LIPOPOLYSACCHARIDE (LPS) STIMULATES THE PRODUCTION OF TUMOR-NECROSIS-FACTOR (TNF)-ALPHA AND EXPRESSION OF INDUCIBLE NITRIC-OXIDE SYNTHASE (INOS) BY OSTEOCLASTS (OCL) IN MURINE BONE-MARROW CELL-CULTURE

Osteoclasts (OCL) resorb bone. They are essential for the development of normal bones and the repair of impaired bones. The function of OCL is presumed to be supported by cytokines and other biological mediator s, including tumor necrosis factor (TNF)-alpha and nitric oxide (NO), Bacterial lipopolysaccharide (LPS) is a potent inducer of TNF-alpha an d inducible nitric oxide synthase (iNOS), which is the specific enzyme for synthesizing NO from L-arginine. To obtain direct evidence on LPS -induced TNF-alpha production and iNOS expression by OCL, OCL-enriched cultures were prepared by 7-day cocultures of bone marrow cells of ad ult BALB/c mice and osteoblastic cells (OBs) derived from calvaria of newborn BALB/c mice, and the generation of TNF-alpha and iNOS in OCL s timulated with LPS was examined immunocytochemically. When the culture d cells were stimulated with 100 ng/ml of LPS, OCL clearly showed TNF- alpha and iNOS expression. Without LPS-stimulation, no expression was observed. TNF activity in the culture supernatants of the OCL-enriched cultures in the presence of LPS was also detected by cytotoxic assay that used TNF-sensitive L929 cells, The dentin resorption activity of OCL was estimated by area and number of pits formed on dentin slices, which were covered by the OCL fraction and cultured in the presence or absence of LPS, sodium nitroprusside (SNP; a NO generating compound), N-G-monomethyl L-arginine acetate (L-NMMA; a competitive inhibitor of NO synthase (NOS)), or LPS plus L-NMMA, Pit formation was obviously i nhibited in the presence of SNP and slightly inhibited in the presence of L-NMMA, but it was not affected in the presence of LPS or LPS plus L-NMMA, These findings indicate that OCL produces TNF and expresses i NOS in response to LPS, but the LPS-activation of OCL scarcely affects pit formation by them.