鼠尾草酚调节AMPK/NLRP3信号通路对H2O2诱导的骨关节炎软骨细胞凋亡、氧化损伤和炎症反应的影响OA

Objective:To explore the effects of carnosol(CAR)on apoptosis,oxidative damage,and inflammatory response of chondrocytes induced by H2O2 in osteoarthritis(OA),and to analyze whether isthey are related to the adenosine monophosphate activated protein kinase(AMPK)/Nod like receptor protein 3(NLRP3)pathway.Methods:Mouse primary chondrocytes were culturedisolated and isolated,cultured,and assigned into control group(CK group),CAR group,H2O2 group,low,medi-um,and high-dose CAR groups(L,M,H-CAR groups),and H-CAR+AMPK inhibitor Compound C group(H-CAR+Compound C group),CK+Compound C group.The prolifer-ation,apoptosis of chondrocytes,and ROS fluorescence intensity were detected using Calcein-AM,Hoechst 33342 stai-ning,and DCFH-DA fluorescence probe method,respectively.The levels of SOD,MDA,TNF-α,IL-6,and IL-8 were meas-ured using ELISA.In addition,Western Blot was used to measure cartilage specific proteins and AMPK/NLRP3 pathway related proteins.Results:The isolation and culture of primary chondrocytes were successfully achieved.There were no significant differences in cell proliferation rate,SOD,Col2α1 expression,p-AMPK/AMPK,cell apoptosis rate,ROS fluorescence intensity,MDA,TNF-α,IL-6,IL-8 levels,and NLRP3 expression between the CAR group,CK+Compound C group and the CK group(P>0.05).The H2O2 group showed decreased cell proliferation rate,SOD level,Col2α1 expression,and p-AMPK/AMPK,while increased apoptosis rate,ROS fluorescence intensity,MDA,TNF-α,IL-6,IL-8 levels,and NLRP3 protein expression than the CK group(P<0.05).Compared with the H2O2 group,the aforementioned indicators in the L-,M-,and H-CAR groups exhibited op-posite changes.Compared with the H2O2 group,the cell proliferation rate,SOD,Col2α1 expression,and p-AMPK/AMPK in the H-CAR group increased from 38.64%±4.12%,(28.16±3.02)U/mL,0.30±0.04,and 0.31±0.04 to 91.34%±8.37%,(62.78±6.46)U/mL,1.07±0.12,and 0.79±0.08,respectively;the cell apoptosis rate,ROS fluorescence intensity,levels of MDA,TNF-α,IL-6,and IL-8,and NLRP3 expression decreased from 31.57%±3.35%,68.15±7.02,(4.78±0.49)nmol/mL,(51.69±0.54)pg/mL,(32.09±3.37)pg/mL,(67.38±0.69)pg/mL,and 1.28±0.14 to 6.78%±0.71%,12.34±1.38,(2.11±0.25)nmol/mL,(22.03±0.26)pg/mL,(10.07±1.10)pg/mL,(34.70±0.38)pg/mL,and 0.51±0.06,respectively.The H-CAR+Compound C group showed decreased cell proliferation rate,SOD,Col2α1 expression,and p-AMPK/AMPK,while increased apoptosis rate,ROS fluorescence intensity,MDA,TNF-α,IL-6,IL-8 levels,and NLRP3 expression than the H-CAR group(P<0.05).Conclusion:CAR may alleviate H2O2 induced apoptosis,oxidative damage,and inflammatory response in OA chondrocytes by regulating the AMPK/NLRP3 pathway,thereby exerting a protective effect on chondrocytes.