Acute Gouty Arthritis Rats Research Articles

Mechanism of Biqi capsules in the treatment of gout based on network pharmacology and experimental verification

Ethnopharmacological relevanceGout is a crystal-related arthropathy caused by monosodium urate (MSU) deposition, resulting from purine metabolism disorders and hyperuricemia (HUA). Gout belongs to the traditional medicine category of Bi syndrome. Biqi capsules (BQ) is a traditional Chinese medicine formula used to treat Bi syndrome. The BQ prescription is derived from the ancient prescription of Hua Tuo, a famous physician in the Han Dynasty. Aim of the studyTo study the effect and mechanism of BQ in treating acute gouty arthritis (AGA) and HUA. Materials and methodsAnalyzing BQ's signaling pathways for gout treatment via network pharmacology. The HUA model was induced orally with adenine and potassium oxonate. The rat AGA model was established by MSU injection. In vitro, MH7A and RAW 246.7 cells were treated with LPS and MSU. Serum uric acid, creatinine, and urea nitrogen levels were evaluated. Kidney and ankle joint pathology was observed via HE staining. Inflammatory signaling pathway proteins, epithelial-mesenchymal transition (EMT) pathway proteins, and uric acid metabolism-related proteins were detected by Western blot. Results1780 potential targets for gout treatment were identified, and 1039 target proteins corresponding to BQ's active ingredients were obtained. Pathway enrichment analysis revealed BQ improved gout mainly through inflammatory pathways. Experimental results showed BQ could reduce serum uric acid level and increase uric acid clearance rate by regulating the expression of adenosine deaminase (ADA), and organic anion transporter 1 (OAT1) and glucose transporter 9 (GLUT9) in HUA mice. BQ could improve renal function and injury by inhibiting the NLRP3 pathway in HUA mice' kidneys. Additionally, BQ could alleviate ankle joint swelling and synovial injury, inhibit the TLR4/NLRP3 pathway, and reduce levels of inflammatory factors including interleukin 6 (IL-6), interleukin 1β (IL-1β), and tumor necrosis factor-alpha (TNF-α) in AGA rats. The main component of BQ, brucine, could inhibit the activation of NLRP3/NF-κB pathway induced by MSU and reduce the expression level of inflammatory factors (IL-6, IL-1β, and TNF-α) in macrophages. Brucine could inhibit the activation of the EMT pathway and reduce the expression level of inflammatory factors (IL-6, TNF-α) in human fibroblast-like synoviocytes (MH7A cells) induced by MSU. ConclusionsBQ effectively reduced serum uric acid levels, improved kidney and joint damage, and ameliorated the inflammatory response caused by MSU. Its main component, brucine, effectively improved the inflammatory response and reduced the invasive ability of synoviocytes induced by MSU.

Anti-inflammatory Properties of Tongfeng Li'an Granules in an Acute Gouty Arthritis Rat Model.

To examine the anti-inflammatory properties and underlying mechanisms of Tongfeng Li'an Granules (TFLA), a traditional medicine, in acute gouty arthritis using a rat model. We identified 55 major compounds in TFLA via ultrahigh-performance liquid chromatography connected to quadrupole time-of-flight mass spectrometry (UPLC-TQF-MS/MS). Databases were employed for the prediction of potential targets, followed by PPI network construction as well as GO and KEGG analyses. After network-pharmacology-based analysis, a rat gouty arthritis model was used to validate the anti-inflammatory mechanism of TFLA. UPLC-TQF-MS/MS and network pharmacology analyses revealed 55 active ingredients and 160 targets of TFLA associated with gouty arthritis, forming an ingredient-target-disease network. The PPI network identified 20 core targets, including TLR2, TLR4, IL6, NFκB, etc. Functional enrichment analyses highlighted the Toll-like receptor signaling pathway as significantly enriched by multiple targets, validated in in vivo experiments. Animal experiments demonstrated that TFLA improved pathological changes in gouty joint synovium, with decreased ankle joint circumference, serum IL6, IL10, and TNFα levels, as well as reduced protein and mRNA expression of NLRP3, TLR2, and TLR4 in ankle joint synovial tissue observed in the middle- and high-dose TFLA and positive control groups compared to the model group (p < 0.05). This research elucidated the pharmacological mechanisms of TFLA against gouty arthritis, implicating various ingredients, targets, and signaling pathways. Animal experiments confirmed TFLA's efficacy in alleviating inflammation in acute gouty arthritis by modulating Toll-like receptor signaling and NLRP3 expression.

To Investigate the Mechanism of Qinpi Tongfeng Formula in Treating Acute Gouty Arthritis by UHPLC-Q-Orbitrap-MS, Network Pharmacology and Experimental Validation.

Acute gouty arthritis (AGA) is characterized by the accumulation of monosodium urate crystals within the joints, leading to inflammation and severe pain. Western medicine treatments have limitations in addressing this condition. Previous studies have shown the efficacy of Qinpi Tongfeng formula (QPTFF) in treating AGA, but further investigation is needed to understand its mechanism of action. We used ultra-high-performance liquid chromatography tandem Q-Exactive Orbitrap high-resolution mass spectrometry (UHPLC-Q-Orbitrap-MS) to identify compounds in QPTFF. Target proteins regulated by these compounds were obtained from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform, Chemistry Database, and Swiss Target Prediction Database. AGA-related targets were searched and screened from various databases, including Genecards, PharmGKB, Drugbank, etc. Intersection targets of QPTFF and AGA were analyzed for protein-protein interaction networks, GO function enrichment, and KEGG pathway enrichment. We then verified QPTFF's mechanism of action using an AGA rat model, assessing pathological changes via H&E staining and target expression via ELISA, RT-qPCR, and Western blot. UHPLC-Q-Orbitrap-MS identified 207 compounds in QPTFF, with 55 selected through network pharmacology. Of 589 compound-regulated targets and 1204 AGA-related targets, 183 potential targets were implicated in QPTFF's treatment of AGA. Main target proteins included IL-1β, NFKBIA, IL-6, TNF, CXCL8, and MMP9, with the IL-17 signaling pathway primarily regulated by QPTFF. Experimental results showed that medium and high doses of QPTFF significantly reduced serum inflammatory factors and MMP-9 expression, and inhibited IL-17A, IL-6, IKK-β, and NF-κB p65 mRNA and protein expression in AGA rats compared to the model group. Key targets of QPTFF include IL-1β, NFKBIA, IL-6, TNF-α, CXCL8, and MMP9. QPTFF effectively alleviates joint inflammation in AGA rats, with high doses demonstrating no liver or kidney toxicity. Its anti-inflammatory mechanism in treating AGA involves the IL-17A/NF-κB p65 signaling pathway.

Sanmiao wan alleviates inflammation and exhibits hypouricemic effect in an acute gouty arthritis rat model

Ethnopharmacological relevanceSanmiao wan (SMW), a classical traditional Chinese medicine (TCM) formula, has been employed to treat gouty diseases in clinic as early as Yuan dynasty. It shows remarkably therapeutic effects in acute gouty arthritis (GA). However, the potential mechanisms of SMW are still not fully revealed. Aim of the studyThe objective of this project is to evaluate the pharmacological effects and possible mechanisms of SMW in a rat model of acute GA. Materials and methodsMonosodium urate (MSU) suspension was injected into the ankle joint of rats to establish acute GA model. The inflammation was evaluated by measuring the posterior ankle diameter. The pathological status of synovial tissue was assessed by hematoxylin eosin (HE), Masson, and picrosirius red staining. The level of IL-6 was measured using ELISA kit. The levels of blood urea nitrogen (BUN), creatinine (CR), UA (uric acid), and xanthine oxidase (XOD) in the serum were measured using standard diagnostic kits. The percentage of Th17 cells in blood samples was performed using flow cytometry. Moreover, RT-qPCR was performed to examine the mRNA level of RANK, RORγt, RANKL, and STAT3 in the synovial tissue. Furthermore, immunofluorescence was carried out to assess the expression of STAT3 in the synovial tissue. ResultsSMW effectively alleviated the inflammation and improved the pathological status of the ankle joint in rats with acute GA. It significantly suppressed the release of proinflammatory cytokine (IL-6). Meanwhile, the levels of UA, BUN, and CR were markedly reduced after SMW treatment. A remarkable reduction of XOD activity was observed in the study. Importantly, SMW treatment significantly reduced the frequency of Th17 cells, decreased the mRNA levels of RANK, RORγt, RANKL, and STAT3 in the synovial tissue. Furthermore, the suppression of STAT3 was also demonstrated using immunofluorescence in SMW-treated group. ConclusionSMW showed significant anti-inflammatory and hypouricemic effects in a rat model of GA. It is an effective TCM formula for GA therapy.

A liquid chromatography-tandem mass spectrometry method for comprehensive determination of metabolites in the purine pathway of rat plasma and its application in anti-gout effects of Lycium ruthenicum Murr.

It has been proved that purine metabolites are implicated in various biological syndromes and disorders. Therefore, the realization of panoramic detection of purine metabolites will be of great significance to the pathogenesis of purine metabolic disorders. In the present study, an ultra-high performance liquid chromatography with tandem mass spectrometry method was developed for the comprehensive quantification of purine metabolites in rat plasma. The 17 purine metabolites were separated and quantified in the short running time of 15min. The proposed method was strictly validated by applying SeraSub solution as a matrix and proved to be linear (R2 ≥ 0.9944), accurate (the recoveries of all analytes ranged from 85.3% to 103.0%, with relative standard deviation values ≤ 9.3%), and precise (the intra- and inter-day precisions were less than 10.8% and 12.4%, respectively). The method was then successfully applied to the qualification of the endogenous purine metabolites in acute gouty arthritis rats, as well as colchicine and anthocyanin-intervened rats. Results showed that uric acid, xanthine, hypoxanthine, and xanthine were considered the key factors of acute gouty arthritis. The established method and measurement of purines in rat plasma might help the investigation of the action mechanisms between purine disorders and related diseases.

Effects of evodiamine on ROS/TXNIP/NLRP3 pathway against gouty arthritis.

Evodiamine (EVO) was tested on acute gouty arthritis rats to investigate its anti-inflammatory effect. Seventy-two male Sprague-Dawley (SD) rats were randomly assigned into the control, model, high, medium, and low dose of EVO groups and colchicine group. The ankle swelling degrees were measured at 2h, 6h, and 24h following sodium urate injection into ankle joint. Histopathological examination was performed 24h after injection. Reactive oxygen species (ROS) content in the ankle joint was detected using chemical fluorescence. Serum interleukin-1β (IL-1β), interleukin-18 (IL-18), and tumor necrosis factor-α (TNF-α) content were determined by ELISA. Serum xanthine oxidase (XOD), superoxide dismutase (SOD), and malondialdehyde (MDA) were determined by spectrophotometry. The expressions of thioredoxin-interacting protein (TXNIP), NOD-like receptor thermal protein domain associated protein 3 (NLRP3), pro-caspase-1, caspase-1, and apoptosis-related spot like protein (ASC) in synovium were detected by Western blot. Evodiamine alleviated the ankle swelling of the affected foot in gouty arthritis rats and reduced inflammatory cell infiltration in joint synovial tissue. Evodiamine also decreased the content of serum inflammatory factors including IL-1β, IL-18, and TNF-α, and increased serum SOD activity, while it decreased serum XOD, MDA activity, and ROS level. Moreover, evodiamine downregulated the protein expression levels of TXNIP, NLRP3, pro-caspase-1, cleavedcaspae-1, and ASC. The mechanism of EVO in treating gouty arthritis is associated with the inhibition of NLRP3 inflammasome by regulating the ROS/TXNIP/NLRP3 signaling pathway.

Cinnamomi cortex extract mitigated monosodium urate-induced acute gouty arthritis in rats through nuclear factor-κB-NOD-like receptor thermal protein domain associated protein 3 signaling pathway.

Cinnamomi cortex was applied to mitigate joint injury since ancient China. However, the effect of Cinnamomi cortex on gouty arthritis (GA) was rarely reported. This study aimed to explore the effect of Cinnamomi cortex on monosodium urate (MSU)-induced acute GA (AGA) in rats, and clarify the underlying mechanism. The results showed that Cinnamomi cortex extract (CE) containing rich polyphenols and flavonoids alleviated joint swelling and inflammation by reducing programmed cell death in MSU-induced AGA rats. Network pharmacology analysis showed that CE's predictive inflammatory pathways included nuclear factor-κB (NF-κB) and necroptosis pathways. CE reduced expression of pyroptosis-related regulators including Gasdermin D and Caspase 1 via regulating NF-κB/NOD-like receptor thermal protein domain associated protein 3 signaling pathway in AGA rats. In conclusion, this study provided a theoretical basis for Cinnamomi cortex applied as a new veterinary medicine to protect against GA.

Tanshinone IIA Improves Acute Gouty Arthritis in Rats through Regulating Neutrophil Activation and the NLRP3 Inflammasome.

To investigate the prevention and treatment effect of tanshinone IIA (TIIA) on acute gouty arthritis (AGA) and its mechanism. The anti-AGA effect of TIIA was observed in vivo and in vitro. Neutrophils were isolated from the abdominal cavity of mice, and the anti-AGA effect of TIIA was investigated in a rat model of MSU-induced AGA. The pathological changes of the ankle joint tissues were assessed by H&E. Cytokine and chemokine expression were determined by ELISA and RT-qPCR. The NLRP3 inflammasome pathway protein levels in the ankle joint tissues were evaluated via western blotting. Neutrophil migration was evaluated in air pouch and transwell assays. Immunohistochemistry and immunofluorescence analysis evaluate the release of myeloperoxidase (MPO), neutrophil elastase (NE), and citrullination of histone H3 (CitH3). Beclin-1 and LC3B expressions were determined using western blotting and immunofluorescence. Key Findings. Treatment with TIIA alleviated synovial hyperplasia and neutrophil infiltration, regulated cytokine and chemokine expressions, and inhibited NLRP3 activation in AGA rats, neutrophil migration, MPO, NE, and CitH3 expression, and LC3B and Beclin-1 protein expression. These results demonstrate that TIIA can effectively enhance AGA by focusing on the neutrophils and NLRP3 inflammasome, demonstrating that TIIA may act as a potential helpful agent for AGA.

Modified Baihu decoction therapeutically remodels gut microbiota to inhibit acute gouty arthritis.

Background: Acute gouty arthritis (AGA) is the most common first symptom of gout, and the development of gout as a metabolic and immune inflammatory disease is also correlated with the gut microbiota. However, the mechanism of the effect of changes in the gut microbiota on AGA remains unclear. The intestinal flora can not only affect purine metabolism or regulate inflammation, but also influence the therapeutic effect of drugs on AGA. The aim of this study was to investigate the exact mechanism of modified Baihu decoction (MBD) in the treatment of AGA and whether it is related to the regulation of the structure of the intestinal flora. Methods: On the 21st day of MBD administration by continuous gavage, a rat acute gouty arthritis model was constructed using sodium urate (0.1mL/rat, 50mg/mL), and the ankle joint swelling was measured before and 4h, 8h, 24h, and 48h after the injection of sodium urate. After 48h of sodium urate injection, serum, liver, kidney, ankle synovial tissue and feces were collected from rats. The collected samples were examined and analyzed using H&E, Elisa, Immunohistochemistry, Histopathology, 16S rDNA, and Biochemical analysis. To investigate the mechanism of MBD to alleviate AGA using pro-inflammatory factors and intestinal flora. Results: MBD (5.84, 35g/kg) was administered orally to AGA rats and diclofenac sodium tablets (DS-tablets) were used as standard treatment control. Serum biochemical assessment confirmed that MBD is a safe drug for the treatment of AGA. In addition, our findings confirmed that MBD relieved AGA-related symptoms, such as toe swelling. Lowering serum levels of uric acid, IL-1β, and TGF-β1 immunohistochemical results also confirmed that MBD reduced the expression of inflammatory elements such as IL-1β, NLRP3, ASC, and Caspase-1 in synovial tissue.Furthermore, compared with control group, the 16s rDNA sequencing of AGA rat faeces revealed an increase in the relative abundance of Lachnospiraceae, Muribaculaceae, and Bifidobacteriaceae species. While the relative abundance of Lactobacillaceae, Erysipelotrichaceae, Ruminococcaceae, Prevotellaceae and Enterobacteriaceae showed a relative decrease in species abundance. Of these, the reduction in species abundance of Enterobacteriaceae was associated with a reduction in amino acid metabolism and environmental perception. After MBD therapeutic intervention, the disturbance of the intestinal flora caused by AGA was restored. Conclusion: In summary, MBD is an effective agent for the treatment of AGA, with the potential mechanism being the regulation of intestinal flora to control inflammation. This would help to promote the therapeutic effect of MBD on AGA.

Revealing the pharmacological effect and mechanism of darutoside on gouty arthritis by liquid chromatography/mass spectrometry and metabolomics.

Darutoside is a diterpenoids compound with significant anti-inflammatory activity, however the pharmacological action and mechanism are still unclear. Metabolomics strategy was used to uncovering the pharmacological action and effective mechanism of darutoside against acute gouty arthritis rats. Liquid chromatography coupled with mass spectrometry technique was performed to explore the serum metabolites and potential pathways. We found that darutoside can up-regulate the level of glutamate, alanine, chenodeoxycholic acid, 1-methyladenosine, aspartic acid, citric acid, and down-regulate the level of valine, isoleucine, glutamine, alanyl-threonine, pyruvic acid, gamma-aminobutyric acid, uric acid. Metabolic pathway analysis showed that the therapeutic effect of darutoside was involved in amino acid metabolism, sugar metabolism, fatty acid metabolism, energy metabolism, purine metabolism and butanoate metabolism. It indicated that darutoside protect against acute gouty arthritis by regulating the expression of the key protein targets. It revealed that the mechanism of darutoside on acute gouty arthritis, which may be leading to the changes of serum metabolites, metabolic pathways and key protein targets to improve immune system response, inhibit oxidative stress and inflammatory response. It provides a novel method for molecular mechanisms of natural product in the disease treatment.

Anti-Gout Effects of the Medicinal Fungus Phellinus igniarius in Hyperuricaemia and Acute Gouty Arthritis Rat Models.

Background: Phellinus igniarius (P. igniarius) is an important medicinal and edible fungus in China and other Southeast Asian countries and has diverse biological activities. This study was performed to comparatively investigate the therapeutic effects of wild and cultivated P. igniarius on hyperuricaemia and gouty arthritis in rat models. Methods: UPLC-ESI-qTOF-MS was used to identify the chemical constituents of polyphenols from wild P. igniarius (WPP) and cultivated P. igniarius (CPP). Furthermore, WPP and CPP were evaluated in an improved hyperuricaemia rat model induced by yeast extract, adenine and potassium oxonate, which was used to examine xanthine oxidase (XO) activity inhibition and anti-hyperuricemia activity. WPP and CPP therapies for acute gouty arthritis were also investigated in a monosodium urate (MSU)-induced ankle swelling model. UHPLC-QE-MS was used to explore the underlying metabolic mechanisms of P. igniarius in the treatment of gout. Results: The main active components of WPP and CPP included protocatechuic aldehyde, hispidin, davallialactone, phelligridimer A, hypholomine B and inoscavin A as identified by UPLC-ESI-qTOF-MS. Wild P. igniarius and cultivated P. igniarius showed similar activities in reducing uric acid levels through inhibiting XO activity and down-regulating the levels of UA, Cr and UN, and they had anti-inflammatory activities through down-regulating the secretions of ICAM-1, IL-1β and IL-6 in the hyperuricaemia rat model. The pathological progression of kidney damage was also reversed. The polyphenols from wild and cultivated P. igniarius also showed significant anti-inflammatory activity by suppressing the expression of ICAM-1, IL-1β and IL-6 and by reducing the ankle joint swelling degree in an MSU-induced acute gouty arthritis rat model. The results of metabolic pathway enrichment indicated that the anti-hyperuricemia effect of WPP was mainly related to the metabolic pathways of valine, leucine and isoleucine biosynthesis and histidine metabolism. Additionally, the anti-hyperuricemia effect of CPP was mainly related to nicotinate and nicotinamide metabolism and beta-alanine metabolism. Conclusions: Wild P. igniarius and cultivated P. igniarius both significantly affected the treatment of hyperuricaemia and acute gouty arthritis models in vivo and therefore may be used as potential active agents for the treatment of hyperuricaemia and acute gouty arthritis.

Studies on the Anti-Gouty Arthritis and Anti-hyperuricemia Properties of Astilbin in Animal Models

AbstractThe purpose of this study was to investigate potential anti-gouty effect of astilbin (AS) and its possible mechanisms. In mice with hyperuricemia induced by potassium oxonate (OXO) and yeast extract powder (YEP), AS and febuxostat (FB) reduced the serum uric acid (UA) and xanthine oxidase (XO). Moreover, AS and FB reduced the levels of reactive oxygen species and increased the content of superoxide dismutase (SOD), glutathione peroxidase and catalase present in the serum. In acute gouty arthritis rats induced by intraarticular monosodium urate crystal injection, AS and Colchicine (COL) alleviated the ankle joints swelling, and reduced the inflammatory cell infiltration. AS also reduced the levels of interleukin 1β, interleukin 6, tumor necrosis factor alpha and monocyte chemoattractant protein 1 in liver. The present study first confirmed the anti-gouty effect of AS in mice with hyperuricemia and rats with acute gouty arthritis, which provides the experimental evidence for further evaluation of AS as a candidate for gout treatment.

Anti-inflammatory effects of luteolin on acute gouty arthritis rats via TLR/MyD88/NF-κB pathway.

To investigate the anti-inflammatory effect of luteolin on the acute gouty arthritis (AGA) rats and the underlying mechanisms. A total of sixty rats were chosen and randomly divided into 5 groups:A control group, a monosodium urate (MSU) group, a colchicine group, 2 luteolin groups (50 mg/kg, 150 mg/kg). The AGA model of rats was established by injecting monosodium urate (MSU) at the concentration of 25 mg/mL into the ankle joint cavity. Changes of joint swelling index at different time points and the levels of interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in serum and synovial were measured. The mRNA expression of TLR2, TLR4, and MyD88 in synovial tissue was detected by real-time PCR, and the protein expression of TLR2, TLR4, MyD88, and NF-κB in synovial tissues was determined by Western blotting. The inflammatory cells of the ankle joint and its surrounding soft tissues were observed after HE staining, and the expression of NF-κB was determined by immunohistochemistry. Compared with the MSU group, the joint swelling indexes of the luteolin group and the colchicine group were significantly decreased (P<0.05), and the levels of IL-1β, IL-6 and TNF-α were also significantly decreased (P<0.01). The mRNA levels of TLR2, TLR4, and MyD88 and the protein levels of TLR2, TLR4, MyD88, and NF-κB were significantly decreased (P<0.01). Luteolin can reduce the inflammatory response of acute gouty arthritis via down-regulating the TLR/MyD88/NF-κB pathway, and it is expected to be an effective drug for the treatment of acute gouty arthritis.

Investigation of the Effects and Mechanisms of Dendrobium loddigesii Rolfe Extract on the Treatment of Gout.

Objective Gout is a chronic disease that causes inflammatory arthritis, which is closely related to urate accumulation induced by a disorder of uric acid metabolism and the consequent deposition of monosodium urate crystals. Dendrobium loddigesii Rolfe is an herbal medicine that has been used in some traditional Chinese medicine formulae in the treatment of gout. This study aimed to explore and verify the antigout activity of Dendrobium loddigesii extract (DLE) on alleviating the hyperuricaemia of mice and the acute gouty arthritis of rats. Methods An animal model of hyperuricaemia was established using potassium oxonate (PO). We analysed the expression of uric acid transporter mRNA in the kidney in the hyperuricaemic mice after treatment with DLE. Simultaneously, a monosodium urate crystal-induced acute gouty arthritis rat model was used to evaluate the effects of DLE, according to the level of ankle swelling, as well as the protein levels of inflammatory receptors and cytokines, as assayed by WB and ELISA. Results DLE alleviated hyperuricaemia in mice and inhibited acute gouty arthritis in rats (P < 0.05). Meanwhile, DLE regulated the levels of uric acid transporters mRNA transcripts, including mouse organic anion transporter 1 (mOAT1), organic anion transporter 3 (mOAT3), urate transporter 1 (mURAT1), and glucose transporter 9 (mGLUT9) in the kidney (P < 0.05), suggesting that DLE promoted uric acid metabolism. Furthermore, DLE significantly suppressed the protein levels of TLRs, MyD88, and NF-κB in the ankle joint's synovium (P < 0.05), and the serum levels of IL-1β, IL-6, and TNF-α were also reduced, which demonstrated the anti-inflammatory effects of DLE. Conclusion DLE alleviates hyperuricaemia by regulating the transcription level of uric acid transporters in the kidney. It also inhibits acute gouty arthritis by inhibiting the pathway of TLRs/MyD88/NF-κB in the ankle joint's synovium. The findings of the present study imply that DLE alleviates gout by promoting uric acid metabolism and inhibiting inflammation related to the TLRs/MyD88/NF-κB pathway.

MiR-146a alleviates inflammation of acute gouty arthritis rats through TLR4/MyD88 signal transduction pathway.

The aim of this study was to explore the effect of micro-ribonucleic acid (miR)-146a on acute gouty arthritis rats through Toll-like receptor-4/myeloid differentiation factor 88 (TLR4/MyD88) signal transduction pathway. A total of 30 clean-grade Sprague-Dawley rats were divided into three groups, including agomiR-146a group (n=10), antagomiR-146a group (n=10) and negative control group (NC, n=10). The model was successfully established via a one-time injection of sodium urate into ankle joint cavity. Subsequently, agomiR-146a (10 μL), antagomiR-146a (10 μL) and normal saline (10 μL) were intrathecally injected into rats in the three groups at 1 h before injection and 12 h, 24 h, 48 h and 72 h after injection, respectively. The ankle joint swelling index, joint dysfunction index and joint inflammation index of rats in the three groups were closely monitored. After 72 h of observation, the rats were euthanized, and synovial tissues were collected from the knee joint. The expression and distribution of nuclear factor-κB (NF-κB) in synovial tissues were detected using the immunohistochemical method. Meanwhile, the expression levels of inflammatory factors, including tumor necrosis factor-α (TNF-α), interleukin-1 (IL-1) and interleukin-6 (IL-6) were detected via enzyme-linked immunosorbent assay. Furthermore, the mRNA and protein expression levels of TLR4 and MyD88 were detected via quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) and Western blotting, respectively. No statistically significant differences in the joint swelling index, joint dysfunction index, joint inflammation index, TLR4 and MyD88 and related inflammatory factors were found between the NC group and antagomiR-146a group. Compared with the NC group, agomiR-146a group showed markedly reduced ankle joint swelling index (p<0.05). Meanwhile, joint landing behavior and inflammatory swelling were significantly relieved in the agomiR-146a group (p<0.05). The mRNA and protein expression levels of TLR4 and MyD88 were remarkably decreased as well (p<0.05). Furthermore, the expression and distribution of NF-κB in synovial tissues of agomiR-146a group was markedly reduced when compared with the NC group (p<0.05). In addition, agomiR-146a group exhibited significantly lower expression levels of inflammatory factors (TNF-α, IL-1 and IL-6) in synovial tissues (p<0.05). MiR-146a alleviates joint inflammation of acute arthritis in rats through the TLR4/MyD88/NF-κB signaling pathway, which may become a new therapeutic target.

Effect of Electroacupuncture on Synovial M 1/M 2 Macrophage Polarization in Rats with Acute Gouty Arthritis

To explore the effect of electroacupuncture (EA) on the expression of synovial AMP-activated protein kinase (AMPK) protein α， arginase-1 mRNA, nitric oxide synthase 2 (NOS 2) mRNA, NOD-like receptor protein 3 (NLRP 3) mRNA, and interleukin-1 β (IL-1 β) mRNA in acute gouty arthritis (AGA) rats, so as to explore its mechanisms underlying improvement of AGA via M 1/M 2 macrophage polarization. Male Wistar rats were randomly divided into normal control, model, medication (colchicine) and EA groups (n＝15 rats in each group). The AGA model was established by injection of sodium urate crystal (MSU) suspension (0.2 mL) into the articular cavity of the left knee. The rats of the normal control group received articular injection of normal saline (0.2 mL) of the left knee, and those of the medication group were treated by gavage of the colchicine (0.3 mg•kg－1•d－1) once daily for 7 days. EA (2 Hz/10 Hz, 1.0 mA) was applied to "Zusanli"(ST 36) and "Sanyinjiao" (SP 6) of the left hind limb for 10 min, once daily for 7 days. The inflammatory conditions of the synovial membrane tissue of the left knee joint were observed by H．E. staining. The expression levels of phosphorylated AMPKα (p-AMPKα) protein, and arginase-1 (a maker of M 2 macrophages) mRNA, NOS 2 (a maker of M 1 macrophages) mRNA, NLRP 3 mRNA, and IL-1 β mRNA in the knee joint synovial tissue were detected by Western blot and quantitative real-time PCR, respectively. Compared with the normal group, the inflammatory cell infiltration of the synovial tissue was more severe, the expression of p-AMPKα protein was significantly decreased (P<0.01), and the expression levels of arginase-1, NOS 2, IL-1 β and NLRP 3 mRNAs were considerably increased in the model group (P<0.01). The expression levels of p-AMPKα protein and arginase-1 mRNA were significantly up-regulated, and those of NOS 2, IL-1 β and NLRP 3 mRNAs obviously down-regulated in both EA and medication groups relevant to the model group (P<0.01, P<0.05), suggesting an increase of M 2 macrophage and a decrease of M 1 macrophage activity after EA. No significant differences were found between the EA and medication groups in up-regulating p-AMPKα expression and in down-regulating NOS 2, IL-1 β and NLRP 3 mRNA expression (P>0.05), except higher up-regulation of arginase-1 mRNA in the medication group (P<0.05)．. EA intervention can up-regulate the expression of arginase-1 mRNA and p-AMPKα protein, and down-regulate the expression of NOS 2, IL-1 β and NLRP 3 mRNAs in synovial tissues in AGA rats, which may contribute to its anti-inflammatory effect by promoting conversion of macrophages from M 1 pro-inflammatory phenotype to M 2 anti-inflammatory phenotype.

Changes of Treg/Th17 Ratio in Spleen of Acute Gouty Arthritis Rat Induced by MSU Crystals.

Acute gouty arthritis is the inflammation of joint tissues in the acute form due to the deposition of monosodium urate (MSU) crystals. Regulatory T cells (Tregs) and Th17 cells play an important role in the development and progression of inflammatory diseases. However, the expression and role of Tregs and Th17 cells are not clear in this disease. Here, we investigated the changes of Tregs, Th17 cells, and Treg/Th17 ratio in spleen, as well as the inflammatory cytokines in blood and joint tissue pathology in acute gouty arthritis rat induced by MSU. We found that both the percentages of Tregs and Th17 cells in spleen increased at an early stage (6h). Tregs decreased at 12 and 24h, and rise again at 48 and 72h. However, Th17 cells reached its peak at 24h, and then decreased after 48h. Treg/Th17 ratio showed an initial decrease and then increase, and further reached its minimum value at 24h. But the ratios of Treg/Th17at all times were lower than that of normal control. The level of serum cytokines (IL-1β, IL-6, IL-17, TNF-α, and IL-10) showed an opposite trend to Treg/Th17 ratio, except the level of TGF-β1 was similar to Tregs. In summary, Tregs and Th17 cells in spleen changed over time during the development of acute gouty arthritis. Decrease of Treg/Th17 ratio was consistent with inflammation development in the joints, suggesting that Treg/Th17 imbalance may involve in pathogenesis of acute gouty arthritis.

Achyranthis bidentatae radix enhanced articular distribution and anti-inflammatory effect of berberine in Sanmiao Wan using an acute gouty arthritis rat model

Ethnopharmacological relevanceSanmiao Wan (SMW) has been a basic prescription employed for the treatment for gout in the clinic since Yuan dynasty. Achyranthis bidentatae radix (ABR) is designed as a lower-guiding drug in SMW to augment the articular accumulation of active ingredients and improve the anti­inflammatory effect. Aim of the studyPresent study was undertaken to investigate the dose-response relationship of berberine in SMW between the articular concentration and anti-inflammatory effect in the knee joint under the lower-guiding of ABR. Materials and methodsRats were divided into control group, model group and SMW without or with low, medium and high doses of ABR groups. Rat model of acute gouty arthritis (AGA) was established by intra-articular injection of 0.2 mL monosodium urate crystal (20 mg/mL) inside knee joint cavity on day 2 during drug treatment slots. Knee joint swelling, synovial hyperplasia and inflammatory cell infiltration were investigated for anti-inflammatory study. The concentrations of berberine in rat plasma and tissues were determined by UPLC–MS/MS method. The effect of ABR on the expression levels of P-glycoprotein (P-gp) and MDR1 mRNA in the synovial tissues of knee joints in AGA rats was examined by Western blot and RT-qPCR assay, respectively. ResultsThe distribution of berberine increased by 6.53%, 44.31% and 212.96% in the knee joint and 474.93%, 631.01% and 1063.3% in the ankle for SMW with low, medium and high doses of ABR groups, compared with SMW without ABR group. Similarly, the plasma level of berberine increased by 19.81%, 143.4% and 681.13%. On the contrary, the distribution of berberine evidently decreased 3.23, 10.61 and 46.21-fold in heart and 3.68, 6.74 and 24.78-fold in lung. SMW with different doses of ABR groups exhibited better efficiency than SMW without ABR group on ameliorating knee joint swelling, inhibiting synovial hyperplasia and alleviating inflammatory cell infiltration of AGA rats. The treatment with ABR could down-regulate the MDR1 mRNA and P-gp expressions of synovial tissues of knee joints in AGA rats. ConclusionsThe enhanced articular distribution of berberine in SMW was attributed to the lower-guiding effect of ABR, which could evidently increase the plasma concentration of berberine, improve the supply of blood of inflamed joint, reduce the distribution of berberine in heart and lung and significantly inhibit the MDR1 mRNA and P-gp expression of synovial tissues of knee joints in AGA rats. The dose-response relationship of berberine between the enhanced articular concentration and improved anti-inflammatory effect in the knee joint under the lower-guiding of ABR was observed for the first time.

Liver metabonomics of acute gouty arthritis treated by Diosocorea nipponica

To explore the prevention and protection effect of Diosocorea nipponica (DNM)) on acute gouty arthritis (AGA) rats based on liver metabonomics, and find potential biomarkers and related pathways. AGA model rats were induced by monosodium urate crystal suspension. UPLC-TOF-MS coupled with pattern recognition technique was employed to find out the potential biomarkers and related metabolic pathways. Eleven common potential biomarkers were identified. Among the potential intervention targets in normal rats given by DNM, 4 biomarkers were up-regulated, and the other 4 targets were down regulated. Among the potential intervention targets in AGA rats given by DNM, 5 metabolites were up-regulated by MSU and 5 metabolites were down regulated. The abnormal expression levels of adenosine monophosphate, 5-methyltetrahydrofolic acid, oxidized glutathione, hypoxanthine, docosahexaenoic acid, glutathione, uridine diphosphate glucose and inosine could be corrected by DNM extract. Three pathways were founded with greatest correlation, including purine metabolism, starch and sucrose metabolism and glutathione metabolism. Therefore, it could be inferred that D. nipponica has the effect for anti-acute gouty arthritis by intervening endogenous metabolites from the liver under physiological condition and acute gouty arthritis condition.

Hypothermia induced by adenosine 5′-monophosphate attenuates early stage injury in an acute gouty arthritis rat model

To investigate whether the hypothermia induced by Adenosine 5'-Monophosphate (5'-AMP) could attenuate early stage injury in a rat acute gouty arthritis model. Ankle joint injection with monosodium urate monohydrate crystals (MSU crystals) in hypothermia rat model which was induced by 5'-AMP and then observe whether hypothermia induced by 5'-AMP could be effectively inhibit the inflammation on acute gouty arthritis in rats. AMP-induced hypothermia has protective effects on our acute gouty arthritis, which was demonstrated by the following criteria: (1) a significant reduction in the ankle swelling (p < 0.001); (2) a significant decrease in the occurrence of leukocyte infiltration and mild hemorrhage; (3) a significant reduction in the presence of serum Interleukin-1β (IL-1β, p < 0.001) and metalloproteinase-9 (MMP-9, p < 0.001); and (4) a significant inhibition in the Nuclear Factor -κappaB (NF-κB) activity (p < 0.001). AMP-induced hypothermia could inhibit acute inflammation reaction and protect the synovial tissue against acute injury in a rat acute gouty arthritis model.