Study On The Mechanism Of Zhuanggu Huoxue Decoction On Papain-induced Inflammation And Cartilage Matrix In KOA Rats

Objective:1.To observe the effects of papain on the knee joint diameter,pain threshold,the changes of IL-1 ?,TNF-? in serum and the injury degree of articular cartilage in rats,and to evaluate whether the model is successful.2.To observe the changes of knee joint diameter,pain threshold and articular cartilage morphology in rats with papain induced osteoarthritis.The content of inflammatory factors and comp in serum and the expression of wnt16a protein,?-Catenin protein,type ? collagen protein and MMP-13 protein in articular cartilage were detected to explore the mechanism of the formula.Method:1.Establishment and identification of rat knee osteoarthritis model induced by papain injection in the knee joint cavityTwenty SD rats weighing approximately 250 g were randomly divided into a blank group and a papain injection group,with 10 rats in each group.In the model group,after being anesthetized with 10%chloral hydrate,papain was injected into the knee joint cavity to induce the SD rat knee osteoarthritis model,which was injected twice a week for four consecutive weeks.After the modeling was completed,the degree of swelling and pain threshold of the knee and toes of the rats were measured.Then the blood was taken from the abdominal aorta and the rats were sacrificed.The articular cartilage of the rats was dissected to observe the morphology of the articular cartilage and taken photos.Inflammatory factors TNF-?,IL-1? content to identify whether the KOA model was successfully prepared.2.Research on the mechanism of self-made Huoxue Zhuanggu decoction40 SD rats with a body weight of about 250g were randomly divided into blank control group,model group,self-made Huoxue Zhuanggu decoction group with low,high and glucosamine hydrochloride positive In the control group,there were 8 rats in each group.Except for the blank control group,the remaining four groups were induced by papain to establish KOA model rats.After the modeling was completed,the degree of swelling and pain threshold of the knee and toes of the rats were measured.The self-made Huoxue Zhuanggu decoction group and the positive drug group were given self-made Huoxue Zhuanggu decoction and glucosamine hydrochloride gavage,respectively,and the blank group and the model group were given the same amount of 0.9%saline.Once a day for four consecutive weeks.After the treatment was completed,the degree of swelling and pain threshold of the knee and toes of the rats were measured.The blood was taken from the abdominal aorta to kill the rats,the knee joint cavity was dissected,the articular cartilage was collected after the articular cartilage was photographed.The serum levels of IL-1?,IL-6,TNF-?,and COMP were measured by ELISA.The articular cartilage tissue was fixed and decalcified and stained with HE,MASON,and toluidine blue.The degree of articular cartilage damage was observed under a light microscope for scoring.Immunohistochemical method was used to detect the expression of MMP-13,Wnt16a,?-catennin and type ? collagen in cartilage.Result:1.Model identification resultsAfter 4 weeks of modeling,the degree of swelling of the knee joints and toes of the rats showed that compared with the blank group,the model group was significantly increased(P<0.05).The pain threshold of rats showed that compared with the blank group,the pain threshold of the model group decreased significantly(P<0.05).The Mankin score of articular cartilage showed that compared with the blank group,the model group was significantly higher(P<0.05).Comparison of IL-1? and TNF-? content:Compared with the blank group,the model group increased significantly(P<0.05).Therefore,no matter from the macroscopic performance,the inflammation level or the degree of articular cartilage damage,it shows that this model was successfully prepared.2.Self-made Huoxue Zhuanggu Decoction mechanism results:The degree of swelling of the knee joint and tissue of rats showed that:compared with the blank group,the model group was significantly increased(P<0.05).The glucosamine hydrochloride group decreased(P<0.05).The measurement of pain threshold in rats showed that compared with the blank group,the model group was significantly reduced(P<0.05).Compared with the model group,the self-made Huoxue Zhuanggu decoction group and the glucosamine hydrochloride group were both increased(P<0.05).The results of Mankin and OARSI scores of articular cartilage tissue showed that compared with the blank group,the degree of cartilage damage in the model group was significantly increased(P<0.05).Compared with the model group,the self-made Huoxue Zhuanggu decoction group and the glucosamine hydrochloride group were reduced(P<0.05).Serum IL-1?,IL-6,TNF-? and COMP content comparison showed that:compared with the blank group,the model group was significantly increased(P<0.05),compared with the model group,the self-made Huoxue Zhuanggu decoction group and hydrochloric acid The glucosamine group decreased(P<0.05).Immunohistochemistry showed that compared with the blank group,the expression of Wnt16a,MMP-13,?-catennin protein in the remaining groups increased,while the expression of type ? collagen decreased.Compared with the model group,the expressions of Wnt16a,MMP-13 and ?-catennin protein in the self-made Huoxue Zhuanggu decoction group and the glucosamine hydrochloride group decreased,and the expression of type ?collagen increased.Conclusion:Papain injection can induce swelling of knee joint,decrease pain threshold,increase IL-1? and TNF-? in serum,and induce the injury of articular cartilage in rats.Koa model can be successfully prepared.Zhuanggu Huoxue Decoction can effectively regulate the koa of rats,and its mechanism may be to reduce the degradation of type ? collagen,inhibit the inflammatory reaction,reduce the swelling degree of knee joint and toe,and improve the pain threshold of rats by inhibiting the expression of MMP-13,wnt16a and ?-Catenin protein,so as to facilitate the recovery of KOA symptoms.