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目的观察嘌呤霉素(PAN)诱导和地塞米松(DEX)干预后足细胞凋亡率及凋亡相关蛋白Bad表达的变化。方法体外培养小鼠足细胞,将其分为对照组、PAN组和DEX组。对照组用含二甲基亚砜的RPM I-1640培养液培养;PAN组加入PAN;DEX组同时加入PAN和DEX。处理8 h、24 h和48 h后,用实时荧光定量聚合酶链式反应及W estern blot分别检测各时间点Bad mRNA和蛋白的表达;用Annexin V-FITC/PI双染色法结合流式细胞仪检测足细胞凋亡率的变化。结果 1.PAN组各时间点Bad mRNA的表达均较对照组升高,并呈上升趋势,24 h和48 h时均显著升高(Pa<0.01);8 h时Bad蛋白表达与对照组比较无明显差异(P>0.05),24 h和48 h时均较对照组明显升高(Pa<0.05);各时间点足细胞凋亡率均较对照组升高,48 h时显著升高(P<0.01)。2.DEX组Bad mRNA和蛋白的表达8 h时与PAN组比较无明显差异,但24 h时明显降低(P<0.05),各时间点足细胞凋亡率均较PAN组降低,24 h时显著降低(P<0.01)。结论 PAN诱导足细胞后其凋亡率明显升高,可能与Bad mRNA及蛋白的表达增高有关,而DEX可能通过降低Bad mRNA及蛋白的表达减少足细胞的凋亡而抑制PAN的诱导,起到保护足细胞的作用。
Objective To observe the changes of podocyte apoptosis rate and apoptosis-related protein Bad expression induced by puromycin (PAN) and dexamethasone (DEX). Methods Mouse podocytes were cultured in vitro and divided into control group, PAN group and DEX group. The control group was cultured in RPM I-1640 medium containing dimethyl sulfoxide; the PAN group was added to PAN; the DEX group was added with both PAN and DEX. After treatment for 8, 24 and 48 h, the expression of Bad mRNA and protein were detected by real-time fluorescence quantitative polymerase chain reaction and Western blot respectively. Flow cytometry was performed with Annexin V-FITC / PI double staining The change of podocyte apoptosis rate was detected by the instrument. The expression of Bad mRNA in the PAN group was higher than that in the control group at each time point, and was increased at 24 and 48 h (P <0.01). Compared with the control group, Bad mRNA expression at 8 h (P <0.05). Compared with the control group, the apoptosis rate of podocytes at 24 h and 48 h was significantly higher than that of the control group (P <0.05) P <0.01). Bad expression of Bad mRNA and protein in DEX group had no significant difference compared with PAN group at 8 h, but decreased significantly at 24 h (P <0.05), and podocyte apoptosis rate at each time point was lower than that of PAN group. At 24 h Significantly lower (P <0.01). Conclusions The apoptosis rate of podocytes induced by PAN is obviously increased, which may be related to the increase of Bad mRNA and protein expression. However, DEX may inhibit the induction of PAN by decreasing the expression of Bad mRNA and protein, thereby decreasing the podocyte apoptosis Protect podocytes.