摘要
目的探究miR-30b对子宫腺肌症(AM)模型小鼠子宫内膜间质细胞(ESCs)的影响,并进一步探究其可能的潜在调控机制。方法选取24只ICR雌性小鼠,制备AM动物模型,另选取4只ICR小鼠作为正常对照;HE染色观察造模小鼠子宫病理学变化;反转录荧光定量-聚合酶链反应(qRT-PCR)检测AM模型小鼠及正常小鼠子宫组织中miR-30b表达情况。分离并培养AM模型小鼠ESCs,随机分为对照组(Control组)、miR-30b过表达组(miR-30b组)和miR-30b过表达阴性对照组(miR-NC组);qRT-PCR检测各组细胞中miR-30b表达;MTT法、Transwells法和流式细胞术分别检测细胞增殖、迁移、凋亡能力;蛋白质印迹(Western blot)检测细胞中趋化因子受体4(CXCR4)和血管内皮生长因子(VEGF)蛋白表达;双荧光素酶报告验证miR-30b和CXCR4的靶向关系。结果24只小鼠中AM造模成功22只(91.67%)。与正常小鼠相比,AM模型小鼠子宫扭曲变形且明显粗壮,表面可见明显突出的腺肌病结节,子宫肌层发育紊乱,内膜间质受到浸润,内膜间质细胞侵入肌肉内层。miR-30b在AM模型小鼠子宫组织中显著低表达(P<0.05),而过表达miR-30b可以降低模型小鼠ESCs的增殖能力和侵袭能力、加速细胞凋亡(P<0.05),且过表达miR-30b可显著降低ESCs中VEGF、CXCR4蛋白表达水平(P<0.05)。miR-30b直接靶向CXCR4。结论miR-30b可能通过直接靶向调控CXCR4表达,进而促进ESCs的增殖和侵袭,抑制细胞凋亡,参与AM发病。
Objective:To explore the effects of miR-30b on endometrial stromal cells(ESCs)in mice with adenomyosis(AM)as well as its potential regulatory mechanism.Methods:A total of 24 ICR female mice were selected to make AM animal model,and 4 ICR mice were selected as normal controls.HE staining was used to observe the pathological changes of the uterus,and reverse transcription fluorescence quantification polymerase chain reaction(qRT-PCR)was used to detect the expression of miR-30b in the uterus of AM model mice and normal mice.ESCs of AM model mice were isolated and cultured,and randomly divided into the control group,miR-30b overexpression group(miR-30b group)and negative control group with miR-30b overexpression(miR-NC group).The expression of miR-30b was detected by qRT-PCR,and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)assay,transwells assay and flow cytometry were used to detect cell proliferation,migration and apoptosis,respectively.Western blot was used to detect the expression levels of CXC chemokine receptor 4(CXCR4)and vascular endothelial growth factor(VEGF)in cells.The targeting relationship between miR-30b and CXCR4 was verified by dual luciferase report.Results:Among the 24 mice,22(91.67%)were successful in AM modeling.Compared with normal mice,the uterus of AM model mice was distorted and strong,with prominent adenomyopathy nodules on the surface,uterine myometrium development disorder,endometrial interstitial infiltration,and endometrial interstitial cells invaded the inner muscle layer.The expression of miR-30b was significantly lower in the uterine tissue of AM model mice(P<0.05),while overexpression of miR-30b reduced the proliferation and invasion abilities of ESCs and accelerated cell apoptosis of model mice(P<0.05).Furthermore,overexpression of miR-30b significantly decreased the expression levels of VEGF and CXCR4 protein in ESCs(P<0.05).It was found that miR-30b directly targeted CXCR4.Conclusions:miR-30b may directly regulate CXCR4 expression,resulting in the promotion of
作者
许丹丹
郑艳莉
郭娟
XU Dan-dan;ZHENG Yan-li;GUO Juan(The Second Affiliated Hospital of Nantong University/The First People’s Hospital of Nantong City,Nantong 226000;Department of Obstetrics&Gynecology,Yancheng Third People’s Hospital,Yancheng 224000)
出处
《生殖医学杂志》
CAS
2024年第10期1357-1363,共7页
Journal of Reproductive Medicine
基金
南通市科技项目(MS22020004)。
