OBJECTIVE: To evaluate the indicators of an animal model of gouty arthritis(GA) with dampness heat accumulation and the intervention effect of Tongfeng Qingxiao formula(痛风清消方, TFQXF). METHODS: Seventy-two healthy...OBJECTIVE: To evaluate the indicators of an animal model of gouty arthritis(GA) with dampness heat accumulation and the intervention effect of Tongfeng Qingxiao formula(痛风清消方, TFQXF). METHODS: Seventy-two healthy adult Sprague-Dawley male rats were selected and randomly divided into a normal group, model group, low-dose group, mediumdose group, high-dose group, and diclofenac group using a random number table method, with 12 rats in each group. After group intervention, the general condition of the rats in each group was monitored and recorded, and the swelling index was measured. After separating the serum, the changes in glutamic pyruvic transaminase (ALT), glutamic oxaloacetic transaminase(AST), carbamide(UREA), creatinine(CREA), triglyceride(TG), total serum cholesterol(TC), low density lipoprotein cholesterol(LDL-C), and high density lipoprotein cholesterol(HDL-C) in the serum of the rats in each group were measured using an automatic biochemical analyzer. The levels of motilin(MTL), gastrin(GAS), endothelin(ET), calcitonin gene-related peptide(CGRP), heat shock protein 70(HSP70), interleukin(IL)-1β, and nuclear factor kappa-B(NF-κB) in the serum of the rats in each group were evaluated using enzyme-linked immunosorbent assay(ELISA) kits. Kidney tissues were used to evaluate the protein and m RNA expression of aquaporin(AQP) 1 and AQP2. Colon tissue was used to evaluate the protein and m RNA expression of AQP3 and AQP4 by Western blotting(WB) assay and real-time quantitative polymerase chain reaction(RT-q PCR). The levels of ALT, AST, UREA, and CREA were used to evaluate the liver and kidney function of rats. The levels of MTL and GAS were used to evaluate the gastrointestinal function of rats. The levels of TG, TC, LDL-C, HDL-C, AQP1, AQP2, AQP3, and AQP4 were used to evaluate the "dampness" syndrome performance in rats. The levels of ET, CGRP, and HSP70 were used to evaluate the "heat" syndrome performance in rats. The levels of IL-1β and NF-κB were used to evaluate the degree of inflammation in rats. The pathological changes in synovial and colonic tissues were observed by hematoxylin and eosin staining. RESULTS: Except for the normal group, after modeling treatment, the ankle joint of rats in both the model group and drug treatment groups gradually swelled, reaching a peak at 12 h, and then gradually began to subside. The results of biochemical analyzer detection indicated that the serum ALT, AST, UREA, CREA, TG, TC and LDL-C levels were significantly higher, but the HDL-C level was significantly lower in the rats of the model group than in the rats of the normal group(P < 0.05). The serum ALT, AST, UREA, CREA, MTL, TC and LDL-C levels were significantly lower, but the HDL-C level was significantly higher in the rats of all drug treatment groups than in the rats of the model group(P < 0.05). The results of ELISA detection indicated that the MTL, GAS, ET, HSP70, IL-1β,and NF-κB levels were significantly higher, but the CGRP level was significantly lower in the rats of the model group than in the rats of the normal group(P < 0.05). The levels of MTL, GAS, ET, HSP70, IL-1β, and NF-κB were significantly lower, but CGRP was significantly higher in the rats of the drug treatment groups than in the rats of the model group(P < 0.05). The results of WB and RTq PCR indicated that compared to the normal group, the levels of AQP1 and AQP2 in the model group were significantly higher in the kidney tissue, whereas the levels of AQP3 and AQP4 were significantly lower in the colon tissue(P < 0.05). Compared to those in the model group, the levels of AQP1 and AQP2 in the drug treatment groups were significantly lower in the kidney tissue, whereas the levels of AQP3 and AQP4 were significantly higher in the colon tissue(P < 0.05). In the model group, erosion of the colonic mucosal surface and inflammatory exudate occurred. Some mucosal epithelium had fallen off, the number of glands in the lamina propria was lower, many inflammatory cells infiltrated the interstitial layer, the connective tissue in the submucosa became loose and edematous, and lymph follicles developed. We found a significant proliferation of synovial cells in the ankle joint, an increase in cell density and neovascularization, and visible infiltration of inflammatory cells. The cartilage surface was not smooth. However, each drug group could improve the pathological changes in intestinal and synovial tissues to varying degrees. CONCLUSIONS: Blood lipid metabolism indexes and AQPs could be used as objective evaluation indexes for the "dampness" syndrome performance of damp-heat accumulation type GA. ET, HSP70 and CGRP could be used as objective evaluation indexes for the "heat" syndrome performance, and the immune inflammation index could be used as objective evaluation indexes for the inflammation degree. The overall efficacy of TFQXF in the treatment of damp-heat accumulation-type GA could be determined by adjusting the above objective evaluation indexes. It provided some ideas and directions for clinical risk assessments and drug development of GA.展开更多
基金the National Natural Science Foundation of China:a Study on the Use of Tongfengqingxiao for Treating Gouty Arthritis and Neutrophil Extracellular Traps Formation based on the Transforming Growth Factor-β1 Signaling Pathway (No. 81860857)a Study on the Mechanism by which Tongfeng Qingxiao Regulates the Toll-like receptors-myeloid Differentiation Factor 88-nuclear Factor Kappa-B Signaling Pathway in Gouty Arthritis Rats with Gut Dysbiosis and Dampness-Heat Syndrome based on the Gut-enteric-microbiota Axis (No. 82060871)National Natural Science Foundation of Jiangxi Province:a Study on the Regulatory Mechanism of the Toll-like receptors-Myeloid differentiation factor 88-nuclear factor kappa-B Signaling Pathway Related to Dysbacteriosis of the Intestinal Flora with Gouty Arthritis in Rats Mediated by Tongfengqingxiao,based on the “Gut-Microbiota” Axis (No. 20202BAB206071)。
文摘OBJECTIVE: To evaluate the indicators of an animal model of gouty arthritis(GA) with dampness heat accumulation and the intervention effect of Tongfeng Qingxiao formula(痛风清消方, TFQXF). METHODS: Seventy-two healthy adult Sprague-Dawley male rats were selected and randomly divided into a normal group, model group, low-dose group, mediumdose group, high-dose group, and diclofenac group using a random number table method, with 12 rats in each group. After group intervention, the general condition of the rats in each group was monitored and recorded, and the swelling index was measured. After separating the serum, the changes in glutamic pyruvic transaminase (ALT), glutamic oxaloacetic transaminase(AST), carbamide(UREA), creatinine(CREA), triglyceride(TG), total serum cholesterol(TC), low density lipoprotein cholesterol(LDL-C), and high density lipoprotein cholesterol(HDL-C) in the serum of the rats in each group were measured using an automatic biochemical analyzer. The levels of motilin(MTL), gastrin(GAS), endothelin(ET), calcitonin gene-related peptide(CGRP), heat shock protein 70(HSP70), interleukin(IL)-1β, and nuclear factor kappa-B(NF-κB) in the serum of the rats in each group were evaluated using enzyme-linked immunosorbent assay(ELISA) kits. Kidney tissues were used to evaluate the protein and m RNA expression of aquaporin(AQP) 1 and AQP2. Colon tissue was used to evaluate the protein and m RNA expression of AQP3 and AQP4 by Western blotting(WB) assay and real-time quantitative polymerase chain reaction(RT-q PCR). The levels of ALT, AST, UREA, and CREA were used to evaluate the liver and kidney function of rats. The levels of MTL and GAS were used to evaluate the gastrointestinal function of rats. The levels of TG, TC, LDL-C, HDL-C, AQP1, AQP2, AQP3, and AQP4 were used to evaluate the "dampness" syndrome performance in rats. The levels of ET, CGRP, and HSP70 were used to evaluate the "heat" syndrome performance in rats. The levels of IL-1β and NF-κB were used to evaluate the degree of inflammation in rats. The pathological changes in synovial and colonic tissues were observed by hematoxylin and eosin staining. RESULTS: Except for the normal group, after modeling treatment, the ankle joint of rats in both the model group and drug treatment groups gradually swelled, reaching a peak at 12 h, and then gradually began to subside. The results of biochemical analyzer detection indicated that the serum ALT, AST, UREA, CREA, TG, TC and LDL-C levels were significantly higher, but the HDL-C level was significantly lower in the rats of the model group than in the rats of the normal group(P < 0.05). The serum ALT, AST, UREA, CREA, MTL, TC and LDL-C levels were significantly lower, but the HDL-C level was significantly higher in the rats of all drug treatment groups than in the rats of the model group(P < 0.05). The results of ELISA detection indicated that the MTL, GAS, ET, HSP70, IL-1β,and NF-κB levels were significantly higher, but the CGRP level was significantly lower in the rats of the model group than in the rats of the normal group(P < 0.05). The levels of MTL, GAS, ET, HSP70, IL-1β, and NF-κB were significantly lower, but CGRP was significantly higher in the rats of the drug treatment groups than in the rats of the model group(P < 0.05). The results of WB and RTq PCR indicated that compared to the normal group, the levels of AQP1 and AQP2 in the model group were significantly higher in the kidney tissue, whereas the levels of AQP3 and AQP4 were significantly lower in the colon tissue(P < 0.05). Compared to those in the model group, the levels of AQP1 and AQP2 in the drug treatment groups were significantly lower in the kidney tissue, whereas the levels of AQP3 and AQP4 were significantly higher in the colon tissue(P < 0.05). In the model group, erosion of the colonic mucosal surface and inflammatory exudate occurred. Some mucosal epithelium had fallen off, the number of glands in the lamina propria was lower, many inflammatory cells infiltrated the interstitial layer, the connective tissue in the submucosa became loose and edematous, and lymph follicles developed. We found a significant proliferation of synovial cells in the ankle joint, an increase in cell density and neovascularization, and visible infiltration of inflammatory cells. The cartilage surface was not smooth. However, each drug group could improve the pathological changes in intestinal and synovial tissues to varying degrees. CONCLUSIONS: Blood lipid metabolism indexes and AQPs could be used as objective evaluation indexes for the "dampness" syndrome performance of damp-heat accumulation type GA. ET, HSP70 and CGRP could be used as objective evaluation indexes for the "heat" syndrome performance, and the immune inflammation index could be used as objective evaluation indexes for the inflammation degree. The overall efficacy of TFQXF in the treatment of damp-heat accumulation-type GA could be determined by adjusting the above objective evaluation indexes. It provided some ideas and directions for clinical risk assessments and drug development of GA.
