Cellular localization and expression of insulin-like growth factors (IGFs)and IGF binding proteins within the epiphyseal growth plate of the ovine fetus: possible functional implications

The insulin-like growth factors (IGFs) are important in the regulation of n ormal fetal musculoskeletal growth and development, and their actions have been shown to be modulated by IGF binding proteins (IGFBPs). Because the an atomical distribution of IGFBPs is likely to dictate IGF bioavailability, w e determined the cellular distribution and expression of IGF-I, IGF-II, and IGFBP-1 to IGFBP-6 in epiphyseal growth plates of the fetal sheep, using i mmunocytochemistry and in situ hybridization. Little mRNA for IGF-I was det ectable within the growth plates, but mRNA for IGF-II was abundant in germi nal and proliferative chondrocytes, although absent from some differentiati ng chondrocytes and hypertrophic cells. Immunohistochemistry for IGF-I and IGF-II showed a presence of both peptides in all chondrocyte zones, includi ng hypertrophic cells. Immunoreactive IGFBP-2 to -5 were localized within t he germinal and proliferative zones of chondrocytes, but little immunoreact ivity was present within the columns of differentiating cells. IGFBP immuno reactivity again appeared in hypertrophic chondrocytes. IGFBP mRNA in chond rocytes of the epiphyseal growth plate was below the detectable limit of in situ hybridization. However, low levels of mRNAs for IGFBP-2 to -6 were de tected by the reverse transcriptase polymerase chain reaction. A co-localiz ation of IGFBPs with IGF peptides in intact cartilage suggests that they ma y regulate IGF bioavailability and action locally. To test this hypothesis, monolayer cultures of chondrocytes were established from the proliferative zone of the growth plate, and were found to release immunoreactive IGF-II and to express mRNAs encoding IGFBP-2 to -6. Exogenous IGFBP-3, -4, and -5 had an inhibitory action on IGF-II-dependent DNA synthesis. IGFBP-2 had a b iphasic effect, potentiating IGF-II action at low concentrations but inhibi ting DNA synthesis at equimolar or greater concentrations relative to IGF-I I. Long R3 IGF-I, which has a reduced binding affinity for many IGFBPs, was more potent than native IGF-I in promoting DNA synthesis by chondrocytes. Our findings suggest that locally produced IGF-II and IGF-I derived from th e circulation can influence fetal epiphyseal chondrogenesis, and that this may be modulated locally by multiple IGFBP expression.