Hyperglycemia Alters the Responsiveness of Smooth Muscle Cells to Insulin-Like Growth Factor-I

Abstract

IGF-I stimulation of smooth muscle cell (SMC) migration and proliferation requires alphaVbeta3 ligand occupancy. We hypothesized that changes in the levels of extracellular matrix proteins induced by alterations in glucose concentrations may regulate the ability of SMCs to respond to IGF-I. IGF-I stimulated migration and proliferation of SMCs that had been maintained in 25 mM glucose containing media, but it had no stimulatory effect when tested using SMCs that had been grown in 5 mM glucose. IGF-I stimulated an increase in Shc phosphorylation and enhanced activation of the MAPK pathway in SMCs grown in 25 mM glucose, whereas in cells maintained in 5 mM glucose, IGF-I had no effect on Shc phosphorylation, and the MAPK response to IGF-I was markedly reduced. In cells grown in 25 mM glucose, the levels of alphaVbeta3 ligands, e.g. osteopontin, vitronectin, and thrombospondin, were all significantly increased, compared with cells grown in 5 mM glucose. The addition of these alphaVbeta3 ligands to SMCs grown in 5 mM glucose was sufficient to permit IGF-I-stimulated Shc phosphorylation and downstream signaling. Because we have shown previously that alphaVbeta3 ligand occupancy is required for IGF-I-stimulated Shc phosphorylation and stimulation of SMC growth, our data are consistent with a model in which 25 mM glucose stimulates increases in the concentrations of these extracellular matrix proteins, thus enhancing alphaVbeta3 ligand occupancy, which leads to increased Shc phosphorylation and enhanced cell migration and proliferation in response to IGF-I.

Insulin-like Growth Factor-I-stimulated Insulin Receptor Substrate-1 Negatively Regulates Src Homology 2 Domain-containing Protein-tyrosine Phosphatase Substrate-1 Function in Vascular Smooth Muscle Cells

“…Previously, we showed that IGF-I-stim- ulated IRS-1 phosphorylation and subsequent Grb2 binding were markedly down-regulated in VSMCs that had been maintained in high glucose (e.g. 25 mM) (29). The results from the current study confirm the previous observation that VSMCs in high glucose had a significant reduction in total IRS-1 protein levels at 12, 24, and 48 h compared with cells exposed to normal glucose.…”

“…In VSMCs, Shc phosphorylation in response to IGF-I depends on SHPS-1 phosphorylation and subsequent SHPS-1 complex assembly (2,29). Although some studies have shown that IRS-1 and Shc are competitive substrates for receptor tyrosine kinase (8,9,64), our results show that competition occurs between SHPS-1 and IRS-1, and this leads to impaired SHPS-1 phosphorylation.…”

“…Our prior studies have shown that when IRS-1 levels are substantially increased (e.g. VSMCs maintained in 5 mM glucose), SHPS-1 phosphorylation, SHP-2 transfer, and Shc/MAPK activation are suppressed (29,70). Consequently, although the protein synthesis response to IGF-I is maintained, the cell migration and proliferation responses are significantly decreased.…”

“…Previously, we reported that VSMCs maintained in high glucose concentrations (Ͼ12 mM) had a significant decrease in IRS-1. We also showed that the IRS-1 tyrosine phosphorylation response and subsequent Grb2 binding are significantly reduced in response to IGF-I (29). These studies were undertaken to determine the significance of the reduction in IRS-1 and whether failure to reduce the IRS-1 concentration alters assembly of the SHPS-1 complex, thus leading to altered cell proliferation and migration in response to IGF-I.…”

“…To determine phosphorylation of SHPS-1/CD, the final product was immunoblotted using an anti-Tyr(P) 99 antibody. Cell Culture-Porcine SMCs were isolated from the porcine aortic explants according to the protocol described by Ross (74) and were maintained as described previously (2,29). Cells were maintained in DMEM-high glucose growth medium (HG) or DMEM-normal glucose growth medium (NG) with 10% fetal bovine serum (Hyclone, Logan, UT), streptomycin (100 ng/ml), and penicillin (100 units/ml).…”