瘦素调节人卵巢黄素化颗粒细胞功能的体外研究

Study of effects of leptin on cultured human luteinized granulosa cell function

目的 探讨瘦素、促卵泡激素 (FSH)和胰岛素样生长因子Ⅰ (IGF Ⅰ )对人卵巢黄素化颗粒细胞雌二醇 (E2 )、孕酮 (P)生成的影响及可能的作用机制。方法 培养人黄素化颗粒细胞 ,分别以瘦素 (3 0ng/ml)、FSH (1 0ng/ml)、IGF Ⅰ (30 0ng/ml)以及相同终浓度的瘦素 +FSH、瘦素 +IGF Ⅰ、FSH +IGF Ⅰ、瘦素 +FSH +IGF Ⅰ对其刺激 2 4h ,对照组不加任何药物。对药物作用后的黄素化颗粒细胞行形态学观察、细胞计数 ;用放射免疫法检测培养液中E2 和P水平 ;同时用逆转录聚合酶链反应法对黄素化颗粒细胞行瘦素受体mRNA检测。结果 瘦素、FSH和IGF Ⅰ对黄素化颗粒细胞生长无影响。瘦素对黄素化颗粒细胞E2 生成量无影响 ,对FSH刺激E2 的生成也无影响 ,E2 水平作用前分别为 (0 10 3± 0 0 36 )pmol/10 0 0细胞、(0 32 3± 0 0 4 2 )pmol/10 0 0细胞 ,作用后分别为 (0 12 0± 0 0 0 8)pmol/10 0 0细胞、(0 343± 0 0 34)pmol/10 0 0细胞 ;而对IGF Ⅰ、FSH +IGF Ⅰ刺激黄素化颗粒细胞生成E2 有显著的抑制作用 ,E2 水平作用前后分别为 (0 318± 0 0 37)pmol/10 0 0细胞与 (0 4 93± 0 0 36 )pmol/10 0 0细胞、(0 193± 0 0 2 5 )pmol/10 0 0细胞与 (0 2 5 1± 0 0 33)pmol/10 0 0细胞 (P <0 0 5 。

Objective To assess the effects of leptin on follicle stimulating hormone (FSH) and insulin like growth factor Ⅰ(IGF Ⅰ) induced production of estradiol(E 2) 17β and progesterone(P) in cultured human luteinized granulosa cell(GC) Methods Human luteinized GCs were obtained from pre ovulatory follicles in an in vitro fertilization program, and were cultured for 24 hours in the presence of leptin (3 0 ng/ml) or FSH( 1 0 ng/ml) or IGF Ⅰ(30 0 ng/ml) alone, leptin with FSH or IGF Ⅰor both , and FSH with IGF Ⅰ The conditioned media were aspirated for measuring E 2 and P content Luteinized GC were also counted for cell number and detected the expression of the leptin receptor by reverse transcription polymerase chain reaction analysis Results In cultured luteinized GC system, leptin or FSH or IGF Ⅰalone did not affect the growth of cultured human luteinized GC Leptin alone or in the presence of FSH had no effect on E 2 production E 2 levels in culture media without leptin or FSH were (0 103±0 036) pmol/ 1 000 cells, (0 323±0 042) pmol/ 1 000 cells respectively When cultured with leptin alone or with leptin and FSH, E 2 levels were (0 120±0 008) pmol/ 1 000 cells, (0 343±0 034) pmol/ 1 000 cells Leptin significantly inhibited FSH+IGF Ⅰ or IGF Ⅰ induced E 2 production E 2 levels decreased from (0 318±0 037)pmol/1 000 cells to (0 193±0 025)pmol/1 000 cells(IGF Ⅰ)( P <0 05) and from (0 493±0 036)pmol/1 000 cells to (0 251±0 033)pmol/1 000 cells (FSH+IGF Ⅰ)( P <0 01) However, the production of P did not change The leptin receptor expression was demonstrated in luteinized GC Conclusions Leptin may directly attenuate the IGF Ⅰ or IGF Ⅰand FSH stimulated E 2 synthesis in cultured human luteinized GC by the leptin receptor mechanism