氧化型低密度脂蛋白诱导肝窦内皮细胞植物血凝素样氧化型低密度脂蛋白受体1的表达

摘要

目的：探讨植物血凝素样氧化型低密度脂蛋白（ox-LDL）受体1（LOX-1）在肝窦内皮细胞（HSECs）中的表达和ox-LDL对其表达的调控作用。方法使用实时定量聚合酶链反应（RT-PCR）和Western blotting法从基因和蛋白水平检测未经处理HSECs 中LOX-1的表达。应用不同浓度ox-LDL（0、20、40、60、80和100 mg/L）对HSECs作用24 h并应用80 mg/L ox-LDL对HSECs作用不同时间（0、12、24和48 h），作用后实时定量PCR检测HSECs内LOX-1 mRNA的表达水平，Western blotting法检测细胞内LOX-1蛋白表达。给予80 mg/L ox-LDL干预组多聚肌苷酸250 mg/L作用24 h后，测定LOX-1 mRNA和蛋白的表达。采用单因素方差分析及t检验进行数据分析。结果 LOX-1 mRNA和蛋白在HSECs中均有表达。20～80 mg/L ox-LDL组HSECs中LOX-1 mRNA、蛋白表达水平随ox-LDL剂量增加而升高，与剂量有明显相关性（F＝38.7、3.43，均P＜0.05）。与80 mg/L ox-LDL组相比，100 mg/L ox-LDL 组 LOX-1 mRNA、蛋白表达下降，差异有统计学意义（ t ＝23.75、18.26， P ＜0.05）。80 mg/L ox-LDL对HSECs作用时间在0～24 h时，随着时间延长，LOX-1 mRNA、蛋白表达递增，与ox-LDL作用时间有明显相关性（F＝2.36、0.33，均P＜0.05）。与作用24 h相比，作用48 h组HSECs中LOX-1 mRNA、蛋白表达下降（t＝69.21、36.27，均P＜0.05）。与80 mg/L ox-LDL组相比，多聚肌苷酸组中LOX-1 mRNA和蛋白表达降低，两组差异均有统计学意义（ t＝54.93、28.19，均P＜0.05）。结论LOX-1存在于HSECs。在一定浓度和时间范围内，ox-LDL对HSECs LOX-1的调控作用具有时间和浓度依赖性，而多聚肌苷酸可部分抑制这种效应。%Objective To investigate the presence of lectin-like oxidized low-density lipoprotein (ox-LDL) receptor-1 (LOX-1) in cultured hepatic sinusoidal endothelial cells (HSECs), and meanwhile discuss the regulation of its mechanisms under ox-LDL injury conditions.Methods The mRNA and protein expression of LOX-1 was testified by real-time quantitative polymerase chain reaction ( RT-PCR) and Western blotting methods.Meanwhile HSECs were stimulated with different concentration of ox-LDL (0, 20, 40, 60, 80 and 100 mg/L) for 24 h, and HSECs were stimulated with 80 mg/L for different times (0, 12, 24 and 48 h).Then expression of LOX-1 mRNA was measured by RT-PCR, and the protein expression of LOX-1 was testified by using Western blotting.Data were analyzed with one-way ANOVA and t test.Results LOX-1 exists in HSECs.After intervened with 20-80 mg/L ox-LDL for 24 h, LOX-1 mRNA and protein levels increased significantly in a dose-dependent manner (F=38.7, 3.48, both P <0.05).However, when concentration of ox-LDL reached 100 mg/L, the LOX-1 mRNA and protein expression decreased significantly compared with those intervened with 80 mg/L ox-LDL ( t =23.75,18.26, both P<0.05 ).Meanwhile, after intervened with 80 mg/L ox-LDL at different time (0-24 h), the expression of LOX-1 mRNA and protein significantly increased in a time-manner ( F =2.36, 0.328, both P <0.05 ).And compared with those in cells intervened for 24 h, the mRNA and protein expression of LOX-1 decreased significantly when intervened for 48 h(t=69.21, 36.27, both P<0.05).However, compared with the cells incubated with 80 mg/L ox-LDL, the mRNA and protein expression of LOX-1 significantly decreased in HSECs treated with polyinosinic cytidylic acid ( t=54.93, 28.19, both P<0.05 ).Conclusion LOX-1 exists in HSECs and ox-LDL could regulate it in a certain reach of concentration and time.However , polyinosinic cytidylic acid can partly restrained ox-LDL-induced LOX-1 expression.