Ciereszko R E, Petroff B K, Ottobre A C, Guan Z, Stokes B T, Ottobre J S
Department of Animal Sciences, Ohio State University, Columbus, Ohio, 43210, USA.
J Endocrinol. 1998 Nov;159(2):201-9. doi: 10.1677/joe.0.1590201.
Previously, we reported that administration of prolactin (PRL) during the early luteal phase in sows increases plasma progesterone concentrations. In the current study, we searched for the mechanisms by which PRL exerts this luteotrophic effect. The objectives of the study were (1) to examine the effect of PRL and/or low-density lipoproteins (LDL) on progesterone production by porcine luteal cells derived from early corpora lutea, and (2) to assess the ability of PRL to activate phosphoinositide-specific phospholipase C (PI-PLC) and protein kinase C (PKC) in these luteal cells. Ovaries with early corpora lutea (day 1-2 of the oestrous cycle) were obtained from the slaughterhouse. Progesterone production by dispersed luteal cells was measured after treatment with PRL, phorbol 12-myristate 13-acetate or inhibitors of PKC in the presence or absence of LDL. LDL increased progesterone concentration in the incubation medium (304.5 vs 178.6 ng/ml in control, P<0.05). PRL augmented LDL-stimulated progesterone secretion by luteal cells (to 416 ng/ml, P<0.05), but PRL alone did not affect progesterone production (209.6 ng/ml, P>0.05). Staurosporine, a PKC inhibitor, inhibited progesterone secretion stimulated by the combined action of LDL and PRL; however, such inhibition was not demonstrated when cells were treated with the PKC inhibitor, H-7. PKC activation was assessed by measuring the specific association of [H]phorbol dibutyrate (H-PDBu) with luteal cells after treatment with PRL or ionomycin (a positive control). PRL and ionomycin increased H-PDBu-specific binding in early luteal cells by 28+/-5.5% (within 5 min) and 70.2+/-19.3% (within 2 min) over control binding respectively (P<0.05). In addition, PRL did not augment the LDL-stimulated progesterone production in PKC-deficient cells. In contrast with PKC, total inositol phosphate accumulation, as well as intracellular free calcium concentrations, were not affected by PRL in the current study. We conclude that PRL, in the presence of LDL, stimulates progesterone production by early corpora lutea in vitro. Moreover, PRL appears to activate PKC, but not PI-PLC, in these cells. Thus intracellular transduction of the PRL signal may involve activation of PKC that is not dependent on PI-PLC.
此前，我们报道过在母猪黄体期早期给予催乳素（PRL）可提高血浆孕酮浓度。在本研究中，我们探寻了PRL发挥这种促黄体作用的机制。本研究的目的是：（1）检测PRL和/或低密度脂蛋白（LDL）对源自早期黄体的猪黄体细胞孕酮生成的影响；（2）评估PRL激活这些黄体细胞中磷酸肌醇特异性磷脂酶C（PI-PLC）和蛋白激酶C（PKC）的能力。从屠宰场获取处于发情周期第1 - 2天、带有早期黄体的卵巢。在用PRL、佛波醇12 - 肉豆蔻酸酯13 - 乙酸酯或PKC抑制剂处理分散的黄体细胞后，检测其孕酮生成情况，处理过程中存在或不存在LDL。LDL使孵育培养基中的孕酮浓度升高（对照组为178.6 ng/ml，处理组为304.5 ng/ml，P<0.05）。PRL增强了LDL刺激的黄体细胞孕酮分泌（达到416 ng/ml，P<0.05），但单独使用PRL对孕酮生成没有影响（209.6 ng/ml，P>0.05）。PKC抑制剂星形孢菌素抑制了LDL和PRL联合作用刺激的孕酮分泌；然而，当用PKC抑制剂H - 7处理细胞时，未观察到这种抑制作用。通过测量用PRL或离子霉素（阳性对照）处理后黄体细胞中[H]佛波醇二丁酸酯（H - PDBu）的特异性结合来评估PKC的激活情况。与对照结合相比，PRL和离子霉素分别使早期黄体细胞中H - PDBu特异性结合在5分钟内增加了28±5.5%，在2分钟内增加了70.2±19.3%（P<0.05）。此外，PRL在PKC缺陷细胞中并未增强LDL刺激的孕酮生成。与PKC不同，在本研究中，PRL对总肌醇磷酸积累以及细胞内游离钙浓度没有影响。我们得出结论，在存在LDL的情况下，PRL在体外刺激早期黄体的孕酮生成。此外，PRL似乎激活了这些细胞中的PKC，但未激活PI - PLC。因此，PRL信号的细胞内转导可能涉及不依赖PI - PLC的PKC激活。
