Growth Factor-Induced Proliferation of Osteoblasts Measured by Bromodeoxyuridine Immunocytochemistry

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Abstract
Immuno-localization of BUdr was used to identify DNA synthesis in vitro in chicken embryonic bone cells stained positively or negatively for alkaline phosphatase activity. The results were similar to, but more sensitive than, our standard bioassay which assesses 3H-thymidine incorporation into DNA by liquid scintillation counting, and more rapid than autoradiographic localization of 3H-thymidine. SGF/IGF-II and bFGF stimulate cellular proliferation equally in ALP(+) and ALP(-) cells. In contrast, IGF-I and TGF-/J stimulate proliferation more in the ALP(-) than ALP(+) cells. The greatest increase in DNA replication of ALP(-) cells occurred following incubation with SGF/IGF-II or TGF-/3, and in the ALP(+) cells with SGF/IGF-II or bFGF. TGF-/3 stimulated cellular proliferation at the lowest dose (1 ng/ml). The differential effect of the growth factors on each population of cells indicates that all these bone-matrix derived growth factors may play different roles in the local regulation of skeletal metabolism.