青蒿琥酯下调人喉癌细胞所致T细胞免疫抑制的体外实验研究

Down-regulating effect of Artesunate on T lymphocytes' immunosuppression in human laryngocarcinoma cells in vitro

目的体外实验研究青蒿琥酯(ART)对人喉癌细胞所致T细胞免疫抑制的影响。方法制备经ART预处理的人喉癌Hep-2细胞培养上清(ART-S),收集连续2次再培养的上清(分别称为ART-S1、ART-S2),以不经ART作用的Hep-2及其上清作对照;分析对人外周血单个核细胞经光镜观察、噻唑蓝(MTT)法测定的植物血凝素(PHA)诱导淋巴细胞转化增殖反应,以及流式细胞计数(FCM)法检测的T淋巴细胞表面CD3ε+、CD3ε+ζ+、IL-2Rα+表达的影响。结果 Hep-2对5项免疫指标的抑制率分别为(52.58±6.13)%、(50.34±7.26)%、(45.28±5.38)%、(39.74±5.27)%、(64.25±9.37)%。ART作用后,可显著下调Hep-2所致T细胞免疫抑制(P<0.01);第1次再培养后,对淋巴细胞转化、诱导增殖、IL-2Rα表达抑制的逆转均明显降低,与ART-S比较差异有统计学意义(P<0.01),对CD3ε+及CD3ε+ζ+表达抑制的逆转持续增强,与ART-S比较差异有统计学意义(P<0.05);第2次再培养后,对淋巴细胞转化、诱导增殖、IL-2Rα表达抑制的逆转继续保持,与ART-S1比较差异无统计学意义(P>0.05),对CD3ε+、CD3ε+ζ+表达抑制的逆转明显下降回复至ART-S水平,差异无统计学意义(P>0.05)。结论人喉癌Hep-2细胞可产生T细胞免疫抑制作用,ART可对其产生稳定的逆转作用。下调肿瘤免疫抑制是ART的重要抗瘤机制之一。

Objective To investigate the effects of Artesunate （ ART ） on T lymphocytes ’ immunosuppression in human laryngocarcinoma cells in vitro .Met hods The supernatants from Hep-2 （ human laryngocarcinoma cell line ） were pretreated with ART （ ART-S） ,then the supernatants cultured for the first time and the second time were collected （ ART-S1 and ART-S2,respectively）,and the corresponding supernatants and Hep-2 without ART treatment were served as control group.The effects of different supernatants on immune parameters in human peripheral blood mononuclear cells were analyzed , including the lymphocytes proliferation induced by phytohemagglutinin （PHA）,which was detected by MTT and microscope observation,and expressions of CD3ε＋,CD3ε＋ζ＋and IL-2Rα＋were measured by flow cytometry （FCM）.Results The inhibitory rates of five immune parameters caused by Hep -2 cells were52.58%±6.13%,50.34%±7.26%,45.28%± 5.38%,39.74%±5.27%,64.25%±9.37%,respectively.After pretreated with ART,the inhibition effects caused by Hep-2 were significantly decreased （ P 0.05）,nor were the reversing effects on the inhibition of CD3ε＋and CD3ε＋ζ＋expressions between ART-S2 group and ART-S1 group （ P 〉0.05）.Conclusion Human laryngocarcinoma Hep-2 cells have T lymphocytes immunosuppression effects .However Artesunate can steadily reversethe immunosuppression effects.Down-regulating tumor immunosuppression effect may be one of important anti -tumor actionmechanisms of Artesunate.