Objective To observe the effect of angiotensin Ⅱ(Ang Ⅱ) on the expression of CTGF mRNA in cultured HPTC.Methods The cell line of HPTC were cultured in Dulbeccos modified Eagle medium (DMEM) containing 10% fetal calf serum.After incubation with serum free DMEM for 24 h,cells were incubated in absence or prescence of various concentrations(0,10 -9,10 -7,10 -5 mol·L -1) of Ang Ⅱ for 48 h,or incubated with Ang Ⅱ(10 -7 mol·L -1) for different period (12?h,24?h,48?h,72?h).Total RNA was isolated from cultured HPTC by TRIZOL method.RT?PCR was performed using access RT?PCR kit.The reaction products were measured by 2% agarose gel electrophoresis.All experiments were performed in triplicate.Results CTGF mRNA expression was elevated 12 h after cells exposed to 10 -7?mol·L -1 Ang Ⅱ and reached its peak level at the end of 72 h.Compared to control group,it increased to 6.3 folds after 48 h incubation(0.097±0.015 vs 0.631?7±0.041,P<0.01).When different concentration(0,10 -9,10 -7,10 -5mol·L -1) of Ang Ⅱ were added to the medium for 48 h,CTGF mRNA expression increased 1.5, 5.5 and 7.4 folds respectively (P<0.01).The consequence analysis for the PCR products demonstrated the same as human CTGF cDNA in gene bank.Conclusion Ang Ⅱ induced a time-and dose-dependent expression of CTGF mRNA in HPTC,which implied a profibrotic role of Ang Ⅱ in mediating tubulointerstitial fibrosis possibly through stimulating the production of CTGF. |
