槐定碱、苦参碱和苦豆碱对LPS诱发结肠上皮细胞炎症模型中细胞因子IL-6和TNF-α水平的影响

Effects of Sophoridine, Matrine and Aloperine on the Contents of Cytokine IL-6 and TNF-α in LPS-induced IEC-6 Cells Inflammatory Model

摘要:

目的：构建结肠上皮细胞炎症模型，探讨槐定碱、苦参碱和苦豆碱对细胞因子白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）表达的影响。方法：将细胞接种12孔板中，每孔接种肠上皮细胞株IEC-6 2×10⁵个细胞。细菌脂多糖（LPS，1 mg·L^-1）干预生长良好的IEC-6细胞12 h，建立细胞炎症模型。四甲基偶氮唑盐（MTT）法检测3种生物碱不同浓度对炎症细胞增殖的影响；ELISA法检测给药后细胞上清中IL-6，TNF-α的变化；免疫印迹法检测细胞内IL-6及TNF-α蛋白表达水平。结果：与正常组比较，模型组细胞上清中IL-6，TNF-α水平显著升高，LPS （1 mg·L^-1）干预12 h后，IL-6，TNF-α含量分别为（1 899.68±45.80），（148.67±5.93） ng·L^-1。苦豆碱、槐定碱和苦参碱均可有效减弱LPS诱导的结肠上皮细胞损伤。当生物碱浓度为100 μmol·L^-1时，槐定碱、苦参碱和苦豆碱均可提高LPS损伤细胞的活性，细胞存活率分别从80.47%提高为91.96%，92.40%，97.23%。此外，这3种生物碱可降低IL-6和TNF-α的含量及其在细胞内的表达。结论：槐定碱、苦参碱和苦豆碱3种生物碱均可通过降低IL-6和TNF-α活性发挥抗炎作用。其中，苦豆碱的抗炎作用最佳。

Abstract:

Objective: To observe the protective effects of sophoridine, matrine and aloperine on lipopolysaccharide (LPS)-induced IEC-6 cells inflammatory model. Method: IEC-6 cells were seeded into 12-well plates at the density of 2×10⁵/well and treated with LPS for 12 h. Then, the inflammatory model was constructed by detecting the content of interleukin(IL)-6 and tumor necrosis factor-α(TNF-α). MTT assay was used to determine the activities of aloperine, sophoridine and matrine on LPS-injured IEC-6 cells. Enzyme-linked immunosorbant assay(ELISA) was used to detect the content of IL-6 and TNF-α in cell supernatant and Western blotting analysis was employed to detect the protein expression levels of IL-6 and TNF-α. Result: Compared with the control group, levels of IL-6 and TNF-α of inflammatory model group were significantly upregulated. Aloperine, sophoridine and matrine could significantly reduce the cytotoxicities of IEC-6 cells induced by LPS. The cell viabilities were increased from 80.47% to 91.96%, 92.40% and 97.23% after sophoridine, matrine and aloperine treatment at the concentration of 100 μmol·L^-1, respectively. Moreover, ELISA and Western blotting analysis results demonstrated that these compounds could attenuate the content and expression of IL-6 and TNF-α. Conclusion: Aloperine, sophoridine and matrine exhibit anti-inflammatory effects though decreasing the activities of IL-6 and TNF-α. Among them, aloperine shows the best activities.