DING, J; DUAN, C; Univ. of Michigan, Ann Arbor: IGF binding protein-5 is required for the pharyngeal skeleton formation in vivo

IGFBPs are a conserved family of proteins that bind to IGFs and modulates IGF bioactivity. Some IGFBPs may also possess intrinsic biological activities that are IGF-independent. Their in vivo physiological functions of IGFBPs, however, are unknown. In this study, we have elucidated the physiological function of IGFBP-5 using a teleost model organism, the zebrafish. Zebrafish IGFBP-5 is highly similar to its human homologue in primary structure and binds to human as well as fish IGFs with high affinity and specificity. IGFBP-5 mRNA expression began in somites and branchial mesenchyme at 24 hours post fertilization (hpf). By 48 hpf, its expression became restricted in the newly forming craniofacial cartilage tissues. This pattern remained until juvenile stage. To investigate its physiological function, we ablated the IGFBP-5 gene product by injecting morpholino-based antisense oligos into fertilized eggs. The IGFBP-5 morphants had little or no craniofacial skeletal tissues. In addition, the size of pedal fins and otolith was also reduced. The lack of cartilage tissue was further confirmed by Alcian blue staining. To determine whether the lack of the head skeletal tissues was due to defects in tissue formation and/or altered tissue growth/survival, we analyzed the expression of several dlx genes, which are key transcriptional factors expressed in the pharryngeal cartilage precursor cells. Dlx gene expression was greatly diminished in the IGFBP-5 morphants, suggesting that ablation of IGFBP-5 disrupts the tissue formation. This effect was specific to craniofacial skeleton because ablation of IGFBP-5 had little effect on the expression of brain and neuronal marker genes such as Emx-1 and Rx-1. This study provides the first compelling in vivo evidence that IGFBP-5 plays a critical role in the formation of craniofacial skeletal tissues in vivo.