Protease-Resistant Insulin-Like Growth Factor (IGF)-Binding Protein-4 Inhibits IGF-I Actions and Neointimal Expansion in a Porcine Model of Neointimal Hyperplasia

Abstract

IGF-I has been shown to play a role in the progression of atherosclerosis in experimental animal models. IGF-binding protein-4 (IGFBP-4) binds to IGF-I and prevents its association with receptors. Overexpression of a protease-resistant form of IGFBP-4 has been shown to inhibit the ability of IGF-I to stimulate normal smooth muscle cell growth in mice. Based on these observations, we prepared a protease-resistant form of IGFBP-4 and infused it into hypercholesterolemic pigs. Infusion of the protease-resistant mutant inhibited lesion development by 53.3 +/- 6.1% (n = 6; P < 0.01). Control vessels that received an equimolar concentration of IGF-I and the protease-resistant IGFBP-4 showed no reduction in lesion size compared with control lesions that were infused with vehicle. Infusion of a nonmutated form of IGFBP-4 did not significantly inhibit lesion development. Proliferating cell nuclear antigen analysis showed that the mutant IGFBP-4 appeared to inhibit cell proliferation. The area occupied by extracellular matrix was also reduced proportionally compared with total lesion area. Immunoblotting revealed that the mutant IGFBP-4 remained intact, whereas the wild-type IGFBP-4 that was infused was proteolytically cleaved. Further analysis of the lesions revealed that a marker protein, IGFBP-5, whose synthesis is stimulated by IGF-I, was decreased in the lesions that received the protease-resistant, IGFBP-4 mutant, whereas there was no change in lesions that received wild-type IGFBP-4 or the mutant protein plus IGF-I. These findings clearly illustrate that infusion of protease-resistant IGFBP-4 into the perilesion environment results in inhibition of cell proliferation and attenuation of the development of neointima. The findings support the hypothesis that inhibiting IGFBP-4 proteolysis in the lesion microenvironment could be an effective means for regulating neointimal expansion.

Expression of a protease-resistant insulin-like growth factor-binding protein-4 inhibits tumour growth in a murine model of breast cancer

“…We believe that our approach using a protease-resistant IGFBP4 to block IGF activity is superior to those described above, as it inactivates IGF1, is small enough (24 -32 kDa) to cross the endothelial barrier and should have a long half-life in vivo due its resistance to the PAPP-A protease. In addition to the data presented here, a protease-resistant IGFBP4 was recently shown to inhibit IGF and therefore neointimal expansion in a model of neointimal hyperplasia (Nichols et al, 2007). It is a relatively small molecule compared with antibody therapies, and hence may penetrate tumours more effectively.…”

Structural, gene expression, and functional analysis of the fugu (Takifugu rubripes) insulin-like growth factor binding protein-4 gene

“…Several studies (26,36,44,46) have shown that targeted overexpression of IGFBP-4 in transgenic mice resulted in decreased tissue growth. For example, overexpression of IGFBP-4 in osteoblasts reduces bone formation and severely impairs skeletal growth and causes global postnatal growth retardation in mice (43).…”

“…For example, overexpression of IGFBP-4 in osteoblasts reduces bone formation and severely impairs skeletal growth and causes global postnatal growth retardation in mice (43). Targeted overexpression of IGFBP-4 in smooth muscle cells led to smooth muscle tissue hypoplasia in transgenic mice and inhibited neointimal expansion in a porcine model of neointimal hyperplasia (26,36,44). Additionally, overexpressing IGFBP-4 using a lymphoid tissue promoter (H-2Kb) decreases thymus growth in transgenic mice (46).…”

IGF-1, oxidative stress and atheroprotection

“…These models attempt to mimic the vascular repair responses to angioplasty, but several important differences must be recognized; for instance, the injury is in general to non-diseased vessels with no pre-existing neointimal cell populations and thus responses come predominantly from proliferating VSMC. Since IGF-1 is a potent VSMC mitogen, the majority of studies exploring mechanical injury models and direct [25,26] or indirect [27–30] mechanisms to alter vascular IGF-1 signaling reported that increased IGF-1 or IGF-1 signaling correlates with increased neointimal burden, suggesting that IGF-1 promotes vascular hyperplasia. In particular, targeted overexpression of IGF-1 in VSMC increases neointimal area size [25].…”