Novel Insights into the Insulin-Like Growth Factor 1 System Revealed by Studies of New Mutant Mouse Models

The Insulin-like growth factor system (IGF) is a major growth regulatory signaling network comprised of two-ligands IGF-I and IGF-II, two receptors IGF type I and II (IGF-IR and IGFII-R), and regulatory proteins consisting of the acid-labile subunit (ALS) and six binding proteins (IGFBP-1-6). IGF-I binding stimulates autophosphorylation of the receptor, creating binding sites for the insulin like substrates (IRS-1-4) that act as docking proteins transducing signals down a number of pathways most notably the RAS/Raf-1/MAPK and PI3K/PDK/AKT pathways. The molecular and phenotypic investigations of two mouse models with genetic mutations in IGFBP-2 and IRS-1 are reported here revealing novel insights into the IGF-I pathway. The first model studied, Igfbp2 null (Igfbp2- -) mice have been reported previously to have no major physiological defects. Developmental data reported herein revealed that male Igfbp2- - were heavier, had shorter femurs, and smaller spleens than controls. Serum IGF-I was increased in Igfbp2- - with two fold increases in hepatic Igfbp3 and Igfbp5 mRNA transcripts. The skeletal phenotype of the null was gender specific; female Igfbp2- - had increased cortical bone mass, while male Igfbp2- - had reductions in both cortical and trabecular bone mass compared to controls. Serum osteocalcin was reduced in male Igfbp2- -. In vitro and in vivo, osteoblasts, osteoclasts, and bone formation rates were reduced in male Igfbp2- -. The phosphatase and tensin homolog (PTEN) protein levels were increased in the null mice pointing to a potential mechanism for decreased IGF activity and bone formation. The second model investigated, a spontaneous mouse mutant, designated \u27small\u27 (sml), genetically mapped to the Chromosome 1 region containing the Irsl gene. Sequencing this mutant gene revealed a single nucleotide deletion resulting in a premature stop codon. Despite normal mRNA levels, Western blot analysis revealed no detectable protein in mutant liver lysates. The Irslsml/Irslsml (Irslsml sml) mice are small, lean, hearing impaired, have 20% less serum IGF-I, are hyperinsulinemic and are mildly insulin resistant compared to control littermates. Irslsml sml mice have low bone mineral density, reduced trabecular and cortical thicknesses and low bone formation rates, while osteoblast and osteoclast numbers were increased in the females but not different in the males compared to controls. Irslsmlsml bone marrow stromal cell cultures showed decreased pre-osteoblasts and normal numbers of osteoclasts. Irslsml sml stromal cells treated with IGF-I exhibited a 50% decrease in AKT phosphorylation, and increased IRS-2 phosphorylation. Similarities between engineered knockouts and the spontaneous mutation of Irsl were identified as well as significant differences with respect to heterozygosity and gender. Thus, studies with these two mouse mutant models suggest a new role for IGFBP-2 in bone development modified by gender having implications for the gender-biased progression of osteoporosis in humans. Furthermore, the demonstration of an unanticipated heterozygous phenotype in the Irslsml mice suggests that similar mutations in IRS-1 in humans may be responsible for short stature and/or osteoporosis. In summary, systematic investigation of the Igfbp2- - and Irslsml sml mice demonstrate the power of mouse mutant gene models to reveal novel insights into the biological actions of the IGF-I system