Vos Q, Ortaldo J R, Conan-Cibotti M, Vos M D, Young H A, Anderson S K, Witherspoon K, Prager I, Snapper C M, Mond J J
Department of Medicine, Uniformed Services University of the Health Sciences, Bethesda, MD 20814, USA.
Int Immunol. 1998 Aug;10(8):1093-101. doi: 10.1093/intimm/10.8.1093.
NK cells not only function as cytotoxic effector cells, but also have immunoregulatory roles including the enhancement of Ig secretion. To have a stable and uniform population of NK cells to study their role in Ig secretion, we generated murine NK clones. Thus, culture of splenocytes from mice that were homozygous for a mutation in the p53 tumor suppressor gene (p53-KO) with IL-2 and poly(IC) resulted in a long-term NK line, from which four stable clones were derived. This approach also yielded a long-term NK line from splenocytes of normal C57BL/6 mice. Identification of the clones as members of the NK lineage was based on large granular morphology, expression of NK-TR and absence of TCR gene rearrangement. Flow cytometry revealed that all clones expressed IL-2R alpha and beta, chains and B220, but no CD3, NK1.1, DX5 or Ly-49. RT-PCR analysis showed heterogeneity in NK1.1 gene expression, and demonstrated expression of perforin and several granzymes in all clones. Three out of four clones lysed YAC-1, but not P815 target cells, corresponding to a pattern of NK specificity. All NK clones enhanced Ig secretion in an in vitro model for T cell-independent type 2 antigens, albeit to varying degrees. We found no correlation between the degree of helper activity of the NK clones and the level of their cytotoxic activity on YAC-1 targets. Thus, we established murine NK clones, and show that they mediate both cytotoxicity and enhancement of Ig secretion.
自然杀伤（NK）细胞不仅作为细胞毒性效应细胞发挥作用，还具有免疫调节作用，包括增强免疫球蛋白（Ig）分泌。为了获得稳定且均一的NK细胞群体以研究其在Ig分泌中的作用，我们构建了小鼠NK克隆。因此，用白细胞介素-2（IL-2）和聚肌胞苷酸（poly(IC)）培养p53肿瘤抑制基因纯合突变小鼠（p53基因敲除小鼠，p53-KO）的脾细胞，得到了一个长期存活的NK细胞系，并从中衍生出四个稳定的克隆。这种方法也从正常C57BL/6小鼠的脾细胞中得到了一个长期存活的NK细胞系。根据大颗粒形态、NK-TR的表达以及T细胞受体（TCR）基因重排的缺失，将这些克隆鉴定为NK细胞谱系的成员。流式细胞术显示，所有克隆均表达IL-2Rα链和β链以及B220，但不表达CD3、NK1.1、DX5或Ly-49。逆转录聚合酶链反应（RT-PCR）分析显示NK1.1基因表达存在异质性，并证明所有克隆均表达穿孔素和几种颗粒酶。四个克隆中有三个能裂解YAC-1细胞，但不能裂解P815靶细胞，这与NK细胞的特异性模式相符。在一个针对2型非T细胞依赖性抗原的体外模型中，所有NK克隆均能增强Ig分泌，尽管程度有所不同。我们发现NK克隆的辅助活性程度与其对YAC-1靶细胞的细胞毒性活性水平之间没有相关性。因此，我们构建了小鼠NK克隆，并表明它们既能介导细胞毒性，又能增强Ig分泌。
