Analysis of human cytomegalovirus replication in primary cultured human corneal endothelial cells.
Analysis of human cytomegalovirus replication in primary cultured human corneal endothelial cells.
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DOI：
10.1136/bjophthalmol-2014-306486
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发表时间：
2015-11
期刊：
影响因子：
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通讯作者：
Isomura H
中科院分区：
文献类型：
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作者：
Hosogai M;Shima N;Nakatani Y;Inoue T;Iso T;Yokoo H;Yorifuji H;Akiyama H;Kishi S;Isomura H
自首例人巨细胞病毒（HCMV）诱导的角膜内皮炎病例使用PCR从患者体液中检测到HCMV DNA以来，HCMV内皮炎的临床证据一直在积累。然而，HCMV能否在角膜内皮细胞中高效复制仍有待证实。因此，我们试图确定原代培养的人角膜内皮细胞（HCECs）是否支持HCMV复制。人包皮成纤维细胞（HFFs）已被证明完全允许HCMV复制，并且通常被用作HCMV裂解复制的体外模型。因此，原代培养的HCECs或HFFs感染血管内皮性HCMV菌株TB40/E或实验室菌株Towne。然后，我们比较了TB40/ e感染和towne感染的HCECs和HFFs之间的病毒mRNA和蛋白表达、基因组复制和生长。当HCECs感染TB40/E或Towne时，检测到类似猫头鹰眼睛的圆形细胞和病毒抗原。在感染TB40/E或Towne的HCECs和HFFs中，病毒mRNA的合成和蛋白的表达在高感染多重性（MOI）下仍能有效进行。同样，病毒基因组也被有效复制，在HCECs细胞核中观察到ul44 -一种病毒DNA聚合酶加工因子灶。在高MOI和低MOI条件下，HCECs感染TB40/E后均产生大量感染性病毒粒子。在高MOI和低MOI条件下，原代培养的HCECs都能有效地支持HCMV感染后的复制。
Since the first case of human cytomegalovirus (HCMV)-induced corneal endotheliitis in which HCMV DNA was detected from the patient's aqueous humour using PCR, the clinical evidence for HCMV endotheliitis has been accumulating. However, it remains to be confirmed whether HCMV can efficiently replicate in corneal endothelial cells. We, therefore, sought to determine whether primary cultured human corneal endothelial cells (HCECs) could support HCMV replication. Human foreskin fibroblasts (HFFs) have been shown to be fully permissive for HCMV replication, and are commonly used as an in vitro model for HCMV lytic replication. Therefore, primary cultured HCECs or HFFs were infected with the vascular endotheliotropic HCMV strain TB40/E or laboratory strain Towne. We then compared viral mRNA and protein expression, genome replication and growth between the TB40/E-infected and Towne-infected HCECs and HFFs. When HCECs were infected with TB40/E or Towne, rounded cells resembling owl's eyes as well as viral antigens were detected. Viral mRNA synthesis and protein expression proceeded efficiently in the HCECs and HFFs infected with TB40/E or Towne at a high multiplicity of infection (MOI). Similarly, the viral genome was also effectively replicated, with UL44—a viral DNA polymerase processivity factor—foci observed in the nuclei of HCECs. HCECs produced a substantial number of infectious virions after infection with TB40/E at both a high and low MOI. Primary cultured HCECs could efficiently support HCMV replication after infection at both a high and low MOI.