丙戊酸钠在γδT细胞对肺癌A549细胞免疫中的作用

The effect of sodium valproate in γδT cells on lung cancer A549 lines cells

目的:研究丙戊酸钠(Sodium Valproate,VPA)作为组蛋白脱乙酰酶抑制剂对肺癌A549细胞MICA蛋白的影响,并探讨丙戊酸钠在人外周血γδT细胞对肺癌A549细胞免疫中的作用。方法:用Ficoll密度梯度离心法分离健康人外周血单核细胞(PBMC),用流式细胞仪分选纯化γδT细胞,用IL-2及唑来磷酸刺激γδT细胞扩增。用不同浓度的丙戊酸钠处理肺癌A549细胞,培养24小时后用PCR及Western blot检测MICA的变化。将分离纯化的γδT细胞与处于对数期生长的肺癌A549细胞混合培养,并加入VPA、MICA抗体处理,24小时后用LDH法检测γδT细胞的细胞毒性。结果:加入VPA后,PCR及Western blot结果显示肺癌A549细胞MICA蛋白在转录及表达上均增强;分离纯化的γδT细胞经体外扩增后,对肺癌A549细胞有较强的细胞毒性作用;加入VPA后γδT细胞对肺癌A549细胞的细胞毒性作用较未加入组增强(P<0.05)。结论:体外扩增的γδT细胞对肺癌A549细胞产生较强的细胞毒性杀伤作用;组蛋白脱乙酰酶抑制剂-丙戊酸钠可上调A549细胞MICA的表达,并增强γδT细胞对肺癌A549细胞的杀伤作用。

Objective ： To research the effects of histone deacetylase inhibitor Sodium Valproate （VPA） on the protein MICA of lung cancer A549 lines cells, and also to detect the cytotoxic effects of peripheral bloodγδT cells on lung cancer A549 lines cells. Methods ： Peripheral blood mononuclear cells （PBMC） were collected from health human blood with the ficoll density gradient centrifu- gation methods, and the γδTcells were separated from the PBMC with Flow cytometry. The interleukin-2 （IL-2） and zoledronic acid （ZA） were used to stimulate and expand theγδT cells. A549 lines cells were cultured with different concentration of VPA, and the change of MICA was analyzed by the methods of PCR and Western blot after 24 hours cultured. Purification γδT cells and exponential phase A549 lines ceils were co-cultured with VPA and MICA-Ab, then the cytotoxicity of γδT ceils against A549 ceils was tested with LDH method after 24 hours cultured. Results ：The results of PCR and Western blot showed that the VPA could enhance the expression of MICA, The purification γδT cells stimulated and expanded by IL-2 and ZA in vitro showed strong cytotoxic effects towards lung canc- er A549 lines cells ,the cytotoxicity of group with VPA was stronger than the group without VPA （P 〈 0. 05 ）. Conclusion：γδT ceils stimulated and expanded in vitro showed strong cytotoxic on lung cancer A549 lines cells ; the histone deacetylase inhibitor VPA can en- hance the expression of MICA, and also increase the cytotoxic of γδT ceils on lung cancer A549 lines cells.