TNF-α对雌性大鼠生殖轴调控作用的研究

The Research on the Regulating Effect of TNF-αon the Reproductive Axis of Female Rat

目的 通过动物实验探讨肿瘤坏死因子 (TNF α)在生殖轴调控中的作用及作用部位。方法 情前期成年雌性S D大鼠 4 4只 ,取下丘脑、腺垂体及卵巢分别进行原代细胞培养 ,贴壁后 ,于腺垂体及卵巢培养液中分别加入TNF α (10 0ng/ml) ,孵育 2 4h后 ,进行顺序灌流共 5次 ,将培养液冷冻干燥后按放免试剂盒说明测FSH、LH、E2 及P ,并收集各组培养卵巢颗粒细胞 ,用流式细胞仪检测颗粒细胞凋亡率 ,观察TNF α对生殖轴调控作用及卵巢颗粒细胞增殖的作用的影响。结果 (1)TNF α (10 0ng/ml)明显抑制原代培养的雌性动情前期大鼠卵巢组织E2 及P的分泌量。 (2 )TNF α (10 0ng/ml)明显抑制原代培养的雌性动情前期大鼠腺垂体细胞LH、FSH的分泌量。 (3)FCM检测结果 :实验组卵巢颗粒细胞凋亡率为 (37 84± 3 84 ) %与对照组 (2 6 3±0 37) %相比有显著性差异 (P <0 0 1) ,说明TNF α能够抑制颗粒细胞增殖 ,并促进其凋亡。结论 TNF α在腺垂体及卵巢水平上同时发挥作用 ,抑制FSH、LH、E2 、P的合成 ,同时抑制颗粒细胞的增殖 ,促进其凋亡。

Objective The main idea of this study was to investigate the effect and the position of TNF α on the reproductive axis of female rat in vitro.Methods 44 adult female Sprague Dawley rats were decapitated.All hypothalami,anterior pituitaries and ovaries from them were harvested and primary monolayer cultures were made during the morning of protestrus as determined by vaginal smears.When these cells had been attached to the wells,TNF α was added to pituitary and ovary after 24h incubation period.Sequence perfusion in hypothalamus,anterior pituitary and ovary technique were used every 24h.LH and FSH in the anterior pituitary culture fluid was detected by RIA,so did as P and E 2 in ovary culture fluid.At the same time,all granulosa cells were harvested.FCM was used to measure the ratio of granulosa apoptosis.Results (Ⅰ)The results showed that TNF α(100ng/ml)significantly inhibited P and E 2 release relative to control.(Ⅱ)The results also showed that TNF α played a inhibited role in LH and FSH release in primary pituitary cells from female protestrus rats relative to control.(Ⅲ)The results of flow cytometry:the ratios of apopotis granulosa cells were(37 84±3 84)% The difference was significant( P< 0 01)relative to control well(2 63±0 37)%.Conclusions TNF α inhibits E 2,P,LH and FSH release in the level of ovary and pituitary inhibits the reproduction of granulosa cell and as the same time stimulates the apoptis of granulosa cell.