Monosodium urate, hydroxyapatite, and calcium pyrophosphate crystals induce tumor necrosis factor-alpha expression in a mononuclear cell line

Objective. Tumor necrosis factor-alpha (TNF-alpha) is thought to be important in chronic inflammation of joints characteristic of crystal induced arthritis. Monocytes are instrumental in maintaining that inflammation. We investigated production of mRNA and protein for TNF-alpha in vitro in a murine mononuclear cell line, after exposure to relevant crystal types. Methods. Using the cell Line designated RAW 264.7, cells were grown in standard medium and exposed to varying amounts of monosodium urate (MSU), hydroxyapatite (HA), and calcium pyrophosphate dihydrate (CPPD) crystals for differing time periods. Analysis of TNF-alpha mRNA induced by such exposure was by Northern hybridization; analysis of TNF-alpha protein was by ELISA. Results. RNA analyses of cells treated with various levels of MSU, HA, and CPPD crystals showed strong induction of TNF-alpha transcripts. ELISA on culture supernatants confirmed high level TNF-alpha. peptide secretion resulting from that transcriptional upregulation. Time course studies showed peak accumulation of TNF-alpha mRNA 1-6 h post-treatment. Study of the signalling pathway involved in TNF-alpha transcriptional upregulation indicated that increased phospholipase A(2) activity was required. Conclusion. These observations suggest that crystals in joints can directly stimulate production of TNF-alpha, and that the source of that cytokine may be the monocytes known to be present and playing an important role in chronic joint disease.