Electroacupuncture Ameliorates Knee Osteoarthritis In Rats Via Inhibiting NLRP3 Inflammasome And Reducing Pyroptosis

Objective:Knee Osteoarthritis(KOA),is the most common joint disease worldwide.The pathogenesis of KOA is complex and electroacupuncture(EA)is an effective therapy for KOA,but the mechanism remains unclear.In this study,we aim to investigate the potential therapeutic effect of EA on the rat model of KOA induced by monosodium iodoacetate(MIA)and its relationship with NLRP3 inflammasome.Methods:KOA was induced by intra-articular injection of MIA(3mg/50μL)into the right knee joint of rats.Forty-five male rats weighing 250–300 g were randomly divided into 3groups: control group,KOA group,and KOA + electroacupuncture group(KOA+EA).EA treatment lasted for 2 weeks(6 times a week).Paw withdrawal threshold tests were used to assess mechanical allodynia once a week.Safranin O/Fast Green and hematoxylin and eosin(H&E)staining were used to assess the damage to cartilage,synovium,and subpatellar fat pad(IFP).Immunohistochemistry was used to observe NLRP3 inflammasome-associated protein-positive cells in the same field of view and western blot was used to detect the expression of the associated protein in cartilage tissue.Results:1.The results of mechanical allodynia pain values in rats showed that the ipsilateral paw withdrawal threshold(PWT)was significantly reduced in the KOA group and the KOA+EA group after monosodium iodoacetate injection.After two weeks of treatment,the KOA+EA group showed different degrees of improvement compared with the KOA group,and the KOA+EA group was superior to the KOA group with significant differences.2.The results of joint swelling showed that the knee joints of the Ctrl group had no redness and swelling,normal flexion and extension functions,transparent light white articular cartilage,and neat joint edges with no deformation.In the KOA group,there was obvious redness,swelling and deformation,congestion and thickening of synovial tissues,and impaired flexion and extension functions,with obvious bone flaps visible at the edges.the KOA+EA group had no obvious tissue redness and swelling,no deformation of knee joints,and normal flexion and extension functions.The function was normal.3.The results of Safranin O/Fast Green staining showed that the Ctrl group showed intact articular cartilage tissue structure,a smooth cartilage surface,sharp and uniform coloration of the whole layer,a clear boundary between cartilage and bone,normal chondrocyte morphology.In the KOA+EA group,the degree of structural damage to cartilage tissue was significantly improved compared with the KOA group,and the structure was intact,with loss of coloration of the cartilage surface layer only.the OARSI score of the right knee joint in the KOA group was significantly higher than that of the control group,and the score of the KOA+EA group was significantly lower compared with that of the KOA group.4.The results of H&E staining showed that the cartilage tangent layer was continuous and intact in the Ctrl group,with uniform distribution of cytoplasmic and matrix components,whereas the cartilage tangent layer fibers were thinned and the cartilage destruction was severe in the KOA group,with complete disappearance of the tide line.the cartilage surface was continuous and more regular in the KOA+EA group,with the neater arrangement of chondrocytes and clear structure of the radiolucent and calcified layers.5.Immunohistochemical results showed that in cartilage and synovial tissues,CD68,TNF-α,MMP13,NLRP3,Caspase-1,ASC,IL-1β,and IL-18 staining was heavier,and positive cell expression was significantly higher in the KOA group compared with the Ctrl group,and positive expression was significantly decreased in the KOA+EA group,with statistically significant differences.6.Western Blot results showed that NLRP3,caspase-1,ASC,IL-1β,GSDMD,and GSDMD-N-terminal expression were significantly increased in cartilage tissue in the KOA group,and the KOA+EA group showed a decreasing trend compared with the KOA group,and the differences were statistically significant.Conclusion:In summary,the purpose of this study was to explore the role of EA in the treatment of KOA and found that EA could inhibit NLRP3 inflammasome and reduce reduce pyroptosis,thereby protecting cartilage tissue.It provides reliable evidence for the development of EA in the treatment of KOA and the clinical application of acupuncture.