乏氧对人外周血NK细胞NKG2A、NKG2D及CD44表达的影响

目的： 观察乏氧微环境对人外周血自然杀伤细胞(NK)表面自然杀伤细胞2族成员A(NKG2A)、自然杀伤细胞2族成员D(NKG2D)及CD44分子表达的影响，探讨乏氧抑制NK细胞杀伤活性的分子机制。方法: 采用密度梯度离心法分离健康人外周血单个核细胞（PBMC），贴壁去除单核细胞获得外周血淋巴细胞（PBL），分别置常氧（21％O2）、乏氧（1％O2）以及有或无人重组白细胞介素2(rhIL-2)（1×106 U/L）刺激条件下培养16 h，流式细胞术（FCM）检测不同 NK细胞亚群 NKG2A、NKG2D以及CD44分子的表达。结果: 常氧条件，人外周血CD3-CD56+NK细胞NKG2A、NKG2D表达的阳性率分别为16.16%和78.45%，乏氧条件下二者表达的阳性率分别为15.16%和71.08%；rhIL-2上调NKG2A和NKG2D的表达，乏氧不影响 rhIL-2对NKG2D、 NKG2A的上调作用；rhIL-2显著上调NK细胞CD44的表达，乏氧抑制CD44的表达（P<0.05）。结论: 乏氧下调外周血NK细胞表面受体NKG2D及CD44的表达，但对NKG2A的表达无显著影响。由此提示，NKG2D及CD44分子可能在乏氧引起的NK细胞杀伤活性抑制中具有重要作用。

Effects of hypoxia on the expression of NKG2A, NKG2D and CD44 by human PBMC derived natural killer cells

AIM: To examine the surface expressions of natural killer group 2, member A, D (NKG2A, NKG2D) and CD44 on human peripheral blood mononuclear cells (PBMC) derived nature killer (NK) cells under different oxygen conditions and to investigate the molecular mechanisms of hypoxia inhibiting the cytotoxicity of NK. METHODS: The PBMC from healthy people were isolated by density gradient centrifugation and the peripheral blood lymphocytes (PBL) were acquired through removing the nonadherent cells. The PBL under different oxygen tension were cultured with or without recombinant human interleukin 2 (rhIL-2) for 16 h, then the expressions of NKG2A and NKG2D on the surface of CD3-CD56＋NK subset and CD44 on CD56+CD16+ NK subset respectively were examined by flow cytometry.RESULTS: The NKG2A and NKG2D expressions under normoxia condition were 16.16% and 78.45%, respectively. Hypoxia down-regulated the expression of NKG2D (P<0.05) but did not change the level of NKG2A. The effect of hypoxia on the expressions of NKG2A and NKG2D was not found with addition of rhIL-2. All the human PBMC derived NK cells highly expressed CD44, and rhIL-2 significantly up-regulated CD44 expression on CD56+CD16+ NK subset. Hypoxia inhibited the expression of CD44 (P<0.05). CONCLUSION: Hypoxia inhibits the expression of NKG2D and CD44 on PBMC derived NKs while have no effect on the expression of NKG2A, indicating the important roles of NKG2D and CD44 in the hypoxia induced inhibition of NKs cytotoxicity.