Effects of growth factors on chondrocyte proliferation and differentiation

Abstract

Longitudinal bone growth is regulated at the level of growth plate chondrocytes by circulating or locally produced hormones and growth factors. Present investigation focused on the effects of growth hormone (GH) and growth factors on chondrocyte proliferation and differentiation during pre- and postnatal cartilage development. Rat rib growth plate chondrocytes and murine limb bud mesenchymal cells were used in vitro, and murine limb buds in vivo.

The mitogenic effects of GH and growth factors were found to be strongly age-dependent. Basic fibroblast growth factor (bFGF) was shown to be the most potent mitogen before day 16 in rat, and insulin-like growth factor-I (IGF-I) later, indicating increased importance of IGF-I during prepubertal growth. IGF-I stimulated synthesis of cartilage matrix. The effect of IGF-I on cell proliferation was potentiated by platelet-derived growth factor (PDGF). The mitogenic effect of GH and the growth factors were found to vary between the subpopulations of cells in the growth plate. The two PDGF isoforms (PDGF AA and PDGF BB) were involved in the regulation of postnatal cartilage growth: PDGF AA in chondrocyte maturation and PDGF BB in chondrocyte proliferation and differentiation. bFGF treatment of serum-free chondrocyte cultures caused phenotypic modulations, characteristic of immature chondroblasts. bFGF induced expression of the proto-oncogene c-fos, downregulated the expression of cartilage specific genes and modulated the effects of the IGFs.

Changes in gene and protein expression were characterized in micromass cultures of mesenchymal cells isolated from El 1.5 Iimb buds. The expression of cartilage specific genes was related to the expression of DNA-binding proteins, and revealed increased expression of c-fos and FS-I at the onset of chondrogenic differentiation. Upon cartilage nodule formation, an increase in proteoglycan synthesis and alkaline phosphatase activity was observed. It was also shown that in analogy with limb buds in vivo, the initially widespread expression of the intermediate filament nestin is downregulated during chondrogenesis and upregulated in myogenesis, which could be modulated by the addition of growth factors. In the limb bud, the expression of nestin mRNA was confined to the ends of the muscles, whereas protein was uniformly distributed along the muscle fibers.

Changes in the endogeneous expression of growth factors in the developing limb in vivo were shown to be related to the expression of cartilage differentiation markers. In micromass cultures the same patterns of growth factors expression were established, and the events of chondrogenesis were reproduced by addition of growth factors stimulating chondrogenesis. bFGF-caused inhibition of chondrogenesis was associated with lowered expression of IGF-II indicating that these growth factors modulate each other’s actions. In conclusion, rib growth plate cartilage and limb buds have proven to be useful models for studies concerning the mechanisms of action of GH and growth factors on the regulation of chondrogenesis and longitudinal bone growth.