| Objective:To investigate the protective effect of pancreatic kininogenase(PK)on renal fibrosis induced by unilateral ureteral obstruction(UUO)in rats.Methods:In this study,21 clean grade male SD rats aged 7 weeks were randomly divided into Sham group,UUO group and unilateral ureteral obstruction group(7.2U/kg/day)were injected with pancreatic kininogenase(UUO+PK group).During the experiment,the body weight(BW)of the rats in each group was measured daily.At the end of the experiment,the urine of each group was collected for 24h,and the 24-hour urinary protein excretion(UPE)was detected by urinalysis.Blood urea nitrogen(BUN)and serum creatinine(Scr)were detected by automatic serum biochemical analyzer.Serum and urine levels of 8-hydroxydeoxyguanosine(8-OHdG)were determined by enzyme-linked immunosorbent assay(ELISA).The changes of renal fibrosis in each group were observed by Masson trichrome staining.The expression of ED-1 and 8-OHdG in renal tissues of each group was observed by immunohistochemistry.The expression of connective tissue growth factor(CTGF),transforming growth factor-?1(TGF-?1),matrix metalloproteinase-2(MMP-2),interleukin-6(IL-6)and Cleaved cysteine aspartate protease 3(Cleaved Caspase-3)were detected by western blot.TUNEL in situ staining was used to observe the apoptosis of renal tissue.Results:1.Visual inspection Compared with Sham group,the kidneys of UUO group were pale in color,larger in size,thinner in renal parenchyma,and disorganized in tissue structure.In section,dilatation of renal pelvis and calices was observed,and the boundaries of medullary atrophy of renal parenchyma were blurred.2.The basic and biochemical indexes Compared with Sham group,UUO group showed no significant changes in BW,UV,BUN,Scr and UPE.There were no significant changes in BW,UV,BUN,Scr and UPE between UUO group and UUO+PK group.3.Enzvme-linked immunosorbent assav of 8-OHdG The serum and urine levels of 8-OHdG in UUO group were significantly higher than those in Sham group.Compared with UUO group,8-OHdG in serum and urine of UUO+PK group was significantly decreased,and the differences were statistically significant(all P<0.05).4.Immunohistochemical staining results The expression of 8-OHdG and the expression of ED-1 positive cells were significantly increased in UUO group compared with Sham group(P<0.05).After PK treatment,the expression of 8-OHdG and the expression of ED-1 positive cells decreased in the UUO group(P<0.05).5.Masson trichrome staining results Compared with Sham group,ECM deposition and diffuse glomerulosclerosis were observed in UUO group(P<0.05).Compared with UUO group,the degree of ECM deposition in renal tubule interstitial was decreased in UUO+PK group,and the degree of diffuse glomerulosclerosis was significantly improved(P<0.05).6.Results of western blot analysis The expressions of TGF-1,CTGF,MMP-2,IL-6 and Cleaved Caspase-3 in UUO group were significantly higher than those in Sham group,and the expression of MnSOD was significantly decreased.Compared with UUO group,the expressions of CTGF,TGF-1,IL-6,MMP-2 and Cleaved Caspase-3 in renal tissues of UUO+PK group were significantly decreased,and the expressions of MnSOD were significantly increased,which were statistically analyzed(P<0.05).7.TUNEL satining results There were few apoptotic cells in renal tissue of Sham group,and the number of TUNEL positive cells in UUO group was significantly higher than that in Sham group.Compared with UUO group,the number of TUNEL positive cells in renal tissue of UUO+PK group was significantly reduced,and the difference was statistically significant(P<0.05).Conclusion:Pancreatic Kininogenase has protective effect on renal fibrosis in UUO model rats,and its mechanism may be related to the improvement of degree of fibrosis by pancreatic kininogenase,inhibition of inflammatory reaction,anti-oxidative stress and regulation of apoptosis. |
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