甘木通总黄酮对氧化型低密度脂蛋白诱导血管上皮细胞凋亡的影响及其相关通路分析  

Effect of TFC on apoptosis of vascular epithelial cells induced by ox-LDL and its relevant pathway

作　　者：王莹[1] 王丽[1] 黄宇涵 李晓[2] Wang Ying;Wang Li;Huang Yuhan;Li Xiao(Department of Pharmacy,Shangqiu Medical College,Shangqiu 476QQQ,Henan Province,China)

机构地区：[1]商丘医学高等专科学校药学教研室,476000 [2]河南省生物技术开发中心

出　　处：《中华老年心脑血管病杂志》2021年第6期637-641,共5页Chinese Journal of Geriatric Heart,Brain and Vessel Diseases

基　　金：河南省高等学校重点科研项目(17B360006)。

摘　　要：目的探讨甘木通总黄酮(total flavonoids of clematis filamentosa,TFC)对氧化型低密度脂蛋白(oxidized low-density lipoprotein,ox-LDL)诱导血管上皮细胞凋亡的影响。方法将人脐静脉内皮细胞(HUVEC)随机分为对照组、ox-LDL组、低剂量组(TFC 25μg/ml)、中剂量组(TFC 50μg/ml)、高剂量组(TFC 100μg/ml)。检测细胞存活率和细胞凋亡率,采用二氢乙啶-活性氧荧光探针法检测活性氧水平,采用酶联免疫吸附法检测超氧化物歧化酶(SOD)、丙二醛和一氧化氮(NO)水平,蛋白印迹分析法检测Bax、半胱氨酸天冬氨酸蛋白酶3(Caspase-3)、Bcl-2、磷酸化磷脂酰肌醇3激酶(p-PI3K)、磷酸化蛋白激酶B(p-Akt)和沉默信息调节因子1(SIRT1)表达。结果与对照组比较,ox-LDL组HUVEC存活率、SOD、NO、p-PI3K、p-Akt、Bcl-2及SIRT1蛋白表达水平显著降低,HUVEC凋亡率、Bax、Caspase-3蛋白表达、活性氧及丙二醛水平显著升高(P<0.05)。与ox-LDL组比较,低、中、高剂量组HUVEC存活率依次升高[(68.55±1.35)%、(76.63±2.34)%、(87.01±2.32)%vs(53.37±2.19)%,P<0.05],HUVEC凋亡率依次降低[(16.08±0.43)%、(9.96±0.30)%、(3.77±0.26)%vs(23.49±0.87)%,P<0.05]。与ox-LDL组比较,低、中、高剂量组细胞内Bax、Caspase-3蛋白表达、活性氧及丙二醛水平水平依次降低,Bcl-2蛋白、SOD、NO、SIRT1蛋白、p-PI3K和p-Akt水平依次升高(P<0.05)。结论TFC能够抑制ox-LDL诱导的HUVEC凋亡,可能与激活PI3K/Akt/SIRT1通路相关。Objective To study the effect of total flavonoid from Clematis filamentosa(TFC)on apoptosis of vascular epithelial cells induced by ox-LDL.Methods Human umbilical vein endothelial cells(HUVEC)were randomly divided into control group,ox-LDL group,ox-LDL+TFC group,low TFC dose group,medium TFC dose group and high TFC dose group.The survival rate of HUVEC was detected by methyl thiazolyl tetrazolium assay.The apoptosis rate of HUVEC was detected by annexin V-isothiocyanate fluorescein/propidium iodide assay.The level of reactive oxygen species in HUVEC was detected with a dihydroacetamide reactive oxygen species fluorescence probe.The serum levels of superoxide dismutase(SOD),malondialdehyde(MDA)and nitric oxide(NO)in HUVEC were measured by ELISA.The expressions of Bax,cysteinine,Caspase 3,Bcl-2,p-PI3 K,p-Akt and SIRT1 in HUVEC were detected by Western blot.Results The survival rate of HUVEC and expression levels of SOD,NO,p-PI3 K,p-Akt,Bcl-2 and SIRT1 were significantly lower while the apoptosis rate of HUVEC and expression levels of Bax,Caspase 3,active oxygen and MDA were significantly higher in ox-LDL group than in control group(P<0.05).The survival rate of HUVEC was significantly higher while the apoptosis rate of HUVEC was significantly lower in low,medium and high TFC dose groups than in ox-LDL group(68.55%±1.35%,76.63%±2.34%,87.01%±2.32%vs 53.37%±2.19%,P<0.05;16.08%±0.43%,9.96%±0.30%,3.77%±0.26%vs 23.49%±0.87%,P<0.05).The expression levels of Bax,Caspase 3,active oxygen and MDA were significantly lower while those of Bcl-2,SOD,NO,SIRT1,p-PI3 K and p-Akt were significantly higher in low,medium and high TFC dose groups than in ox-LDL group(P<0.05).Conclusion TFC inhibits the ox-LDL-induced apoptosis of HUVEC by activating the PI3 K/Akt/SIRT1 pathway.

关 键 词：脂蛋白类 LDL 动脉粥样硬化 超氧化物歧化酶 半胱氨酸天冬氨酸蛋白酶3 内皮细胞 细胞凋亡 黄酮 

分 类 号：R285.5[医药卫生—中药学]