Poff J P, Fairchild D L, Condon W A
Department of Animal and Nutritional Sciences, University of New Hampshire, Durham 03824.
J Reprod Fertil. 1988 Jan;82(1):135-43. doi: 10.1530/jrf.0.0820135.
Corpora lutea were removed from regularly cycling dairy cows, dissociated with collagenase and cultured for 8 or 10 days in Ham's F-12 medium. In Exp. 1 treatment with insulin, or an insulin-transferrin-selenium combination (ITS), increased progesterone production from basal levels on Day 4 of culture to 234% (P less than 0.01) above controls on Day 10. LH alone increased progesterone production 45% above controls on Day 10 (P greater than 0.05). When LH was combined with insulin or ITS, progesterone production was stimulated to an average of 1802% (P less than 0.01) above controls on Day 10 of culture. Transferrin or selenium without insulin did not allow LH to stimulate progesterone synthesis. In Exp. II, LH alone or LH plus gentamicin or penicillin-streptomycin increased progesterone production from basal levels on Day 2 steadily to an average of 468% (P less than 0.01) above controls (no antibiotics) by Day 8 of culture. The addition of amphotericin-B, alone or in combination with the other antibiotics, inhibited all LH-stimulated progesterone synthesis, but did not affect basal progesterone levels. We conclude that insulin is essential for maximal steroidogenesis in a bovine luteal cell culture system, and that LH-stimulated progesterone production is inhibited in the presence of amphotericin-B, but is not inhibited by gentamicin or penicillin-streptomycin. The elimination of amphotericin-B, coupled with the addition of insulin to the cell culture system increased the responsiveness of the cells to LH. These culture conditions represent the first report in which LH increased total progesterone production for 10 days, maintaining luteal function in a chemically-defined culture system.
从处于正常发情周期的奶牛体内取出黄体，用胶原酶进行解离，并在哈姆氏F-12培养基中培养8天或10天。在实验1中，用胰岛素或胰岛素-转铁蛋白-硒组合（ITS）处理，可使培养第4天的基础孕酮产量在培养第10天增加至比对照组高234%（P<0.01）。单独使用促黄体生成素（LH）可使培养第10天的孕酮产量比对照组高45%（P>0.05）。当LH与胰岛素或ITS联合使用时，培养第10天的孕酮产量被刺激至比对照组平均高1802%（P<0.01）。没有胰岛素的转铁蛋白或硒不能使LH刺激孕酮合成。在实验II中，单独使用LH或LH加庆大霉素或青霉素-链霉素可使培养第2天的基础孕酮产量在培养第8天稳步增加至比对照组（无抗生素）平均高468%（P<0.01）。单独添加两性霉素B或与其他抗生素联合添加，均抑制所有LH刺激的孕酮合成，但不影响基础孕酮水平。我们得出结论，在牛黄体细胞培养系统中，胰岛素对于最大程度的类固醇生成至关重要，并且在两性霉素B存在的情况下，LH刺激的孕酮产生受到抑制，但不受庆大霉素或青霉素-链霉素的抑制。去除两性霉素B并在细胞培养系统中添加胰岛素，可增加细胞对LH的反应性。这些培养条件代表了首次报道LH在化学限定的培养系统中可使总孕酮产量增加10天，并维持黄体功能。
