原花青素对人肝癌细胞株sEH表达及活性的影响

摘要

目的 观察原花青素(PC)对人肝癌细胞株可溶性环氧化物水解酶(sEH)表达及活性的影响,为研发sEH抑制剂提供依据.方法 取对数生长期人肝癌细胞株Huh7,分别予0.1％的DMSO、10 μmol/L的sEH抑制剂12-(3-金刚烷-脲基)月桂酸衍生物(AUDA)及25 ～ 100 μmol/L的PC处理,培养30 min和24 h后分别采用酶活性测定试剂盒测定细胞内sEH活性;培养后24 h,采用RT-PCR检测sEH mRNA表达,Western blot检测sEH蛋白表达.结果 25～ 100 μmol/L的PC处理后24 h,sEH活性明显受抑,且呈浓度依赖性;100 μmol/L的PC处理后30 min,sEH活性无明显受抑;100 μmol/L的PC处理后24h,sEH mRNA及蛋白表达均明显受抑.结论 PC对sEH的直接活性无明显影响,其可能主要通过抑制sEH表达、降低细胞sEH的酶量发挥作用;本实验为从天然药物中开发sEH抑制剂提供了新线索.%Objective To investigate the effect of procyanidins (PC) on the expression and activity of soluble epoxide hydrolase (sEH) in human hepatocellular carcinoma cell line,which will provide the basis for research and development of sHE inhibitors.Methods The logarithmic phase human hepatocellular carcinoma cell line was cultured in DMEM medium and treated with 0.1％ DMSO,10 μmol/L sEH inhibitor 12-(3-adamantyl-ureido)-dodecanoic acid (AUDA) and PC (25,50,75 and 100 μmol/L) for 30 minutes and 24 hours respectively.The sEH activity in cells was measured by a commercialized kit.After treatment for 24 h,the expression levels of sEH mRNA and protein were detected by RT-PCR and Western blot respectively.Results The sEH activity was inhibited by 25-100 μmol/L PC after being incubated for 24 h with a dose-dependent manner.The sEH activity was not inhibited by 100 μmol/L PC after being incubated for 30 min.The sEH mRNA and protein expression was significantly inhibited by 100 μmol/L PC after being incubated for 24 h.Conclusions PC cannot inhibit sEH activity directly,but it can obviously down-regulate the sEH mRNA and protein expression.Our results suggest that sEH inhibitors may be developed from natural medicinal plant.