Establishment of the human papillomavirus type 16 (HPV-16) life cycle in an immortalized human foreskin keratinocyte cell line
Establishment of the human papillomavirus type 16 (HPV-16) life cycle in an immortalized human foreskin keratinocyte cell line
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DOI：
10.1006/viro.1999.9868
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发表时间：
1999-09-30
期刊：
影响因子：
3.7
通讯作者：
Lambert, PF
中科院分区：
文献类型：
--
作者：
Flores, ER;Allen-Hoffmann, BL;Lambert, PF
人乳头瘤病毒（HPV）在细胞培养中的研究一直受到阻碍，因为难以重建病毒赖以完成其生命周期的上皮的三维结构。此外，由于早期传代人包皮角质形成细胞（HFK）的寿命有限，使用目前的将分子克隆的HPV基因组转染到早期传代人包皮角质形成细胞（HFK）中的方法难以研究HPV基因组中的遗传突变及其对病毒生命周期的影响。除非将HPV基因组转染到早期传代的HFK中延长细胞的寿命，否则稳定转染子的分析变得困难。在这项研究中，我们使用BC-1-Ep/SL细胞，一种永生化的人包皮角质形成细胞系，来重建HPV-16的生命周期。该细胞系表现出早期传代HFKs的许多特征，包括在器官型筏培养系统中分层和终末分化的能力。由于它们与早期传代的HFKs相似，这些细胞被测试了它们支持HPV-16生命周期的能力。BC 1-Ep/SL细胞可稳定维持HPV-16和HPV-31 b两种基因型。此外，当用HPV-16稳定转染BC-1-Ep/SL细胞系并使用器官型筏培养系统（筏）培养时，其维持HPV-16生命周期。HPV-16生命周期的生产阶段的证据提供了：DNA原位杂交证明HPV-16 DNA扩增的筏，L1的免疫组化染色显示存在的衣壳蛋白在筏的基底层，和电子显微镜表明存在的病毒样颗粒（VLP）在细胞核中的筏的分化层。(C)北京：科学出版社.
The study of human papillomaviruses (HPVs) in cell culture has been hindered because of the difficulty in recreating the three-dimensional structure of the epithelium on which the Virus depends to complete its life cycle. Additionally the study of genetic mutations in the HPV genome and its effects on the viral life cycle are difficult using the current method of transfecting molecularly cloned HPV genomes into early-passage human foreskin keratinocytes (HFKs) because of the limited life span of these cells. Unless the HPV genome transfected into the early-passage HFK extends the life span of the cell, analysis of stable transfectants becomes difficult In this study, we have used BC-1-Ep/SL cells, an immortalized human foreskin keratinocyte cell line, to recreate the HPV-16 life cycle. This cell line exhibits many characteristics of the early-passage HFKs including the ability to stratify and terminally differentiate in an organotypic raft culture system. Because of their similarity to early-passage HFKs, these cells were tested for their ability to support the HPV-16 life cycle. The BC1-Ep/SL cells could stably maintain two HPV genotypes, HPV-16 and HPV-31b, episomally. Additionally, when the BC-1-Ep/SL cell line was stably transfected with HPV-16 and cultured using the organotypic raft culture system (rafts), it sustained the HPV-16 life cycle. Evidence for the productive stage of the HPV-16 life cycle was provided by: DNA in situ hybridization demonstrating HPV-16 DNA amplification in the suprabasal layers of the rafts, immunohistochemical staining for L1 showing the presence of capsid protein in the suprabasal layers of the rafts, and electron microscopy indicating the presence of virus like particles (VLPs) in nuclei from cells in the differentiated layers of the rafts. (C) 1999 Academic Press.