Isolation, characterization, and culture of cell subpopulations forming the pregnant rat corpus luteum.
Isolation, characterization, and culture of cell subpopulations forming the pregnant rat corpus luteum.
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DOI：
10.1210/endo.130.2.1733737
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发表时间：
1992-02
期刊：
影响因子：
4.8
通讯作者：
S. Nelson;M. McLean;P. Jayatilak;G. Gibori
中科院分区：
文献类型：
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作者：
S. Nelson;M. McLean;P. Jayatilak;G. Gibori
本研究的目的是分离、鉴定和培养形成妊娠大鼠黄体的大小黄体细胞亚群。由于大的黄体细胞非常脆弱，不能在标准细胞分散中存活，因此开发了一种允许小细胞和大细胞在无血清培养基中存活和长期培养的方法。这两个黄体细胞群不仅在大小上不同，而且在培养物中的形态也不同。小黄体细胞（直径12-20 μ）的特征是核大，卵圆形，含有少量脂滴，呈星状。相比之下，大的黄体细胞具有较小的球形核，高脂质含量，并且在培养中不完全变平，最可能是由于脂滴丰富。两种黄体细胞类型都表达参与类固醇生成的3 β HSD和细胞色素P450酶。然而，充满脂质的大黄体细胞分泌最多的孕酮、雄激素和雌二醇;表达更大量的P450 SCC和P450 AROM;并具有更多的PRL和LH受体。尽管LH受体在大黄体细胞中的表达更高，但小黄体细胞和大黄体细胞对LH的反应是类固醇生成量的同等增加。在无血清培养中，黄体细胞产生孕酮长达20天;然而，外源性胆固醇来源是最大孕酮分泌的先决条件。小黄体细胞和大黄体细胞培养物的孕酮分泌模式与混合细胞群的孕酮分泌模式明显不同。当非类固醇黄体细胞与大黄体细胞共培养时，观察到孕酮分泌增加了几倍。这种刺激发生，即使当细胞在没有外源性来源的胆固醇共培养。综上所述，我们成功地建立了一种分离、分离和独立培养大鼠黄体细胞的方法。结果表明，这两个细胞群的类固醇生成能力的显着差异是由于，在很大程度上，在他们的大小，而不是他们的起源在卵泡的差异。此外，这些结果揭示了形成黄体的非类固醇生成细胞对黄体细胞类固醇生成的重要影响。
The aim of this investigation was to isolate, characterize, and culture the small and large luteal cell subpopulations forming the corpus luteum of the pregnant rat. Since the large luteal cells are extremely fragile and do not survive standard cell dispersion, a method which allows the survival and the long-term culture in serum-free media of small and large cells was developed. The two luteal cell populations differed not only by their size but also by their morphology in culture. The small luteal cells (12-20 mu in diameter) are characterized by a large oval nucleus, contain few lipid droplets and have a stellate shape. In contrast, the large luteal cells have a smaller spherical nucleus, high lipid content, and do not flatten out completely in culture, most probably due to the abundance of lipid droplets. Both luteal cell types express 3 beta HSD and the cytochrome P450 enzymes involved in steroidogenesis. However, it is the lipid filled large luteal cells that secrete the most progesterone, androgen, and estradiol; express greater amounts of P450scc and P450AROM; and possess more PRL and LH receptors. Despite the greater expression of LH receptor in the large luteal cells, small and large luteal cells responded to LH with equal increase in steroidogenic output. In serum free culture, luteal cells produced progesterone for up to 20 days; however, an exogenous source of cholesterol was a prerequisite for maximal progesterone secretion. The pattern of progesterone secretion by cultures of small and large luteal cells differed remarkably from that of mixed cell population. When nonsteroidogenic corpus luteum cells were cocultured with the large luteal cells, a severalfold increase in progesterone secretion was observed. This stimulation occurred even when cells were cocultured in the absence of exogenous source of cholesterol. In summary, a successful method was developed to disperse, isolate, and independently culture the two luteal cell populations forming the rat corpus lutem. The results indicate that the marked difference in the steroidogenic capacity of these two cell populations is due, in large part, to the difference in their size rather than to their origin in the follicle. In addition, the results have revealed an important effect of the nonsteroidogenic cells forming the corpus luteum on luteal cell steroidogenesis.