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目的探讨化痰通络方对IL-1β刺激的类风湿关节炎滑膜成纤维细胞(rheumatoid arthritis synovial fibroblast,RASFB)增殖及肿瘤坏死因子-α(TNF-α)和酸性成纤维细胞生长因子(acidic fibroblast growth factor,a FGF)分泌的影响。方法体外培养RASFB细胞株,加入终浓度为1、5、10、20μg/L的IL-1β24、48 h以WST-1法检测RASFB增殖率;然后选择20μg/L的IL-1β为诱导剂量为IL-1β组。在此基础上加入终浓度(V/V)为5%、2%、1%的高、中、低浓度化痰通络方水煎液为高、中、低浓度化痰通络方组,培养24、48 h,并设空白对照组。比较RASFB的增长率。ELISA法检测各组TNF-α和a FGF含量,RT-PCR检测TNF-α和a FGF mRNA表达。结果24、48 h时,与IL-1β1μg/L比较,IL-1β5、10、20μg/L RASFB增殖率升高(P<0.01);与IL-1β5μg/L比较,IL-1β10、20μg/L RASFB增值率升高(P<0.01),且IL-1β20μg/L RASFB增值率高于IL-1β10μg/L(P<0.01)。48 h时各浓度的RASFB增值率高于24 h(P<0.01)。与高浓度化痰通络方组比较,中、低浓度化痰通络方组RASFB增殖率降低,且中浓度化痰通络方组对IL-1β刺激的RASFB增殖率明显降低(P<0.01)。与空白对照组比较,IL-1β组24、48 h时TNF-α、a FGF mRNA表达及含量均升高(P<0.05);与IL-1β组比较,除24 h低浓度化痰通络方组TNF-αmRNA表达外,24、48 h高、中、低浓度化痰通络方TNF-α、a FGF mRNA表达及含量均降低(P<0.05);与高浓度化痰通络方组比较,24、48 h时中、低浓度化痰通络方TNF-α、a FGF mRNA表达升高,24 h低浓度化痰通络方TNF-α含量以及24、48 h中、低浓度化痰通络方TNF-α、a FGF含量升高(P<0.05)。结论化痰通络方治疗RA的机理可能涉及抑制RASFB的增殖,降低TNF-α和a FGF mRNA的表达及其蛋白的分泌。
Objective To investigate the effects of Huatan Tongluo Prescription on the proliferation of rheumatoid arthritis synovial fibroblast (RASFB) stimulated by IL-1β and the effects of TNF-α and acidic fibroblast growth factor acidic fibroblast growth factor, a FGF) secretion. Methods RASFB cell lines were cultured in vitro. IL-1β24 at a final concentration of 1, 5, 10 and 20 μg / L was added to the cells for 48 and 48 h, respectively. The proliferation rate of RASFB was detected by WST-1 method. Then, IL- IL-1β group. On this basis, the high, middle and low concentrations of Huatan Tonglu Prescription Decoction with final concentration (V / V) of 5%, 2% and 1% Culture 24,48 h, and set a blank control group. Compare RASFB growth rates. The contents of TNF-α and a FGF in each group were detected by ELISA. The expression of TNF-α and a FGF mRNA was detected by RT-PCR. Results Compared with IL-1β1μg / L, the proliferation rate of IL-1β5,10,20μg / L RASFB increased at 24 and 48 hours (P <0.01). Compared with IL-1β5μg / L, IL- RASFB increased (P <0.01), and the added value of IL-1β20μg / L RASFB was higher than that of IL-1β10μg / L (P <0.01). The RASFB value of each concentration at 48 h was higher than that of 24 h (P <0.01). Compared with high-concentration Huatan Tongluo Recipe group, the proliferation rate of RASFB in middle and low concentration Huatan Tongluo Recipe group was decreased, and the proliferation rate of RASFB stimulated by IL-1β was significantly lower in Huatan Tongluo Recipe group (P <0.01) ). Compared with the blank control group, the expression and content of TNF-α and a FGF mRNA in IL-1β group increased significantly at 24 and 48 h (P <0.05). Compared with IL-1β group, The expression of TNF-α and a FGF mRNA in high, middle and low concentration of Huatan Tongluo Recipe at 24h and 48h were significantly decreased (P <0.05), except for the expression of TNF-αmRNA, Compared with 24 and 48 h, the expression of TNF-α and a FGF mRNA in low-concentration Huatan Tongluo Tongluo recipe increased, and the content of TNF-α in low-density Huatan Tongluo Recipe at 24 h and 24 h and 48 h The levels of TNF-α and a FGF in Sputum Tongluo Recipe increased (P <0.05). Conclusion The mechanism of Huatan Tongluo Recipe in treating RA may involve inhibiting the proliferation of RASFB and decreasing the expression of TNF-α and a FGF mRNA and the secretion of its protein.