Involvement of ligand occupancy in Insulin‐like growth factor‐I (IGF‐I) induced cell growth in osteoblast like MC3T3‐E1 cells

Abstract Growth factors and matrix proteins regulate the proliferation and differentiation of osteoblasts. The insulin‐like growth factor (IGF) system comprises IGF‐I, IGF‐II, and six high‐affinity IGF‐binding proteins (IGFBPs). IGFs stimulate cell growth in many types of tissue; IGF‐binding proteins regulate cellular actions and can affect cell growth. IGF‐I is involved in differentiation, proliferation, and matrix formation in osteoblasts; IGFBP‐5 is associated with the extracellular matrix (ECM) and can potentiate the actions of IGF‐I. We investigated the effect of ECM proteins on the responses of MC3T3‐E1 osteoblast cells to IGF‐I and IGFBP‐5. In addition, because extracellular signal‐regulated kinases 1 and 2 (Erk 1/2) affect cell growth, we evaluated the effects of IGFBP‐5 on Erk 1/2 phosphorylation in MC3T3‐E1 cells. IGF‐I caused an increase in IGFBP‐5 expression in cultured MC3T3‐E1 cells, and IGF‐I plus IGFBP‐5 significantly increased cell growth. Likewise, the addition of IGF‐I and IGFBP‐5 to cultured MC3T3‐E1 cells increased the synthesis of the ECM proteins osteopontin (OPN) and thrombospondin‐1 (TSP‐1), which can bind to αVβ3 integrin receptors on the cell surface. By contrast, the addition of an antibody against ECM proteins inhibited the effects of OPN and TSP‐1 on IGFBP‐5 expression. The stimulatory effect of IGFBP‐5 was mediated via Erk 1/2 activation. These data suggest that IGFBP‐5 regulates Erk 1/2 phosphorylation in cultured MC3T3‐E1 cells via ECM proteins that may ultimately stimulate the growth of osteoblasts. We determined whether occupation of the αVβ3 integrin receptor affects IGF‐I receptor (IGF‐IR)‐mediated signaling and function in MC3T3‐E1 osteoblast cells. Occupation of the αVβ3 integrin receptor with ECM proteins induced IGF‐I‐stimulated IGF‐IR phosphorylation. Conversely, in the presence of the αVβ3‐specific disintegrin echistatin, IGF‐I‐stimulated IGF‐IR activation was inhibited. IGF‐I‐stimulated IGF‐IR phosphorylation was accompanied by IRS‐1 phosphorylation and MAPK activation. However, these effects were attenuated by echistatin. Thus, occupancy of the αVβ3 disintegrin receptor modulates IGF‐I‐induced IGF‐IR activation and IGF‐IR‐mediated function in MC 3T3‐E1 osteoblasts.