Differential effects of transforming growth factors beta 1, beta 2, beta 3 and beta 5 on chondrogenesis in mouse limb bud mesenchymal cells.

The present study was performed to determine whether mammalian TGF-beta isoforms and Xenopus TGF-beta 5 elicit a differential chondrogenic response on mesenchymal cells during mouse limb development. Results showed that TGF-beta isoforms produced a distinct chondrogenic pattern depending on embryonic stage. When they were applied to 5 day micromass cultures of limb mesenchymal cells from embryonic stages 19, 20 and 21, a differential response to all four TGF-beta isoforms assayed was observed. By stage 19 the cells formed a uniform sheet of cartilage cells; by stage 20, mesenchymal cells were more responsive to TGF-beta 1 and TGF-beta 5 than at stages 19 and 21, showing an entire cell layer of chondrogenic cells with higher accumulation of extracellular matrix. The diminished effect of TGF-beta 2 and TGF-beta 3 at stages 20 and 21 was accompanied by a nodular pattern of chondrogenic cells rather than by a uniform sheet, as seen at stage 19. At stage 20 TGF-beta 1 and TGF-beta 5 enhanced the expression of sulfated proteoglycans, type II collagen, cartilage link protein and alkaline phosphatase activity. In contrast, TGF-beta 2 and TGF-beta 3 caused less expression in the same parameters. Only a transient exposure to TGF-beta isoforms at days 1 and 2 of culture stimulate chondrogenesis, indicating that TGF-beta isoforms could regulate chondrogenesis at early stages of chondrocyte differentiation. However, when TGF-beta isoforms were applied to low density cultures of mesenchymal cells, chondrogenesis was enhanced only by 25%, suggesting that TGF-beta isoforms enhanced cartilage differentiation to higher levels in micromass cultures than in situations in which little or no chondrogenic differentiation normally occurs.