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目的观察阿米洛利对脂多糖诱导的足细胞活动力的影响,并探讨其机制。方法足细胞分为3组:正常对照组,脂多糖组(50mg·L~(-1)脂多糖诱导损伤足细胞24h),脂多糖+阿米洛利组(50 mg·L~(-1)脂多糖与50 mg·L~(-1)阿米洛利共同作用于足细胞24h)。利用免疫荧光共聚焦、流式细胞术、实时荧光定量PCR、转板迁移实验、划痕实验等技术,检测足细胞uPAR蛋白和Plaur mRNA的表达情况以及足细胞活动力的改变。结果脂多糖组足细胞uPAR蛋白及PlaurmRNA的表达均高于正常对照组和脂多糖+阿米洛利组(P<0.05),正常对照组和脂多糖+阿米洛利组无显著差异(P>0.05)。转板迁移、划痕实验中脂多糖组每个视野足细胞数[(324±15),(43±7)个]高于正常对照组[(249±10),(15±5)个]和脂多糖+阿米洛利组[(274±8),(18±6)个](P<0.01),正常对照组和脂多糖+阿米洛利组无显著差异(P>0.05)。结论阿米洛利可能通过抑制uPAR的表达,降低足细胞活动力,起到稳定、保护足细胞的作用。
Objective To observe the effect of amiloride on lipopolysaccharide-induced podocyte viability and to explore its mechanism. Methods Podocytes were divided into 3 groups: normal control group, lipopolysaccharide group (50 mg · L -1 lipopolysaccharide induced injury podocyte 24h), lipopolysaccharide + amiloride group (50 mg · L -1 ) Lipopolysaccharide and 50 mg · L -1 amiloride act on podocytes 24h). Immunofluorescence confocal, flow cytometry, real-time fluorescence quantitative PCR, transfer plate migration assay, scratch test and other techniques were used to detect the expression of uPAR protein and Plaur mRNA in podocytes and the changes of podocyte viability. Results The expressions of uPAR protein and Plaur mRNA in podocytes were higher than those in normal control group and lipopolysaccharide plus amiloride group (P <0.05), and there was no significant difference between normal control group and lipopolysaccharide plus amiloride group (P > 0.05). The numbers of podocytes in each field of the LPS group were (324 ± 15) and (43 ± 7), respectively, higher than those in the normal control group [(249 ± 10) and (15 ± 5)] (P <0.01). There was no significant difference between normal control group and lipopolysaccharide + amiloride group (P> 0.05). Conclusion Amiloride may inhibit the expression of uPAR, reduce the activity of podocytes, and play a stabilizing role in protecting podocytes.