| Objective:The objective of the experimental study is to explore the mechanism of Modified Xuanbi decoction in treating acute gouty arthritis,by evaluating the inflammation index,dysfunction index,skin temperature and foot swelling degree of left ankle joint in rats with acute gouty arthritis,measuring the levels of serum UA,IL-1β,TNF-α and CRP,observing the histomorphological features of ankle synovium,detecting and analyzing the mRNA expression of TLR4,MyD88 and IRAK4 and taking TLR4/MyD88/IRAK signaling pathway as the entry point.Methods:After one week of adaptive feeding,a total of 60 Wistar rats were randomly divided into normal group,model group,colchicine group and low,medium and high dose groups of Modified Xuanbi decoction,with 10 rats in each group,fed with ordinary feed and distilled water every day.Hyperuricemia(HUA)model combined with acute gouty arthritis(AGA)model was prepared:adenine and ethambutol mixed solution was used at 10mL/kg for 21 days,once a day,to establish the HUA model.The normal group was given equal amount of distilled water by gavage.After the establishment of the hyperuricemia model on the 15 th day,the inflammatory symptoms of the left ankle joints of the rats in each group were evaluated,and 0.2ml sodium urate suspension was injected into the left ankle joints of each rat to prepare the AGA model.Meanwhile,the normal group was injected with an equal amount of sterile saline.The inflammatory symptoms of ankle in rats were evaluated again within 4 hours after modeling,and the same method was used before daily drug intervention.The evaluation of joint inflammatory symptoms was performed for a total of 8 times.After the model was established successfully,colchicine and different doses of Modified Xuanbi decoction were administered in groups once a day for 7 consecutive days.Blood,left ankle synovial fluid,ankle joint and its surrounding soft tissue were collected from all rats after 3 hours of the administration of drugs on the last day.The morphological features of synovial membrane of the ankle joint were observed by HE staining.Serum concentrations of UA,IL-1β,TNF-α and CRP were detected,and mRNA expression levels of TLR4,MyD88 and IRAK4 in synovial fluid were detected by real-time quantitative PCR.Results:1.Assessment results of ankle inflammatory symptoms: Starting from 24 hours after modeling was established(administer the drug once),the skin temperature of the low-dose group of Modified Xuanbi decoction,and the inflammation index,dysfunction index,skin temperature and foot swelling degree of rats in the medium and high dose group of Modified Xuanbi decoction were lower than those in the model group(P<0.05).From 48 hours after the model was established(two times of drug instillation),the inflammation index,dysfunction index and foot swelling degree of the low-dose rats of Modified Xuanbi decoction were lower than those of the model group(P<0.05).2.Results of serum UA concentration: Compared with the model group,the serum UA level of rats in the low-dose,medium-dose and high-dose groups of Modified Xuanbi decoction was significantly lower(P<0.05),and the serum UA level in the high-dose group of Modified Xuanbi decoction was closest to that of the normal group.3.Serum IL-1β,TNF-α and CRP contents: Compared with the model group,the levels of serum IL-1β,TNF-α and CRP were significantly lower in the low,medium and high dose groups of Modified Xuanbi decoction and Colchicine group(P< 0.05).The serum levels of IL-1β,TNF-α and CRP in the medium and high dose groups were lower than those in the Colchicine group(P<0.05).4.Pathological morphological results of HE staining of the synovial tissue of the ankle joint: The surface layer of the synovial tissue of the model group was damaged;the synovial cells were proliferated;the inflammatory cells were closely clustered and disorderly;the stroma is edematous;congestion were found in the endovascular and lumen deformation were increased.Rats’ ankle synovial tissue were more loosely arranged in Colchicine group and the low,medium and high dose groups of Modified Xuanbi decoction,with less inflammatory cells and significantly less vascular congestion in the same multiple field of vision.The pathological morphology of synovial tissue in the Modified Xuanbi decoction high dose group was the closest to that in the normal group.5.RT-qPCR results: Compared with the model group,the mRNA expression levels of TLR4,MyD88 and IRAK4 in the rats of Colchicine group and Modified Xuanbi decoction groups were down-regulated to different degrees(P<0.05).In addition,the down-regulation effect of TLR4,MyD88 and IRAK4 mRNA in the medium and high dose groups of the Modified Xuanbi decoction was better than that in the Colchicine group.Conclusion:1.Modified Xuanbi decoction can effectively improve the general situation,acute inflammatory symptoms of ankle joint(inflammation index,dysfunction index,skin temperature,foot swelling degree),high serum uric acid level and synovial tissue inflammatory lesions of HUA combined with AGA model in rats,and has a certain degree of anti-inflammatory,analgesic,antipyretic,swelling and uric acid lowering effect,in which the anti-inflammatory effect is significantly reduced in response to the serum CRP level.2.Modified Xuanbi decoction significantly reduced the expression levels of TLR4,MyD88 and IRAK4 mRNA in the synovial fluid of AGA rats’ ankle joints.3.Modified Xuanbi decoction inhibits the activation of TLR4,MyD88 and IRAK4,blocks the signal transduction of the mediated signal pathway,leads to the decrease and release of inflammatory factors TNF-α and IL-1β,and plays a role in the treatment of AGA,which may be one of the anti-AGA mechanisms of Modified Xuanbi decoction. |
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