EPIGENETIC SELECTION AS A POSSIBLE COMPONENT OF TRANSDIFFERENTIATION - FURTHER STUDY OF THE COMMITMENT OF HYPERTROPHIC CHONDROCYTES TO BECOME OSTEOCYTES

Transdifferentiation of hypertrophic chondrocytes into osteogenic cell s was induced in 14 day chick embryo femurs by cutting through the reg ion of hypertrophic cartilage. The process was studied in organ cultur e, using electron microscopy, staining for alkaline phosphatase, immun ocytochemistry of collagen type I and proliferative cell nuclear antig en, and in situ localization of DNA strand-breaks. In addition, DNA an d RNA synthesis were studied by (3)[H]-T and (3)[H]-U radioautography. Loss of ECM components from the cut edge occurred in culture. During the 12 day period necessary for transdifferentiation we observed pheno typic instability and bi-potentiality, the death of some cells and the gradual promotion of the osteoblastic phenotype in the survivors. Tra nsition from chondrocytic to osteoblastic phenotype progressed stepwis e, through variable mosaic intermediates, and involved a few cell cycl es including asymmetric (differential) divisions. Proliferating and ap optotic cells were found in close proximity. As judged by the relative proportion of apoptotic cells and composition of the surrounding intr alacunar matrix, negative selection of intermediate cell types display ing chondrocytic and altered mosaic phenotypes occurred. When the oste oblastic lineage was finally established, apoptotic cells were no long er present. Our hypothesis is that after disruption of cell-cell or ce ll-matrix interactions and lack of growth factors certain cells are se lected and channelled through proliferation into the new stable phenot ype. This process is targeted by the environment through a set of pre- determined steps.