Gregoraszczuk E L, Bylica A, Gertler A
Department of Animal Physiology, Institute of Zoology, Jagiellonian University, Ingardena 6, 30-060, Krakow, Poland.
Anim Reprod Sci. 2000 Feb 28;58(1-2):113-25. doi: 10.1016/s0378-4320(99)00083-4.
In Experiment 1, the influence of exogenous GH on steroid secretion by granulosa and theca interna cells recovered from small (1-3 mm), medium (4-6 mm) and large (8-12 mm) follicles was tested. In the second experiment, theca cells (Tc) and granulosa cells (Gc) obtained from large follicles were cultured separately or in two types, Tc/Gc co-culture, where both types of cells were mixed in one well or Gc and Tc were separated by cell culture membrane inserts. In the third experiment, the influence of GH on the morphology of Gc and Tc cells and activity of Delta(5),3beta-hydroxysteroid dehydrogenase (3beta-HSD) was studied. Cells were grown in the control medium (M199+5% of calf serum) or supplemented with 100 ng/ml GH. Testosterone (10(-7) M) was added as the aromatase substrate to granulosa cells cultures. The media were assayed after 48 h of culture for progesterone and oestradiol by RIA. GH added to the culture media had no effect on oestradiol and progesterone secretion by granulosa cells isolated from small and medium follicles while it stimulated both oestradiol and progesterone secretion by Gc isolated from large preovulatory follicles. A stimulatory effect on oestradiol secretion by Tc isolated from all size follicles was observed. GH did not stimulate progesterone secretion by Tc isolated from small follicles but stimulated progesterone secretion by Tc isolated from medium and large preovulatory follicles. Both co-culture systems exhibited synergistic effect on oestradiol secretion. The stimulatory effect on progesterone secretion under the influence of GH was observed in Gc cultured alone and Tc cultured alone. In contrast, the secretion of progesterone was attenuated in both co-culture systems and the addition of GH further augmented this attenuation. A statistically significant increase in oestradiol secretion was observed in all culture conditions. The addition of GH to the culture medium stimulated the activity of 3beta-HSD compared with the control culture from both types of cells. In conclusion, the present studies indicate that there are direct and follicular development stage dependent actions of GH on steroidogenesis of porcine follicular cells.
在实验1中，测试了外源性生长激素（GH）对从小（1 - 3毫米）、中（4 - 6毫米）和大（8 - 12毫米）卵泡中回收的颗粒细胞和卵泡内膜细胞类固醇分泌的影响。在第二个实验中，从大卵泡中获得的卵泡内膜细胞（Tc）和颗粒细胞（Gc）分别培养，或进行两种类型的共培养，即Tc/Gc共培养，两种细胞在一个孔中混合，或Gc和Tc通过细胞培养膜插入物分开培养。在第三个实验中，研究了GH对Gc和Tc细胞形态以及δ⁵,3β - 羟基类固醇脱氢酶（3β - HSD）活性的影响。细胞在对照培养基（M199 + 5%小牛血清）中生长，或补充100 ng/ml GH。将睾酮（10⁻⁷ M）作为芳香化酶底物添加到颗粒细胞培养物中。培养48小时后，通过放射免疫分析法（RIA）测定培养基中的孕酮和雌二醇。添加到培养基中的GH对从小卵泡和中卵泡分离的颗粒细胞的雌二醇和孕酮分泌没有影响，而它刺激了从大的排卵前卵泡分离的Gc的雌二醇和孕酮分泌。观察到对从所有大小卵泡分离的Tc的雌二醇分泌有刺激作用。GH不刺激从小卵泡分离的Tc的孕酮分泌，但刺激从中等大小和大的排卵前卵泡分离的Tc的孕酮分泌。两种共培养系统对雌二醇分泌均表现出协同作用。在单独培养的Gc和单独培养的Tc中观察到在GH影响下对孕酮分泌的刺激作用。相反，在两种共培养系统中孕酮分泌均减弱，并且添加GH进一步增强了这种减弱。在所有培养条件下均观察到雌二醇分泌有统计学意义的增加。与两种细胞的对照培养相比，向培养基中添加GH刺激了3β - HSD的活性。总之，本研究表明GH对猪卵泡细胞类固醇生成有直接且依赖卵泡发育阶段的作用。
