Use of bone cell cultures to study skeletal pathology

We describe procedures for the isolation, culture, and analysis of neonatal osteoblasts from osteopetrotic (toothless (tl) and osteopetrosis [op]) rats and normal littermates. Normal osteoblasts produce and mineralize an extracellular matrix indistinguishable from that of well-characterized fetal rat osteoblasts in vitro. Mutant (tl and op) cultures show an early abnormal pattern of cell proliferation and a later premature, extensive mineralization which mimic the mutant phenotype in vivo. In cocultures with normal osteoclasts, mutant (tl) osteoblasts also show a greatly reduced ability to orchestrate bone resorption, as revealed by pit formation in bone slices, in response to physiologic mediators. These phenomena in vitro are consistent with the behavior of mutant osteoblasts and osteoclasts in vivo and suggest that more definitive microscopic analyses of osteoblasts from each mutation in vitro will provide insights on the roles of osteoblasts in the compromised bone resorption which characterizes the osteopetroses as well as their role in osteoclast ontogeny. This study shows that when their behavior is confirmed in vivo, bone cell cultures offer rigorous systems for understanding skeletal cell dysfunction in normal and pathological development.