巨噬细胞集落刺激因子对卵泡颗粒黄体细胞性激素分泌的影响及其受体的测定

Effects of macrophage colony-stimulating factor on production of estradiol and progesterone by human luteinized granulosa cells in vitro and detection of macrophage colony-stimulating factor receptor on human luteinized granulosa cells

目的 了解人卵泡颗粒黄体细胞中巨噬细胞集落刺激因子 (M CSF)受体的表达 ,M CSF在体外对人卵泡颗粒黄体细胞产生雌、孕激素的影响。方法 采用免疫细胞化学染色法 ,测定 2 0例行卵母细胞浆内单精子注射治疗患者的卵泡颗粒黄体细胞的M CSF受体 ;采用酶免疫分析法 ,测定卵泡颗粒黄体细胞在M CSF(浓度分别为 0、10、2 5、5 0、10 0、2 5 0ng/ml)单独及与促卵泡激素 (FSH ,浓度75IU/ml)联合作用 72h后的上清液中雌二醇 (E2 )和孕酮 (P)的浓度。结果 约 80 %的颗粒黄体细胞膜上M CSF受体阳性。在无FSH存在时 ,M CSF空白对照颗粒黄体细胞培养上清液中E2 浓度为(2 185± 189)pmol/L ,P浓度为 (315 7± 4 0 1)nmol/L ;M CSF为 10～ 10 0ng/ml时 ,E2 浓度在 (2 789± 36 5 )～ (42 82± 318)pmol/L之间 ,P浓度在 (42 5 6± 5 95 )～ (7789± 82 8)nmol/L之间。在有FSH作用时 ,M CSF空白对照的E2 和P浓度分别为 (5 0 4 5± 4 86 )pmol/L和 (86 6 7± 92 3)nmol/L ;M CSF为 10～ 10 0ng/ml时 ,E2 浓度在 (6 5 6 7± 6 73)～ (8373± 935 )pmol/L之间 ,P浓度在 (10 999± 985 )～ (14 990±115 8)nmol/L之间。M CSF促颗粒黄体细胞E2 和P分泌的作用随浓度增加而增强 (P均 <0 0 5 ) ;M

Objective To investigate the effects of macrophage colony stimulating factor (M CSF) on estradiol and progesterone production by human luteinized granulosa cells in vitro and to detect M CSF receptor in human granulosa cells Methods Human luteinized granulosa cells (LGC) were isolated from follicular fluid of superovulated infertile patients undergoing intracytoplasmatic sperm injection Some of the LGC were used for detecting M CSF receptor by immunocytochemical staining (ABC method) Most of them were cultured with HAM′s F 10 medium plus various concentration of M CSF (0, 10, 25, 50, 100, 250 ng/ml) in the presence or absence of follicle stimulating hormone (FSH) Media were collected at 72 hours after culture and estradiol (E 2) and progesterone (P) in media were measured by enzyme immunoassays Results About 80% of the LGC presented a positive M CSF receptor staining with the immunochemical signal on cell membrane After 72 hours culture of LGC, in absence of FSH, the baseline concentration of E 2 and P were (2 185± 189) pmol/L, (3 157±401) nmol/L respectively; with the increasing dose of M CSF from 10 to 100 ng/ml, the concentrations of E 2 increased from (2 789± 365)pmol/L to (4 282±318) pmol/L, and those of P increased from (4 256± 595) nmol/L to (7 789± 828) nmol/L In presence of FSH (75 IU/ml) the baseline concentrations of E 2 and P were (5 045± 486) pmol/L and (8 667± 923) nmol/L respectively; while the 10～100 ng/ml M CSF was added the concentrations of E 2 and P were from (6 567± 673) to (8 373± 935) pmol/L and from (10 999± 985) to (14 990± 1 158) nmol/L respectively Thus M CSF caused a significant dose dependent increase of estradiol and progesterone production ( P <0 05), and M CSF+ FSH further stimulated higher production of E 2 and P by LGC than M CSF or FSH alone ( P <0 05). Conclusions M CSF receptor was expressed on membrane of human LGC M CSF enhances estradiol and progesterone production by LGC and FSH has an additional or synergetic effect with M CSF