(5R)-5-Hydroxytriptolide Attenuated Collagen-Induced Arthritis in DBA/1 Mice via Suppressing Interferon-γ Production and Its Related Signaling

(5R)-5-Hydroxytriptolide (LLDT-8) displays strong immunosuppressive activities both in vitro and in vivo in our previous studies. This study aims to investigate whether LLDT-8 has antiarthritic potential in a murine model of type II bovine collagen (CII)-induced arthritis (CIA) and to show the mechanism(s) of LLDT-8 action. DBA/1 mice were immunized with CII to induce arthritis and administered with LLDT-8. The severity of arthritis was evaluated according to the clinical score and joint damage. The effects of LLDT-8 on immune responses were determined by measurement of serum antibody levels, lymphocyte proliferation assay, cytokine assay, nitric oxide (NO) production, arginase activity assays, fluorescence-activated cell sorting analysis of splenic Mac-1⁺ cells, as well as polymerase chain reaction analysis for interferon-γ (IFN-γ)-related gene expression. We showed that LLDT-8 treatment significantly reduced the incidence and severity of CIA. The preventive and therapeutic effects of LLDT-8 are associated with 1) reduction of serum anti-CII immunoglobulin (Ig) G, IgG2a, and IgG1 levels; 2) inhibition of CII-specific lymphocyte proliferation, IFN-γ and interleukin-2 production; 3) blockade of gene expressions in IFN-γ signaling, including IFN-γ production pathways [signal transducer and activator of transcription (STAT) 1, T-box transcription factor, interleukin 12Rβ2, and STAT4] and IFN-γ-induced chemokine transcription [macrophage inflammatory protein (Mip)-1α, Mip-1β, regulated on activation normally T cell expressed and secreted, and inducible protein 10]; and 4) retardation of the abnormal increase of NO via IFN-γ/STAT1/interferon regulatory factor 1/inducible nitric-oxide synthase pathway and arginase activity. Moreover, the mRNA transcription of chemokine receptors was also suppressed [including C-C chemokine receptor (CCR) 1, CCR5, and C-X-C chemokine receptor 3]. In conclusion, our data suggest that the antiarthritic effect of LLDT-8 is closely related to the blockade of IFN-γ signaling. LLDT-8 may have a therapeutic value in the treatment of rheumatoid arthritis.