Shioda Setsuko, Kasai Fumio, Ozawa Midori, Hirayama Noriko, Satoh Motonobu, Kameoka Yousuke, Watanabe Ken, Shimizu Norio, Tang Huamin, Mori Yasuko, Kohara Arihiro
Japanese Collection of Research Bioresources (JCRB) Cell Bank, Laboratory of Cell Cultures, National Institutes of Biomedical Innovation, Health and Nutrition, 7-6-8 Saito-Asagi, Ibaraki, Osaka, 567-0085, Japan.
A-CLIP Institute, Chiba, Japan.
Cytotechnology. 2018 Feb;70(1):141-152. doi: 10.1007/s10616-017-0119-y. Epub 2017 Jul 28.
Human herpes virus 6 (HHV-6) is a common human pathogen that is most often detected in hematopoietic cells. Although human cells harboring chromosomally integrated HHV-6 can be generated in vitro, the availability of such cell lines originating from in vivo tissues is limited. In this study, chromosomally integrated HHV-6B has been identified in a human vascular endothelial cell line, HUV-EC-C (IFO50271), derived from normal umbilical cord tissue. Sequence analysis revealed that the viral genome was similar to the HHV-6B HST strain. FISH analysis using a HHV-6 DNA probe showed one signal in each cell, detected at the distal end of the long arm of chromosome 9. This was consistent with a digital PCR assay, validating one copy of the viral DNA. Because exposure of HUV-EC-C to chemicals did not cause viral reactivation, long term cell culture of HUV-EC-C was carried out to assess the stability of viral integration. The growth rate was altered depending on passage numbers, and morphology also changed during culture. SNP microarray profiles showed some differences between low and high passages, implying that the HUV-EC-C genome had changed during culture. However, no detectable change was observed in chromosome 9, where HHV-6B integration and the viral copy number remained unchanged. Our results suggest that integrated HHV-6B is stable in HUV-EC-C despite genome instability.
人类疱疹病毒6型(HHV-6)是一种常见的人类病原体,最常在造血细胞中被检测到。虽然携带染色体整合型HHV-6的人类细胞可以在体外产生,但源自体内组织的此类细胞系的可用性有限。在本研究中,在源自正常脐带组织的人血管内皮细胞系HUV-EC-C(IFO50271)中鉴定出了染色体整合型HHV-6B。序列分析表明,病毒基因组与HHV-6B HST株相似。使用HHV-6 DNA探针的荧光原位杂交(FISH)分析显示每个细胞中有一个信号,在9号染色体长臂的远端检测到。这与数字PCR分析结果一致,证实了病毒DNA的一个拷贝。由于HUV-EC-C暴露于化学物质中不会导致病毒重新激活,因此对HUV-EC-C进行了长期细胞培养以评估病毒整合的稳定性。生长速率根据传代次数而改变,并且在培养过程中形态也发生了变化。单核苷酸多态性(SNP)微阵列图谱显示低代和高代之间存在一些差异,这意味着HUV-EC-C基因组在培养过程中发生了变化。然而,在9号染色体上未观察到可检测到的变化,HHV-6B的整合和病毒拷贝数保持不变。我们的结果表明,尽管基因组不稳定,但整合型HHV-6B在HUV-EC-C中是稳定的。