Demethoxycurcumin Delay The Progression Of Osteoarthritis Via The NRF2-ARE/AGE-RAGE Axis

Osteoarthritis(OA)is a chronic degenerative disease of the joint,mainly characterized by articular cartilage degeneration,subchondral bone sclerosis,synovial hyperplasia,and the formation of osteophytes at the edge of the joint.Many endogenous adverse factors,including inflammation,oxidative stress,aging,and hyperglycemia,lead to the deposition of advanced glycation end products(AGEs)in the body.AGEs activate the AGE-RAGE signaling pathway by binding to the receptor RAGE on the cell membrane,thereby upregulating related enzymes such as NADPH oxidase 4(NOX4).With the excessive production of reactive oxygen species(ROS)in mitochondria,cells appear oxidative stress,while mitogen-activated protein kinase(MAPK)and nuclear factor kappa-B(NF-κB)signaling pathways are activated,thereby upregulating the expression of proapoptotic proteins and inflammatory factors.Under the action of matrix metalloproteinases(MMPs)and a disintegrin and metalloproteinasease with thrombospondin(ADAMTS),the cartilage extracellular matrix(ECM)is lost largely,which ultimately leads to the occurrence of OA.Demethoxycurcumin(DMC)is a natural curcumin(CUR)analogue.Compared with CUR,DMC lacks methoxy combined with benzene ring,and has better solubility and bioavailability.However,the biological characteristics of DMC in the treatment of OA are not yet clear.Therefore,this study revealed the feasibility of DMC in treating OA through in vitro and in vivo experiments,elucidated the mechanism of action of DMC,and explored a new pathway for the treatment of OA.Part I Demethoxycurcumin delay the progression of osteoarthritis by inhibiting the degradation of cartilage extracellular matrixObjective: To detect the effect of DMC on cartilage ECM and evaluate the efficacy of DMC in treating OA.Methods: Through network pharmacology analysis,a "drug-disease-pathwaytarget" network was constructed to predict the potential of DMC in treating OA.The optimal concentration range of DMC was screened through the measure of cell viability.The expression of ECM was detected by using techniques such as Western blotting(WB),immunofluorescence(IF)staining,establishment of rat OA model,knee joint cavity injection,knee joint function testing,histological evaluation,and OARSI scoring.Results: DMC is significantly correlated with biological functions such as oxidative stress,inflammatory response and apoptosis.For chondrocytes,the optimal concentration range of DMC is 0.01-1 μM。 In vitro studies,the expression of ACAN,COL2A1,SOX9,and PRG4 was upregulated after DMC intervention,while the expression of ADAMTS5 and MMP13 was downregulated.In vivo studies,after DMC intervention,the response time to pain for rats was prolonged,the difference in weight between both hind limbs was reduced,the OARSI score of the knee joint for rats was reduced,the loss of proteoglycans,chondroitin sulfate and hyaluronic acid was reduced,the number of ACAN and COL2A1 positive cells increased,while the number of ADAMTS5 and MMP13 positive cells decreased.Conclusion: DMC promotes the synthesis of cartilage ECM,inhibits the degradation of cartilage ECM,alleviates the pain manifestations of OA,and delays the development of OA.Part II Demethoxycurcumin inhibits oxidative stress,inflammatory response and apoptosis induced by the deposition of advanced glycation end productsObjective: To elucidate the mechanism of DMC delaying the progression of OA.Methods: The effects of DMC on chondrocytes were detected by transcriptome sequencing,molecular docking,WB analysis,IF staining and flow cytometry(FCM).Results: ECM related biological functions as well as AGE-RAGE,MAPK and NF-κB signaling pathways in chondrocytes were significantly enriched.The nuclear translocation of NRF2 increased after DMC intervention,the expression of HMOX1,NQO1,GCLC,SOD2,GPX1 and CAT was upregulated,the expression of NOX4 was downregulated,and the production of ROS was reduced.Meanwhile,the expression of BCL2 was upregulated,while the expression of IL6,TNF,COX2,NOS2,BAX,CCASP3 and C-PARP1 was downregulated after DMC intervention.In addition,the phosphorylation modification of ERK,JNK,p38,p65 and IκBα was inhibited.Conclusion: DMC activates NRF2-ARE signaling pathway and inhibits AGE-RAGE,MAPK and NF-κB signaling pathways,eliminates oxidative stress,inflammatory reaction and apoptosis caused by the deposition of AGEs.