Ghrelin参与FSH对牛卵丘细胞体外增殖的调节  

Ghrelin Involvements in FSH Induced Bovine Cumulus Cell Proliferation in vitro

作　　者：米焱[1] 张伶俐[1] 李海军[1] 杜晨光[1] 曹贵方[1] 王秀梅[1] 

机构地区：[1]内蒙古农业大学兽医学院,呼和浩特010018

出　　处：《畜牧兽医学报》2013年第8期1236-1243,共8页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金(30860201);内蒙古自然科学基金(2012MS0411;2010BS0508)

摘　　要：为了解牛卵丘-卵母细胞复合体(COCs)体外成熟过程中Ghrelin对卵丘细胞增殖的可能调节及其与促卵泡素(Follicle-stimulating hormone,FSH)在卵丘细胞增殖过程中可能的调控关系,本研究运用实时定量RT-PCR技术首先检测了体外不同成熟时间(0、8、16和24h)牛卵丘细胞中Ghrelin mRNA的相对表达差异,然后在体外成熟8h,探讨了FSH与FSH受体结合抑制剂(FSH binding inhibitor,FSHBI)对Ghrelin mRNA表达的影响,以及FSH与Ghrelin受体抑制剂([D-Lys3]-GHRP-6)对增殖相关基因PCNA,BCL-2和BAX mRNA表达可能的调节作用;最后利用细胞增殖活性测定技术检测了FSH与[D-Lys3]-GHRP-6对卵丘细胞增殖活性的影响。结果显示:(1)Ghrelin mRNA相对水平在体外成熟8h卵丘细胞中最高,显著高于0和24h组(P<0.05);(2)0.01IU·mL-1FSH添加组与对照组相比,Ghrelin mRNA表达显著升高(P<0.05);在添加400ng·mL-1 FSHBI后,GhrelinmRNA表达下降且差异显著(P<0.05);(3)添加FSH后,PCNA和BCL-2mRNA表达以及卵丘细胞增殖活性显著升高(P<0.05);添加[D-Lys3]-GHRP-6后,上述基因表达水平与增殖活性显著降低(P<0.05)。上述结果表明,Ghrelin参与FSH对体外成熟过程卵丘细胞增殖的调节,本研究结果有助于进一步了解FSH调控卵泡发育的机制。To explore the possible effects of Ghrelin and certain relationship between Ghrelin and FSH（Follicle-stimulating hormone） in regulating the proliferation of cumulus cell freed from bovine cumulus-oocyte complex（COCs） in vitro maturation process,the Ghrelin mRNA levels within bovine cumulus cells varied depending on the oocyte maturation stage,the effect of FSH and FSHBI（FSH binding inhibitor） on Ghrelin mRNA expression and the effect of FSH and Ghrelin receptor antagonist（[D-Lys3]-GHRP-6） on regulating the expression of PCNA,BCL-2 and BAX mRNA at 8 h in vitro were detected by relative real-time RT-PCR experiments.And cumulus cell proliferative activity affected by FSH and [D-Lys3]-GHRP-6 was determined by cell proliferation activity assay technology during bovine COCs maturation.The results showed that the expression of Ghrelin mRNA within cumulus cell was the highest at 8 h and the expressed difference among 8,0 and 24 h was significant（P0.05）.The expression of Ghrelin mRNA in cumulus cell was prominently elevate when addition of 0.01 IU·mL-1 FSH（P0.05）,while the Ghrelin mRNA levels was significantly decrease affected by 400 ng·mL-1 FSHBI.The expression of PCNA and BCL-2 genes and the proliferative activity of cumulus cell were observably promoted by 0.01 IU·mL-1 FSH（P0.05）,but were markedly reduced after adding [D-Lys3]-GHRP-6（P0.05）.The results suggested that Ghrelin involves in regulating cumulus cell proliferation triggered by FSH and that this research could help in understanding the mechanisms behind follicular development regulated by FSH.

关 键 词：牛卵丘细胞 增殖 FSH GHRELIN 实时定量PCR 

分 类 号：S826[农业科学—畜牧学] S814.8[农业科学—畜牧兽医]