Availability of NK cell expansion agent combined with recombinant IL-2 and IL-15 stimulation on the expansion and high-purity of NK cells in patients with immune-related pancytopenia in vitro
Availability of NK cell expansion agent combined with recombinant IL-2 and IL-15 stimulation on the expansion and high-purity of NK cells in patients with immune-related pancytopenia in vitro
复制标题
NK细胞扩增剂联合重组IL-2和IL-15刺激对免疫相关性全血细胞减少症患者NK细胞体外扩增和高纯度的有效性
DOI：
10.3892/mmr.2019.10654
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发表时间：
2019-11-01
影响因子：
3.4
通讯作者：
Fu, Rong
中科院分区：
文献类型：
--
作者：
Li, Yang;Liu, Bingnan;Fu, Rong
自然杀伤（NK）细胞是一组大颗粒淋巴细胞，在先天免疫和适应性免疫中起重要调节作用。免疫相关性全血细胞减少症（Immune-related pancytoproteinesis，IRP）是由于骨髓造血细胞被自身抗体破坏或抑制而引起的全血细胞减少症。IRP的具体机制尚不清楚。在本研究中，它被确定为外周血淋巴细胞中的NK细胞的百分比降低，在IRP患者。采用免疫磁珠分选法、磁激活细胞分选法从6例未经治疗的IRP患者中提取高纯度的NK细胞，培养于含20%FBS的RPMI-1640培养基中。在重组IL-2的基础上，使用含或不含重组白细胞介素（IL）-15的NK细胞扩增剂扩增高纯度NK细胞。流式细胞仪检测培养前后CD 56（+）NK细胞活化受体NKG 2-D II型膜整合蛋白（NKG 2D）、自然杀伤细胞p46相关蛋白（NKp 46）和抑制性受体CD 158 a、NKG 2-A/NKG 2-B II型膜整合蛋白（NKG 2A）的表达。观察到用NK细胞扩增剂与重组IL-2和重组IL-15的刺激组合的处理可以增加NK细胞的数量，同时保持NK细胞的纯度。IRP患者NK细胞培养前后NKG 2D、NKp 46、NKG 2A和CD 158 a的表达无明显变化。这种新的扩增方法为临床NK细胞免疫治疗和抗肿瘤应用奠定了基础。
Natural killer (NK) cells are a group of large granular lymphocytes that play an important regulatory role in innate immunity and adaptive immunity. Immune-related pancytopenia (IRP) is a type of pancytopenia resulting from bone marrow hematopoietic cells that were destroyed or suppressed by auto-antibodies. The specific mechanism of IRP is not clear. In the present study, it was identified that the percentage of NK cells in peripheral blood lymphocytes was decreased in patients with IRP. Subsequently, high purity NK cells were extracted from 6 untreated patients with IRP using the immunomagnetic beads sorting, magnetic-activated cell-sorting method, which were then cultured then in RPMI-1640 medium containing 20% FBS. NK cell expansion agents, with or without recombinant interleukin (IL)-15, were used to amplify high-purity NK cells on the basic of recombinant IL-2. Expression of the activated receptors NKG2-D type II integral membrane protein (NKG2D) and natural killer cell p46-related protein (NKp46), and the inhibitory receptors CD158a and NKG2-A/NKG2-B type II integral membrane protein (NKG2A), in CD56(+) NK cells were detected by flow cytometry before and after cell culture. It was observed that treatment with an NK cell expansion agent combined with the stimulation of recombinant IL-2 and recombinant IL-15 could increase the number whilst maintaining the purity of NK cells. There were no significant changes in the expression of NKG2D, NKp46, NKG2A and CD158a in patients with IRP before and after NK cell culture. This new amplification method lays a foundation for clinical NK cell immunotherapy and anti-tumor applications.