Insulin-like growth factor-binding protein 5 (Igfbp5) compromises survival, growth, muscle development, and fertility in mice

Abstract

The insulin-like growth factors (IGFs) are essential for development; bioavailable IGF is tightly regulated by six related IGF-binding proteins (IGFBPs). Igfbp5 is the most conserved and is developmentally up-regulated in key lineages and pathologies; in vitro studies suggest that IGFBP-5 functions independently of IGF interaction. Genetic ablation of individual Igfbps has yielded limited phenotypes because of substantial compensation by remaining family members. Therefore, to reveal Igfbp5 actions in vivo, we generated lines of transgenic mice that ubiquitously overexpressed Igfbp5 from early development. Significantly increased neonatal mortality, reduced female fertility, whole-body growth inhibition, and retarded muscle development were observed in Igfbp5-overexpressing mice. The magnitude of the response in individual transgenic lines was positively correlated with Igfbp5 expression. Circulating IGFBP-5 concentrations increased a maximum of only 4-fold, total and free IGF-I concentrations increased up to 2-fold, and IGFBP-5 was detected in high M(r) complexes; however, no detectable decrease in the proportion of free IGF-I was observed. Thus, despite only modest changes in IGF and IGFBP concentrations, the Igfbp5-overexpressing mice displayed a phenotype more extreme than that observed for other Igfbp genetic models. Although growth retardation was obvious prenatally, maximal inhibition occurred postnatally before the onset of growth hormone-dependent growth, regardless of Igfbp5 expression level, revealing a period of sensitivity to IGFBP-5 during this important stage of tissue programming.

Fig. 1.

Igfbp5-myc expression is widespread but at different levels in the three lines. (A) Schematic of the Igfbp5-myc construct. CMV-IE, immediate early cytomegalovirus. (B) Hybridization of Igfbp5 cDNA probe to mRNA isolated from hemizygous skeletal muscle at 8 wk (mice from WT females × Tg males). (C) Endogenous and Tg mRNA levels from key tissues of line 3 mice at 8 wk.

Fig. 2.

Reduced litter size in Igfbp5-myc hemizygous females. Gray columns, mice generated from WT siblings; left shaded columns, mice generated from WT mothers and hemizygous fathers; right shaded columns, mice derived from hemizygous mothers and fathers. **, P < 0.01 Tg versus WT litters; mean ± SEM of four to ten litters per line.

Fig. 3.

Growth inhibition in Igfbp5-myc-overexpressing mice. (A) Birth weights of nongenetically manipulated WT, WT siblings (-/-), hemizygous (-/+), and homozygous (+/+) Igfbp5-myc-expressing mice; mean ± SEM of three to eight mice per group. (B) Whole-body weight gain of female WT and hemizygous siblings derived from a WT mother and Igfbp5 hemizygous father. *, P < 0.05; ***, P < 0.001; mean ± SEM of 4–16 mice per group.

Fig. 4.

Igfbp5-myc overexpression exerts differential effects on fractional tissue weight. Fractional tissue weights for female nonmanipulated WT, WT sibling (-/-), hemizygous (-/+), and homozygous (+/+) mice. *, P < 0.05; **, P < 0.01; ***, P < 0.001; mean ± SEM of five to ten mice per group.

Fig. 5.

Histology of Igfbp5 transgenic embryos at e18.5. (A) Hematoxylin/eosinstained sagittal sections of a line 3 homozygote (+/+) and WT sibling derived from hemizygous parents. (B) Diaphragm muscle. Insets demonstrate diaphragm width. (C) Lung.

Fig. 6.

Igfbp5-myc overexpression changes serum IGFBP and IGF-I concentrations postnatally. (A) Ligand blot to display IGFBP levels in serum derived from 8-wk-old hemizygous Igfbp5-overexpressing mice (Tg) and WT littermates. (B) Ligand blot to demonstrate changes in IGFBP levels between birth and 8 wk in serum derived from line 3 hemizygous and WT littermates. (C) Serum total IGF-I concentrations in 8-wk-old hemizygotes and WT siblings. *, P < 0.05; ***, P < 0.001 compared with WT; mean ± SEM of six to seven mice per group. Serum was collected from offspring of WT mothers and hemizygous fathers.