CAJ | 학술논문

目的检测重型再生障碍性贫血(SAA)患者外周血自然杀伤(NK)细胞体外对慢性髓细胞白血病细胞株(K562细胞)的杀伤活性,探讨NK细胞在SAA免疫发病机制中的作用.方法应用免疫磁珠分选法(MACS)分选2013年6月至2014年8月天津医科大学总医院血液科收治的12例SAA患者(SAA组)及10名健康人(健康对照组)外周血NK细胞(CD3-CD56+ CD16+)作为效应细胞,以K562细胞作为靶细胞,按照不同的效靶细胞比(1∶1、5∶1、10∶1)混合培养,培养后应用膜联蛋白V (Annexin-V)和碘化丙啶(PI)双标法标记细胞,流式细胞术检测K562细胞的凋亡率,并对两组结果进行比较.结果健康人与SAA患者外周血NK细胞分别与K562细胞以相同效靶比(1∶1)培养,SAA组K562细胞凋亡率(10.30％±6.08％)高于健康对照组(6.30％±3.46％),两者比较差异有统计学意义(P＜0.05);SAA患者NK细胞分别与K562细胞以不同效靶比(1∶1、5∶1、10∶1)共培养后,K562细胞凋亡率分别为(10.30％±6.08％、16.47％±8.29％、25.45％±9.88％),效靶比越高,K562细胞凋亡比例越高,各组比较差异有统计学意义(P＜0.05).结论 SAA患者外周血NK细胞体外杀伤功能增强,可能在SAA免疫耐受机制异常中发挥了重要作用.
목적 검측중형재생장애성빈혈(SAA)환자외주혈자연살상(NK)세포체외대만성수세포백혈병세포주(K562세포)적살상활성,탐토NK세포재SAA면역발병궤제중적작용.방법 응용면역자주분선법(MACS)분선2013년6월지2014년8월천진의과대학총의원혈액과수치적12례SAA환자(SAA조)급10명건강인(건강대조조)외주혈NK세포(CD3-CD56+ CD16+)작위효응세포,이K562세포작위파세포,안조불동적효파세포비(1∶1、5∶1、10∶1)혼합배양,배양후응용막련단백V (Annexin-V)화전화병정(PI)쌍표법표기세포,류식세포술검측K562세포적조망솔,병대량조결과진행비교.결과 건강인여SAA환자외주혈NK세포분별여K562세포이상동효파비(1∶1)배양,SAA조K562세포조망솔(10.30％±6.08％)고우건강대조조(6.30％±3.46％),량자비교차이유통계학의의(P＜0.05);SAA환자NK세포분별여K562세포이불동효파비(1∶1、5∶1、10∶1)공배양후,K562세포조망솔분별위(10.30％±6.08％、16.47％±8.29％、25.45％±9.88％),효파비월고,K562세포조망비례월고,각조비교차이유통계학의의(P＜0.05).결론 SAA환자외주혈NK세포체외살상공능증강,가능재SAA면역내수궤제이상중발휘료중요작용.
Objective To observe the in vitro killing functions of natural killer (NK) cells for K562 cells in peripheral blood of severe aplastic anemia (SAA) patients and explore the role of NK cells in the immunological pathogenesis of SAA.Methods NK cells (CD3-CD56 + CD16 +) as effector cells were sorted from peripheral blood mononuclear cells (PBMNC) of 12 SAA patients and 10 normal controls by magnetic activated cell sorting system (MACS)from June 2013 to August 2014.K562 cells as target cells were co-cultured with the effector cells (1 ∶ 1,1 ∶ 5,1 ∶ 10).Using Annexin V and PI double staining markers,the apoptotic rate of K562 ceils were measured by flow cytometry after culturing.Results The apoptotic rate of K562 cells in SAA patients was (10.30％ ± 6.08％) according to the ratio of effector-target (1 ∶ 1).And it was significantly higher than that in normal controls (6.30％ ± 3.46％) (P ＜ 0.05).The apoptotic rate of K562 cells in SAA patients were 10.30％ ± 6.08％,16.47％ ± 8.29％ and 25.45％ ± 9.88％ respectively according to the ratio of effector-target(1∶ 1,5∶ 1,10∶1).The higher ratio of effectortarget,the higher apoptotic rate of K562 cells.There were statistic differences among three groups (P ＜ 0.05).Conclusion There are enhanced in vitro functions of NK cells in peripheral blood of SAA patients NK cells may play an important role in abnormal immune tolerance mechanisms of SAA patients.