Replication of human cytomegalovirus in human retinal glial cells.
OBJECTIVE
The purpose of these studies was to characterize the replication cycle of human cytomegalovirus (HCMV) in human retinal glial cells in vitro.


METHODS
Cultured human retinal glial cells were exposed to HCMV strain AD169 or low-passage clinical isolates for a 2-hour adsorption period and then incubated in the appropriate growth medium at 37 degrees C. Cultures were examined by microscopy for cytopathic effect and by immunofluorescence staining using monoclonal antibodies directed against immediate-early, early, and late HCMV proteins. Viral DNA was analyzed by field inversion gel electrophoresis and detected using Southern blot analysis or the polymerase chain reaction.


RESULTS
Immunocytochemical staining revealed that the glial cells expressed all three classes of HCMV proteins and that infectious virus could be transferred from the medium of the infected cultures to susceptible MRC-5 cell monolayers. Less than 1% of the glial cells expressed the S-phase enzyme, thymidine kinase, at the time of infection compared to MRC-5 fibroblasts, of which 81% expressed it. Progeny virus was found to be highly cell associated in glial cells (80%) at peak virus titer compared to MRC-5 cells (39% cell associated at peak titer). Four low-passage clinical isolates of HCMV from patients with acquired immune deficiency virus also productively infected cultures of human retinal glial cells. Field inversion gel electrophoresis of HCMV-infected glial cell lysates was performed to identify the replicative forms of DNA. Southern blots probed with HCMV-specific probes showed that HCMV DNA replication proceeds through high molecular weight intermediates before forming the 230-kb unit length genome.


CONCLUSIONS
The full permissive replication of HCMV in human retinal glial cells indicates that glial cells are a likely site of HCMV replication in the retina and thus may play an important role in the pathogenesis of HCMV retinitis.
Burd
E M
EM
Department of Ophthalmology, Medical College of Wisconsin, Milwaukee 53226-4812, USA.
Pulido
J S
JS
Puro
D G
DG
O'Brien
W J
WJ
D000293:Adolescent; D000368:Aged; D000369:Aged, 80 and over; D000911:Antibodies, Monoclonal; D015139:Blotting, Southern; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D003588:Cytopathogenic Effect, Viral; D004261:DNA Replication / Q000502:physiology; D004279:DNA, Viral / Q000032:analysis; D004587:Electrophoresis, Agar Gel; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D008875:Middle Aged; D009457:Neuroglia / Q000821:virology*; D016133:Polymerase Chain Reaction; D012160:Retina / Q000166:cytology / Q000821:virology*; D014764:Viral Proteins / Q000032:analysis; D014779:Virus Replication / Q000502:physiology*
D000911:Antibodies, Monoclonal; D004279:DNA, Viral; D014764:Viral Proteins