体外扩增NK细胞中非克隆性染色体异常的特征及意义

The characteristics and significance of non-clonal chromosome aberrations in the in vitro expanded NK cells

背景:非克隆性染色体异常在干细胞培养中较为常见且研究较多,而在体外扩增的NK细胞中研究甚少。目的:探讨体外扩增NK细胞中非克隆性染色体异常的特征及意义。方法:分离肿瘤患者和健康志愿者外周血单个核细胞,应用人工化抗原递呈细胞刺激NK细胞增殖,流式细胞术鉴定其表型,并测定其产生γ-干扰素的功能。传统G显带法分析NK细胞染色体异常。对两组NK细胞的非克隆性染色体异常发生率、分泌γ-干扰素的功能及两者相关性进行分析。结果与结论:(1)7例肿瘤患者的NK细胞样本中,6例检测出非克隆性染色体异常,而在12例健康志愿者中,6例存在非克隆性染色体异常;(2)肿瘤组481个中期分裂相,其中17个含有非克隆性染色体异常;对照组785个中期分裂相,其中7个含有非克隆性染色体异常[3.53%(17/481)vs.0.89%(7/785),Z=-2.212,P=0.028];(3)4例有放化疗病史的患者,其NK细胞中非克隆性染色体异常频率明显高于对照组(P=0.003)及初诊肿瘤患者(P=0.034),而初诊患者与对照组并无明显差别(P=0.819);(4)肿瘤组分泌γ-干扰素的NK细胞比例与对照组无明显差别(P=0.371)。NK细胞产生γ-干扰素水平与非克隆性染色体异常发生率亦无明显相关性;(5)实验首次探讨了以人工化抗原递呈细胞扩增的NK细胞中非克隆性染色体异常的特征,并初步研究了其与NK细胞功能之间的关系。

BACKGROUND:The significance of non-clonal chromosome aberrations(NCCAs)in in vitro expanded stem cells has been discussed in many studies;however,it is not well documented in the expanded NK cells.OBJECTIVE:To investigate the characteristics and significance of NCCAs in the in vitro expanded NK cells.METHODS:Peripheral blood mononuclear cells were isolated from patients with tumors and healthy controls.Artificial antigen presenting cells were used to stimulate the proliferation of NK cells.On the 7th day of culture,the cells were collected for phenotype identification and interferon-γsecretion test by flow cytometry and for cytogenetic analysis by the conventional G-banding technique.The difference in the frequency of NCCAs and interferon-γsecretion between the two groups and the correlation between NCCAs and interferon-γsecretion were studied.RESULTS AND CONCLUSION:NCCAs were found in 6 of 7 samples from the tumor patient group and 6 of 12 samples from the healthy controls.In the tumor patient group,17 metaphases containing NCCAs were found out of 481 metaphases,and in the control group,7 NCCA-containing metaphases were detected out of 785 metaphases[3.53%(17/481)vs.0.89%(7/785),Z=-2.212,P=0.028].Four patients with a history of chemoradiotherapy showed higher NCCA frequencies than those patients undergoing chemoradiotherapy(P=0.034)and the healthy controls(P=0.003).No difference was found between the newly diagnosed tumor patients and the healthy controls(P=0.819).The ratios of interferon-γsecreting NK cells in the two groups were not significantly different(P=0.371)and the correlation between the frequency of NCCAs and the interferon-γsecreting level was not identified.It was the first time that the characteristics of NCCAs in the artificial antigen presenting cells-expanded NK cells and the relationship between NCCAs and the NK cell function were preliminarily studied.The effect of NCCAs on the function and survival of NK cells in vitro and in vivo needs further investigations.