摘要
目的 观察细胞因子诱导的杀伤细胞（CIK细胞）对白血病细胞凋亡的影响,探讨miR-155在此过程中的作用.方法 MTT法检测CIK细胞对白血病NALM-6和Jurkat细胞的细胞毒作用,实时定量反转录PCR检测白血病细胞miR-155表达,流式细胞术检测miR-155对CIK细胞诱导白血病细胞凋亡的影响,构建包含miR-155作用位点的CEBP/β基因3＆#39;-UTR区的质粒,转染白血病细胞,双荧光素酶报告基因试剂盒检测白血病细胞CEBP/β荧光素酶活性.结果 CIK细胞对NALM-6和Jurkat细胞具有强大的细胞毒作用,且呈时间与剂量依赖性（P＜0.05）,同时CIK细胞能上调NALM-6和Jurkat细胞中miR-155的表达,其中NALM-6细胞上调（2.87±0.19）倍（t=2.787,P＜0.05）,Jurkat细胞上调（1.98±0.25）倍（t＝ 3.513,P＜ 0.05）.另外,miR-155 mimics能促进CIK细胞对NALM-6和Jurkat细胞诱导的凋亡作用（t=4.239,P＜0.05;t=3.565,P＜0.05）,而miR-155抑制剂能够拮抗此过程（t=3.772,P＜ 0.05;t=4.017,P＜ 0.05）.miR-155直接作用于CEBP/β基因3＆#39;-UTR区,且CIK细胞降低白血病细胞荧光素酶活性,其中NALM-6细胞下降（42.89±2.07）％（t=3.578,P＜0.05）,Jurkat细胞下降（37.02±1.95）％（t=4.393,P＜ 0.05）.结论 CIK细胞通过上调miR-155促进白血病细胞凋亡,这为CIK细胞治疗白血病提供新的数据.
Objective To observe the effect of cytokine-induced killer （CIK） cells on the apoptosis of human leukemia cells,and explore the role of miR-155 in this process.Methods The cytotoxicity of CIK cells against a variety of leukemic cell lines （NALM-6,Jurkat） was investigated by MTT technique,miR-155 was determined by real time quantitative PCR,and the apoptosis was detected by flow cytometry in NALM-6 and Jurkat cells induced by CIK cells.Psi CHECK2-CEBP/β 3＇-UTR containing the binding site of miR-155 was constructed,and then it was transfected into NALM-6 and Jurkat cells.Luciferase activity of CEBP/β （CCAAT/enhancer binding protein beta） was determined with the assistance of dual luciferase report system.Results CIK cells possessed strong cytotoxicity against NALM-6 and Jurkat cells,which was time-dependent and dose-dependent （P 〈 0.05）.CIK cells could increase the expression of miR-155 in NALM-6 cells by （2.87±0.19） fold （t =2.787,P 〈 0.05）,and in Jurkat cells by （1.98±0.25） fold （t =3.513,P 〈 0.05）.Moreover,miR-155 mimics could promote the apoptosis of NALM-6 and Jurkat cells induced by CIK cells （t =4.239,P 〈 0.05;t =3.565,P 〈 0.05）.However,miR-155 inhibitor might block this process （t =3.772,P 〈 0.05;t =4.017,P 〈 0.05）.MiR-155 targeted at the site of CEBP/β3＇-UTR,and CIK cells could decrease the luciferase activity of NALM-6 cells by （42.89±2.07） % （t =3.578,P 〈 0.05）,meanwhile,in Jurkat cells by （37.02±1.95） % （t =4.393,P 〈 0.05）.Conclusion CIK cells could enhance human leukemia NALM-6 and Jurkat cells apoptosis by upregulating miR-155,which may provide a new database to elucidate leukemia cell therapy using CIK cells.
出处
《白血病．淋巴瘤》
CAS
2015年第6期328-333,共6页
Journal of Leukemia & Lymphoma
基金
湖北省自然科学基金（2014CFB395）
武汉大学自主科研项目（2042014kf0113）
武汉大学人民医院国家自然科学基金孵化项目（2013RMFH002）
