| Objective: To explore the effects of fluvastatin with or without losartan on the expression of vascular endothelial growth factor (VEGF) and the role of ERK signaling pathway in normal immortalized human podocytes induced by angiotensinⅡ(AngⅡ).Methods: The normal immortalized human podocytes were cultured at 33℃and allowed to differentiate at 37℃for 2 weeks when the podocytes were in exponential phase of growth.The differentiated podocytes were incubated with serum-free medium for 24 hours to synchronize the cell growth. Podocytes were divided into several groups:①Podocytes were incubated with different concentrations of AngⅡ(10-9, 10-8, 10-7, 10-6, 10-5 mol/L) for 24 h.②AngⅡ(10-7 mol/L) was given on different time courses (0 h, 6 h, 12 h and 24 h).③The podocytes were divided into five groups: normal medium, AngⅡ(10-7 mol/L), AngⅡ(10-7 mol/L) plus three different concentrations of fluvastatin (10-7 mol/L, 10-6 mol/L, 10-5 mol/L) respectively. These groups were incubated for 24 h.④The podocytes were divided into five groups: normal medium, AngⅡ(10-7 mol/L), AngⅡ(10-7 mol/L) plus three different concentrations of losartan (10-7 mol/L, 10-6 mol/L, 10-5 mol/L) respectively. These groups were incubated for 24 h.⑤Then podocytes were divided into six groups: normal medium, AngⅡ, AngⅡplus the ERK specific inhibitor PD98059 (5×10-5 mol/L), AngⅡplus fluvastatin (10-6 mol/L), AngⅡplus losartan (10-6 mol/L), AngⅡplus fluvastatin (10-6 mol/L) combined with losartan (10-6mol/L). The concentration of AngⅡwas 10-7mol/L and these groups were incubated for 24 h. The protein expressions of VEGF and p-ERK1/2 in podocytes were detected by western blot.Results: 1. Compared with other concentrations of AngⅡ, the group of 10-7mol/L AngⅡincreased the expressions of VEGF in podocytes most significantly (p<0.05 or p<0.01). 2. Compared with 0h group, AngⅡ(10-7 mol/L) increased the expressions of VEGF and p-ERK1/2 in podocytes in a time-dependent manner (p<0.01). 3. Compared with AngⅡ(10-7mol/L) group, fluvastatin suppressed the expression of VEGF protein obviously, the group of 10-5 mol/L fluvastatin had the most significant effect (p<0.01). 4. Compared with AngⅡ(10-7 mol/L) group, losartan suppressed the expression of VEGF protein obviously, the group of 10-5 mol/L losartan had the most significant effect (p<0.01). 5. Compared with AngⅡ(10-7 mol/L) group, the group of PD98059 suppressed the expressions of VEGF and p-ERK1/2 obviously (p<0.01), the group of fluvastatin (10-6 mol/L) combined with losartan (10-6 mol/L) suppressed the expressions of VEGF and p-ERK1/2 obviously and more siginificantly than the effect of the two respectively (p<0.01).Conclusion: AngⅡincreases the expressions of VEGF and p-ERK1/2 in human podocytes in a time dependent manner. Fluvastatin and losartan can inhibit this effect respectively and they have synergistic inhibitory effects via inhibiting ERK signaling pathway. The combination of fluvastatin and losartan may exert a protective effect on podocyte and then has the effects of prevention and treatment on diabetic nephropathy.
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