Coronavirus (JHM) Replication within the Retina: Analysis of Cell Tropism in Mouse Retinal Cell Cultures

Abstract

The murine coronavirus, mouse hepatitis virus, JHM strain, induces a retinal degenerative disease in adult BALB/c mice. Coronavirus infections are highly species specific with virus exhibiting a strong tissue and cell specificity. In this report we evaluated the cellular basis of JHM virus retinal tropism. Retinal cultures and retinal pigment epithelial (RPE)-retinal mixed cell cultures were prepared from eyes obtained from Balb/c mice. The ability of JHM virus to infect and replicate in these retinal cultures was evaluated by light microscopy, immunofluorescent staining, electron microscopy, and virus isolation. Cytopathology was not observed and virus could not be detected in supernatant fluid in retinal cultures. However, low levels of infectious virus could be detected within the cells for the first 4 days. This observation suggested that cell-to-cell interactions may be critical since virus particles and virus antigens can be seen in vivo within the neural retina and the RPE. In contrast to the retinal cultures, retinal-RPE mixed cultures were supportive to JHM virus replication. Syncytial cytopathology was observed for the first 4 days and virus was isolated from supernatant fluids. By electron microscopy, virus was found intracellularly within vacuoles and extracellularly at the plasma membrane. After Day 4, a persistent virus infection was established in which cells produced virus for 5 weeks without cytopathic effects or cell death. Double-labeling immunofluorescent studies of retinal-RPE mixed cultures showed that the virus antigen was co-expressed with a Muller cell marker, glutamine synthetase. This cell is the most prominent glial element in the retina. These studies demonstrate that JHM virus is capable of establishing a persistent virus infection in mixed retinal (Muller)-RPE cell cultures. Moreover, these data suggest that cell-to-cell interactions influence the establishment of coronavirus infections in the retina.

The RPE Cell and the Immune System

“…In addition, the RPE cells were morphologically abnormal with foci of RPE cell swelling, proliferation and cell atrophy [67,68]. During this infection, we also noted that the RPE cell was the first cell infected by the virus [64,65,69,70]. This was not unexpected since the RPE cell serves as an excellent reservoir for numerous pathogens.…”

Infections Associated with Retinal Autoimmunity

“…This cell is also the first cell to express new viral antigens during infection in vivo and is persistently infected in vitro. 18 It is critically important to point out that this RPE cell has been shown to process and present retinal and non-retinal antigens to sensitized T cells, and it is up-regulated to express MHC class II molecules during retinal autoimmune and degenerative processes. 19,20…”

Infections Associated with Retinal Autoimmunity

“…It was noted that the first cell to express these MHC molecules was the RPE cell. This cell is also the first cell to express new viral antigens during the infection in vivo and is persistently infected in vitro [ 17]. It is critically important to point out that this RPE cell has been shown to process and present retinal and non-retinal antigens to sensitized T cells and is upregulated to express MHC class II molecules during retinal autoimmune and degenerative processes [ 18,19].…”