The Mechanism Research Of FH535 Regulating Knee Osteoarthritis By Blocking Wnt/β-catenin Signaling Pathway

Objective:To explore the therapeutic effect of FH535 on osteoarthritis in rats,and to detect the structural changes of knee joint,pathological changes,expression of related pathways and changes of related inflammatory indexes in order to understand the role of FH535 in osteoarthritis model and its related regulatory mechanism.Methods:(1)Chondrocytes of newborn rats were digested and extracted,and chondrocytes were stimulated by interleukin-1β(IL-1β)to construct an inflammatory model,and the model was intervened by FH535 with different drug concentrations.After the intervention,RNA and related proteins were extracted for real-time quantitative PCR(QPCR).Western blot(WB)was used to detect the protective effect of FH535 on IL-1β-induced model,and Immunofluorescence technique(IF)was used to detect the expression levels of inflammatory factors and protective factors and the expression of β-catenin related signal pathway.(2)Six-week-old mice were divided into blank control group,arthritis model group,low-concentration drug intervention group and high-concentration drug intervention group,and the animal model of knee osteoarthritis in mice was established by destabilization of the medial meniscus(DMM).After dissolving FH535 with cosolvent,the animal model of mouse knee joint was intervened by intraperitoneal injection at regular intervals,and the isolated knee joint was reconstructed by micro-computed tomography(Microct).The degree of knee joint degeneration in each group was analyzed from two-dimensional and three-dimensional perspectives,and the therapeutic effect of the drug was analyzed by measuring bone mineral density and other related indicators.At the same time,the protective effect of the drug on articular cartilage of DMM osteoarthritis rats and its resistance to cartilage degeneration were observed from the histological morphology and immunofluorescence.Results:(1)In vitro cell experiment,compared with arthritis model group,the inflammatory indexes of FH535 on chondrocytes in experimental group decreased and the protective factors increased obviously;(2)In the animal experiment of knee osteoarthritis in mice intervened by FH535,after FH535 treatment,the expression of joint-related inflammation in mice decreased,while the expression of related protective proteins increased,and the morphological manifestations were reduced cartilage wear of knee joint,subchondral sclerosis and osteophyte formation.(3)IF results showed that the expression of β-catenin decreased,the expression of P-β-catenin increased and the expression of Axin2 increased in3)FH535-treated mice.Conclusion:(1)FH535 has a strong function of protecting cartilage,delaying the process of cartilage fibrosis and resisting joint degeneration.(2)Inhibition of Wnt/β-catenin signaling pathway can effectively delay the occurrence of OA.FH535 can reduce the degradation of Axin2,thus inhibiting the expression of β-catenin and down-regulating Wnt/β-catenin signaling pathway,thus achieving the purpose of improving OA.