Therapeutic Effect Of Acute And Chronic Treatments With Koumine On Rheumatoid Arthritis In Lewis Rats

Rheumatoid arthritis is a chronic systemic disease characterized by synovitis. Clinical treatment of rheumatoid arthritis is mainly using drugs. Generally speaking, early administration of RA drugs can achieve a better therapeutic effect and prognosis. The disease whether to be diagnosis correctly and treated promtly is the key to determine the patient’s arthritis to be recover or disability. Commonly used drugs for RA are NSAIDs, DMARDs, glucocorticoid etc. But the side effects of these drugs make them be limited on clinical application. So a new type of high efficiency and low toxicity for rheumatoid arthritis drugs need to be developed. Our previous study found that koumine which is the highest concentration and low toxicity Gelsemium alkaloid from Chinese Gelsemium may have anti-rheumatoid arthritis effect on collagen-induced arthritis model. But the model could not reflect the treatment on the acute phase and the treatment characteristics of RA. In view of this, this paper choose the adjuvant-induced rheumatoid arthritis rat model to administrate koumine on acute phase and chronic phase respectively to further clear the therapeutic effect of koumine in acute and chronic phase in RA. Explore the efficacy of the different dosing regimens of pros and cons, providing a scientific basis for koumine developing into a new drug for RA. The main contents are as follows. 1、Established the adjuvant-induced arthritis model in Lewis rats.For AA induction, female Lewis rats were immunized by injecting into the right hind footpad 100 ul of Complete Freund’s Adjuvant(CFA) prepared as a suspension of 10mg/ml heat-killed Mycobacterium tuberculosis in paraffin oil. The results showed significant joint swelling and MWT reducing, the appearance of rats consistent with the literature description, which can be divided into acute phase(D0-7), remission phase(D8-14) and chronic progressive phase(D15-28). Mainly symptom in acute phase were ipsillteral foot swelling within 24 hours and decreasing MWT; During remission phase, 6 the ipsillteral foot swelling continue to deteriate, but the growth speed slow down; When the rats enter the chronic progression phase, the contralateral foot and forelimbs of rats began to develop subcutaneous erythema, joint swelling and deformity erosion, base of tail and body appear subcutaneous nodules. 2、Effect of acute treatments with koumine in AA rats60 Lewis rats were randomly assigned in acute treatments(0.6, 3.0, or 15mg/kg) with koumine test, indomethacin(2.5mg/kg) test, model or normal group. Acute treatments with koumine were administered beginning 1hour prior to CFA administered and continuing daily for ten days. The model group was given normal saline. 10 non-immunized rats as normal groups were oral administration of an equal volume normal saline. After the inducing, each group continued gavage once a day until the ninth day after the injection. Monitored the change in MWT, hind paw volume, AI score, WBC and ESR to the day 23. Whole rats were sacrificed on 23 th day, and their thymus, liver and spleen were taken carefully to weight, ankles were taken for pathological examination to observe the therapeutic effect of acute treatments with koumine in AA rats. Compared with controls, the MWT in AA models were reduced significantly; Hind paw volume, arthritis index, WBC and ESR were elevated. HE staining showed hind ankle joints had obvious inflammatory cell infiltration, and visible joints structure damage and osteoporosis. The acute treatment showed that all three doses of koumine(0.6, 3, and 15 mg/kg) increased mechanical paw withdraw threshold, inhibited the acute onset of ipsillateral joint swelling within 24 hours, decreased WBC and ESR during the acute phase. After rats entering chronic systemic disease, when acute treatment was terminate for 1 week or longer, it still has a certain protective effect in MWT, WBC and erythrocyte sedimentation rate and outside joint performance, such as rheumatoid nodules form. Acute treatment with koumine still can restrain central immune organ thymus and liver enlargement, improved the structure of the joint inflammation cell infiltration and joint damage in AA rats. 3、Effect of chronic treatments with koumine in AA rats60 Lewis rats were grouped and modeled as former method. Chronic treatments with koumine were commenced 14 th days after CFA administered and continuing daily for ten days. Observed the features of AA in mechanical paw withdraw threshold, hind paw volume, arthritis index, white blood counts and erythrocyte sedimentation rate from day 0 to day 23 after which rats were sacrificed. At the end of the study, the thymus, spleen, and the liver weights were measure, the ankle joints of rats were taken to investigate the change of pathology. The chronic treatment showed that all three doses of koumine(0.6, 3, and 15 mg/kg) increased both foot MWT, inhibited the ipsillteral joint swelling, decreased ESR, improved AI score and the structure of the joint inflammation cell infiltration and joint damage in AA rats during the chronic phase. Analysis of the two treatments, the acute dosing regimen has a better effect in changes of WBC, organ index, et al.In conclusion, our study results have 2 notifications:(1) Acute treatment with koumine can significantly relieve acute and chronic pain and swelling, reduce WBC, ESR, inhibite thymus and liver increases, improve the pathological change of ankles. Koumine may have significant therapeutic effects on rheumatoid arthritis with treatments during acute or chronic phases.(2) Chronic treatment with koumine for RA may still have a treatment effect in chronic phase. But the acute treatment have a better effect on pathological changes, and the chronic dosing regimen can not inhibit model rats increasing thymus and liver, failed to significantly lower elevated WBC. It notified that acute administration may be superior to chronic administration.