-1 -AduCPI2 alleviates MSU-induced acute gouty arthritis in mice by inhibiting cathepsin S and the C5a-C5aR1 axis

Sijing Liu¹Yongli SITU^2*Zheng Shao²Li DENG²Meng Liang²Bin Yu¹

• ¹Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong Province, China
• ²Guangdong Medical University, Zhanjiang, Guangdong, China

Background: Gout, a prevalent and severe form of arthritis precipitated by hyperuricemia, has been the subject of extensive research. However, the intricate underlying mechanisms governing gout-related inflammation are still only partially elucidated. Ancylostoma duodenale cysteine protease inhibitor 2 (AduCPI2, abbreviated as CPI2, GenBank No. JQ762417) is a protein isolated from Ancylostoma duodenale by our research group in the early stage. CPI2 can effectively inhibit the activity of cathepsin S and reduce the level of C5a. In this study, we aimed to investigate the effect of CPI2 on acute gouty arthritis.The recombinant CPI2 fusion protein was purified. the inhibitory effect of CPI2 on cathepsin S was detected. In an attempt to mimic human gouty arthritis, suspensions of MSU crystals were injected into the right foot pads of (WT) mice and C5aR1 -/-mice. After the injection of MSU crystals, the mice were intravenously administered CPI2 at doses of 0.5, 1, and 2 mg/kg body weight.The effects of CPI2 on mice with MSU-induced gouty arthritis were evaluated by -3 -measuring the degree of paw swelling, observing the histopathological changes in paw tissues using HE staining, detecting the levels of inflammatory cytokines, C5a, and cathepsin S by ELISA, determining the activities of GSH-Px and SOD as well as the level of MDA by chemiluminescence assay, and observing the expression level of C5aR1 protein in paw tissues by immunohistochemistry. CPI2 with a purity of over 98% was finally obtained. CPI2 ameliorated the inflammation of MSU-induced gouty arthritis in a dose-dependent manner. It reduced joint swelling, decreased the infiltration of inflammatory cells, lowered the levels of IL-1β, IL-6, and TNF-α, increased the level of the anti-inflammatory cytokine IL-10, enhanced the activities of SOD and GSH-Px, while reducing the content of MDA, decreasing the levels of C5a and cathepsin S, and down-regulating the expression level of C5aR1 protein. The knockout of the C5aR1 gene was beneficial for inhibiting the inflammation of MSU-induced gouty arthritis. Conclusions: CPI2 alleviates MSU-induced acute gouty arthritis in mice through the inhibition of cathepsin S and the C5a -C5aR1 axis. CPI2 may represent a potential candidate for the treatment of gouty arthritis.