Expression And Significance Of Micro RNA-200b In Fibroblasts Of The Uterosacral Ligament In Patients With Pelvic Organ Prolapse

Background:pelvic organ prolapse?POP?refers to pelvic organ prolapse in the vagina outside or inside of the vagina,seriously affect the patients quality of life.POP is a disease with multiple causes,such as the number of vaginal births,age,and body mass index.The structural and functional abnormalities of the extracellular matrix of the pelvic floor caused by genetic defects are important molecular basis for the development of female POP.Collagen fiber and elastic fiber,as important components of extracellular matrix,provide tensile strength to pelvic floor tissues in the form of fascia and ligament,and play an important role in maintaining the normal position and function of pelvic floor organs.The major cells of the pelvic floor ligaments and fascia are fibroblasts.Micro RNA?miRNA?is a non-coding RNA with a length of about 19-22 nucleotides,which regulates gene expression after transcription.Mir-200b has been implicated in a variety of fibrosis diseases,such as pulmonary fibrosis,renal fibrosis,and cardiac fibrosis.Objective:to investigate the expression of microRNA-200b in fibroblasts of the uterosacral ligament in primary and subcultured pelvic organ prolapse patients in vitro,so as to provide feasible theoretical basis and method guidance for the pathogenesis and future treatment of pelvic organ prolapse.Methods:Collected for pelvic organ prolapse?quantitative framework based on pelvic prolapse stage II or above?uterus resection in patients with 10 cases of specimens of the sacral near the ligament of the uterus,as experimental group,collection of patients with pelvic organ prolapse uterus resection specimens from 10 cases of the sacral beside ligament of the uterus as normal control group,using the improved digestion of type I collagenase,for pelvic organ prolapse group and no pelvic organ prolapse of the normal control group parametric sacral ligament fibroblasts were primitive and subculture in vitro.Morphological observation,immunohistochemistry and immunofluorescence were used to identify the types and purity of the fibroblasts from the uterosacral ligament in the two groups of cultured cells,so as to ensure the number of cells,and to compare the differences between the two groups of samples.SPSS25.0 software was used to analyze and compare the clinical characteristics of the experimental group and the control group in terms of age,body mass index,family history of pelvic organ prolapse,menopause and normal natural delivery.The expression levels of microRNA-200b gene in the normal control group without pelvic organ prolapse and in the paracral ligament fibroblasts of the experimental group without pelvic organ prolapse were detected by real-time fluorescence quantitative PCR method.The relative expression levels of the two groups of microRNA-200b gene were calculated by 2^-??Ct method.??Ct value=(Ct_{microRNA-200b}-Ct _U6)experimental group-(Ct_{microRNA-200b}-Ct _U6)control group.Finally,SPSS25.0 software was used for statistical analysis of our results.Results:The primitive and subculture in vitro for P3 generation of parametric sacral ligament cells in morphology observation under inverted phase contrast microscope,cell formed a long spindle,connected to each other,densely packed are arranged spiral or radial,state clearly that the cell body is bigger,transparent cytoplasm and nucleus is larger,more oval,in line with the morphological characteristics of fibroblasts,compared with the normal control of fiber cells,pelvic organ prolapse of fiber cells quantity is less,relatively sparse;Immunohistochemical expression of vimentin was positive,which proved that the cultured cells were fibroblasts.Immunofluorescence vimentin showed diffuse green fluorescence,which further verified that the cultured cells were fibroblasts.Using SPSS25.0 software of the experimental group?pelvic organ prolapse patients?and control group?no patients with pelvic organ prolapse group?for age,body mass index,family history of pelvic organ prolapse proportion,the proportion of menopause and comparison between normal natural birth,there was no statistically significant difference?P>0.05?,shows good match between the group of 20 subjects.Using 2^-??Ct to represent the microRNA-200b gene expression differences in experimental group and normal control group,2^-??Ct>1,said microRNA expression levels rise,on the other hand,2^-??Ct<1,said microRNA expression levels drop,the experimental results of 2^-??Ct>1,the said,compared to control group and no pelvic organ prolapse microRNA-200b gene sacral beside the pelvic organ prolapse of uterus ligament fibroblasts in expression level to rise,and P<0.01,the difference was statistically significant.Conclusion:compared with normal uterosacral ligament fibroblasts,the expression level of microRNA-200b gene in uterosacral ligament fibroblasts of patients with pelvic organ prolapse was significantly increased,suggesting that microRNA-200b gene may be involved in the pathogenesis of pelvic organ prolapse.