西达本胺联合干扰素α对人皮肤T细胞淋巴瘤细胞系Hut78的协同作用及其分子机制的研究

Synergistic effect of chidamide and interferon alpha on mycosis fungoides and Sézary syndrome-related cell lines and its possible molecular mechanism

目的研究西达本胺联合干扰素α(IFN-α)对蕈样肉芽肿及Sézary综合征相关的人皮肤T细胞淋巴瘤细胞系Hut78的协同作用及可能的分子机制,进而为治疗蕈样肉芽肿及Sézary综合征提供一种更有效的治疗方案。方法用不同浓度的西达本胺和不同浓度的干扰素(IFN)-α以及西达本胺联合IFN-α分别作用Hut78细胞系24、48、72 h后,利用MTS[四唑化合物,3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium]法检测Hut78细胞的生存率,利用甲基纤维素半固体培养基(CFC)检测西达本胺、IFN-α单独以及两者联合作用Hut78细胞24 h后的体外克隆形成能力,并用流式细胞仪检测西达本胺、IFN-α以及两者联合作用细胞24 h后,Hut78细胞的凋亡情况。采用实时荧光定量聚合酶链反应(real-time PCR)检测西达本胺、IFN-α以及两者联合作用Hut78细胞24 h后凋亡相关基因Fas、FasL、Caspase3及Caspase8 mRNA的表达情况。结果西达本胺和IFN-α均能有效抑制Hut78细胞的增殖,且这种作用呈剂量依赖性和时间依赖性。西达本胺联合IFN-α比单药对Hut78细胞的增殖抑制、克隆形成、凋亡诱导作用更明显。西达本胺、IFN-α能使Fas、Caspase8、Caspas3表达明显增加,且西达本胺联合IFN-α比单药对上述基因表达增加作用更加明显,但对FasL的表达没有影响。结论西达本胺通过抑制细胞增殖及克隆形成和促进细胞凋亡对蕈样肉芽肿及Sézary综合征起到治疗作用。与单独使用IFN-α相比,西达本胺联合IFN-α能明显提高蕈样肉芽肿及Sézary综合征的治疗效果,减少IFN-α的使用量。

Objective To investigate the synergistic effect and possible molecular mechanism of chidamide combined with interferon alpha(IFN-α)on mycosis fungoides and Sézary syndrome-related cell line Hut78,so as to provide a more effective treatment for mycosis fungoides and Sézary syndrome.Methods Hud78 cell line was treated with different concentrations of chidamide,IFN-αand the mixture of the above two components respectively and the survival rates of Hut78 cells were measured by MTS method after 24,48,and 72 hours of coculture.The colony forming ability of Hut78 cells was detected by methylcellulose semi-solid medium(CFC),as well as the apoptosis of Hut78 cells was detected by flow cytometry,and the expression of Fas,FasL,Caspase3,and Caspase8 mRNA in Hut78 cells was measured by real-time PCR after 24 hours of coculture.Results Both chidamide and IFN-αcould effectively inhibit the proliferation of Hut78 cells in a dose-dependent and time-dependent manner.Chidamide combined with IFN-αshowed a more obvious effect on the inhibition of proliferation,colony formation,and apoptosis induction of Hut78 cells than that of the single drug.Chidamide and IFN-αcould significantly increase the expression of Fas,Caspase8 and Caspas3,and the combination of chidamide and IFN-αhad a more significant effect on the expression of the genes mentioned above than that of the single drug,but had no effect on the expression of FasL.Conclusion Chidamide may play a therapeutic role in mycosis fungoides and Sézary syndrome by inhibiting cell proliferation and colony formation,as well as promoting apoptosis.To be compared with the use of IFN-αalone,the combination of chidamide and IFN-αcan significantly improve the treatment effect of mycosis fungoides and Sézary syndrome,meanwhile reduce the dosage of IFN-α.