Epstein-Barr virus infection of human natural killer cell lines and peripheral blood natural killer cells
Epstein-Barr virus infection of human natural killer cell lines and peripheral blood natural killer cells
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DOI：
10.1158/0008-5472.can-03-1562
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发表时间：
2004-03-15
期刊：
影响因子：
11.2
通讯作者：
Oshimi, K
中科院分区：
文献类型：
--
作者：
Isobe, Y;Sugimoto, K;Oshimi, K
尽管相当一部分自然杀伤（NK）细胞肿瘤具有EBV基因组，但目前还没有直接证据表明EBV在体外感染人类NK细胞。在这项研究中，我们使用重组 EBV 证明了 EBV 能够进入 NK 细胞，该重组 EBV 的基因组中含有增强型绿色荧光蛋白 (EGFP) 基因 (EGFP-EBV)。暴露于 EGFP-EBV 48 小时后，我们在三名健康志愿者的 30% 的 NK-92 和 NKL 细胞以及 > 40% 的外周血 NK 细胞中检测到了类似的 EGFP 信号。对各种 EBV 相关基因的逆转录 PCR 分析证实了 EBV 感染。 EBER 和 BHLF 的原位杂交表明，两种 NK 细胞系在 EBV 感染的早期阶段潜伏性感染和裂解性感染同时存在。虽然早期裂解期的 BHLF 阳性细胞呈圆形，但潜伏 EBV 感染的 EBER 阳性细胞往往呈现出奇怪的形状。对EGFP-EBV暴露的NK细胞系的流式细胞术分析表明，大多数EBV感染的细胞在EBV暴露72小时后进入早期凋亡，这解释了建立携带EBV的NK克隆的困难。流式细胞术和逆转录-PCR 分析表明，两种 NK 细胞系通过与 CD21 不同的分子与病毒结合后，可能会使用 HLA II 类与 EBV 融合。我们建立了两个携带 EBV 的 NKL 克隆，显示潜伏期 I 型和 II 型，这两种克隆都在 EBV 相关 NK 细胞肿瘤中被识别。由于 EBV 感染的 NKL 细胞在感染早期仅表现出 I 型潜伏期，因此潜伏基因表达的时间特征与 T 细胞相似。我们首先报道了体外 EBV 感染人类 NK 细胞并建立了携带 EBV 的 NK 克隆，这将有助于阐明 EBV 在 NK 细胞肿瘤发展中的作用。
Although considerable part of natural killer (NK) cell neoplasms possess EBV genome, there has been no direct evidence that EBV infects human NK cells in vitro. In this study, we demonstrated EBV entry into NK cells using a recombinant EBV, which contains enhanced green fluorescent protein (EGFP) gene in its genome (EGFP-EBV). After 48 h of exposure to EGFP-EBV, we detected EGFP signals in similar to30% of NK-92 and NKL cells and >40% of peripheral blood NK cells from three healthy volunteers. Reverse transcription-PCR analysis of various EBV-associated genes confirmed EBV infection. In situ hybridization for EBERs and BHLFs showed that latent and lytic infections coexisted at the early phase of EBV infection in two NK cell lines. Although BHLF-positive cells in the early lytic phase were round-shaped, EBER-positive cells in latent EBV infection tended to show a bizarre shape. Flow cytometric analysis of EGFP-EBV-exposed NK cell lines showed that most of EBV-infected cells entered early apoptosis after 72 h of EBV exposure, which explains the difficulties to establish EBV-carrying NK clones. Flow cytometry and reverse transcription-PCR analysis indicated that two NK cell lines may fuse with EBV using HLA class II after binding to the virus through a distinct molecule from CD21. We established two EBV-carrying NKL clones showing latency types I and II, both of which are recognized in EBV-associated NK cell neoplasms. Because EBV-infected NKL cells showed only type I latency during the early phase of infection, the temporal profile of latent gene expression is similar to that of T cells. We first report in vitro EBV infection of human NK cells and establishment of EBV-carrying NK clones, which should contribute to elucidate the role of EBV in the development of NK cell neoplasms.