Cells and Materials


Rat bone marrow derived cells were cultured using a-Minimal Essential Medium supplemented with antibiotics, ascorbic acid and !3-glycerphosphate in the presence of 10-8M dexamethasone, on polystyrene and hydrophilic fluorocarbon substrata for periods of 2 - 4 weeks. During this time, a large yield of bone nodules was achieved and the elaborated tissue was examined by both scanning and transmission electron microscopy. The matrix produced by the cells contacting the underlying substratum was an afibrillar, globular, calcified material which formed a layer approximately 0.5μm thick. The calcium and phosphorus content of this material was confirmed by energy dispersive X-ray dot mapping analysis. The collagenous matrix of the forming bone nodules was intimately associated with, and anchored to, this layer. The bulk of the bone nodule, above the interfacial zone, was of a normal appearance with osteocytes buried in a collagenous matrix exhibiting spheritic foci of mineralization. The cells, but not the extracellular matrix, of this culture were then removed using a trypsin citrate saline solution and the dishes containing these nodules reseeded with fresh bone marrow cells. These second stage cultures were maintained in supplemented medium, without dexamethasone. During this second period, osteoclasts resorbed both the afibrillar and collagen containing calcified matrices laid down in the first stage of the culture, producing characteristic scalloped osteoclast resorption lacunae.