Purchase this article with an account.
S. S. Atherton, M. Zhang; Herpes Simplex Virus Type 1 (HSV-1) Infection and Autophagy in Organotypic Retinal Cultures. Invest. Ophthalmol. Vis. Sci. 2010;51(13):3811.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Organotypic retinal cultures were used to determine the pattern of HSV-1 infection and whether autophagy is induced during HSV-1 retinal infection.
The retina from a C57BL/6 mouse was attached to a filter which was then mounted on a coverslip with a drop of Matrigel. The coverslip with attached retina was inserted in a culture tube in 1 ml of culture medium and incubated at 37°C. Some retinas were infected with HSV-1 and some were also treated with cloroquine. At 12, 24, 48, and 72 hrs p.i., the retinal cultures were collected and prepared for examination by EM, for double staining for retinal cell antigens and HSV-1 antigens, or for western blot for protein light chain 3-I (LC3-I) and LC3-II.
Replicating virus was recovered and viral antigens or viral particles were observed in all layers of the HSV-1 infected retinal cultures. Similar to the observation in animal model, ganglion cells were the first targets of HSV-1 infection and virus then gradually spread to the inner nuclear layer and the outer nuclear layer. HSV-1 infected ganglion cells, glia, GABA+ or glycine+ amacrine cells, horizontal cells, bipolar cells and photoreceptors. EM examination revealed not only apoptotic cells, but also the presence of more autophagosomes and autolysosomes in HSV-1 infected retinas. When infected cultured retinas were treated with cloroquine, an increase in the amount of LC3-II was observed due to an increase in the number of autophagosomes that were unable to fuse with lysosomes to form autolysosomes. Interestingly however, this increase correlated with reduced virus titer (average ± SEM: 3.68 × 105 ± 2.28 × 105 vs. 3.53 × 104 ± 3.12 × 104, untreated vs. chloroquine treated).
The pattern of HSV-1 infection of cultured retinas appears to be similar to that observed in the HSV-1 infected retina of the uninjected contralateral eye after uniocular anterior chamber injection of HSV-1. The observation that reduced autophagy in chloroquine treated cultures correlated with an increase in apoptosis and a decrease in virus titer suggests that if apoptosis is increased, there are fewer retinal cells that are available to be infected with/replicate virus. Although the precise role of autophagy in the pathogenesis of HSV-1 infection of the retina remains to be deciphered, the results of these studies suggest that both autophagy and apoptosis play a role.
This PDF is available to Subscribers Only