Boulle N, Gicquel C, Logié A, Christol R, Feige J J, Le Bouc Y
Laboratoire d'Explorations Fonctionnelles Endocriniennes, Hôpital Trousseau, Paris, France.
Endocrinology. 2000 Sep;141(9):3127-36. doi: 10.1210/endo.141.9.7632.
The IGF system is thought to play a major role in adrenocortical tumorigenesis. In this study, we used the NCI H295R cell line as a model to investigate the effects of fibroblast growth factor-2 (FGF-2), a potent mitogen for normal adrenal cells, on the proliferation and on the expression of the IGF system in cultured adrenocortical tumor cells. Three immunoreactive FGF-2 isoforms of molecular masses 18, 22, and 24 kDa were detected in H295R cell extracts. Recombinant human FGF-2 stimulated the proliferation of adrenocortical tumor cells in a dose- and time-dependent manner, with a maximal effect at a concentration of about 1 ng/ml. Treatment of H295R cells with 10 ng/ml FGF-2 for 7 days had no significant effect on IGF-II messenger RNA levels. However, a marked increase in levels of intracellular IGF-II protein was detected by immunoblotting. In contrast, FGF-2 induced a marked decrease in the amount of IGF-II protein secreted, with the disappearance of mature IGF-II and secretion of higher molecular forms of the growth factor, suggesting modifications of IGF-II processing. Cell cultures in the presence of brefeldin A (1 microg/ml), a specific inhibitor of protein secretion, suggested that FGF-2 did not increase IGF-II synthesis but instead inhibited the secretion of pro-IGF-II from H295R cells, thereby impairing the final steps of IGF-II processing to the mature 7.5-kDa peptide. At the same concentrations, FGF-2 also decreased both IGFBP-2 messenger RNA and secreted protein, which might increase IGF-II bioavailability. No proteolysis of IGFBP-2 was detected in FGF-2-conditioned medium. Altogether, these results indicate that FGF-2 is mitogenic for NCI H295R tumor cells and regulates the expression of both IGF-II and IGFBP-2 in this tumor model. Moreover, this study shows a novel effect of FGF-2, by which this growth factor modulates the processing of pro-IGF-II.
胰岛素样生长因子（IGF）系统被认为在肾上腺皮质肿瘤发生过程中起主要作用。在本研究中，我们以NCI H295R细胞系为模型，研究成纤维细胞生长因子-2（FGF-2，一种对正常肾上腺细胞有强大促有丝分裂作用的因子）对培养的肾上腺皮质肿瘤细胞增殖及IGF系统表达的影响。在H295R细胞提取物中检测到三种免疫反应性FGF-2同工型，分子量分别为18、22和24 kDa。重组人FGF-2以剂量和时间依赖性方式刺激肾上腺皮质肿瘤细胞增殖，在浓度约为1 ng/ml时达到最大效应。用10 ng/ml FGF-2处理H295R细胞7天，对IGF-II信使核糖核酸水平无显著影响。然而，通过免疫印迹检测到细胞内IGF-II蛋白水平显著升高。相反，FGF-2导致分泌的IGF-II蛋白量显著减少，成熟IGF-II消失，生长因子出现更高分子形式的分泌，提示IGF-II加工过程发生改变。在布雷菲德菌素A（1 μg/ml，一种蛋白质分泌特异性抑制剂）存在的情况下进行细胞培养表明，FGF-2并未增加IGF-II的合成，而是抑制了H295R细胞中前体IGF-II的分泌，从而损害了IGF-II加工为成熟7.5 kDa肽的最后步骤。在相同浓度下，FGF-2还降低了IGFBP-2信使核糖核酸和分泌蛋白水平，这可能会增加IGF-II的生物利用度。在FGF-2条件培养基中未检测到IGFBP-2的蛋白水解。总之，这些结果表明FGF-2对NCI H295R肿瘤细胞有促有丝分裂作用，并在该肿瘤模型中调节IGF-II和IGFBP-2的表达。此外，本研究显示了FGF-2的一种新作用，即这种生长因子调节前体IGF-II的加工过程。
