KE-298 and its active metabolite KE-758 suppress nitric oxide production by murine macrophage cells and peritoneal cells from rats with adjuvant induced arthritis

Objective. To analyze the effects of KE-298 and KE-758 on lipopolysaccharid e (LPS) induced nitric oxide (NO) production by the RAW264.7 murine macroph age cell line, and the effect of KE-758 on spontaneous NO production by per itoneal cells from rats with adjuvant induced arthritis. Methods. The amount of NO was determined using Griess reagents. The protein s for inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) Were detect ed by Western blot, then mRNA for interferon-beta (IFN)-beta, IFN regulator y factor-1 (IRF-1), and iNOS were detected by RT-PCR. Degradation of iNOS m RNA was analyzed using Northern blot. Nuclear factor-kappaB (NF-kappaB) in nuclear extracts was determined by EMSA. Adjuvant arthritis in rats was ind uced by inoculating heat killed Mycobacterium butyricum SC in the tail. Results. KE-298 and KE-758 suppressed NO production by LPS activated RAW264 .7 cells by inhibiting iNOS gene expression. Neither LPS induced NF-kappaB activation nor degradation of iNOS mRNA was affected by KE-758 treatment. L PS induced IFN-beta and IRF-1 gene expression were markedly suppressed by K E-758. In rats with adjuvant induced arthritis, enhanced NO and iNOS produc tion by cultured peritoneal cells and the development of arthritis were sup pressed by KE-758. Conclusion. KE-758 suppressed LPS induced iNOS gene expression by murine ma crophage cells by inhibiting IFN-beta /IRF-1 expression. The potential of K E-758 to inhibit iNOS production might partly explain its efficacy on adjuv ant induced arthritis in rats.