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[目的]研究钙网织蛋白在阿霉素诱导的凋亡人肺癌A549细胞免疫原性中的作用。[方法]体外培养人肺癌A549细胞,将阿霉素加入A549细胞培养液中,使其终浓度分别为1×10-3g/L、3×10-3g/L、5×10-3g/L,12 h后以吖啶橙/溴化乙锭(AO/EB)双染法检测细胞凋亡;应用激光共聚焦显微镜观察凋亡A549细胞表面钙网织蛋白暴露情况。阿霉素3×10-3g/L作用12 h后诱导的凋亡细胞为抗原组(A组),A组加入钙网织蛋白阻断蛋白(A-Ca组),分别刺激外周血单个核细胞,72h后以MTT法检测其对培养的A549细胞的杀伤效果。[结果]阿霉素3×10-3g/L终浓度作用A549细胞12h后,凋亡率为62.93%±1.70%。经激光共聚焦显微镜检测凋亡A549细胞表面可见钙网织蛋白抗体荧光。外周血单个核细胞杀伤实验MTT检测OD值:A-Ca组0.537±0.056,A组0.437±0.023(P<0.05)。[结论]阿霉素诱导凋亡的A549细胞表面有钙网织蛋白的暴露。阿霉素诱导的凋亡A549细胞经添加钙网织蛋白阻断蛋白后,其免疫原性下降,表明钙网织蛋白在其免疫原性中起重要作用。
[Objective] To investigate the role of calcium reticulin in the immunogenicity of adriamycin-induced apoptosis in human lung cancer A549 cells. [Method] Human lung cancer A549 cells were cultured in vitro. Adriamycin was added to A549 cell culture medium to make final concentration of 1 × 10-3g / L, 3 × 10-3g / L, 5 × 10-3g / L Apoptosis was detected by acridine orange / ethidium bromide (AO / EB) staining 12 h later. The exposure of apoptotic A549 cells to Ca2 + was observed by laser confocal microscopy. The apoptotic cells induced by doxorubicin 3 × 10-3g / L for 12 h were antigen group (group A), and group A was treated with A-Ca, and stimulated peripheral blood mononuclear cells After 72h, the cytotoxicity against A549 cells was detected by MTT assay. [Result] The apoptosis rate of A549 cells treated with doxorubicin at a concentration of 3 × 10-3g / L for 12 hours was 62.93% ± 1.70%. Laser confocal microscopy was used to detect the fluorescence of calcium reticulin antibody on the surface of apoptotic A549 cells. Peripheral blood mononuclear cell killing test MTT OD value: A-Ca group 0.537 ± 0.056, A group 0.437 ± 0.023 (P <0.05). [Conclusion] Adriamycin induced the apoptosis of A549 cells with the presence of calcium reticulin exposure. Adriamycin-induced apoptosis A549 cells after addition of calcium reticulin blocking protein, its immunogenicity decreased, indicating that calcium reticulin plays an important role in its immunogenicity.