Inhibition of chondrogenesis by integrin antibody in vitro.

Integrins mediate cell attachment to a variety of extracellular matrix proteins. These interactions play an important role in morphogenesis and differentiation. The mediating functions of integrins during chondrogenesis in vitro were investigated by using mesenchymal cells from limb buds of day 12 mouse embryos. The cells were treated with anti-beta 1, -alpha 1, and -alpha 5 integrin antibodies (a) from day 1 to day 3 and (b) from day 3 to day 7 of cultivation. The total culture period was 7 days. The presence of exogenous anti-beta 1, but not -alpha 1 and -alpha 5 integrin antibodies, from day 1 to 3 completely inhibited the differentiation of blastemal cells to chondroblasts and the formation of cartilage matrix. On the other hand, the presence of exogenous anti-beta 1, -alpha 1, and -alpha 5 integrin antibodies from day 3 of cultivation onwards had no effect. Immunoblotting and immunomorphological findings in the cultures treated with anti-beta 1 antibody from day 1 to day 3 revealed a pattern of integrins and collagen composed of beta 1, alpha 1, alpha 5 beta 1 integrins and collagen type I. The cartilage-specific chondroitin sulfate proteoglycan (CSPG) could not be demonstrated in these cultures. The cultures treated later (day 3 to day 7) showed a pattern of beta 1, alpha 3, alpha 5 beta 1, and alpha v beta 3 integrins, collagen types I and II, and CSPG identical to that of the untreated controls. These findings indicate that beta 1-integrins play a crucial role in early cartilage differentiation and point to a possible important cell-matrix interaction in the induction of chondrogenesis.