Thompson M A, Cox A J, Whitehead R H, Jonas H A
University of Melbourne Department of Medicine, Victoria, Australia.
Endocrinology. 1990 Jun;126(6):3033-42. doi: 10.1210/endo-126-6-3033.
We have shown that a pleomorphic cell line of abnormal human karyotype derived from a stomach carcinoma (LIM-1839) proliferates in serum-free medium, expresses insulin-like growth factor II (IGF-II) mRNA, and secretes IGF-II (up to 56 ng/ml in serum-free conditioned medium, as measured in a rat liver RRA. No detectable levels of IGF-I can be measured in serum-free conditioned medium by RIA. These cells also secrete IGF-binding proteins, detected by a charcoal adsorption assay. The release of IGF-II and IGF binding proteins into serum-free conditioned medium (1.7 pmol/10(6) cells.24 h and 0.8 pmol binding sites/10(6) cells.24 h for 3 days, respectively) is inhibited 80% by cycloheximide (10 micrograms/ml). The LIM-1839 cells have type I and type II IGF receptors, determined by affinity cross-linking and competition binding studies. These cells proliferated 1.6-fold over 4 days in serum-free medium, with fresh medium changes on days 0 and 2: their growth was inhibited 56% by 40 micrograms/ml Sm 1.2, a monoclonal antibody which recognizes IGF-I and IGF-II. The addition of 20 and 50 ng/ml multiplication stimulating activity (rat IGF-II) caused 1.8- and 1.7-fold increases in cell growth between days 0 and 4 compared to controls, while [Thr59]IGF-I, at 20 and 50 ng/ml, caused 1.6- and 2.0-fold increases. Insulin, at 2 and 10 micrograms/ml, had no significant effect. The stimulatory effects of endogenous and exogenous IGFs on LIM-1839 cell proliferation were inhibited by a monoclonal antibody to the type I IGF receptor, alpha IR-3. These results suggest that the LIM-1839 cells are biologically responsive to endogenously produced IGF-II, and may thereby provide an in vitro model for autocrine regulation of human tumor growth by IGF-II.
我们已证明，源自胃癌的具有异常人类核型的多形细胞系（LIM-1839）能在无血清培养基中增殖，表达胰岛素样生长因子II（IGF-II）mRNA，并分泌IGF-II（在无血清条件培养基中高达56 ng/ml，通过大鼠肝脏放射受体分析测定）。通过放射免疫分析在无血清条件培养基中检测不到IGF-I的水平。这些细胞还分泌IGF结合蛋白，通过活性炭吸附分析检测到。IGF-II和IGF结合蛋白释放到无血清条件培养基中（分别为1.7 pmol/10⁶细胞·24小时和0.8 pmol结合位点/10⁶细胞·24小时，持续3天），被放线菌酮（10微克/毫升）抑制80%。通过亲和交联和竞争结合研究确定，LIM-1839细胞具有I型和II型IGF受体。这些细胞在无血清培养基中4天内增殖了1.6倍，在第0天和第2天更换新鲜培养基：它们的生长被40微克/毫升的Sm 1.2抑制了56%，Sm 1.2是一种识别IGF-I和IGF-II的单克隆抗体。添加20和50 ng/ml的增殖刺激活性（大鼠IGF-II）导致与对照相比，在第0天到第4天细胞生长分别增加了1.8倍和1.7倍，而20和50 ng/ml的[Thr59]IGF-I导致增加了1.6倍和2.0倍。2和10微克/毫升的胰岛素没有显著影响。I型IGF受体的单克隆抗体αIR-3抑制了内源性和外源性IGF对LIM-1839细胞增殖的刺激作用。这些结果表明，LIM-1839细胞对内源性产生的IGF-II具有生物学反应性，因此可能为IGF-II对人类肿瘤生长的自分泌调节提供一个体外模型。
