Monniaux D, Pisselet C, Fontaine J
INRA, Station de Physiologie de la Reproduction des Mammifères Domestiques, URA CNRS 1291, Nouzilly, France.
J Endocrinol. 1994 Sep;142(3):497-510. doi: 10.1677/joe.0.1420497.
Granulosa cells of ovarian follicles both proliferate and undergo differentiation. In vivo, an inverse relationship between proliferation and steroidogenesis is observed. However, both processes can be enhanced by insulin-like growth factor-I (IGF-I) in vitro. Studies were undertaken in the ewe to understand the mechanisms controlling the balance between proliferation and differentiation in cultured granulosa cells from antral follicles better. For this purpose, granulosa cells from ovine small follicles (1-3 mm in diameter) and large follicles (5-7 mm in diameter) were compared for progesterone secretion, cytochrome P450 side-chain cleavage (P450scc) expression and their proportions of non-proliferating (G0) cells, in response to IGF-I and FSH stimulation in vitro. IGF-I mainly enhanced the proliferation of granulosa cells from small follicles but it strongly increased progesterone secretion and P450scc expression in granulosa cells from large follicles, in synergy with FSH. Blocking granulosa cell proliferation by the administration of colcemid or aphidicolin had no effect or a weak stimulating effect on progesterone secretion. At the beginning of the culture period, the proportion of non-proliferating cells, estimated by continuous [3H]thymidine labelling experiments, was clearly higher in large than in small follicles (91% vs 30%, P < 0.001). For both cell types, treatment with IGF-I in vitro reduced the proportion of non-proliferating cells at 72 h of culture (40% vs 70% respectively in IGF-I-stimulated and unstimulated cells from large follicles, P < 0.001, and 17% vs 30% respectively in IGF-I-stimulated and unstimulated cells from small follicles, P < 0.001). Treatment with FSH had no effect on the proportion of non-proliferating cells. As revealed by immunohistochemistry experiments, IGF-I, in synergy with FSH, clearly increased the percentage of cells expressing P450scc enzyme and the intensity of staining in granulosa cells from large follicles. Unexpectedly, heavily stained cells in mitosis were observed in IGF-I-stimulated cells from large follicles after 96 h of culture, suggesting that dividing cells might also produce progesterone. Overall, these results support the hypothesis that the growth-promoting and the cytodifferentiative effects of IGF-I are clearly distinct. Moreover, they suggest that uncoupling between proliferation and steroidogenesis may occur in cultured ovine granulosa cells. The loss of proliferative activity accompanying terminal follicular growth in vivo could be reversed in vitro. During terminal follicular growth in vivo, the existence of an active mechanism inhibiting granulosa cell proliferation, and unrelated to terminal differentiation, is therefore strongly suspected.
卵巢卵泡的颗粒细胞既能增殖又能进行分化。在体内，观察到增殖与类固醇生成之间呈负相关。然而，在体外，胰岛素样生长因子-I（IGF-I）可增强这两个过程。在母羊身上进行了研究，以更好地了解控制体外培养的窦状卵泡颗粒细胞增殖与分化平衡的机制。为此，比较了来自绵羊小卵泡（直径1 - 3毫米）和大卵泡（直径5 - 7毫米）的颗粒细胞在体外对IGF-I和促卵泡激素（FSH）刺激的孕酮分泌、细胞色素P450侧链裂解酶（P450scc）表达以及非增殖（G0）细胞比例。IGF-I主要增强小卵泡颗粒细胞的增殖，但与FSH协同作用时，它能强烈增加大卵泡颗粒细胞的孕酮分泌和P450scc表达。通过给予秋水仙酰胺或阿非迪霉素阻断颗粒细胞增殖，对孕酮分泌没有影响或有微弱的刺激作用。在培养期开始时，通过连续[3H]胸腺嘧啶核苷标记实验估计，大卵泡中非增殖细胞的比例明显高于小卵泡（91%对30%，P < 0.001）。对于两种细胞类型，体外使用IGF-I处理可降低培养72小时时非增殖细胞的比例（大卵泡中IGF-I刺激组和未刺激组分别为40%对70%，P < 0.001；小卵泡中IGF-I刺激组和未刺激组分别为17%对30%，P < 0.001）。FSH处理对非增殖细胞比例没有影响。免疫组织化学实验显示，IGF-I与FSH协同作用，明显增加了大卵泡颗粒细胞中表达P450scc酶的细胞百分比和染色强度。出乎意料的是，培养96小时后，在大卵泡的IGF-I刺激细胞中观察到有丝分裂期染色深的细胞，这表明正在分裂的细胞也可能产生孕酮。总体而言，这些结果支持以下假设：IGF-I的促生长作用和细胞分化作用明显不同。此外，它们表明在培养的绵羊颗粒细胞中可能发生增殖与类固醇生成的解偶联。体内卵泡终末生长伴随的增殖活性丧失在体外可能被逆转。因此，强烈怀疑在体内卵泡终末生长过程中存在一种抑制颗粒细胞增殖且与终末分化无关的活跃机制。
