Three isolation techniques for primary culture of human osteoblast-like cells - A comparison

The culture of osteoblast-like cells of human origin has become an importan t experimental model in bone biology. We report here a comparison and evalu ation of three of the most widely used systems available today: bone marrow stroma cell cultures (BMSC), human osteoblast explant cultures (hOB) and o steoblast explant cultures from collagenase-treated bone (hOB(col)), Cultur es from 16 bone specimens obtained from various donors were established and their expression of the osteoblast phenotype were then compared in seconda ry cultures by use of biochemical markers. BMSC had the highest basal and 1 ,25-dihydroxyvitaminD(3) (1,25(OH)(3)D-3)-induced alkaline phosphatase acti vities in all cell isolations, with levels approximately twice those in exp lant cultures. nasal osteocalcin secretion was low-to-undetectable in all c ell cultures but was detected in 1,25(OH)(2)D-3-stimulated cultures. BMSC p roduced half of the amount of osteocalcin synthesized in explant cultures. The BMSC cultures also synthesized the lowest amounts of type I collagen, w hereas collagen type III synthesis did not differ significantly among the v arious cultures. When secondary cultures were treated with 100 nM dexametha sone in the presence of ascorbic acid (50 mu g/ml) and beta-glycerophosphat e (10 mM), cultures deposited calcium mineral into the cell layer within 2- 4 weeks. PTH-induced cAMP formation was detected in only 5 of 15 isolations and no consistent isolation-dependent response pattern was seen. We conclu de that BMSC cultures differ significantly from explant cultures obtained f rom the same bone specimen. However, all cultures represent cells which can differentiate further and induce mineralization of the extracellular matri x in response to osteoinductive drugs.