| Background: Osteoarthritis(OA)is a highly prevalent disease worldwide,which affects all mammals and is an important cause of disability(1).With the development trend of the disease,it will slowly endanger the normal function of animal bone and joint and reduce the quality of life.Drug therapy is the most common treatment used in clinical practice,which can control the disease and delay the development of the disease by managing the pain of sick animals and reducing or relieving the clinical symptoms.However,the drugs used in clinical practice have certain limitations or adverse reactions,so it is very necessary to excavate the drugs with good therapeutic effect,strong application,small side effects,no addiction and low cost for clinical treatment.Koumine and gelsemine are natural drugs extracted from green plants,which has the effects of anti-infection,pain relief,cartilage tissue protection,treatment of cardiovascular diseases and anti-cancer drugs,and has the advantages of small therapeutic dose,less side effects,no dependence,and common sources.Objective: To establish a drug induced model in this experiment.The drug was a mixture of 2% papain and 0.03 mol/L L-cysteine,which was injected into the articular cavity of rats.After the model was successfully identified,Meloxicam was used as a positive control,i.e.The model rats were treated with different doses of koumine and gelsemine to explore the therapeutic effect of koumine and gelsemine on osteoarthritis,and the most appropriate therapeutic dose was selected to provide reference for clinical treatment of osteoarthritis.Methods: First,12 male 180-220 g SD rats were divided into two groups,6 rats in the blank control model group.Osteoarthritis models were induced by intra-articular injection of0.2 ml 2% papain with 0.03 mol/L L-cysteine in rats in the model group on days 1,4,and 7,respectively.Two weeks later,the rat osteoarthritis model was judged to be successful by the results of X-ray photography of the rat knee joint and Safranin O fast green staining and Hamp E staining of the knee joint followed by Mankin’s score.Secondly,42 male SD rats were divided into blank group(KB),model group(M),koumine low-dose group(ZD),koumine high-dose group(ZG),koumine A low-dose group(JD),koumine A high-dose group(JG)and meloxicam control group(MX)with six rats each.Except for the blank group,each group was first modeled by the above method.After successful modeling,according to the set dose,the low-dose group 0.5 mg/kg and the high-dose group 1 mg/kg were orally gavaged with 1 m L of the drug in the test group,the blank control group and the model group were gavaged with the same amount of 1 m L of normal saline.The positive control meloxicam group was gavaged with 2 mg/kg meloxicam on the first day,and then the maintenance dose was 1 mg/kg.The rats were intragastrically administered once daily,and sampling was performed after 2 weeks of continuous oral administration.The rats were fasted for 12 hours before sampling,and general anesthesia was performed with sulfamethoxazole for X-ray shooting.The scoring was performed according to the X-ray scoring table modified by the canine osteoarthritis scoring system.Serum was collected,and histological pathological sections were made from the knee joints of rats and stained with Hamp E.After light microscopic observation,they were scored according to the Mankin’s score table,and the results of the X-ray score table and Mankin’s score were compared to verify the feasibility of the modified X-ray score table.Enzyme-linked immunosorbent assay(ELISA or ELASA)was used to detect the serum levels of interleukin-1 β(IL-1β),tumor necrosis factor-α(TNF-α),cyclo-Oxygen-Ase-2(COX-2)and prostaglandin E2(PGE2).Results: The results of pathological section showed that compared with group M,the scores of JG group and ZG positive control group were decreased(p<0.05),and the scores of JD and ZD groups were also decreased(p>0.05);compared with the positive control group,the scores of koumine high dose group were decreased(p>0.05).These results indicated that the high and low doses of koumine A had a down-regulation effect on proinflammatory factors in the serum of rat G osteoarthritis model,with the down-regulation effect being most significant in the high dose group of koumine A(1 mg/kg).ELISA results showed that the levels of inflammatory factors were decreased in all drug groups,but TNF-α and PGE2 were not statistically significant in each drug group(p>0.05),and the results of IL-1β factor showed that the down-regulation of IL-1β was statistically significant in the ZG JD group(p<0.05),with the down-regulation of IL-1β being more significant in the ZG group.The results of COX-2 factor showed that the COX-2 levels in each drug group were significantly decreased(p<0.05),with the down-regulation of COX-2being more significant in the ZG group.Radiographic X-ray results indicate that Comparing with the model group,the X-ray scores of bone and joint injury in the ZG,JD,and JG groups were significantly reduced(p<0.05),with the effect of koumine high-dose group in improving OA model injury being the most significant.The high dose of gelsemine was the second,and the therapeutic effect of the low dose of koumine was weak.Conclusion:(1)The rat osteoarthritis model can be successfully established two weeks later by injecting the drug — 2% papain + 0.03 mol/L L-cysteine mixture into the knee joint cavity by drug induction in rats on days 1,4,and 7.(2)There was a high agreement between the X-ray scoring results and Mankin’s scoring results,indicating that the modified X-ray scoring system on rat osteoarthritis can be used as a clinical reference.(3)Koumine and gelsemine reduced the contents of proinflammatory cytokines IL-1β,TNF-α,COX-2 and PGE2,relieved pain,reduced the clinical symptoms of osteoarthritis,and controlled the development of the course of osteoarthritis in a rat model of OA.(4)1 mg/kg dose of koumine had the most significant effect on the rat osteoarthritis model. |
