Shimada Masayuki, Nishibori Masahide, Isobe Naoki, Kawano Natsuko, Terada Takato
Laboratories of Animal Reproduction, Graduate School of Biosphere Science, Hiroshima University, Hiroshima, Japan.
Biol Reprod. 2003 Apr;68(4):1142-9. doi: 10.1095/biolreprod.102.010082. Epub 2002 Nov 13.
We investigated the formation of LH receptor (LHR) in cumulus cells surrounding porcine oocytes and the role of LHR in meiotic maturation of oocytes. At least three splice variants of LHR mRNA were detected in cumulus cells, in addition to the full-length form. Low levels of three types of products were seen in cumulus cells from cumulus oocytes complexes (COCs), whereas the full-length form was significantly increased by 12-h cultivation with FSH. The addition of FSH also significantly increased the binding level of biotinylated hCG to COCs. The formation of LHR in FSH-stimulated cumulus cells was not affected by additional 0.5 mM phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX), and the oocytes were synchronized to the germinal vesicle (GV) II stage by exposure to 0.5 mM IBMX and FSH for 20 h. The binding of LH to its receptor induced a further increase in cAMP level and progesterone production and acceleration of meiotic progression to the metaphase I stage. The oocytes cultured with LH for 24 h following cultivation with FSH and IBMX were used for in vitro fertilization. At 6 days after in vitro fertilization, blastocyst rate in oocytes matured under these conditions was significantly higher than that of oocytes cultured in the absence of LH. Treatment of oocytes with FSH and 0.5 mM IBMX to express LH receptor in cumulus cells while holding oocytes at the GV II stage is a very beneficial way to produce in vitro-matured oocytes, which have high developmental competence.
我们研究了猪卵母细胞周围卵丘细胞中促黄体生成素受体(LHR)的形成以及LHR在卵母细胞减数分裂成熟中的作用。除全长形式外,在卵丘细胞中检测到至少三种LHR mRNA的剪接变体。在卵丘卵母细胞复合体(COC)的卵丘细胞中可见三种类型产物的低水平表达,而通过与促卵泡素(FSH)培养12小时,全长形式显著增加。添加FSH也显著增加了生物素化人绒毛膜促性腺激素(hCG)与COC的结合水平。FSH刺激的卵丘细胞中LHR的形成不受额外添加的0.5 mM磷酸二酯酶抑制剂3-异丁基-1-甲基黄嘌呤(IBMX)的影响,并且通过暴露于0.5 mM IBMX和FSH 20小时,卵母细胞同步到生发泡(GV)II期。促黄体生成素(LH)与其受体的结合导致环磷酸腺苷(cAMP)水平进一步升高、孕酮产生增加以及减数分裂进程加速至中期I期。在用FSH和IBMX培养后用LH培养24小时的卵母细胞用于体外受精。体外受精6天后,在这些条件下成熟的卵母细胞的囊胚率显著高于未添加LH培养的卵母细胞。用FSH和0.5 mM IBMX处理卵母细胞以在卵丘细胞中表达LH受体,同时将卵母细胞维持在GV II期,是产生具有高发育能力的体外成熟卵母细胞的非常有益的方法。