miR-130b-3p和其靶基因在PCOS患者颗粒细胞中的表达研究-现代医学

miR-130b-3p和其靶基因在PCOS患者颗粒细胞中的表达研究

单位：秦皇岛市妇幼保健院生殖医学科, 河北秦皇岛 066000

分类号：R711.75

出版年·卷·期（页码）：2021·49·第九期（1022-1029）

摘要：

目的：探讨miR-130b-3p和其靶基因在多囊卵巢综合征（PCOS）患者颗粒细胞中表达的意义。方法：选取2019年10月至2020年6月于我院就诊且行体外受精/卵泡浆内单精子显微注射-胚胎移植（IVF/ICSI-ET）助孕治疗的35例PCOS患者作为PCOS组。另招募年龄、体质量指数（BMI）等相匹配的卵巢功能正常因男方因素或者单纯输卵管因素接受IVF/ICSI-ET助孕治疗的30例患者作为对照组。采用密度梯度离心法收集纯化颗粒细胞；利用生物信息学技术预测miR-130b-3p的下游靶基因；利用双荧光素酶实验验证miR-130b-3p的靶基因，在转染实验中将细胞分为miR-130b-3p模拟物组、miR-130b-3p抑制物组和各自的对照组；采用CCK8法检测细胞的增殖；采用流式细胞仪检测人卵巢颗粒细胞瘤细胞（KGN细胞）的凋亡指数；采用蛋白印迹法检测Smad4的表达水平。结果：PCOS组miR-130b-3p的表达水平（1.45±0.20）显著高于对照组（0.82±0.17）（P<0.05）；KGN细胞miR-130b-3p的表达水平（2.30±0.19）显著高于人正常卵巢上皮细胞（ISOE80细胞）（0.98±0.05）（P<0.05）；miR-130b-3p模拟物组的KGN细胞增殖活性显著高于其他3组，而凋亡率显著低于其他3组，均P<0.05。双荧光素酶实验证实，miR-130b-3p与Smad4具有靶向调节关系。PCOS组的Smad4蛋白的表达水平显著低于对照组，KGN细胞Smad4的蛋白表达水平显著低于ISOE80细胞，均P<0.05。结论：miR-130b-3p调节PCOS患者颗粒细胞的增殖和凋亡，参与PCOS的发病。

Objective: To investigate the expression of miR-130b-3p and its target genes in granulosa cells ofpolycystic ovary syndrome (PCOS) patients. Methods: From October 2019 to June 2020, 35 PCOS patients who received in vitro fertilization/intra-cytoplasmic sperm injection and embryo transfer(IVF/ICSI-ET) treatment in our hospital were selected as the PCOS group. In addition, 30 patients with normal ovarian function matched by age and body mass index(BMI) who received IVF/ICSI-ET due to male factors or pure fallopian tube factors were recruited as the control group. Density gradient centrifugation was used to collect and purify granulosa cells; Bioinformatics technology was used to predict the downstream target genes of miR-130b-3p; Dual luciferase experiment was used to verify the target genes of miR-130b-3p;In the transfection experiment, the cells were divided into miR-130b-3p mimics group,miR-130b-3p inhibitors groupand their respective control groups;CCK8 method was used to examed cell proliferation; Flow cytometry was used to detect the apoptosis index of KGN cells; Western blot was used to detect the expression of Smad4. Results: The expression levels of miR-130b-3p in PCOS patients and KGN cells were significantly higher than those in the control group and ISOE80 cells(P<0.05); The proliferation activity of KGN cells in the miR-130b-3p mimic group was higher and the apoptosis rate was significantly lower than the other three groups(P<0.05).Dual luciferase experiments confirmed that miR-130b-3p and Smad4 have a targeted regulatory relationship. The expression level of Smad4 protein in the PCOS group was significantly lower than that in the control group, and the protein expression level of Smad4 in KGN cells was significantly lower than that in ISOE80 cells(P<0.05). Conclusion: miR-130b-3p regulates the proliferation and apoptosis of granulosa cells in patients with PCOS, and participates in the pathogenesis of PCOS.

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