Effects of rapamycin on puromycin aminonucleoside induced podocyte injury

Abstract

Objective To study the protective effect of mpamycin on puromycin aminonucleoside (PAN) induced podocyte injury and its possible mechanisms.Methods (1) The synchronized podocytes were divided into four groups:the control group (RPMI 1640 culture medium without serum) and the 20,50,100 mg/L PAN group (a final concentration of 20,50,100 mg/L PAN RPMI 1640 culture medium without serum).The cells were collected after 24 h before Western blot assay was performed to assess relative expression of Nephrin protein.(2) The synchronization podocytes were assigned to the model group and 10,100,1 000μg/L mpamycin group.All the cells were added to a final concentration of 50 mg/L PAN RPMI 1640 serum-free culture medium,incubated for 24 h.The model control group was added to dissolve the same amount of mpamycin solution,and the remaining three groups were added to a final concentration of 10,100,1 000μg/L rapamycin training.After 24 and 48 h,Western blot assay was used to test the relative expression of Nephrin protein.(3) The synchronization podocytes were assigned to thee normal control group (serum-free RPMI 1640 culture solution),model control group (the final concentration of 50 mg/L PAN + serum-free RPMI 1640 culture hquid) and mpamycin group (final concentration of 100 μg/L rapamycin + 50 mg/L PAN + serum free RPMI 1640 medium).The cells were cultured for 24 and 48 h,and their morphology was observed.Western blot was used to detect p-P70S6K relative expression.Results (1) Compared with the control group,Nephrin protein expression was significantly decreased in 50 or 100 mg/L PAN group (P ＜ 0.05).No significant difference in nephrin protein expression was found between 50 or 100 mg/L PAN group.(2) Compared with 10 or 1 000μg/L group,Nephrin protein expression was significantly increased in 100 μg/L mpamycin group (P ＜ 0.05).(3) Compared with the normal control group,Nephrin protein expression did not significantly change in the PAN or mpamycin group.Conclusions 50 mg/L PAN could damage the optimal concentration of PAN in vitro cell model.100 μg/L mpamycinon may protect podocytes from PAN induced injury,which may contribute to the inhibition of mTOR. Key words: Rapamycin; Podocytes; p-p70S6K; Nephrin