MiR-30c-2-3p对足细胞活力及Nephrin和Podocin蛋白表达的影响

Effects of MiR-30c-2-3p on podocyte viability and expression of Nephrin and Podocin

目的探讨MiR-30c-2-3p对足细胞活力及Nephrin和Podocin蛋白的表达影响。方法采用嘌呤霉素(PAN)诱导的足细胞体外模型,将足细胞分成4组,即空白组、PAN组、阴性质粒+PAN组、阳性质粒+PAN组。通过LipofectamineTM2000瞬时转染MiR-30c-2-3p mimic。通过Cell Counting Kit-8(CCK-8)法检测细胞活力、实时荧光定量PCR(qRT-PCR)检测细胞中MiR-30c-2-3p表达、Western blot检测Nephrin和Podocin蛋白的表达。结果(1)CCK-8结果提示PAN组足细胞活力明显低于空白组,差异有高度统计学意义(P<0.01),阴性质粒+PAN组较PAN组差异无统计学意义(P>0.05),阳性质粒+PAN组足细胞活力与PAN组和阴性质粒+PAN组相比均有所提高,差异有统计学意义(P<0.05或P<0.01)。(2)qRT-PCR显示PAN组MiR-30c-2-3p的表达较空白组明显下降,差异有高度统计学意义(P<0.01);阴性质粒+PAN组较PAN组表达差异无统计学意义(P>0.05),而阳性质粒+PAN组MiR-30c-2-3p的表达较PAN组和阴性质粒+PAN组均提高,差异有统计学意义(P<0.05或P<0.01)。(3)Western blot提示PAN组Nephrin和Podocin蛋白的表达较空白组明显下降,差异有高度统计学意义(P<0.01);与PAN组相比,阴性质粒+PAN组两者蛋白的表达差异无统计学意义(P>0.05),而阳性质粒+PAN组Nephrin和Podocin蛋白的表达较单纯PAN组和阴性质粒+PAN组均增加,差异有统计学意义(P<0.05)。结论 PAN对足细胞的活力,MiR-30c-2-3p含量及Nephrin和Podocin蛋白的表达有抑制作用,而通过外源性增加MiR-30c-2-3p表达后可提高足细胞的活力,同时可增加Nephrin和Podocin蛋白的表达。

Objective To investigate the effect of MiR＇30c-2-3p on podocyte viability and Nephrin and Podocin protein expression. Methods The model of mice podocyte injury was induced by puromycin aminonucleoside （PAN） in vitro, the podocytes were divided into four groups： blank group, PAN group, negative plasmids＋PAN group, positive plasmids＋PAN group. The MiR-30c-2-3p mimics were transiently transfected with LipofectamineTM 2000, the cell viability was detected by Cell Counting Kit-8 （CCK-8）, the expression of MiR-30c-2-3p was detected by real-time quantitative PCR （qRT-PCR）, the expression of Nephrin and Podocin protein was detected by Western blot. Results ①CCK-8 showed that the viability of PAN group was significantly lower than that of the blank group, the differences were statistically significant （P 〈 0.01）. There was nO significant difference between the negative plasmid＋PAN group and the PAN group （P 〉 0.05）. The viability of positive plasmid＋PAN group was significantly increased than that of the PAN group and the negative plasmid＋PAN group, the differences were statistically significant （P 〈 0.05 or P 〈 0.01）. ②The expression of MiR-30c-2-3p in the PAN group was significantly lower than that in the blank group, the differences were statistically significant （P 〈 0.01）. There was no significant difference between the negative plasmid group and the PAN group （P 〉 0.05）. The content of MiR-30c-2-3p in the positive plasmid＋PAN group was significantly higher than that of the PAN group and the negative plasm id＋PAN group, the differences were statistically significant （P 〈.0.05 or P 〈 0.01）. ③The expression of Nephrin and Podocin in the PAN group were significantly lower than those in the blank group, the differences were statistically significant （P 〈 0.01）. There was no significant difference between the negative plasmid＋PAN group and the PAN group （P 〉 0.05）. Compared with the PAN group and the negative plasmid＋PAN group, the expression of Nephrin and Podocin in the positive plasmid＋PAN group increased, the differences were statistically significant （P 〈 0.05）. Conclusion The PAN intervention inhibits the podocyte activity, MiR-30c-2-3p content, and the Nephrin and Podocin expression, but the exogenous expression of MiR-30c-2-3p can improve the viability of podocytes and increase the expression of Nephrin and Podocin protein.