敲减miR-145增强骨髓间充质干细胞对大鼠盆底功能障碍的修复作用

Knockdown of miR-145 Enhances Repair Effect of Bone Marrow Mesenchymal Stem Cells on Pelvic Floor Dysfunction in Rats

目的探讨敲减miR-145对骨髓间充质干细胞(BMSC)修复大鼠盆底功能障碍(PFD)的影响。方法分离大鼠BMSC,并鉴定;将miR-NC或miR-145-shRNA慢病毒载体转入BMSC后,采用CCK-8法检测细胞活力。将32只大鼠按照随机对照表法分为对照组(Con)、PFD组、miR-NC治疗组(miR-NC)和miR-145-shRNA治疗组(miR-145-shRNA)。通过阴道扩张法建立PFD大鼠模型,miR-NC和miR-145-shRNA组大鼠分别给予注射已转染miR-NC和miR-145-shRNA慢病毒载体的BMSC,14 d后,进行尿流动力学(膀胱容积、排尿时膀胱内压、漏尿点压)测试,并用western blot法检测各组大鼠盘底支持组织中Ⅰ型胶原(CollagenⅠ)和弹性蛋白(Elastin)的表达水平。结果成功分离大鼠BMSC;与miR-NC组比较,miR-145-shRNA组BMSC的细胞活力明显增强(P<0.001)。与Con组比较,PFD组大鼠膀胱容积、排尿时膀胱内压、漏尿点压以及盘底支持组织中CollagenⅠ和Elastin的表达水平均降低(均P<0.01);与PFD组比较,miR-NC组和miR-145-shRNA组大鼠中上述指标均增加(均P<0.05),且miR-145-shRNA组优于miR-NC组(均P<0.05)。结论敲减miR-145能增强BMSC对大鼠盆底功能障碍的修复作用。

Objective To investigate the effect of miR-145 knockdown on the repair of pelvic floor dysfunction(PFD)with bone marrow mesenchymal stem cells(BMSCs)in rats.Methods Rat BMSCs were isolated and identified.Lentiviral vectors carrying miR-NC or miR-145-shRNA were transfected into BMSCs,and cell viability was detected by CCK-8 assay.Thirty-two rats were randomly divided into control(Con)group,PFD group,miR-NC treatment group,and miR-145-shRNA treatment group.The rat model of PFD was established by vaginal dilatation method.Rats in miR-NC and miR-145-shRNA groups were injected with BMSCs transfected with miR-NC and miR-145-shRNA lentiviral vectors,respectively.After 14 days,urodynamic test was performed.Western blot was used to detect the expression of typeⅠcollagen and elastin in the pelvic floor supporting tissues.Results Rat BMSCs were isolated successfully.Compared with miR-NC group,the cell viability increased in miR-145-shRNA group(P<0.001).Compared with Con group,bladder volume,bladder pressure during urination,leak point pressure and expression of typeⅠcollagen and elastin in the pelvic floor supporting tissues decreased in PFD group(P<0.01).However,these indexes increased in miR-NC and miR-145-shRNA groups when compared to PFD group(P<0.05).Furthermore,the efficacy in miR-145-shRNA group was superior to that in miR-NC group(P<0.05).Conclusion Knockdown of miR-145 can enhance the repair effect of BMSCs on pelvic floor dysfunction in rats.