HRZ三联抗结核药/TGF-β1 siRNA纳米脂质体对体外BCG感染的人巨噬细胞中Ag85A及TGF-β1表达的影响

Effects of HRZ triple antituberculosis drug/TGF-β1 siRNA nanoliposomes on the expression of Ag85A and TGF-β1 in human BCG-infected human macrophages

中文关键词:  异烟肼  利福平  吡嗪酰胺  三联抗结核药/表皮转化生长因子-β1 siRNA纳米脂质体  巨噬细胞  抗结核  靶向基因沉默

英文关键词:Isoniazid/rifampicin/pyrazinamide(HRZ)  Triple antituberculosis drug/transforming growth factor-β1 siRNA nanoliposomes  Macrophage  Antituberculosis  Targeted gene silencing

中文摘要:

  【摘要】 目的：探讨异烟肼、利福平、吡嗪酰胺（HRZ）三联抗结核药/表皮转化生长因子（TGF）-β1 siRNA纳米脂质体对体外BCG感染的人巨噬细胞中Ag85A及TGF-β1表达的影响。方法：体外培养人单核细胞株THP-1诱导分化成巨噬细胞，分为4组，空白组为单纯巨噬细胞培养；模型组为巨噬细胞与卡介苗（bacillus calmette-guerin，BCG）以1∶5的比例共培养3h，制备BCG感染的巨噬细胞模型；对照组为BCG感染的巨噬细胞与HRZ三联抗结核药纳米脂质体共培养；实验组为BCG感染的巨噬细胞与三种不同浓度的HRZ三联抗结核药/TGF-β1 siRNA纳米脂质体（C1组为35mg/ml，C2组为40mg/ml，C3组为50mg/ml）共培养。采用RT-PCR检测各组人巨噬细胞中Ag85A及TGF-β1的mRNA表达量，Western-blot方法检测Ag85A及TGF-β1的蛋白表达量。结果：与空白组相比，模型组Ag85A和TGF-β1的mRNA及蛋白量表达明显上调（P<0.05）；对照组中Ag85A的mRNA及蛋白量和TGF-β1的mRNA水平与模型组相比明显下调（P<0.05）；实验组3种不同浓度（C1组、C2组、C3组）HRZ三联抗结核药/TGF-β1 siRNA纳米脂质体处理后，Ag85A和TGF-β1的mRNA及蛋白表达量与模型组和对照组相比进一步下调（P<0.05），随着纳米脂质体浓度的增加，Ag85A和TGF-β1 mRNA及蛋白表达量有明显下降趋势，C1组与C3组相比Ag85A mRNA及蛋白表达量差异有统计学意义（P<0.05）；C1组与C2组、C3组相比TGF-β1 mRNA及蛋白量表达差异有统计学意义（P<0.05）；C2组与C3组相比Ag85A mRNA和TGF-β1蛋白表达量差异有统计学意义（P<0.05）。结论：HRZ三联抗结核药/TGF-β1 siRNA纳米脂质体对体外BCG感染的人巨噬细胞具有明显的下调Ag85A基因表达作用及TGF-β1基因沉默作用。

英文摘要:

  【Abstract】 Objectives: To investigate the effects of isoniazid/rifampicin/pyrazinamide(HRZ) triple anti-tuberculosis/epidermal transforming growth factor(TGF)-β1 siRNA nanoliposomes on the expression of Ag85A and TGF-β1 in human macrophages in vitro. Methods: The human mononuclear cell strain THP-1 was induced to differentiate into macrophages in vitro. The experiment was divided into 4 groups. The blank group was cultured as macrophage alone. In the model group, macrophage and bacillus calmette-guerin(BCG) were cocultured at the ratio of 1∶5 for 4h to prepare a BCG-infected macrophage model. In the control group, BCG-infected macrophages and HRZ triple antituberculosis nanoliposomes were cocultured. In the experimental group, BCG-infected macrophages and HRZ triple anti-tuberculosis drug/TGF-β1 siRNA nanoliposome(C1 35mg/ml, C2 40mg/ml, and C3 50mg/ml) were cococultured. The mRNA expression levels of Ag85A and TGF-β1 in human macrophages were detected by RT-PCR. The protein expression levels of Ag85A and TGF-β1 were detected by western-blot method. Results: RT-PCR and western-blot results showed that the mRNA and protein levels of Ag85A and TGF-β1 were significantly up-regulated in the model group(BCG-infected macrophage model) compared with the blank group(macrophage culture alone)(P<0.05). The mRNA and protein expressions of Ag85A and TGF-β1 in the control group(cocultured with BCG-infected macrophages and HRZ triple anti-tuberculosis nano-liposomes) were significantly down-regulated compared with the model group(P<0.05); Compared with the model group and the control group, the mRNAs and protein expressions of Ag85A and TGF-β1 treated with HRZ triple antituberculosis drugs/TGF-β1 siRNA nanoliposomes in three different concentrations(C1 group, C2 group and C3 group) in the experimental group was further down-regulated(P<0.05). With the increase of nano-liposome concentration(35mg/ml, 40mg/ml, 50mg/ml), the expressions of TGF-β1 mRNA and protein decreased significantly(the difference between C1 group and C2 group and C3 group was statistically significant, P<0.05). Conclusions: The self-developed HRZ triple anti-tuberculosis drug/TGF-β1 siRNA nano-liposome has obvious down-regulation of Ag85A gene expression and TGF-β1 gene silencing.