| Objective:On the basis of the preliminary study we concluded that 6.75g/kg’s Xiaoyaosan was able to fight against the liver toxicity.So, the project can be used to study the effect of Xiaoyao San against liver injury, at the same time, and whether there is effection on the efficacy of Tripterygium wilfordii or not, which could be the foundation for triptolide treatment of rheumatoid arthritis and other autoimmune diseases with using Xiaoyaosan in prevention and treatment of liver toxicity.Methods:Study one: the toxic effect evaluation of the combination of Xiaoyaosan with Tripterygium wilfordii for the treatment of rheumatoid arthritis model rats:Based on the CIA model, the therapeutic effects and the liver injury of the combined treatment of rheumatoid arthritis model rats were compared between Tripterygium wilfordii and the the combination of Xiaoyaosan with Tripterygium wilfordii.Rats were divided into 5 groups: normal control(NC) group 、 Model(CIA) group, Tripterygium wilfordii(TW) group, Xiaoyaosan low dose(XYSL) group, and high dose(XYSH)group.In NC group and CIA group, the rats were fed with normal saline. The rats in TW group were given saline morning, afternoon were irrigated TW solution(dose 3.75g/kg).The XYSL, XYSH groups of rats in the morning were irrigated Xiaoyaosan solution(crude drug 1.69g/kg, 6.75 g / kg), afternoon were irrigated TW solution(crude drug 3.75g/kg).Continuous administration for 28 days.During the experiment, we should weekly record rats weight, arthritis score and volume and calculate the degree of swelling, And after killing animals, we could detecte ALT, AST, TNF-α and IFN-γ, IL-1, IL-10, IL-17, PGE2 form the rats’ serum, and weight the rats’ liver, spleen, thymus and calculate the organ their index. In the end,we needed to observe and make an value for pathological sections of the liver, ankle joint, knee joint.Study two: the effect of the combination of Xiaoyaosan and Tripterygium wilfordii on the analgesic effect of the Tripterygium wilfordii:1 Torsion body method:Forty BALB / c mice were divided into the following four groups, with 10 rats in each group, half male and half female: normal control(NC) group,Tripterygium wilfordii(TW) group, Xiaoyaosan(XYS) group and Aspirin(AS) group.The NC group according to the weight of the animals in the morning and afternoon were given a certain amount of normal saline. In the morning, the TW group rats was fed with normal saline, and the 5.625g/kg solution was administered in the afternoon.In the morning, the XYS group was irrigated with 10.125g/kg Xiaoyaosan solution, and the 5.625g/kg TW solution was irrigated in the afternoon. Meanwhile, the AS group was irrigated with normal saline in the morning, and in the afternoon with 500mg/kg aspirin. All of they,which for 7consecutive days.In the 60 th minutes after the last administration, the rats was injected intraperitoneally with 0.6% acetic acid solution. After administration of 15 minutes, the latent period and the number of twisting body were recorded. Then the analgesic rate was calculated.2 Hot plate method:there were 50 BALB/c female mice, 40 qualified mice were selected. The method of grouping and administration was the same as that of the Torsion body experiment. Last time to be irrigateed after 60 min, we put the mice on the metal plate,which is constant temperature, then recorded their licking time and calculated the pain threshold, pain threshold increase value, rate of analgesia.Results:Study one: the toxic effect evaluation of the combination of Xiaoyaosan with Tripterygium wilfordii for the treatment of rheumatoid arthritis model rats:1 Hepatic toxicity: ALT and AST levels in TW group and XYSL group were higher than that in NC group(P < 0.01). Compared with group XYSL and group XYSH, group TW was able to decrease ALT and AST levels(P < 0.01). Compared with the TW group,the liver index of XYSH group was decreased(P < 0.05). By observing the liver injury under light microscope, we found that the liver pathological score of XYSH group was lower(P < 0.01) than the TW group and the XYSL group.2 In the treatment of rheumatoid arthritis clinical indicators: compared with NC group, in the CIA group, the left and right foot joint volume of rats, right foot swelling and arthritis index were increased(P < 0.01).However, compared with CIA group, the TW,XYSL, XYSH group those indexes had became reduced(P < 0.01). Compared with the NC group, the levels of TNF-α, IFN-γ and IL-1, IL-17 and PGE2 of CIA rats were increased(P < 0.01), instead IL-10 content was decreased(P < 0.01). However, with CIA group comparison, in the TW, XYSL, XYSH group, they were reduced instead of the level of IL-10 of growing(P < 0.01). Compared with the NC group, the thymus index and spleen index of CIA group were decreased(P < 0.01). Nevertherless, when we compared the CIA group with the rest groups,which except for NC group, it is contrary that these indexes were decreased(P < 0.01). Under the light microscopeto observe the knee and ankle joint pathology, we found that TW group, group XYSL, XYSH rats synovial proliferation and inflammatory cell infiltration, pannus formation, bone destruction pathological score were more decreased than CIA group(P < 0.01).Study two: the effect of the combination of Xiaoyaosan and Tripterygium wilfordii on the analgesic effect of theTripterygium wilfordii:1 Torsion body method:Compared with the NC group, TW group, XYS group, AS group of mice in the number of the reaction times can be proved that was reduced, instead the analgesic rate was increased(P < 0.01). when we compared TW group and XYS group with each other, there was no significant difference in the rate of pain(P > 0.05).2 Hot plate method:Compared with the NC group, TW group, XYS group, AS group of mice in the time of licking can be extended, the analgesic rate increased(P <0.01).However, there was no statistical significance between TW group and XYS group(P > 0.05).Conclusion: The high dose of Xiaoyaosan(6.75 g / kg) can be effective in prevention and treatment of Tripterygium wilfordii on rat liver injury and will not affect the Tripterygium wilfordii treatment of rheumatoid arthritis and analgesic pharmacological effect meanwhile.The low dose of Xiaoyaosan(the effect of 1.69g/kg) does not affect the Tripterygium wilfordii extract for treating rheumatoid arthritis model rats, although, can not prevent the liver injury at the same time. |
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