1.Study on the Effect of Lentivirus-Mediated NLRP3 Overexpression on Levels of Anti-CCP and RF in Synovial Fluid of Experimental RA Rabbits Regulated by Moxibustion
Xin YANG ; Yi WANG ; Shenqiao YANG ; Wenbin MA ; Fei WU ; Donglai XI ; Xiaoqing LU ; Jun CHEN ; Haiyan ZHOU
World Science and Technology-Modernization of Traditional Chinese Medicine 2023;25(6):2089-2096
Objective Observation of the anti-inflammatory and detumescence effects on experimental rheumatoid arthritis(RA)rabbits by moxibustion,With lentiviral vector mediated NLRP3 overexpression in RA animal models,to investigate the effect of moxibustion on anti-CCP/RF levels in synovial fluid of RA rabbit joints and its regulation mechanism of NLRP3.Methods Randomly divided thirty Japanese big-ear white rabbits into 5 groups:group of blank,group of model,group of moxibustion,NLRP3 overexpression group and NLRP3 overexpression negative group,with 6 rabbits in each group.Both joint of the model group,and the groups with moxibustion was injected into FCA(0.5 mL·kg-1),the blank group used the same method to inject sterile saline.On the 3rd day and the 10th day after treatment,10 μL of NLRP3 overexpression lentiviral vector was injected into the joint cavity of the NLRP3 overexpression group of rabbits.The rabbits in the NLRP3 overexpression negative group were injected with the same amount of LvGFP as Contrast.moxibustion bilateral"BL23","ST 36"of the groups with moxibustion(5 cones with one point in a day.6 days for 1 course,1 day for rest,3 courses of treatment in total.)After treatment,observe the circumference of each rabbit's left and right knee joints,the content of RF,anti-CCP,IL-18 in synovial fluid was detected by ELISA.Results Compared with the blank group,the left and right knee joint circumferences in the model group increased,and the anti-CCP,RF,and IL-18 contents in the synovial fluid of the knee joint increased significantly(P<0.01);compared with the modle group,The circumference of the knee joint decreased,and the content of anti-CCP,RF and IL-18 decreased significantly(P<0.01);compared with the moxibustion group,the circumference of the left and right knee joints in the NLRP3 overexpression group increased(P<0.05),the content of anti-CCP,RF and IL-18 increased(P<0.05).Conclusions Moxibustion has anti-inflammatory effects on experimental RA rabbits.Moxibustion at Shenshu(BL23)and Zusanli(ST36)can significantly reduce the level of anti-CCP,RF the immune effector,also IL-18,the in synovial fluid of RA rabbits knee joint,yet this effect can be declined significantly under NLRP3 overexpression,suggesting that moxibustion performed its anti-inflammatory regulation by suppressing abnormal immune function in experimental RA rabbits,which may be closely related to the NLRP3 expression.
2.Effects of Moxibustion on NLRP3 Inflammasome and Caspase-12 in Synovium of Experimental RA Rabbits
Xiaoqing LU ; Huahui LIU ; Jun CHEN ; Donglai XI ; Yi WANG ; Shenqiao YANG ; Haiyan ZHOU ; Wenbin MA ; Fei WU ; Xin YANG
World Science and Technology-Modernization of Traditional Chinese Medicine 2023;25(9):2975-2982
Objective To observe the effects of moxibustion on the expression of Nucleotide binding oligomeric domain like receptor thermoprotein domain associated protein 3(Nod-like receptor protein 3,NLRP3)inflammasome and Cysteine aspartate protease 12(Caspase-12)in synovial tissue of experimental Rheumatoid Arthritis(RA)rabbits,and explore the anti-inflammatory mechanism of moxibustion on RA rabbits.Methods Twenty-four Japanese big-eared white rabbits were randomly divided into 3 groups according to body weight and sex:the control group,the model group and the moxibustion group.The Freund's Complete Adjuvant(FCA)was injected into the joint cavities of two knee joints of rabbits at the dose of 0.5 mL·kg-1 in the model group and the moxibustion group,and the control group was injected with the same amount of normal saline as control.For the moxibustion group,scar free moxibustion was operated at bilateral"BL 23"and"ST 36"acupoints,moxibustion for 3 times per acupoint per day,continuous treatment for 6 days,rest for 1 day,as a course of treatment,a total of 3 courses of treatment.The histological changes of synovium samples of rabbit knee joints were observed under light microscope,real-time quantitative polymerase chain reaction(RT-qPCR)was used to detect the expression of NLRP3,Cysteine aspartate protease 1(Caspase-1),Caspase-12 in synovial tissues and Enzyme-linked immuno sorbent assay(ELISA)was used to measure the change of Interleukin-1β(IL-1β)content in the synovial fluid of experimental animals.Results Compared with the control group,the degree of synovial lesions(inflammatory cell infiltration,synovial tissue hyperplasia,fibrous tissue hyperplasia,macrophage hyperplasia,etc.)and pathological score were increased(P<0.05),the expression of NLRP3,Caspase-1 mRNA were increased(P<0.05),the expression of Caspase-12 mRNA was significantly downregulated(P<0.01)and the expression content of IL-1β was significantly up-regulated(P<0.01)in the model group;Compared with the model group,the synovial lesion degree and pathological score were decreased(P<0.05),the expression of NLRP3,Caspase-1 mRNA were decreased(P<0.05),the expression of Caspase-12 mRNA was significantly increased(P<0.01)and the expression of IL-1β was decreased(P<0.05)in the moxibustion group.Conclusion Moxibustion can significantly inhibit the pathological process of knee synovitis in RA rabbits;Moxibustion at"Shenshu(BL 23)"and"Zusanli(ST 36)"acupoints can significantly up-regulate the expression of Caspase-12 and then inhibit the NLRP3 inflammasome,which may be one of mechanisms of moxibustion to reduce RA inflammatory response.