁盛雪 刘文强 郑晴予 刘盈睿 左中夫 DING Jiayuan;YU Hongdan;YU Shengxue;LIU Wenqiang;ZHENG Qingyu;LIU Yingrui;ZUO Zhongfu(Department of Human Anatomy,Jinzhou Medical University;Liaoning Key Laboratory of Diabetic Cognitive and Perceptive Dysfunction,Jinzhou Medical University;the Third Affiliated Hospital of Jinzhou Medical University;the First Clinical Medical School,Jinzhou Medical University,Jinzhou 121000 China)

机构地区：[1]锦州医科大学人体解剖学教研室 [2]辽宁省糖尿病感知功能障碍重点实验室 [3]锦州医科大学附属第三医院 [4]锦州医科大学第一临床医学院,辽宁锦州121000

出　　处：《锦州医科大学学报》2024年第6期14-21,30,共9页Journal of Jinzhou Medical University

基　　金：辽宁省教育厅基金项目,项目编号:LJKMZ20221241;辽宁省自然科学基金项目,项目编号:2023-MS-312。

摘　　要：目的探讨红景天苷(salidroside,SAL)对高糖条件下HT22海马区神经元细胞凋亡的影响,并探讨NF-κB信号通路参与其中的机制。方法将小鼠HT22海马区神经元置于含10%胎牛血清(fetal bovine serum,FBS)和1%青-链霉素双抗的DME/F12培养基中,放置于37℃、5%CO_(2)条件的培养箱中进行体外培养。不同浓度葡萄糖(25、50、100 mM)以及不同浓度SAL(37.5、75、150、300μM)作用海马区神经元细胞24、48、72 h。增强型细胞活力(cell counting kit-8,CCK-8)试剂盒检测HT22细胞增殖活力,以筛选适宜高糖及SAL药物浓度;流式细胞仪检测HT22海马区神经元细胞凋亡率;Hoechst 33342染色检测HT22海马区神经元细胞凋亡情况;活性氧试剂盒检测各组细胞活性氧(reactive oxygen species,ROS)含量的变化;JC-1线粒体膜电位试剂盒(JC-1 mitochondrial membrane potential assay kit,JC-1)检测线粒体膜电位变化;Western Blot法分析各组细胞内磷酸化NF-κB p65(p-p65)、cleaved Caspase-3、Bax和Bcl-2蛋白的表达。结果通过CCK-8实验确定高糖浓度为50 mM,红景天苷浓度为300μM进行后续实验。与对照组相比,高糖诱导HT22海马区神经元细胞48 h后,细胞增殖活力明显下降(P<0.01),细胞凋亡率明显增加(P<0.01),细胞内ROS含量明显增加(P<0.01),线粒体膜电位明显下降(P<0.01),p-p65、cleaved Caspase-3及Bax蛋白表达增加(P<0.01),Bcl-2蛋白表达明显降低(P<0.01),与高糖组相比,高糖+SAL组可以明显逆转以上结果。结论SAL可抑制高糖条件下HT22细胞凋亡,其机制可能与抑制氧化应激、抑制NF-κB p65磷酸化、恢复线粒体膜电位、上调Bcl-2蛋白表达、下调Bax及cleaved Caspase-3蛋白表达有关。Objective To investigate the effect of salidroside(SAL)on the apoptosis of neurons in the hippocampal area of HT22 under high glucose conditions and to explore the mechanism of NF-κB signaling pathway involved in it.Methods Mouse HT22 hippocampal area neurons were placed in DME/F12 medium containing 10%fetal bovine serum(FBS)and 1%penicillin-streptomycin double antibody,and placed in an incubator at 37℃and 5%CO_(2) for in vitro culture.HT22 cells were treated with different concentrations of glucose(25,50,100 mM)and SAL(37.5,75,150,300μM)for 24,48,and 72 h.The proliferative activity of HT22 cells was detected by CCK-8 reagent to screen for the appropriate concentrations of glucose and SAL;the apoptotic rate of HT22 cells was detected by flow cytometry;and the apoptotic rate of HT22 cells was detected by Hoechst 33342 staining.Hoechst 33342 staining was used to detect HT22 cell apoptosis;reactive oxygen species kit was used to detect changes in reactive oxygen species(ROS)content of cells in each group;mitochondrial membrane potential kit(JC-1)was used to detect changes in mitochondrial membrane potential;Western Blot was employed to analyze the expression of phosphorylated NF-κB p65(p-p65),cleaved Caspase-3,Bax and Bcl-2 protein within the cells of each group.Results The high glucose concentration was determined as 50 mM and the SAL concentration was 300μM through the CCK-8 assay for subsequent experiments.Compared with the control group,after HT22 hippocampal neuron cells were induced by high glucose for 48 h,the cell proliferation viability decreased significantly(P<0.01),the cell apoptosis rate increased significantly(P<0.01),the intracellular ROS content increased markedly(P<0.01),the mitochondrial membrane potential decreased conspicuously(P<0.01),the expression of p-p65,cleaved Caspase-3,and Bax proteins increased(P<0.01),and the expression of Bcl-2 protein decreased significantly(P<0.01).Compared with the high glucose group,the high glucose+SAL group could reverse the above results significantly.Conc

关 键 词：SAL 糖尿病脑病 HT22 NF-ΚB 凋亡 

分 类 号：R587.2[医药卫生—内分泌]