CHARACTERIZATION OF BONE-CELLS ISOLATED ON DISCONTINUOUS PERCOLL GRADIENTS - DISTRIBUTION IN SEQUENTIALLY DERIVED POPULATIONS

Bone cell populations obtained by sequential digestion of newborn mouse calvariae remain morphologically heterogeneous despite well-documented biochemical differences. Fractionation of these populations on Percoll gradient reveal three major cell groups of low, intermediate, and high buoyant density (1.056, 1.070, and 1.095 g/ml) that are present in different ratios in early and late released populations. Cells of low and intermediate density dominate in early released populations. In contrast, late released populations contain mostly high-density cells. Basal levels of alkaline phosphatase are highest in cells of intermediate buoyant density. All cells respond to PTH with cAMP production and morphologic transformation, but biochemical responses to PTH, such as secretion of insulin-like growth factor I (IGF-I) and stimulation of alkaline phosphatase activity, occur mostly in cells of intermediate density. These data suggest that (1) subclasses of osteoblasts can be further separated by density and (2) PTH effects on alkaline phosphatase activity and IGF-I secretion are probably expressed by osteoblasts of a certain subclass and/or stage of development.