CHARACTERIZATION OF HUMAN BONE-MARROW STROMAL CELLS WITH RESPECT TO OSTEOBLASTIC DIFFERENTIATION

Human bone marrow was harvested by means of iliac crest aspiration and cultured under conditions that promote an osteoblastic phenotype. Hum an bone marrow aspirates from 30 normal subjects, ages 8-80 years, wit h no systemic illness, yielded a mean of 92 +/- 65 x 10(6) nucleated c ells per 2 mi of aspirate. The prevalence of potential osteoblastic pr ogenitors was estimated by counting the number of alkaline phosphatase -positive colonies. This assay demonstrated a mean of 43 +/- 28 alkali ne: phosphatase-positive colonies per 10(6) nucleated cells. which was about one per 23,000 nucleated cells. The prevalence of these colonie s was positively correlated with the concentration of nucleated cells in the original aspirate (p = 0.014) and was negatively correlated wit h donor age (p = 0.020). The population of alkaline phosphatase-positi ve colonies in this model sequentially exhibited markers of the osteob lastic phenotype: essentially all colonies (more than 99%) stained pos itively for alkaline phosphatase on day 9, Matrix mineralization, whic h was associated with the synthesis of bone sialoprotein, was demonstr ated on day 17 with alizarin red S staining. On day 45, cells that wer e stimulated with 1,25-dihydroxyvitamin D-3 synthesized and secreted o steocalcin at concentrations consistent with known osteoblastic cell l ines. This model provides a useful method for the assay of progenitors of connective tissue from human subjects, examination of the effects of aging and selected disease states on this progenitor population, an d investigation into the regulation of human osteoblastic differentiat ion.