The Experimental Study Of Interference Expression Vector ShRNA-ADAM8/EGFR Transfection The Animal Model Of Rat Arthritis

Objective : Based on molecular experiments,shADAM8 and shEGFR are evaluated in articular cartilage damage from the overall level of animals.The therapeutic effect of repair,verify the molecular mechanism of targeted inhibition of ADAM8 and EGFR to improve cartilage degeneration,and lay a drug foundation for the next step of arthritis treatment.Methods:1.Establishment of arthritis model2.Transfection of rat arthritis model(1)Normal joint group: no special treatment(2)Simple iodoacetic acid(blank control group): 0.2ml 0.9% Nacl solution was injected into the joint cavity(3)rAd-col2A1-shRNA-ADAM8 interference group;the right posterior knee joint was injected with 1.6×108 pfu/ml rAd-col2A1-shRNA-ADAM8 recombinant adenovirus 0.2ml.(4)rAd-col2A1-shRNA-EGFR interference group;the right posterior knee joint was injected with 1.6×108 pfu/ml rAd-col2A1-shRNA-EGFR recombinant adenovirus 0.2 ml.(5)Empty virus injection group(negative control group);intraocular cavity injection of1.6×108 pfu/ml adenovirus empty vector 0.2 ml in the right posterior knee joint after the model was taken.3.Indicator detection:(1)General observation;animal vital signs,living habits and gait changes,observe the healing of the incision,with or without redness and infection.(2)General specimen observation;knee joint capsule with or without sputum,adhesion and new organism formation;synovial membrane with or without congestion,edema and synovial fluid;cartilage with or without erosion,articular surface edge with orbital formation,meniscus injury,etc.Osteoarthritis changes;repairs the flatness,gloss,color and binding of surrounding tissues.(3)Pathological examination;staining of hematoxylin-eosin staining(HE),observing the histological morphology of synovial membrane and cartilage,whether it was infiltrated byinflammatory cells;staining with toxin 0-fixed green and toluidine The staining of chondrocytes and the pathological mechanism of osteoarthritis were observed by blue staining.The expression of Collagen II,Aggrecan and MMP-9 was observed by immunohistochemical staining(IHE).(4)QPCR detection: At gene level,the expression of ADAM8,EGFR,MMP-9,Collagen II and Aggrecan were detected by QPCR,and the relationship between them was analyzed.The molecular mechanism of shADAM8 and shEGFR in improving cartilage degeneration was validated.SPSS1 9.0 software was used to analyze the gene level,and the differences among the groups were compared by one-way ANOVA.The statistical significance was analyzed by P < 0.05.RESULTS: The pathological mechanism of osteoarthritis was that the degradation of extracellular matrix was greater than that of the synthesis.The ADAM8,EGFR,and MMP-9levels were lower in the drug-administered group than in the blank control group,and the Arrencan and Collagen II levels were elevated.And statistically significant(P <0.05).Conclusion: 1.In the model of osteoarthritis,the expression of ADAM8,EGFR and MMP-9 increased,which was positively correlated with the severity of osteoarthritis.The results were consistent with molecular experiments.2.The new pathways found in previous molecular experiments were validated in animal models.The ligand produced by ADAM8 combined with EGFR regulated the production of MMP-9 through p38 MAPK pathway,which degraded the extracellular matrix and induced the apoptosis of chondrocyte.3.Inhibition of ADAM8 or EGFR may delay the development of osteoarthritis.