微RNA-145抑制剂靶向CRKL调节促分裂原活化的蛋白激酶通路对卵巢颗粒细胞激素合成的影响  

Effects of miRNA-145 Inhibitor Targeting CRKL Regulation of Mitogen-Activated Protein Kinase Pathway on Hormone Synthesis of Ovarian Granulosa Cells

作　　者：张远远 胡昌华[1] 杨书红[2] 王峥[3] ZHANG Yuanyuan;HU Changhua;YANG Shuhong;WANG Zheng(Dept. of Obstetrics and Gynecology, Hanyang Hospital Affiliated to Wuhan University, Hubei Wuhan 430000, China;Dept. of Obstetrics and Gynecology, Tongji Hospital Affiliated to Huazhong University of Science and Technology, Hubei Wuhan 430000, China;Dept. of Obstetrics and Gynecology, the Second People's Hospital of Jingzhou, Hubei Jingzhou 434000, China)

机构地区：[1]武汉科技大学附属汉阳医院妇产科,湖北武汉430000 [2]华中科技大学附属同济医院妇产科,湖北武汉430000 [3]荆州市第二人民医院妇产科,湖北荆州434000

出　　处：《中国医院用药评价与分析》2020年第3期273-279,共7页Evaluation and Analysis of Drug-use in Hospitals of China

基　　金：国家自然科学基金资助项目(No.81501227)。

摘　　要：目的:研究微RNA(microRNA,miRNA)-145在卵巢早衰中的作用,并对其可能的作用机制进行初步探讨。方法:采用miRNA芯片预测并筛选在不同日龄雌鼠中差异性表达的miRNA,采用TargetScan预测miRNA靶基因,通过DAVID数据库对靶基因进行信号通路及生物学功能的富集分析,筛选在不同时期卵巢颗粒细胞中发挥重要调控作用的miRNA;采用MTT法检测卵巢颗粒细胞中miRNA-145抑制后生存率;采用Q-PCR检测miRNA-145及CRKL mRNA的表达水平;采用酶联免疫吸附法检测CRKL、雌二醇、卵泡刺激素、黄体生成素、p-p38丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)及p-ERK1/2水平。结果:miRNA芯片预测差异性表达miRNA 23个,靶基因多与细胞增殖、分化有关,且多集中于MAPK通路;综合结合生物信息预测结果,筛选出与CRKL能够靶向结合的miRNA-145;miRNA-145与CRKL在颗粒细胞中变化水平呈正相关,miRNA-145在卵巢细胞中被抑制后,CRKL水平、颗粒细胞凋亡及MAPK通路的激活均受抑制,但激素分泌趋于正向调节。结论:miRNA-145抑制剂能够作用于MAPK通路抑制卵巢颗粒细胞的凋亡,调节激素的合成,其作用机制可能与miRNA-145靶向CRKL调节MAPK通路有关。OBJECTIVE:To study the role of microRNA(miRNA)-145 in premature ovarian failure(POF),and to probe into the possible mechanism.METHODS:miRNA microarray was used to predict and screen differentially expressed miRNA in different day-aged female mice,TargetScan was used to predict miRNA target genes,and DAVID database was used to analyze the enrichment of signaling pathways and biological function of target genes.miRNA that played important regulatory roles in ovarian granulosa cells at different stages was screened.MTT method was used to detect the survival rate after miRNA-145 inhibition in ovarian granulosa cells.The expression of miRNA-145 and CRKL mRNA in ovarian granulosa cells were detected by Q-PCR,and the levels of CRKL,estrogen,follicle stimulating hormone,luteinizing hormone,p-p38 mitogen-activated protein kinase(MAPK)and p-ERK1/2 were detected by ELISA.RESULTS:miRNA microarray predicted 23 miRNAs were expressed differently,most of the target genes were related to cell proliferation and differentiation and enrich in MAPK pathway.Combined the prediction results of biological information,miRNA-145 that can target CRKL was screened out.miRNA-145 was positively correlated with the changes of level of CRKL in granulosa cells,after miRNA-145 was inhibited in ovarian cells,the CRKL level,apoptosis of granulosa cells,activation of MAPK pathway were inhibited,but the hormone secretion tended to be positively regulated.CONCLUSIONS:miRNA-145 inhibitors can act on the MAPK pathway to inhibit the apoptosis of ovarian granulosa cells and regulate the synthesis of hormones.The mechanism may be related to the action on MAPK pathway by miRNA-145 targeting CRKL.

关 键 词：卵巢早衰 卵巢颗粒细胞 miRNA-145 MAPK通路 

分 类 号：R96[医药卫生—药理学]