牡荆素调控PI3K/Akt通路对IL-1β诱导的关节软骨细胞凋亡的影响  

Effect of vitexin on IL-1β-induced apoptosis of articular chondrocytes by regulating the PI3K/Akt pathway

作　　者：罗鹏 吴钒 陈佳伟 万震宇 LUO Peng;WU Fan;CHEN Jia-wei;WAN Zhen-yu(Department of Orthopedics,Hubei Hospital of Integrated Traditional Chinese and Western Medicine,Wuhan,Hubei Province 430000,China)

机构地区：[1]湖北省中西医结合医院骨科,湖北武汉430000

出　　处：《解剖学研究》2024年第5期451-455,468,共6页Anatomy Research

摘　　要：目的探讨牡荆素对白介素-1β(IL-1β)诱导的关节软骨细胞凋亡的影响及对磷脂酰肌醇3-激酶(PI3K)/蛋白激酶(AKT)通路的影响。方法体外培养人关节软骨细胞并使用0~200μmol/L的牡荆素处理细胞,筛选最佳浓度;将人关节软骨细胞分为对照组、IL-1β组、牡荆素10μmol/L组、牡荆素50μmol/L组和牡荆素+PI3K抑制剂(LY294002)组,CCK-8法检测各组细胞增殖,流式检测细胞凋亡率,ELISA检测细胞上清液中IL-6和TNF-α的水平,硫代巴比妥酸法测定细胞上清液丙二醛(MDA)的含量,黄嘌呤氧化法测定细胞上清液超氧化物歧化酶(SOD)活性,Western blot法检测细胞中磷酸化(p-)PI3K、PI3K、p-Akt、Akt和剪切的含半胱氨酸的天冬氨酸蛋白水解酶3(C-Caspase-3)蛋白的表达。结果与对照组比较,IL-1β组细胞存活率、SOD活性和p-PI3K/PI3K(0.58±0.06比0.91±0.09)、p-Akt/Akt(0.15±0.03比0.69±0.08)蛋白表达水平降低,细胞凋亡率(38.21%±3.26%比2.88%±0.64%)、IL-6、TNF-α、MDA含量以及C-Caspase-3蛋白表达水平升高(P<0.05);与IL-1β组比较,牡荆素10、50μmol/L组细胞存活率、SOD活性和p-PI3K/PI3K(0.71±0.08、0.87±0.09比0.58±0.06)、p-Akt/Akt(0.32±0.04、0.59±0.07比0.15±0.03)蛋白表达水平逐渐升高,细胞凋亡率(25.36%±2.94%、18.36%±1.95%比38.21%±3.26%)、IL-6、TNF-α、MDA含量以及C-Caspase-3蛋白表达水平逐渐降低(P<0.05);与牡荆素50μmol/L组比较,牡荆素+LY294002组细胞存活率、SOD活性和p-PI3K/PI3K(0.61±0.07比0.87±0.09)、p-Akt/Akt(0.19±0.03比0.59±0.07)蛋白表达水平降低,凋亡率[(26.71%±2.78)%比(18.36%±1.95)%]、IL-6、TNF-α、MDA含量以及C-Caspase-3蛋白表达水平升高(P<0.05)。结论牡荆素可能通过激活PI3K/AKT通路抑制IL-1β诱导的关节软骨细胞的凋亡。Objective To investigate the effect of vitexin on interleukin-1β(IL-1β)-induced apoptosis of articular chondrocytes and its effects on the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(AKT)pathway.Methods Human articular chondrocytes were cultured in vitro and treated with O-200μmol/L vitexin to screen for the optimal concentration;human articular chondrocytes were separated into control group,IL-1β group,10μmol/L vitexin group,50μmol/L vitexin group,and vitexin+PI3K inhibitor(LY294002)group,CCK-8 method was applied to detect cell proliferation in each group,flow cytometry was applied to detect cell apoptosis rate,ELISA was applied to detect the levels of IL-6 and tumor necrosis factor-α(TNF-α)in cell supernatant,thiobarbituric acid method was applied to determine the content of malondialdehyde(MDA)in cell supernatant,xanthine oxidation method was applied to determine the activity of superoxide dismutase(SOD)in cell supernatant,Western blot method was applied to detect the expression of phosphorylated(p-)PI3K,PI3K,p-Akt,Akt,and cleaved cysteine containing aspartate proteolytic enzyme 3(C-Caspase-3)proteins in cells.Results Compared with the control group,the cell survival rate,SOD activity,and the expression levels of p-PI3K/PI3K(0.58±0.06 vs 0.91±0.09)and p-Akt/Akt(0.15±0.03 vs 0.69±0.08)proteins decreased in IL-1β group,the apoptosis rate(38.21%±3.26% vs 2.88%±0.64%),IL-6,TNF-α,MDA contents,and the expression level of C-Caspase-3 protein increased(P<0.05);compared with the IL-1β group,the cell survival rate,SOD activity,and the expression levels of p-PI3K/PI3K(O.71±0.08,0.87±0.09 vs 0.58±0.06)and p-Akt/Akt(0.32±0.04,0.59±0.07 vs 0.15±0.03)proteins in the 10 and 50μmol/L vitexin groups gradually increased,the apoptosis rate(25.36%±2.94%,18.36%±1.95% vs 38.21%±3.26%),IL-6,TNF-α,MDA contents,and the expression level of C-Caspase-3 protein gradually decreased(P<0.05);compared with the 50μmol/L vitexin group,the cell survival rate,SOD activity,and the expression levels of p-PI3K/P

关 键 词：牡荆素 骨关节炎 关节软骨细胞 凋亡 磷脂酰肌醇3-激酶/蛋白激酶 

分 类 号：R285[医药卫生—中药学]