Neely E K, Rosenfeld R G
Division of Pediatric Endocrinology, Stanford University, California 94305.
Endocrinology. 1992 Feb;130(2):985-93. doi: 10.1210/endo.130.2.1370799.
Recent studies have provided a consensus that insulin-like growth factor-I (IGF-I) stimulates IGF-binding protein-3 (IGFBP-3) in vivo and in vitro. While it also appears well established that IGFBP-1 is inversely related to insulin concentrations, evidence regarding regulation of other IGFBP is inconclusive. Using immunoprecipitation and Western ligand blot, we have characterized the IGFBPs released into conditioned medium (CM) by cells from the adult human fibroblast cell line N3652 and the human epidermal squamous cell carcinoma line SCL-1. N3652 cells expressed IGFBP-3, IGFBP-2, a 24-kilodalton (kDa) IGFBP presumed to be IGFBP-4, and IGFBPs at 30 and 28 kDa. SCL-1 expressed IGFBP-3 and a putative IGFBP-4, with intermediate bands at 34 and 30 kDa. As determined by ligand blot of CM from confluent cells 72 h after the addition of peptides to serum-free medium, IGF-I and IGF-II potently stimulated IGFBP-3 in both cell lines, but otherwise IGFBP regulation in the two cells diverged. In N3652 cells, IGFBP-3 concentrations in CM increased to 700% and 800% of basal levels in the presence of IGF-I and IGF-II (at 100 ng/ml; n = 5 experiments), respectively. IGFBP-3 was not affected by insulin up to 10 micrograms/ml. In contrast, IGFBP-4 levels were diminished 54% and 73% by 100 ng/ml IGF-I and IGF-II, respectively, with no response to insulin. In SCL-1 cells, IGF-I and IGF-II were virtually identical in stimulating a mean 200% increase in IGFBP-3 (n = 5 experiments). Insulin was less potent, but caused a significant stimulation of IGFBP-3 levels. IGF-I, IGF-II, and insulin all stimulated an approximately 50% increase in IGFBP-4 concentrations. To test the hypothesis that IGF-induced alterations in IGFBP-3 and IGFBP-4 concentrations were regulated via the type 1 IGF receptor, we attempted to block IGFBP changes with type 1 IGF receptor antibody alpha IR-3 and to induce IGFBP changes with an IGF-II analog, [Leu27]IGF-II, with little affinity for the type 1 receptor. alpha IR-3 failed to block either the IGF-induced rise in IGFBP-3 in each cell line or the decline in IGFBP-4 in N3652 CM. [Leu27]IGF-II was as potent as IGF-II or IGF-I in inducing changes in IGFBP-3 and IGFBP-4 concentrations.(ABSTRACT TRUNCATED AT 400 WORDS)
最近的研究已达成共识，即胰岛素样生长因子-I（IGF-I）在体内和体外均可刺激胰岛素样生长因子结合蛋白-3（IGFBP-3）。虽然IGFBP-1与胰岛素浓度呈负相关这一点似乎也已明确确立，但关于其他IGFBP调控的证据尚无定论。我们运用免疫沉淀和Western配体印迹法，对来自成人人类成纤维细胞系N3652和人类表皮鳞状细胞癌系SCL-1的细胞释放到条件培养基（CM）中的IGFBP进行了表征。N3652细胞表达IGFBP-3、IGFBP-2、一种推测为IGFBP-4的24千道尔顿（kDa）IGFBP，以及30 kDa和28 kDa的IGFBP。SCL-1表达IGFBP-3和一种假定的IGFBP-4，在34 kDa和30 kDa处有中间条带。通过对无血清培养基中添加肽72小时后的汇合细胞的CM进行配体印迹分析确定，IGF-I和IGF-II在两种细胞系中均有力地刺激了IGFBP-3，但除此之外，两种细胞中IGFBP的调控出现了分歧。在N3652细胞中，在存在IGF-I和IGF-II（100 ng/ml；n = 5次实验）的情况下，CM中的IGFBP-3浓度分别增加至基础水平的700%和800%。高达10微克/毫升的胰岛素对IGFBP-3没有影响。相比之下，100 ng/ml的IGF-I和IGF-II分别使IGFBP-4水平降低了54%和73%，对胰岛素无反应。在SCL-1细胞中，IGF-I和IGF-II在刺激IGFBP-3平均增加200%方面几乎相同（n = 5次实验）。胰岛素的作用较弱，但导致IGFBP-3水平有显著刺激。IGF-I、IGF-II和胰岛素均使IGFBP-4浓度增加了约50%。为了检验IGF诱导的IGFBP-3和IGFBP-4浓度变化是通过1型IGF受体调控的这一假设，我们试图用1型IGF受体抗体αIR-3阻断IGFBP变化，并使用对1型受体亲和力较低的IGF-II类似物[Leu27]IGF-II诱导IGFBP变化。αIR-3未能阻断每种细胞系中IGF诱导的IGFBP-3升高或N3652 CM中IGFBP-4的降低。[Leu27]IGF-II在诱导IGFBP-3和IGFBP-4浓度变化方面与IGF-II或IGF-I一样有效。（摘要截于400字）
