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腺病毒E1A/E1B转化的羊水细胞支持人巨细胞病毒复制。

Adenovirus E1A/E1B Transformed Amniotic Fluid Cells Support Human Cytomegalovirus Replication.

作者信息

Krömmelbein Natascha, Wiebusch Lüder, Schiedner Gudrun, Büscher Nicole, Sauer Caroline, Florin Luise, Sehn Elisabeth, Wolfrum Uwe, Plachter Bodo

机构信息

Institute for Virology, University Medical Center of the Johannes Gutenberg-University Mainz, D-55131 Mainz, Germany.

Department of Pediatric Molecular Biology, Charité University Medical Centre Berlin, D-10117 Berlin, Germany.

出版信息

Viruses. 2016 Feb 2;8(2):37. doi: 10.3390/v8020037.

Abstract

The human cytomegalovirus (HCMV) replicates to high titers in primary human fibroblast cell cultures. A variety of primary human cells and some tumor-derived cell lines do also support permissive HCMV replication, yet at low levels. Cell lines established by transfection of the transforming functions of adenoviruses have been notoriously resistant to HCMV replication and progeny production. Here, we provide first-time evidence that a permanent cell line immortalized by adenovirus type 5 E1A and E1B (CAP) is supporting the full HCMV replication cycle and is releasing infectious progeny. The CAP cell line had previously been established from amniotic fluid cells which were likely derived from membranes of the developing fetus. These cells can be grown under serum-free conditions. HCMV efficiently penetrated CAP cells, expressed its immediate-early proteins and dispersed restrictive PML-bodies. Viral DNA replication was initiated and viral progeny became detectable by electron microscopy in CAP cells. Furthermore, infectious virus was released from CAP cells, yet to lower levels compared to fibroblasts. Subviral dense bodies were also secreted from CAP cells. The results show that E1A/E1B expression in transformed cells is not generally repressive to HCMV replication and that CAP cells may be a good substrate for dense body based vaccine production.

摘要

人巨细胞病毒(HCMV)在原代人成纤维细胞培养物中可复制至高滴度。多种原代人细胞和一些肿瘤衍生细胞系也确实支持HCMV的允许性复制,不过水平较低。通过转染腺病毒转化功能而建立的细胞系一直以来对HCMV复制和子代产生具有抗性。在此,我们首次提供证据表明,由5型腺病毒E1A和E1B永生化的永久细胞系(CAP)支持完整的HCMV复制周期并释放有感染性的子代。CAP细胞系先前是从羊水细胞建立的,这些羊水细胞可能源自发育中胎儿的胎膜。这些细胞可以在无血清条件下生长。HCMV能有效穿透CAP细胞,表达其即刻早期蛋白并分散限制性的早幼粒细胞白血病(PML)小体。病毒DNA复制启动,并且通过电子显微镜在CAP细胞中可检测到病毒子代。此外,有感染性的病毒从CAP细胞中释放出来,但与成纤维细胞相比水平较低。亚病毒致密体也从CAP细胞中分泌出来。结果表明,转化细胞中E1A/E1B的表达通常不会抑制HCMV复制,并且CAP细胞可能是基于致密体的疫苗生产的良好底物。

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