Toll样受体4参与介导黄曲霉毒素G1对大鼠肺泡Ⅱ型上皮细胞的致损伤作用

Toll-like receptor 4 mediated inducing-injury role of aflatoxin G1 on rat Type Ⅱ alveolar epithelial cells

目的在基因水平上探讨Toll样受体4(TLR4)参与介导黄曲霉毒素G1(AFG1)对肺泡Ⅱ型上皮细胞的致损伤生物学效应,及其作用机制和途径。方法分离、纯化、培养大鼠肺泡Ⅱ型上皮细胞并配置3个不同浓度的AFG1液体。实验随机分为正常组、实验组、溶剂对照组,每组6个样本。正常组加入同等量的无菌生理盐水;实验组加入不同浓度的AFG1液(高、中、低浓度分组),溶剂对照组加入同等量的二甲基亚砜(DMSO),终浓度为0.4 ml/L;同时处理24 h后在上述5组提取细胞mRNA,采用RT-PCR法检测各组TLR4 mRNA、NF-κBmRNA、TNF-αmRNA和IL-6mRNA的表达水平。结果正常组肺泡Ⅱ型上皮细胞上TLR4 mRNA、NF-κBmRNA、TNF-αmRNA和IL-6mRNA的表达量较低。实验组经AFG1液作用后,各项指标的表达量显著高于正常组(P<0.05)和溶剂对照组(P<0.05),且呈浓度依赖性增高。溶剂对照组各项指标的表达量与正常对照组相比,无显著性差异(P>0.05)。结论 TLR4→NF-κB→炎症介质信号转导通路参与AFG1对AT-Ⅱcells的损伤作用。

Objective On gene level to explore biological effects of Toll-like receptor 4 mediated the injury role of aflatoxin on alveolar type Ⅱ epithelial cells, as well as in this process role mechanisms and pathway. Methods In this experiment 3 groups were randomly designed. Group l（normal group）; Group 2 （the experimental group）, and Group 3 （solvent control group）, 6 samples （i.e isolated, purified and cultured Type 11 alveolar epithelial cells, AT-II cells）, per group. In normal group an equal volume of sterile saline was added; in experimental group （i.e Group 2） added different concetrations of AFG1 solution （i.e 2.0 mg/L, 1.0 mg/L and 0.5 mg/L, as B1, B2, B3 subgroups）; in solvent control group （i.e Group3） added DMSO; in Group 2 and 3, the end concetration of DMSO was 0.4ml/L. Then, after treatment for 24hs, from above-mentioned 5 group extracting mRNA was taken to detect the levels of TLR4 mRNA, NF-κBmRNA, TNF-ctmRNA and IL-6mRNA of each group by using RT-PCR. Results In normal group on alveolar Type U epithelial cells the expression of TLR4 mRNA, NF-κBmRNA, TNF-αmRNA and IL-6mRNA was lower levels. In experimental group ceils after AFG1 solution acting, those related indicator expressions were significantly higher than that in normal group （P〈0.05） and solvent control group （/（P〈0.05）, and a dose dependence increased. Solvent control group, in these related indicator expressions compared with normal control group, had no significant differences （P〉0.05）. Conclusion TLR4-→NF-κB→inflammatory mediators involved in signal transduction pathway of AFG1 inducing injury of AT-11 cells.