INSULIN-STIMULATED GROWTH THROUGH AUTOCRINE IGF-II SECRETION AND THE IGF-I RECEPTOR IN A MOUSE MAMMARY EPITHELIAL-CELL LINE

Growth and differentiation of mammary tissue is thought to be mediated in part by the insulin-like growth factor system. The mouse mammary e pithelial cell line (COMMA-D/MME) synthesizes and secretes IGFBP-2 and ICFBP-3, expresses receptors for IGF-I, ICF-II and insulin and is gro wth responsive to the ICF family of mitogens. This report demonstrates expression and regulation of autocrine IGF-II secretion in the MME ce ll line. RT-PCR analysis of mRNA from MME cells cultured in serum free media (SFM) demonstrates ICF-II expression and RIA analysis demonstra tes immunodetectable ICF-II protein in the corresponding conditioned m edia. Addition of IGF-II mAb (1 or 5 ng/ml) in SFM inhibited cell prol iferation whereas IGF-I pAb had no effect. Growth experiments showed t hat while both IGF-I (10 ng/ml) and insulin (50 and 1000 ng/ml) were m itogenic to MME cells, differential regulation of autocrine IGF-II sec retion was observed. Analysis of CM showed that autocrine IGF-II was m aximally secreted when cells were treated with insulin. In contrast, I GF-II secretion was reduced dramatically when cells were treated with IGF-I. Insulin stimulated MME cell growth was effectively blocked by t he ICF-II mAb at physiological doses of insulin (10 and 50 ng/ml), but not at pharmacological doses of insulin (1000 ng/ml). Treatments with IGF-II analogs: Leucine(27)-IGF-II (high affinity for the ICF-II rece ptor), Arginine(54)Arginine(55)-IGF-II (high affinity for the IGF-I re ceptor), IGF-l or IGF-II (0.01, 0.1 1.0 and 10 nM) demonstrated the me chanism by which IGF-II stimulates autocrine growth in MME cells. MME cells treated with Leucine(27) IGF-II did not increase cell number com pared to serum free controls. In contrast, cells treated with Arginine (54)Arginine(55)-IGF-II demonstrated a 1.6-fold growth increase in cel l proliferation. Thus, MME cell proliferation response is via the IGF- I receptor. These data demonstrate that IGF-II is expressed and secret ed by MME cells. Further, secreted autocrine IGF-II mediates insulin m itogenic effects through the IGF-I receptor.