The Effect Of Fibroblast-myofibroblast Transformation On Neurogenic Bladder Fibrosis In Rats And Its Reversibility

Objective: To observe the difference of bladder fibrosis between normal rats and neurogenic bladder rats,and to investigate the effect of bladder fibroblast-myofibroblast transformation on neurogenic bladder fibrosis in primary cultured bladder fibroblasts.Methods: 1.Experiments in vivo: 30 healthy female SD rats were randomly divided into operation group and control group.Neurogenic bladder rat model was established by lumbosacral nerve transection in operation group whereas the lumbosacral nerve was not transected in control group.Bladder fibrosis was assessed assessed by Masson staining.The expression levels of α-SMA,COL-Ⅰ,COL-Ⅲ,MMP3 and MMP9 in bladder tissues were detected by immunohistochemical staining.The quantitative analysis of optical density was performed by Image J.2.Experiments in vitro: primary fibroblasts were isolated and cultured from the bladder tissues of the operation group and the control group,which were labeled as the NB group and the DZ group,respectively.The cultured primary fibroblasts were identified by immunofluorescence staining of Vimentin and α-SMA.TGF-β1 receptor was inhibited by addition of 10 μM SB-505124 to the medium(TGF-β1inhibition group).The NB group and DZ group were further divided into three subgroups(blank group,TGF-β1 inhibition group,and DMSO group),respectively.The expression levels of α-SMA,COL-Ⅰ,COL-Ⅲ,MMP3,and MMP9 in the cultured cell lysate were detected by Elisa assay after 24 hours and 48 hours of cell culture.Results: 1.Results of experiments in vivo: The bladder tissue fibrosis of rats was significantly increased in the operation group compared with the control group.The expressions of α-SMA,COL-Ⅰ,COL-Ⅲ,MMP3 and MMP9 in the bladder tissues of rats in the operation group were significantly increased compared with the control group(p <0.001),indicating that the fibroblast-myofibroblast transformation and ECM synthesis were increased.The expression levels of extracellular matrix metalloproteinases is increased and a large number of ECM were aggregated;2.Results of experiments in vitro:Compared with DZ group,the expressions of α-SMA,COL-Ⅰ,COL-Ⅲ,MMP3 and MMP9 in NB group were significantly increased(p < 0.001),indicating that the degree of fibroblast-myofibroblast transformation was enhanced.Myofibroblasts were increased,ECM synthesis was increased,and the degree of fibrosis was increased.After 24 hours or48 hours of TGF-β1 inhibition,the expression levels of α-SMA,COL-Ⅰ,COL-Ⅲ,MMP3 and MMP9 in the TGF-β1 inhibition group were significantly lower than those in the blank group and the DMSO group(p < 0.001).It is suggested that the degree of fibroblastmyofibroblast transformation was weakened,the number of myofibroblasts is reduced,the synthesis of ECM is reduced,and the degree of fibrosis is reduced when TGF-β1 is inhibited.Conclusion: 1.It is feasible to establish the model of neurogenic bladder fibrosis in rats by cutting the lumbosacral nerve.The bladder of this model rats is obviously fibrotic;2.Myofibroblast plays an important role in the occurrence and development of neurogenic bladder fibrosis,and fibroblast-myofibroblast transformation promotes the occurrence of bladder fibrosis;3.Under the condition that TGF-β1 is inhibited,the phenotypic transformation of fibroblasts to myofibroblasts is reduced,myofibroblasts are reduced,and the degree of fibrosis is alleviated,which provides a new idea for the clinical treatment of neurogenic bladder fibrosis.