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目的:观察蟾蜍灵对大鼠系膜细胞(mesangial cell,MC)p21表达的影响,探讨其调控系膜细胞增殖的机制。方法:MTT法观察不同浓度蟾蜍灵(0.01、0.04、0.08、0.16μmol/L)对PDGF-BB(20 ng/ml)诱导的MC增殖的影响;流式细胞术测定MC细胞周期的变化;RT-PCR法测定p21基因表达的改变;Western blot法测定p21蛋白水平变化。结果:PDGF-BB刺激MC12 h后,加入不同浓度蟾蜍灵干预24 h,蟾蜍灵从0.04μmol/L至0.16μmol/L的呈浓度依赖抑制PDGF-BB诱导的MC增殖(P<0.05)。0.08μmol/L蟾蜍灵干预24 h后,可使PDGF-BB诱导的MC G0/G1期细胞比例升高,使S期细胞比例下降,且上调P21mRNA和蛋白水平的表达(P<0.05)。0.08μmol/L蟾蜍灵对正常系膜细胞无明显影响(P>0.0.5)。结论:蟾蜍灵可能通过上调细胞周期激酶抑制剂p21的表达来抑制PDGF-BB诱导的MC的增殖。
Objective: To observe the effect of bufalin on the expression of p21 in rat mesangial cell (MC), and to explore its mechanism of regulating the proliferation of mesangial cells. Methods MTT assay was used to observe the effects of different concentrations of bufalin on MC proliferation induced by PDGF-BB (20 ng / ml). Flow cytometry was used to determine the changes of MC cell cycle. RT The expression of p21 gene was determined by PCR and the p21 protein level was determined by Western blot. RESULTS: After exposure to PDGF-BB for 12 h, MCF-induced MC proliferation induced by PDGF-BB was inhibited by bufalin from 0.04 μmol / L to 0.16 μmol / L for 24 h after MC12 h stimulation (P <0.05). After treated with 0.08μmol / L bufalin for 24 h, the proportion of cells in MC G0 / G1 phase induced by PDGF-BB increased, the proportion of S phase cells decreased, and the expression of P21 mRNA and protein increased (P <0.05). 0.08μmol / L of bufalin had no significant effect on normal mesangial cells (P> 0.0.5). Conclusion: Bufalin may inhibit the proliferation of MC induced by PDGF-BB by up-regulating the expression of cell-cycle kinase inhibitor p21.