神经生长因子对颗粒细胞增殖检测方法的探讨  

Effect of nerve growth factor on granulosa cell proliferation

作　　者：于洋[1] 柳春[2] 

机构地区：[1]辽宁医学院基础医学院生理学教研室,辽宁锦州121001 [2]辽宁中医药大学基础医学院生化教研室,辽宁沈阳110032

出　　处：《中国现代医学杂志》2015年第16期15-18,共4页China Journal of Modern Medicine

摘　　要：目的为了解神经生长因子(NGF)对小鼠卵巢腔前颗粒细胞促增殖情况并探索其机制,同时建立方便灵敏、无污染和毒性小的细胞增殖检测方法。方法用NGF刺激培养的小鼠卵巢腔前颗粒细胞,细胞计数试剂盒-8(CCK-8)实验分析,溴脱氧尿嘧啶核苷(Brd U)掺入标记,免疫荧光技术检测颗粒细胞在体外培养中的增殖情况和Brd U标记指数。结果 NGF明显促进颗粒细胞增殖,并且具有一定的时效关系。荧光显微镜下可见Brd U阳性标记的颗粒细胞,提示细胞处于增殖期。NGF组的Brd U标记指数为45.03%,明显高于对照组20.65%(P<0.01)。NGF使增殖期的细胞显著增多及增殖指数明显升高。结论该研究证实NGF能够促进颗粒细胞增殖,CCK-8分析和Brd U标记是高效、灵敏的标记腔前颗粒细胞增殖的方法,NGF促进颗粒细胞增殖与促进颗粒细胞DNA的合成有关系。[ Objective ] To discuss the effect of Nerve Growth Factor （NGF） on the mouse ovarian preantral gran- ulosa cell proliferation and its mechanism, and to establish a speedy, sensitive, accurate, low toxicity and pollution free detection way of cell proliferation. [ Methods ] The mouse ovarian preantral granulosa cells were treated with NGF in vitro, the cells proliferation and labeling index were measured by cell counting kit-8 （CCK-8） assay, bro- modeoxyuridine （BrdU） labeling and immunofluorescence technique. [ Results ] The data showed that NGF signifi- cantly promoted the proliferation of granulosa cells in a time-dependent manner. The BrdU-positive granulosa ceils were visible under fluorescence microscope after NGF treatment, which indicated that cells were in the proliferative phase. BrdU labeling index of the treatment group was 45.03%, which was significantly higher than that of the con- trol group （20.65%, P 〈 0.0l）. The number of cells and the proliferation index increased in the proliferative phase after NGF treatment. [ Conclusions ] The results indicated that CCK-8 assay and BrdU labeling was an effective, speedy and sensitive method of detecting preantral granulosa cell proliferation. The promotion effect of NGF on gran- ulosa cell proliferation might be associated with the promotion of DNA synthesis.

关 键 词：神经生长因子 颗粒细胞 细胞计数试剂盒-8 溴脱氧尿嘧啶核苷 增殖 

分 类 号：Q492[生物学—生理学] R-332[医药卫生]