Abstract
<title>Abstract</title> <p>Objective This study aims to investigate the effects of moxibustion on the Ubiquitin-Specific Protease 19/P62/NOD-like receptor family, pyrin domain containing 3 (USP19/P62/NLRP3) pathway in RA mouse model,in order to explore its role and mechanisms in treating RA synovitis. Methods Male BALB/c mice were randomly divided into three groups: a control group (Ctrl), an RA model group (RA), and a moxibustion group (Mox), with 8 mice in each group. The RA model was established by intradermal injection at the base of the tail with bovine type II collagen emulsified in incomplete Freund's adjuvant. Mice in the Moxibustion group received moxibustion treatment at the bilateral acupoints of "Shenshu" (BL23) and "Zusanli" (ST36). Each acupoint was treated for 15 minutes per session. The treatment comprised 3 courses, with each course consisting of 7 daily sessions. Arthritis severity was assessed using the arthritis index (AI) score. Pathological changes in the ankle joints were examined by Hematoxylin and Eosin (H&E) staining. Immunofluorescence assay was used to detect the expression of proteins related to the USP19/P62/NLRP3 signaling pathway in synovial tissues. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to determine the expression levels of genes associated with the USP19/P62/NLRP3 signaling pathway in synovial tissues. Results After modeling, the mice exhibited marked redness and swelling of the feet, with a concomitant increase in the arthritis index score. Moxibustion treatment alleviated the symptoms of joint redness and swelling and reduced the arthritis index score. Meanwhile, HE staining revealed focal loss of synovial cells, loose arrangement of fibrous connective tissue, and inflammatory cell infiltration in the joints of RA model mice. These pathological changes were significantly ameliorated following moxibustion intervention. The study found that the expression of inflammatory cytokines and macrophage phenotypes was dysregulated in the RA model. Specifically, there was an up-regulation in the expression of the pro-inflammatory cytokines IL-1β and IL-18, as well as the pro-inflammatory macrophage marker CD38. Conversely, the expression of the anti-inflammatory cytokine Arg1, IL-4, and the anti-inflammatory macrophage marker CD163 was down-regulated. Moxibustion treatment effectively reversed these alterations. Furthermore, moxibustion was shown to modulate the expression of key components within the USP19/P62/NLRP3 pathway, including USP19, P62, LC3II, NLRP3, and IRF4, thereby alleviating synovial inflammation in RA. Conclusion Moxibustion treatment can ameliorate inflammatory injury in RA mice, regulate USP19 expression, further modulate autophagy levels and NLRP3 activity, thereby exerting a therapeutic effect on synovial inflammation in RA mice. The underlying mechanism may be mediated by the USP19/P62/NLRP3 pathway.</p>