Study On The Mechanisms Of Renal Fibrosis In Offspring Rats Of Maternal Exposure To DBP

Objective: Maternal exposure to DBP during pregnancy can cause renal dysplasia and renal fibrosis in offspring rats.However,the underlying mechanisms of such phenomena are not known.The purpose of this study is to explore the key aspects of the molecular mechanism that contribute to the renal fibrosis in the offspring rats with maternal exposure to DBP during pregnancy.The results from this study will help better understand,and possibly identify therapeutic targets for the prevention and treatment of DBP-induced renal fibrosis in the future.Methods: Sprague Dawley(SD)rats were used to copulate and get pregnant.Animal model with DBP exposure during pregnancy was established by feeding female rats with DBP dissolved in peanut oil at 850mg/kg(DBP group).Rats fed equal amount of peanut oil without DBP during pregnancy were set as Control group.Six male offspring rats were randomly selected from both DBP and Control group on postnatal day 1.The expression level of RhoA,ROCK1,Ang ?,CTGF,SMAD2 and SMAD3 were detected and measured by real-time PCR,Western blot and immunohistochemistry.The renal tubular epithelial cell line NRK-52 E was used in in vitro experiments.The expression of RhoA,ROCK1,E-cad and ?-SMA in NRK-52 E cells,from both DBP treated and control group,were detected and measured by Western blot and real time PCR.For the DBP treated NRK-52 E cell lines,the changes of cell morphology before and after DBP treatment were also analyzed under microscope.Furthermore,we analyzed the effect of ROCK inhibitor Y-27632 on the morphological changes of cells in DBP group,and the expression level of E-cad and ?-SMA in DBP treated NRK-52 E cells after Y-27632 intervention.Statistical analysis was conducted to analyze the experiments results.Results: HE staining showed that compared with the control group,the number of glomeruli in DBP group was significantly reduced and interstitial tissue was proliferated.Masson staining showed evident renal fibrosis in DBP group.The expression level of RhoA and ROCK1 both for RNA transcription and protein in kidney of DBP group was significantly higher than that of Control group(P < 0.01).In DBP group,the gene expression of Ang ?,CTGF,Smad2,Smad3 were significantly upregulated as well.In in vitro experiments,DBP has also been found to activate the RhoA/ROCK pathway in NRK-52 E cells as indicated by the significant increase in RhoA and ROCK1 gene expression both at protein and RNA transcription level.DBP caused EMT in NRK-52 E cells,i.e.cells with typical characteristics of epithelial cells underwent morphological changes into the cells with characteristics of mesenchymal cells after DBP treatment.Furthermore,DBP reduced the expression of E-cad and increased the expression of ?–SMA in NRK-52 E cells.ROCK inhibitor can prevent and reverse these effects in vitro.Conclusion: Maternal exposure to DBP can lead to renal fibrosis in offspring rats.DBP can activate the RhoA/ROCK pathway in kidney and cause EMT in renal tubular epithelial cells.RhoA/ROCK pathway plays an important role in DBP-induced renal fibrosis.Ang ? and the related molecules also play an important role in DBP-induced renal fibrosis.