阿维A诱导皮肤鳞状细胞癌细胞系SCL-1凋亡机制研究

Mechanisms of apoptosis in cutaneous sqnamous cell carcinoma cell line SCL-1 induced by acitretin

目的探讨阿维A诱导人皮肤鳞状细胞癌细胞系SCL-1细胞凋亡的分子转导机制。方法采用酶联免疫分析法（ELISA）检测凋亡的诱导情况，吖啶橙染色证实凋亡的发生，Western印迹分析Fas、FasL、Fas相关死亡结构域（FADD）、caspase-8、caspase-3、多聚腺苷二磷酸核糖多聚酶（PARP）蛋白含量的变化。用Fas中和抗体ZB4检测Fas在阿维A诱导SCL-1细胞凋亡中的作用。结果阿维A能诱导SCL-1细胞凋亡，并具有浓度及时间依赖性。形态学观察可见，SCL-1细胞呈典型的凋亡征象。阿维A能明显上调Fas、FasL、FADD的蛋白表达，活化caspase-8、caspase-3和PARP。单独应用阿维A的凋亡吸光度为0．78±0．04，与ZB4共同处理SCL-1后，凋亡吸光度下降为0．41±0．03，ZB4显著阻断阿维A诱导的凋亡（P〈0．05）。结论阿维A能诱导皮肤鳞状细胞癌细胞凋亡，其机制与Fas死亡受体信号转导途径有关。

Objective To investigate the molecular transduction mechanisms of apoptosis in cutaneous squamous cell carcinoma cell line SCL-1 induced by acitretin. Methods SCL-1 cells were cultured and continuously treated with various concentrations of acitretin. Apoptosis was assessed by enzyme-linked immunosorbent assay （ELISA） in these cells on day 1, 3 and 5. Apoptotic cells were observed by acridine orange staining on day 5. The protein expressions of Fas, FasL, Fas-associated death domain （FADD）, cas- pase-8, caspase-3 and poly ADP-dbose polymemse （PARP） were examined using Western blot in SCL-1 cells treated with acitretin at 1× 10^-5 mol/L at different time points. Neutralizing anti-Fas antibody （ZB4, 1 μg/mL） was utilized to pretreat SCL-1 cells before the treatment with acitretin, following that, ELISA was done to compare the apoptosis in cells.treated with ZB4, acitretin, or the combination of ZB4 and acitretin, respectively. Results Acitretin induced the apoptosis of SCL-1 cells in a dose- and time-dependent manner. Morphologically, acitretin-treated SCL-1 cells showed a typical characteristic of apoptosis. Significant increase in Fas, FasL, and FADD protein expression, as well as the activation of caspase-8, caspase-3 and PARP were induced by the treatment with acitretin. The apoptosis absorbance value was 0.78 ＋ 0.04 in cells treated with acitretin alone, decreased to 0.41 ± 0.03 in cells treated with ZB4 and acitretin （P 〈 0.05）, suggesting that ZB4 could block the apoptosis of SCL-1 cells induced by acitretin. Conclusion Acitretin could induce the apoptosis of cutaneous squamous cell carcinoma cells likely by Fas signaling pathway.