脂多糖调节SH-SY5Y细胞线粒体融合在阿尔茨海默病中的作用  

The role of lipopolysaccharide in regulating mitochondrial fusion of SH-SY5Y cells in Alzheimer's disease

作　　者：王睿 尹亚萍 李国平 马焜 靖雪妍[1] 陈丽丽[1] WANG Rui;YIN Ya-ping;LI Guo-ping(School of Public Health,Harbin Medical University,Harbin 150081,China)

机构地区：[1]哈尔滨医科大学公共卫生学院,150081

出　　处：《中国实用医药》2024年第9期172-177,共6页China Practical Medicine

基　　金：中国博士后面上基金资助(项目编号:2019M661310);黑龙江省博士后基金资助(项目编号:LBH-Z18225)。

摘　　要：目的 探讨脂多糖(LPS)调节人神经母细胞瘤细胞(SH-SY5Y细胞)线粒体融合在阿尔茨海默病(AD)中的作用。方法 选取SH-SY5Y细胞株进行常规培养,于细胞对数生长期以25、50、100μg/ml的LPS处理SH-SY5Y细胞48 h,分别设置为25μg/ml组、50μg/ml组、75μg/ml组,同时设置空白对照组。采用细胞计数试剂(CCK-8)法检测细胞存活率;光镜检测细胞内线粒体损伤情况;酶联免疫吸附测定法(ELISA)检测超氧化物歧化酶(SOD)、丙二醛(MDA)、一氧化氮(NO)和肿瘤坏死因子-α(TNF-α)变化;采用实时荧光定量PCR(Real-time PCR)法检测动力相关蛋白1 (Drp1)、线粒体融合蛋白1(Mfn1)、线粒体融合蛋白2(Mfn2)和视神经萎缩因子1(Opa1)微小核糖核酸(mRNA)表达和蛋白表达水平。分析LPS对SH-SY5Y细胞存活率和β-淀粉样蛋白(Aβ)含量, SH-SY5Y细胞线粒体形态, SH-SY5Y细胞内SOD、MDA、NO和TNF-α, SH-SY5Y细胞内mRNA表达的影响。结果 CCK-8法检测结果显示,与空白对照组相比,在48 h给药时间下, 25、50、100μg/ml组中SH-SY5Y细胞存活率明显下降, Aβ含量显著升高;随着LPS剂量的增加, SH-SY5Y细胞存活率呈剂量依赖性降低, Aβ含量呈剂量依赖性增长,差异具有统计学意义(P<0.05)。光镜结果显示,与空白对照组相比,在48 h给药时间下, 25、50、100μg/ml组中SH-SY5Y细胞内线粒体出现明显变大变圆,体积增大,线粒体出现肿胀和线粒体空泡化等改变,并伴有线粒体自噬的发生。ELISA结果显示,与空白对照组相比,在48 h给药时间下, 25、50、100μg/ml组中SH-SY5Y细胞内SOD表达分别显著降低24.16%、69.08%和123.78%, MDA表达分别显著增加37.65%、52.13%和101.85%, NO表达分别显著增加54.34%、119.08%和167.78%, TNF-α表达分别显著增加54.34%、119.08%和167.78%,随着LPS剂量的增加, SOD含量变化具有剂量依赖性降低, NO、MDA、TNF-α含量变化呈剂量依赖性增长,差异具有统计学意义(P<0.05)。Real-Time PCR结果显�Objective To investigate the role of lipopolysaccharide(LPS)in regulating mitochondrial fusion of Human neuroblastoma(SH-SY5Y)cells in Alzheimer's disease(AD).Methods SH-SY5Y cell line was cultured routinely.During the logarithmic phase,SH-SY5Y cells were treated with 25,50 and 100μg/ml LPS for 48 h,and were set as 25μg/ml group,50μg/ml group and 75μg/ml group,and a blank control group was set.The cell survival rate was detected by cell counting kit-8(CCK-8)method,the mitochondrial damage was detected by light microscope,the changes of superoxide dismutase(SOD),malondialdehyde(MDA),nitric oxide(NO),and tumor necrosis factor-α(TNF-α)were detected by enzyme linked immunosorbent assay(ELISA),and the expression and protein expression of dynamin-related protein 1(Drp1),mitofusin 1(Mfn1),mitofusin 2(Mfn2)and optic atrophy 1(Opa1)micro ribonucleic acid(mRNA)were detected by real-time PCR method.The effects of LPS on survival andβ-amyloid protein(Aβ)content of SH-SY5Y cells,mitochondrial morphology of SH-SY5Y cells, intracellular SOD, MDA, NO and TNF-α in SH-SY5Y cells, and intracellular mRNA expression in SH-SY5Y cells were analyzed. Results The results of CCK-8 assay showed that compared with the blank control group, the survival rate of SH-SY5Y cells in 25, 50 and 100 μg/ml groups decreased significantly and the Aβ content increased significantly at 48 h of administration. With the increase of LPS dose, the survival rate of SH-SY5Y cells decreased in a dose-dependent manner, and the Aβ content increased in a dose-dependent manner, and the differences were statistically significant (P<0.05). The light microscope results showed that compared with the blank control group, mitochondria in SH-SY5Y cells in the 25, 50 and 100 μg/ml groups showed significant enlargement and roundness, increased volume, swelling and vacuolization, accompanied by the occurrence of mitochondrial autophagy at 48 h of administration. ELISA results showed that compared with the blank control group, SOD expression in SH-SY5Y cells in 2

关 键 词：脂多糖 人神经母细胞瘤细胞 线粒体 阿尔茨海默病 

分 类 号：R749.16[医药卫生—神经病学与精神病学]