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目的观察缬沙坦对尾加压素Ⅱ(urotensinⅡ,UⅡ)对肾小管上皮细胞凋亡的影响。方法将体外培养人肾小管上皮细胞(human kidney proximal tubular cell,HK-2)分为正常对照组、10~(-8)mol/L UⅡ刺激组和10~(-8)mol/L UⅡ+10μmol/L缬沙坦组。采用Armexin V-FITC/PI双染法流式细胞术(flow cytometry,FCM)检测细胞凋亡百分比;Western-Blot检测Bax、Bcl-2的表达;荧光实时定量PCR(Real time-PCR)检测Bax、Bcl-2mRNA水平变化。结果①FCM法检测显示,与正常对照组相比,10~(-8)mol/L UⅡ刺激组细胞凋亡百分比显著升高(P<0.01)。10μmol/L缬沙坦能显著降低10~(-8)mol/L UⅡ诱导细胞凋亡的百分比(P<0.01)。②Western-Blot显示,10~(-8)mol/L UⅡ刺激组,与正常对照组相比,Bcl-2表达显著降低(P<0.05),Bax蛋白表达无明显变化(P>0.05),Bax/Bcl-2比率明显升高(P<0.01)。与UⅡ刺激组相比,缬沙坦使Bcl-2表达显著升高(P<0.01),Bax蛋白表达无明显变化(P>0.05),Bax/Bcl-2比率明显降低(P<0.01)。③Real time-PCR显示,10~(-8)mol/L UⅡ刺激组细胞Bcl-2mRNA表达显著减少(P<0.01),BaxmRNA无明显变化(P>0.05)。与UⅡ刺激组相比,缬沙坦显著升高Bcl-2mRNA水平(P<0.01),BaxmRNA无明显变化(P>0.05)。结论缬沙坦的肾脏保护作用可能部分是通过抑制UⅡ诱导的肾小管上皮细胞凋亡实现的。
Objective To observe the effect of valsartan on the apoptosis of renal tubular epithelial cells induced by urotensin Ⅱ (UⅡ). Methods Human renal proximal tubular cell (HK-2) was divided into normal control group, 10 -8 mol / L UⅡ stimulation group and 10 -8 mol / L UⅡ + 10μmol / L valsartan group. The percentage of apoptotic cells was detected by flow cytometry (FCM) using Armexin V-FITC / PI double staining method. The expression of Bax and Bcl-2 was detected by Western-Blot and Real-time PCR , Bcl-2 mRNA level changes. Results ① FCM assay showed that the percentage of apoptotic cells in 10 -8 mol / L UⅡ stimulation group was significantly higher than that in normal control group (P <0.01). Valsartan at a concentration of 10μmol / L could significantly decrease the percentage of apoptosis induced by 10 ~ (-8) mol / L UⅡ (P <0.01). ②Western-Blot showed that Bcl-2 expression was significantly decreased (P <0.05), Bax protein expression was not significantly changed (P> 0.05), Bax expression was significantly increased in 10 -8 mol / L UⅡ stimulation group compared with normal control group / Bcl-2 ratio was significantly increased (P <0.01). Valsartan significantly increased Bcl-2 expression (P <0.01), Bax protein expression (P> 0.05), and Bax / Bcl-2 ratio decreased significantly compared with UⅡ stimulation group (P <0.01). ③Real time-PCR showed that the expression of Bcl-2 mRNA was significantly decreased (P <0.01) and Bax mRNA was not significantly changed in 10 -8 mol / L UⅡ stimulation group (P> 0.05). Valsartan significantly increased Bcl-2 mRNA level (P <0.01), while Bax mRNA had no significant change (P> 0.05) compared with UII stimulation group. Conclusion The renal protective effect of valsartan may be partly through the inhibition of UⅡ-induced renal tubular epithelial cell apoptosis.