Therapeutic Effect Of Puerarin On ACLT-induced Osteoarthritis Model

Osteoarthritis(OA)is a common degenerative joint disease in veterinary clinics,which seriously affects animal health and is one of the main causes of disability in elderly dogs.The current treatment of OA is mainly focused on relieving symptoms and pain,but there are certain side effects and limitations.Therefore,it is necessary to screen safe,effective,and side effects-free drugs for treating dog OA.Puerarin is the main component of plant pueraria lobata,which has various biological functions such as anti-inflammatory,antioxidant and cartilage protection.It is widely used in the treatment of various diseases,but there are few studies on the treatment of OA.In this experiment,the rat OA model was used to explore the therapeutic effect of different doses of puerarin on OA,and compared with the commonly used clinical drug celecoxib,and the best therapeutic dose of puerarin selected in the rat test was applied to canine OA.Model to observe the therapeutic effect of puerarin on canine OA.Forty 5-6 week old male SD rats were randomly divided into control(C)group,model(M)group,celecoxib(Cele)group,puerarin low dose(L)group and puerarin high dose(H)Groups,8 for each group.Group M,Cele,Group L and Group H induced an OA model through Anterior cruciate ligament transection(ACLT),and Group C underwent sham operation.By observing the joint morphology,joint motion range and gait of the rat,it was preliminarily judged that the OA model was successfully established 4 weeks after operation.From the 5th week after surgery,puerarin was injected into the joint cavity of group L(50 mg/kg)and group H(100 mg/kg),50 ?L/time;the same amount of normal saline was given to the joint cavity of group C and group M;Cele Rats in the group were given 1 m L of celecoxib suspension at 2.86 mg/kg twice a week.Rats in each group were given at the same time for 5 weeks.Seven days after the end of the treatment,the rats were sacrificed by cervical dislocation,and samples of serum,urine,tibia and femur were collected.ELISA method was used to detect the contents of IL-1 ?,IL-6 and TNF-? in serum,and the contents of CTX-? and COMP in urine.Observe the morphology of tibia and femur,and perform HE staining and Mankin score;immunohistochemical method to detect the content of MMP-3,MMP-13,ADAMTS-5 and type ? collagen in cartilage;Western blot method to detect MMP-3 and MMP in cartilage-13 content changes.On this basis,12 healthy dogs aged 8-10 years were selected and randomly divided into a control group,a model group and a drug(2 w,4 w,6 w)group,with 4 dogs in each group.The control group was a sham operation group.The model group and the drug administration group constructed the OA model by the ACLT method.After operation,the dog's claudication grade,gait observation,and X-ray examination were used to evaluate the success of the dog's OA model.After successful modeling,puerarin 30 mg/kg was injected into the joint cavity twice a week,2 m L each time.They were administered on Monday and Thursday respectively for 5 weeks.X-ray film of right knee joint was taken to observe the changes of knee joint morphology,and the severity of OA was evaluated by claudication analysis and X-ray examination.Samples of serum,urine and joint fluid were collected from each group,and the contents of IL-1 ?,IL-6 and TNF-? in serum and the contents of CTX-? and COMP in urine and joint fluid were detected by ELISA.Rat test results showed that compared with group M,the degree of articular surface damage in group H was significantly reduced,the Mankin score was reduced,the content of MMP-3,MMP-13 and ADAMTS-5 in joint tissue was significantly reduced,type ? collagen was significantly increased,serum IL-1 ?,IL-6,TNF-? and urine CTX-?,COMP levels were significantly reduced.Screened the best therapeutic dose of 100 mg/kg for clinical dog treatment.Canine OA model reduced the degree of lameness after treatment with puerarin.X-ray results showed that compared with the control group,the model group suffered severe joint damage,accompanied by osteophyte formation,and the dog's joint damage was relatively alleviated after administration.The levels of IL-1 ?,IL-6 and TNF-? in dog serum,as well as the CTX-? and COMP levels in urine and joint fluid were significantly increased in the model group,and significantly decreased from 2 to 6 weeks after administration.The test results show that:(1)Puerarin can reduce the content of MMP-3,MMP-13 and ADAMTS-5 in the cartilage tissue of OA rats,increase the content of type ? collagen,inhibit the degradation of cartilage matrix,reduce cartilage damage,group H effect Better than group L.(2)Puerarin can reduce the content of inflammatory factors IL-1 ?,IL-6 and TNF-? in the serum of OA rats and dogs,reduce the content of cartilage degradation markers CTX-? and COMP,and relieve the clinical symptoms of canine OA.