CD4^(+)CD25^(+)foxp3^(+)调节性T细胞通过NKG2D增强急性髓性白血病中NK细胞毒性  

CD4^(+)CD25^(+)foxp3^(+)regulatory T cells promote NK cytotoxicity in acute myeloid leukemia via NKG2D

作　　者：薛春娥 赵娟娟 刘韦成 张义亮 马毓晗 李迎巧 Xue Chun'e;Zhao Juanjuan;Liu Weicheng;Zhang Yiliang;Ma Yuhan;Li Yingqiao(Department of Hematology,Langfang Traditional Chinese Medicine Hospital,Langfang 065000 Hebei,China)

机构地区：[1]河北廊坊市中医医院血液病科,廊坊065000

出　　处：《中华细胞与干细胞杂志（电子版）》2024年第4期212-219,共8页Chinese Journal of Cell and Stem Cell（Electronic Edition）

摘　　要：目的CD4^(+)CD25^(+)foxp3^(+)调节性T细胞在急性髓性白血病中调控NKG2D介导NK细胞毒性的研究。方法免疫磁珠法分离急性髓性白血病(AML)和健康对照外周血NK细胞和CD4^(+)CD25^(+)细胞,分析foxp3与NKG2D表达,CD4^(+)CD25^(+)foxp3^(+)细胞的比例与NK细胞的细胞毒作用相关性。Spearman法分析CD4^(+)CD25^(+)foxp3^(+)细胞比例与NK细胞的细胞毒性和NGK2D相关性。foxp3过表达处理AML外周血淋巴细胞后,流式细胞术检测NKG2D,IFN-γ和CD107a表达,LDH法检测NK细胞毒性。GEPIA2数据库分析AML患者NKG2D和foxp3基因表达相关性,TCGA数据库分析AML转录组数据中与NKG2D表达高相关性基因。IL-2(10μg/mL),IL-15(10μg/mL)处理NK细胞24 h后,流式细胞术检测NKG2D和CD107a表达,酶联免疫反应法检测NK细胞分泌的IFN-γ,LDH法检测NK细胞毒性。采用t检验等方法统计分析组间差异及变量相关性。结果与对照比较,AML组NK细胞中NKG2D表达(平均荧光强度MFI:942.25±117.17比3245.28±367.43)降低(P<0.001)。foxp3表达与NKG2D呈正相关性(r=0.590,P<0.001),NKG2D表达百分数与外周血淋巴细胞中CD4^(+)CD25^(+)foxp3^(+)细胞的比例呈正相关(r=0.708,P=0.002),AML患者外周血NK细胞活力也与CD4^(+)CD25^(+)foxp3^(+)细胞的比例呈正相关(r=0.655,P=0.006)。与OE-NC比较,OE-foxp3组细胞中NKG2D的表达水平(MFI:2115.37±269.53比914.28±103.36)、杀伤能力(58.24%±3.17%比24.18%±2.35%)、NK细胞毒性评估指标CD107a(45.43%±1.28%比18.24%±1.49%)和IFN-γ水平(24.16%±1.17%比16.28%±0.95%)升高(P均<0.001)。TCGA数据分析发现,NKG2D表达与CD3G、IL-3、IL-2、CD3E、IL-10、IL-17、IL-7R、IL-15、EZH2等基因相关性较高,流式细胞术检测发现,与对照比较,IL-2和IL-15处理后NK细胞的表面受体NKG2D和CD107a表达、IFN-γ水平、NK细胞毒性均升高。结论AML外周血中,NK细胞表面受体NKG2D低于正常对照,其中CD4^(+)CD25^(+)foxp3^(+)调节性T细胞通过IL-2和IL-15影响NKG2D表达进而调控Objective Exploring the effects and mechanisms of CD4^(+)CD25^(+)foxp3^(+)regulatory T cells in regulating NK cytotoxicity through NKG2D in acute myeloid leukemia.Methods Immunomagnetic bead method was used to isolate peripheral blood NK cells and CD4^(+)CD25^(+)cells from acute myeloid leukemia(AML)and healthy controls.The expression of foxp3 and NKG2D,and the relationship between the proportion of CD4^(+)CD25^(+)foxp3^(+)cells and the cytotoxicity of NK cells were analyzed.Spearman method was used to analyze the correlation between CD4^(+)CD25^(+)foxp3^(+)cell ratio and NK cell cytotoxicity as well as NGK2D.After overexpressed foxp3 in AML peripheral blood lymphocytes,flow cytometry was used to detect the expression of NKG2D,IFN-γand CD107a,LDH assay was used to detect the toxicity of NK cell.The correlation between NKG2D and foxp3 gene expression in AML patients was analyzed by GEPIA2 database,while the genes with high correlation of NKG2D expression in AML transcriptome data was analyzed by TCGA database.After treating NK cells with IL-2(10μg/mL)and IL-15(10μg/mL)for 24 hours,24 hours after treated with IL-2(10μg/mL)and IL-15(10μg/mL)in NK cells,flow cytometry was used to detect the expression of NKG2D and CD107a,enzyme-linked immunosorbent assay was used to detect the secretion of IFN-γby NK cells,LDH assay to detect NK cell toxicity.Statistical analysis of inter group differences and variable correlations using t-tests and other methods.Results Compared to the control group,the expression of NKG2D in NK cells in the AML group was significantly reduced(MFI:942.25±117.17 vs 3245.28±367.43,P<0.001).The expression of foxp3 is positively correlated with NKG2D(r=0.590,P<0.001),and the percentage of NKG2D expression is positively correlated with the proportion of CD4^(+)CD25^(+)foxp3^(+)cells in peripheral blood lymphocytes(r=0.708,P=0.002).The activity of peripheral blood NK cells in AML patients is also positively correlated with the proportion of CD4^(+)CD25^(+)foxp3^(+)cells(r=0.655,P=0.006).Compared t

关 键 词：急性髓性白血病 NK细胞 NKG2D CD4^(+)CD25^(+)foxp3^(+) IL-2 

分 类 号：R733.71[医药卫生—肿瘤]