Epigenetic selection as a possible component of transdifferentiation. Further study of the commitment of hypertrophic...

Transdifferentiation of hypertrophic chondrocytes into osteogenic cells was induced in 14 day chick embryo femurs by cutting through the region of hypertrophic cartilage. The process was studied in organ culture, using electron microscopy, staining for alkaline phosphatase, immunocytochemistry of collagen type I and proliferative cell nuclear antigen, and in situ localization of DNA strand-breaks. In addition, DNA and 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 phenotypic instability and bi-potentiality, the death of some cells and the gradual promotion of the osteoblastic phenotype in the survivors. Transition from chondrocytic to osteoblastic phenotype progressed stepwise, through variable mosaic intermediates, and involved a few cell cycles including asymmetric (differential) divisions. Proliferating and apoptotic cells were found in close proximity. As judged by the relative proportion of apoptotic cells and composition of the surrounding intralacunar matrix, negative selection of intermediate cell types displaying chondrocytic and altered mosaic phenotypes occurred. When the osteoblastic lineage was finally established, apoptotic cells were no longer present. Our hypothesis is that after disruption of cell-cell or cell-matrix interactions and lack of growth factors certain cells are selected and channelled through proliferation into the new stable phenotype. This process is targeted by the environment through a set of pre-determined steps.