Role of 5' HoxD genes in chondrogenesis in vitro.

In the present study, we have examined by in situ hybridization, expression of five 5' HoxD cluster genes (D9, D10, D11, D12 and D13) during chondrogenesis of chick limb bud mesenchymal cells in vitro. After one day in culture, D9 and D13 gene expression was restricted to patches of mesenchymal cells, while expression of D10, Dll, and D12 gene was prominent in all mesenchymal cells. In 3-day cultures, D9 and D13 genes were expressed only in cartilage nodules, while D10, Dll, and D12 genes were expressed in both cartilage nodules and in all mesenchymal cells. These observations indicate two different patterns of expression; one for D9 and D13, and a different one for D10, Dll, and D12. These patterns of expression seem to correlate with patterns of cell proliferation and differentiation to chondrocytes. The role of these HoxD genes was further investigated by employing antisense S-oligomers. We found that oligodeoxynucleotides complementary to HoxD (D10-D 13) mRNAs were capable of inhibiting chondrogenesis. These data suggest that expression of HoxD genes is required for mesenchymal condensation, and differentiation to chondrocytes. This in turn implies that these HoxD genes aside from their role in the patterning of the developing skeletal elements might regulate down-stream factors necessary for cartilage differentiation as well.