NICOTINAMIDE ADENINE-DINUCLEOTIDE PHOSPHATE OXIDASE IN THE FORMATION OF SUPEROXIDE IN OSTEOCLASTS

Osteoclasts use a variety of chemical agents to degrade bone. One impo rtant component of this process is the generation of superoxide. It ha s been reported that nicotinamide adenine dinucleotide phosphate (NADP H) oxidase is the enzyme responsible for superoxide production in phag ocyte; however, the NADPH oxidase present in osteoclasts has not been studied in detail. One of the membrane-bound subunits of the NADPH oxi dase is gp91(phox) which represents the rate-limiting component for th e formation of the NADPH oxidase complex. This study was designed to d emonstrate the presence of gp91(phox) in individual osteoclasts using the RT-PCR technique developed for limited numbers of cells. Compared with white cells, 1.8 times the amount of gp91(phox) mRNA was found in osteoclasts. This difference may be related to the size of the osteoc last and the multiple nuclei present. The presence of gp91(phox) in os teoclasts was confirmed at protein level by immunocytochemistry. Osteo clastic superoxide generation is inhibited by diphenylene iodonium, a specific inhibitor of the NADPH oxidase. These studies suggest that su peroxide generation by osteoclasts correlates with the activity of NAD PH oxidase.