Epitopes of the human erythrocyte Ca²⁺-Mg²⁺ ATPase pump in human osteoblast-like cell plasma membranes

Human osteoblast-like cells were examined for the presence of the Ca²⁺-Mg²⁺ ATPase pump. The osteoblast-like cells had characteristic features of the oesteoblast phenotype, including the presence of osteonectin, bone GLA protein, and type I collagen. The cells were able to mineralize matrix, their production of cAMP increased in response to PTH, and their alkaline phosphatase activity increased in response to 1,25-dihydroxyvitamin D₃. Immunocytochemical staining of the osteoblast-like cells with a monoclonal antibody against human red cell Ca²⁺-Mg²⁺ ATPase demonstrated the presence of an epitope of the Ca²⁺-Mg²⁺ ATPase in these cells; staining of paraffin-embedded osteoblast-like cell sections demonstrated anti-Ca²⁺-Mg²⁺ ATPase staining only in cell plasma membranes. Western blot analysis of osteoblast-like cell homogenates showed that the monoclonal antibody to human erythrocyte Ca²⁺-Mg²⁺ ATPase bound to a major band at 140,000 mol wt, similar to the mol wt of known plasma membrane Ca²⁺-Mg²⁺ ATPases. The presence in the osteoblast-like cells of a Ca²⁺-Mg²⁺ ATPase similar to the human red cell calcium pump suggests that this enzyme may play a role in osteoblast intracellular calcium homeostasis.