摘要
为研究牦牛卵巢组织内miR-96对颗粒细胞凋亡、增殖及激素分泌的影响,本试验通过采集成年牦牛卵巢内颗粒细胞,分离培养后将其分为4个处理组,分别转染miR-96模拟物(mimics)和抑制物(inhibitor)及两者的阴性对照(NC)后培养并检测颗粒细胞受到的影响。使用流式细胞术和CCK-8法对细胞凋亡和增殖情况进行检测,采用qPCR和Western Blot检测凋亡相关基因表达情况,ELISA检测转染后类固醇激素分泌情况。结果表明:miR-96-mimics使颗粒细胞凋亡发生抑制促进细胞增殖,并促使凋亡相关基因Bcl-2表达极显著升高,Bax表达极显著降低;ELISA检测类固醇激素水平变化显示,与mimics NC组相比miR-96-mimics组细胞培养液中孕酮(P4)分泌水平极显著降低,雌二醇(E_(2))分泌水平极显著升高;与inhibitor NC组相比miR-96-inhibitor组孕酮(P4)分泌水平极显著升高,雌二醇(E_(2))分泌水平显著降低。miR-96可促进颗粒细胞增殖并抑制凋亡,同时会对生殖激素分泌产生影响,从而参与调控牦牛卵泡发育与闭锁的过程。
The granulosa cells collected from adult Bos grunniens ovaries are taken as research materials to investigate the effects of miR-96 on the apoptosis,proliferation and hormone secretion of the cells in Bos grunniens ovarian tissue.The cells are divided into four groups after isolation and cultivation,transfected with miR-96 mimics and inhibitors as well as their negative controls(NC).The effects on the cells are cultured and detected as well.Flow cytometry and CCK-8 assay are used to detect cell apoptosis and proliferation.Besides,qPCR and Western Blot are used to detect the expression of apoptosis-related genes.The steroid hormone secretion after transfection is detected by using ELISA.The results show that miR-96-mimics group inhibit granulosa cells apoptosis and promote cell proliferation.In addition,it significantly increases the expression of apoptosis-related gene and significantly decreases the expression of Bax.According to the variations steroid hormone levels detected by ELISA,it can be concluded that compared with mimics NC group,the secretion level of P4 in the cell culture medium of miR-96-mimics group is significantly reduced,while the secretion level of E_(2) is significantly increased.Compared with inhibitor group,the secretion level of P4 of miR-96-inhibitor group is significantly rised,while that of E_(2) is significantly decreased.In summary,miR-96 can promote granulosa cell proliferation,inhibit apoptosis and affect reproductive hormone secretion,thereby participating in the regulation of follicular development and occlusion of Bos grunniens.
作者
贾跃宁
彭巍
徐尚荣
刘海婷
黄荣
舒适
王国文
付长其
张君
JIA Yuening;PENG Wei;XU Shangrong;LIU Haiting;HUANG Rong;SHU Shi;WANG Guowen;FU Changqi;ZHANG Jun(College of Agriculture and Animal Husbandry,Qinghai University,Xining 810016,China;Academy of Animal and Veterinary Sciences,Qinghai University,Xining 810016,China)
出处
《青海大学学报》
2024年第1期50-56,共7页
Journal of Qinghai University
基金
青海省科学技术厅项目(2022-NK-119)
青海省“昆仑英才·高端创新创业人才”计划2021培养领军人才。
关键词
牦牛
miR-96
颗粒细胞
凋亡
增殖
Bos grunniens
miR-96
granulosa cell
apoptosis
proliferation
