Analysis of human cytomegalovirus replication in primary cultured human corneal endothelial cells.
OBJECTIVE
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.


METHODS
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.


RESULTS
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.


CONCLUSIONS
Primary cultured HCECs could efficiently support HCMV replication after infection at both a high and low MOI.
Department of Virology and Preventive Medicine, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan Department of Ophthalmology, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Mayumi
M
Hosogai
Department of Ophthalmology, University of Tokyo Hospital, Tokyo, Japan.
Nobuyuki
N
Shima
Department of Virology and Preventive Medicine, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Yoko
Y
Nakatani
Department of Virology and Preventive Medicine, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan Department of Medicine and Molecular Science, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Teruki
T
Inoue
Department of Medicine and Biological Science, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Tatsuya
T
Iso
Department of Human Pathology, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Hideaki
H
Yokoo
Department of Anatomy, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Hiroshi
H
Yorifuji
Department of Ophthalmology, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Hideo
H
Akiyama
Department of Ophthalmology, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Shoji
S
Kishi
Department of Virology and Preventive Medicine, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan.
Hiroki
H
Isomura
D000956:Antigens, Viral / Q000235:genetics; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000502:physiology*; D004279:DNA, Viral / Q000235:genetics; D004728:Endothelium, Corneal / Q000648:ultrastructure / Q000821:virology*; D005347:Fibroblasts / Q000821:virology; D052816:Foreskin / Q000166:cytology; D061307:Human Umbilical Vein Endothelial Cells / Q000166:cytology / Q000821:virology; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics; D015151:Immunoblotting; D008297:Male; D012333:RNA, Messenger / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D014763:Viral Matrix Proteins / Q000235:genetics; D014779:Virus Replication / Q000502:physiology*
D000956:Antigens, Viral; D004279:DNA, Viral; D017874:Immediate-Early Proteins; D012333:RNA, Messenger; D012367:RNA, Viral; D014763:Viral Matrix Proteins; C044582:immediate-early proteins, cytomegalovirus
Anterior chamber; Aqueous humour; Cornea; Experimental &#8211 laboratory; Infection