Objective: To observe the effects of Chinese medicine(CM) Polygonum cuspidatum(PC) on adenosine 5'-monophosphate-activated protein kinase(AMPK), forkhead box O3α(FOXO3α), Toll-like receptor-4(TLR4), NACHT, LRR a...Objective: To observe the effects of Chinese medicine(CM) Polygonum cuspidatum(PC) on adenosine 5'-monophosphate-activated protein kinase(AMPK), forkhead box O3α(FOXO3α), Toll-like receptor-4(TLR4), NACHT, LRR and PYD domains-containing protein 3(NLRP3), and monocyte chemoattractant protein-1(MCP-1) expression in a rat model of uric acid-induced renal damage and to determine the molecular mechanism. Methods: A rat model of uric acid-induced renal damage was established, and rats were randomly divided into a model group, a positive drug group, and high-, medium-, and low-dose PC groups(n=12 per group). A normal group(n=6) was used as the control. Rats in the normal and model groups were administered distilled water(10 m L·kg^(–1)) by intragastric infusion. Rats in the positive drug group and the high-, medium-, and low-dose PC groups were administered allopurinol(23.33 mg·kg^(–1)), and 7.46, 3.73, or 1.87 g·kg^(–1)·d^(–1) PC by intragastric infusion, respectively for 6 to 8 weeks. After the intervention, reverse transcription polymerase chain reaction, Western blot, enzyme linked immunosorbent assay, and immunohistochemistry were used to detect AMPK, FOXO3α, TLR4, NLRP3, and MCP-1 m RNA and protein levels in renal tissue or serum. Results: Compared with the normal group, the m RNA transcription levels of AMPK and FOXO3α in the model group were significantly down-regulated, and protein levels of AMPKα1, pAMPKα1 and FOXO3α were significantly down-regulated at the 6 th and 8 th weeks(P<0.01 or P<0.05). The m RNA transcription and protein levels of TLR4, NLRP3 and MCP-1 were significantly up-regulated(P<0.01 or P<0.05). Compared with the model group, at the 6 th week, the mRNA transcription levels of AMPK in the high-and medium-dose groups, and protein expression levels of AMPKα1, p AMPKα1 and FOXO3α in the high-dose PC group, AMPKα1 and p AMPKα1 in the mediumdose PC group, and p AMPKα1 in the low-dose PC group were significantly up-regulated(P<0.01 or P<0.05); the m RNA transcription and protein levels of TLR4 and NLRP3 in the 3 CM groups, and protein expression levels of MCP-1 in the medium-and low-dose PC groups were down-regulated(P<0.01 or P<0.05). At the 8 th week, the m RNA transcription levels of AMPK in the high-dose PC group and FOXO3α in the medium-dose PC group, and protein levels of AMPKα1, p AMPKα1 and FOXO3α in the 3 CM groups were significantly up-regulated(P<0.01 or P<0.05); the m RNA transcription levels of TLR4 in the medium-and low-dose PC groups, NLRP3 in the high-and low-dose PC groups and MCP-1 in the medium-and low-dose PC groups, and protein expression levels of TLR4, NLRP3 and MCP-1 in the 3 CM groups were down-regulated(P<0.01 or P<0.05). Conclusion: PC up-regulated the expression of AMPK and its downstream molecule FOXO3α and inhibited the biological activity of TLR4, NLRP3, and MCP-1, key signal molecules in the immunoinflammatory network pathway, which may be the molecularmechanism of PC to improve hyperuricemia-mediated immunoinflammatory metabolic renal damage.展开更多
文摘目的观察秦苓液对尿酸性肾病大鼠过氧化物酶体增殖物激活受体γ辅助激活因子-1α(peroxisome proliferator activated receptor coactivator-1 alpha,PGC-1α)、白介素-1β(interleukin beta,IL-1β)、受激活调节正常T细胞表达和分泌因子(regulated upon activation normal T cell expressed and secreted factor,RANTES)的影响。方法采用腺嘌呤灌胃伴酵母饲料喂养大鼠的方法进行造模,成模后随机分为模型组,阳性药物组,秦苓液大、中、小剂量组,每组12只大鼠。选6只正常大鼠作为正常对照组。各组分别用药连续干预6周和8周后采用实时荧光定量逆转录聚合酶链反应(real time reverse transcription-polymerase chain reaction,RT-PCR)、蛋白免疫印迹法(Western blot)、免疫组化和酶联免疫吸附试验(enzyme linked immunosorbent assay,ELISA)检测大鼠PGC-1α、IL-1β、RANTES基因和蛋白表达水平。结果与正常对照组比较,6周时和8周时模型组PGC-1α蛋白表达水平及8周时PGC-1α基因转录水平显著下调,6周时IL-1β基因转录、蛋白表达水平和RANTES蛋白表达水平及8周时两者水平显著上调(P<0.05,P<0.01)。与模型组比较,6周时秦苓液各剂量组PGC-1α蛋白表达水平及8周时中、小剂量组PGC-1α基因水平及蛋白表达水平显著上调,6周时秦苓液各剂量组IL-1β基因转录及大、中剂量组IL-1β蛋白表达水平以及各剂量组RANTES蛋白表达水平显著下调,8周时秦苓液小剂量组IL-1β及中、小剂量组RANTES基因转录及中、小剂量组IL-1β、RANTES蛋白表达水平显著下调(P<0.05,P<0.01)。结论秦苓液抑制UN模型大鼠免疫炎性损伤,其分子机制可能与上调PGC-1α的水平,抑制IL-1β、RANTES的生物学活性有关。
基金Supported by the National Natural Science Foundation of China(No.81473516 and No.30973918)
文摘Objective: To observe the effects of Chinese medicine(CM) Polygonum cuspidatum(PC) on adenosine 5'-monophosphate-activated protein kinase(AMPK), forkhead box O3α(FOXO3α), Toll-like receptor-4(TLR4), NACHT, LRR and PYD domains-containing protein 3(NLRP3), and monocyte chemoattractant protein-1(MCP-1) expression in a rat model of uric acid-induced renal damage and to determine the molecular mechanism. Methods: A rat model of uric acid-induced renal damage was established, and rats were randomly divided into a model group, a positive drug group, and high-, medium-, and low-dose PC groups(n=12 per group). A normal group(n=6) was used as the control. Rats in the normal and model groups were administered distilled water(10 m L·kg^(–1)) by intragastric infusion. Rats in the positive drug group and the high-, medium-, and low-dose PC groups were administered allopurinol(23.33 mg·kg^(–1)), and 7.46, 3.73, or 1.87 g·kg^(–1)·d^(–1) PC by intragastric infusion, respectively for 6 to 8 weeks. After the intervention, reverse transcription polymerase chain reaction, Western blot, enzyme linked immunosorbent assay, and immunohistochemistry were used to detect AMPK, FOXO3α, TLR4, NLRP3, and MCP-1 m RNA and protein levels in renal tissue or serum. Results: Compared with the normal group, the m RNA transcription levels of AMPK and FOXO3α in the model group were significantly down-regulated, and protein levels of AMPKα1, pAMPKα1 and FOXO3α were significantly down-regulated at the 6 th and 8 th weeks(P<0.01 or P<0.05). The m RNA transcription and protein levels of TLR4, NLRP3 and MCP-1 were significantly up-regulated(P<0.01 or P<0.05). Compared with the model group, at the 6 th week, the mRNA transcription levels of AMPK in the high-and medium-dose groups, and protein expression levels of AMPKα1, p AMPKα1 and FOXO3α in the high-dose PC group, AMPKα1 and p AMPKα1 in the mediumdose PC group, and p AMPKα1 in the low-dose PC group were significantly up-regulated(P<0.01 or P<0.05); the m RNA transcription and protein levels of TLR4 and NLRP3 in the 3 CM groups, and protein expression levels of MCP-1 in the medium-and low-dose PC groups were down-regulated(P<0.01 or P<0.05). At the 8 th week, the m RNA transcription levels of AMPK in the high-dose PC group and FOXO3α in the medium-dose PC group, and protein levels of AMPKα1, p AMPKα1 and FOXO3α in the 3 CM groups were significantly up-regulated(P<0.01 or P<0.05); the m RNA transcription levels of TLR4 in the medium-and low-dose PC groups, NLRP3 in the high-and low-dose PC groups and MCP-1 in the medium-and low-dose PC groups, and protein expression levels of TLR4, NLRP3 and MCP-1 in the 3 CM groups were down-regulated(P<0.01 or P<0.05). Conclusion: PC up-regulated the expression of AMPK and its downstream molecule FOXO3α and inhibited the biological activity of TLR4, NLRP3, and MCP-1, key signal molecules in the immunoinflammatory network pathway, which may be the molecularmechanism of PC to improve hyperuricemia-mediated immunoinflammatory metabolic renal damage.