脐带间充质干细胞对卵巢早衰模型家兔TIMP1、FN和MMP9表达的影响  

Effect of umbilical cord-derived mesenchymal stem cells on the expression of TIMP1,FN and MMP9 in a rabbit model of premature ovarian failure

作　　者：张娟 周丽[2] 段瑶 闫夏贝 庹平 王刚 Juan Zhang;Li Zhou;Yao Duan;Xiabei Yan;Ping Tuo;Gang Wang(Department of Reproductive Medicine,General Hospital of Central Theater Command,Wuhan 430070,China;Hubei University of Traditional Chinese Medicine,Wuhan 430065,China)

机构地区：[1]中国人民解放军中部战区总医院生殖医学科,武汉430070 [2]湖北中医药大学,武汉430065

出　　处：《中华生殖与避孕杂志》2021年第12期1071-1078,共8页Chinese Journal of Reproduction and Contraception

基　　金：湖北省自然科学基金项目(2018CFC806)。

摘　　要：目的研究脐带间充质干细胞改善卵巢早衰家兔卵巢功能的潜在机制。方法将10只家兔按随机数表法分为治疗组和模型组各5只,均采用腹腔注射环磷酸酰胺(50 mg·kg^(-1)·d^(-1),连续2 d)构建卵巢早衰模型。造模1周后,治疗组予以耳缘静脉注射人脐带间充质干细胞(2 mL/d,连续3 d),模型组予以注射等量无菌水。第28日后采集两组家兔卵巢组织标本,通过苏木精-伊红染色法观察两组家兔卵巢组织形态结构,并采用蛋白免疫印迹法、实时荧光定量PCR及免疫组织化学技术检测两组家兔卵巢组织内基质金属蛋白酶组织抑制物1(tissue inhibitor of metalloproteinases 1,TIMP1)、纤连蛋白(fibronectin,FN)、纤维形成相关因子基质金属蛋白酶9(matrix metalloproteinases 9,MMP9)在蛋白及mRNA水平上的表达。结果①模型组家兔卵巢组织内原始卵泡数量少,闭锁卵泡多,各级卵泡颗粒层排列紊乱,卵细胞核出现固缩,发生凋亡,卵巢间质出现纤维化;而治疗组家兔卵巢组织内原始、初级卵泡数量多,闭锁卵泡少,各级卵泡颗粒细胞排列规则。②与模型组相比较,治疗组家兔卵巢组织内TIMP1蛋白(0.546±0.021比0.820±0.085)和FN蛋白表达量增多(0.221±0.065比0.516±0.064)(P均<0.001),而MMP9蛋白表达量减少(0.504±0.049比0.204±0.066,P<0.001)。③与模型组相比较,治疗组家兔卵巢组织内TIMP1 mRNA(0.217±0.024比0.470±0.039)和FN mRNA的表达量增多(0.039±0.006比0.125±0.012,P均<0.001),而MMP9 mRNA的表达量减少(0.009±0.000比0.002±0.000,P<0.001)。④TIMP1和FN主要表达在颗粒细胞和卵细胞膜,TIMP1还表达在卵巢间质组织,治疗组TIMP1和FN的表达多于模型组,差异均具有统计学意义(P<0.001,P=0.005)。MMP9主要表达在颗粒细胞和卵巢间质组织,治疗组MMP9的表达少于模型组,差异具有统计学意义(P<0.001)。结论脐带间充质干细胞能够通过调节卵巢组织内TIMP1、FN和MMP9水平来改善卵巢功能�Objective To explore the potential molecular mechanism of umbilical cord-derived mesenchymal stem cells(UCMSCs)to improve ovarian function in premature ovarian failure(POF)rabbits.Methods Ten rabbits were randomly divided into 2 groups(five rabbits in each),treatment group and model group.A rabbit model of POF was established using an intraperitoneal injection of cyclophosphamide for 2 d(50 mg·kg^(-1)·d^(-1)).After a week,the treatment group received UCMSCs intravenously via a marginal ear vein and model group received equal amounts of sterile water.Ovarian samples were collected on day 28 and ovaries morphology was observed by hematoxylin-eosin staining.Tissue inhibitor of metalloproteinases 1(TIMP1),fibronectin(FN)and matrix metalloproteinases 9(MMP9)protein and mRNA expression were detected by Western blotting,PCR,and immunohistochemistry.Results 1)In model group,ovaries of the ovarian tissue displayed as reduced number of primordial follicles,along with increased number of atretic follicles.The granular layer cells of every follicle grade were arranged in disorder.The oocyte nucleus appeared pyknotic and apoptotic changed.Fibrosis was performed in the ovarian stroma.While in treatment group,the number of primordial and primary follicles in the ovarian tissue was large,and there were few atretic follicles.The granule cells of every follicle grade were arranged regularly.2)Compared with model group,the expressions of TIMP1 protein(0.546±0.021 vs.0.820±0.085)and FN protein(0.221±0.065 vs.0.516±0.064)in treatment group were significantly increased(all P<0.001),while the expression of MMP9 protein(0.504±0.049 vs.0.204±0.066)in treatment group was significantly decreased(P<0.001).3)Compared with model group,the expressions of TIMP1 mRNA(0.217±0.024 vs.0.470±0.039)and FN mRNA(0.039±0.006 vs.0.125±0.012)in treatment group were significantly increased(all P<0.001),while the expression of MMP9 mRNA(0.009±0.000 vs.0.002±0.000)in treatment group was significantly decreased(P<0.001).4)TIMP1 and FN were mainl

关 键 词：间充质干细胞 基质金属蛋白酶组织抑制物 纤连蛋白 基质金属蛋白酶 卵巢早衰 

分 类 号：R711.75[医药卫生—妇产科学]