RESUMEN
Insulin-like growth factor-I (IGF-I)-mediated growth of cells can be modulated by specific IGF binding proteins (IGFBPs) that inhibit or augment IGF-I ligand-receptor interaction. IGFBP expression and production by human intestinal muscle cells in culture was characterized in rapidly growing cells (day 3 of culture), in confluent cells (day 7), and in postconfluent cells (day 14). RT-PCR analysis identified IGFBP-3, IGFBP-4, and IGFBP-5 mRNA during all three phases of growth. The production of IGFBP-3 and IGFBP-5 was regulated in reciprocal fashion. IGFBP-5 production was high on day 3 and decreased two- to fivefold by day 14, and IGFBP-3 production was low on day 3 and increased five- to eightfold by day 14. IGFBP-4 production remained constant. IGFBP-3 inhibited and IGFBP-5 augmented IGF-I-induced proliferation. IGFBP-3 and IGFBP-5 production was regulated in reciprocal fashion by transforming growth factor-beta1 (TGF-beta1). Immunoneutralization of endogenous TGF-beta1 decreased the production of IGFBP-3 and increased the production of IGFBP-5. Addition of exogenous recombinant human TGF-beta1 had the opposite effect. We conclude that the expression and time-dependent production of IGFBP-3, IGFBP-4, and IGFBP-5 and their regulation by endogenous TGF-beta1 represent mechanisms by which human intestinal muscle cells regulate autocrine IGF-I-mediated growth.
Asunto(s)
Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/biosíntesis , Proteína 5 de Unión a Factor de Crecimiento Similar a la Insulina/biosíntesis , Mucosa Intestinal/metabolismo , Músculo Liso/metabolismo , Factor de Crecimiento Transformador beta/fisiología , Western Blotting , División Celular/fisiología , Humanos , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/química , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/fisiología , Proteína 5 de Unión a Factor de Crecimiento Similar a la Insulina/antagonistas & inhibidores , Proteína 5 de Unión a Factor de Crecimiento Similar a la Insulina/química , Proteína 5 de Unión a Factor de Crecimiento Similar a la Insulina/fisiología , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/biosíntesis , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/química , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/fisiología , Factor I del Crecimiento Similar a la Insulina/farmacología , Intestinos/citología , Músculo Liso/citología , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Insulin-like growth factor I (IGF-I), acting via its cognate receptor, plays an autocrine role in the regulation of growth of intestinal muscle cells. In the present study the signaling pathways mediating the growth effects of IGF-I were characterized in cultured human intestinal smooth muscle cells. Growth induced by a maximally effective concentration of IGF-I (100 nM), measured as [3H]thymidine incorporation, was only partially inhibited by LY-294002 [phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor] or PD-98059 [mitogen-activated protein (MAP) kinase kinase (MEK) inhibitor] (86 +/- 7% and 35 +/- 6% inhibition, respectively) alone but was abolished by the two combined (114 +/- 18% inhibition), implying the participation of both pathways. IGF-I elicited time- and concentration-dependent increases in PI 3-kinase activity. This effect was inhibited only by LY-294002 (89 +/- 12%). IGF-I elicited time- and concentration-dependent phosphorylation of p44/p42 MAP kinase and increased MAP kinase activity. These effects were inhibited only by PD-98059 (78 +/- 9% and 98 +/- 7%, respectively). We conclude that in human intestinal muscle cells IGF-I activates distinct PI 3-kinase and MAP kinase signaling pathways, which act in conjunction to mediate growth.