After immunosuppression with methylprednisolone, which depletes T cells as well as macrophages,
19,21 and inoculation of MCMV into the supraciliary space, virus-infected cells were observed in the anterior segment and the RPE layer of both groups of mice at day 3 p.i. (
Fig. 1). At this time, although the titer of MCMV was higher in the injected eyes of TNF-α
−/− mice (average PFU log
10 ± SEM: 3.16 ± 0.15) than in the injected eyes of wild-type mice (2.39 ± 0.09), the difference was not significant. At day 6 p.i., a similar number of virus-infected cells was noted mainly in the inner retina of the injected eyes in both animal models (
Fig. 1). At this time, the difference in virus titer between TNF-α
−/− mice and wild-type mice was also not significant (average PFU log
10 ± SEM: 3.85 ± 0.38 vs. 3.49 ± 0.17, respectively). Apoptotic cells were located in the retina of the injected eye in both TNF-α
−/− mice and wild-type mice. Most TUNEL-positive cells were uninfected. At day 3 p.i., most apoptotic cells were uninfected photoreceptor cells, whereas virus-infected cells were observed mainly in the choroid and RPE. At day 6 p.i., apoptotic cells were present in all layers of the retina, and virus-infected cells were also observed throughout the retina. However, more apoptotic cells were always observed in the retinas of MCMV-infected TNF-α
−/− mice than in the retinas of IS wild-type mice (
Fig. 1). To augment the light microscopy studies, sections of injected eyes were examined by electron microscopy. Virus particles (
Fig. 2) and apoptotic cells (
Fig. 3) were observed in the inner retina of TNF-α
−/− and wild-type mice at day 6 p.i. However, significantly more apoptotic cells were observed in the TNF-α
−/− mice than in the wild-type mice (29.71 ± 27.12 vs. 8.14 ± 4.19;
n = 14;
P < 0.01) in all layers of the inner retina, including photoreceptors (
Fig. 3A), inner nuclear layer (
Fig. 3B), and ganglion cell layer (
Fig. 3C). The apoptotic retinal cells exhibited nuclear shrinkage and strong chromatin condensation.
22 Later stages of apoptosis, characterized by more homogeneous chromatin condensation and increased cell shrinkage, were also observed. Advanced chromatin compaction and nuclear fragmentation
23 were occasionally seen in apoptotic cells in both TNF-α
−/− mice and wild-type mice (
Fig. 3).