槲皮素抑制内皮素-1诱导的人脐动脉平滑肌细胞T型钙通道的表达

摘要

AIM : To investigate the effect of quercetin on endothelin - 1 - induced T - type calcium channel ( TCC ) expression in primary cultured human umhilical arterial smooth muscle cells for exploring the protective role of quercetin in cardiovascular system.METHODS : Human umbilical arterial smooth muscle cells were verified by immunocytochemistry.The cells in 2 - 3 passages were used and randomly divided into control group, quercetin alone group, model group and experimental group.The cells in control group were cultured without any drugs for 24 h.The cells in quercetin alone group were cultured with 80 μmol/L quercetin for 24 h.The cells in model group were cultured with ET - 1 at the concentration of 100 nmol/L for 24 h.The cells in experimental groups were pretreated with quercetin for 1 h, then coincubated with 100 nmol/L ET - 1 for 24 h.The concentrations of quercetin used in this study were 20 , 40and 80 μmol/L, respectively.The expression of α1G , a TCC major subunit, was assayed at mRNA and protein levels by RT - PCR and Western hlotting, respectively.The TCC currents( IcaT ) were detected by the technique of whole - cell patch - clamp.RESULTS : Compared with control and experimental group, ICaT density ( P ＜ 0.01 ) and the expression of α1G at mRNA ( P ＜0.05 ) and protein ( P ＜ 0.01 ) levels in model group were significantly increased.No significant difference in the results of quercetin alone group and control group was observed.CONCLUSION : The protective roles of quercetin in cardiovascular functions are related to the depressive effects of quercetin on ET - 1 - induced increase in both ICaT density and the expression of α1G at mRNA and protein levels in cultured human vascular smooth muscle cells.%目的:通过观察原代培养的人脐动脉平滑肌细胞在内皮素(ET-1)作用下对T型钙通道(TCC)的影响,进一步探讨槲皮素(Que)对心血管的保护作用.方法:原代培养人脐动脉平滑肌细胞,经鉴定于2-3代用于实验.将细胞随机分成对照组、Que组、模型组和实验组.对照组:不加入任何药物;Que组:加入Que 80 μmol/L培养24 h模型组:加入100 nmol/L ET-1培养24 h;实验组:加入Que培养1 h后,再加入100 nmol/L ET-1共同培养24 h,其中Que的浓度为20、40、80 μmol/L.采用RT-PCR和Western blotting检测TCC的主要亚基α1G在mRNA和蛋白水平的表达.利用全细胞膜片钳技术,检测TCC电流(ICaT).结果:模型组α1G mRNA和蛋白的表达均强于对照组和实验组(P＜0.05),模型组ICaT密度明显大于对照组和实验组(P＜0.01),而对照组和Que组的实验结果无明显差别(P＞0.05).结论:ET-1诱导人血管平滑肌细胞中TCC的表达和ICaT的增强,Que能抑制这种增强效应.这可能是Que发挥保护心血管功能的机制之一.