TLR4 in Facilitating HSV-1 Neuronal Spread in Experimental Acute Retinal Necrosis (ARN) Model
M Zheng, MA Fields, Y Liu, HM Cathcart, SS Atherton. Medical College of Georgia, Augusta, GA
Background: ARN is a rare disease usually caused by neurotropic human HSV-1. TLR4 is an innate immune mediator against a variety of pathogens, especially bacteria. Recent studies pointed toward TLR4 playing a role in a growing list of virus induced diseases.
Design: Both TLR4 mutant (mu) and wild type (wt) mice were infected with HSV-1 via anterior chamber (AC) or intravitreal inoculation. At different times post infection (p.i.), mice were sacrificed and the inoculated eyes, visual pathway containing optic nerve, chiasm, tract, and brain tissues containing the superior colliculus (SC) and lateral geniculate nucleus (LGN) were isolated. Immunohistochemistry (IHC), RT PCR, flow cytometry (FC) and virus titration were performed on the samples.
Results: Our previous results showed that after HSV-1 AC inoculation, few uninoculated, contralateral eyes of mu mice developed ARN, while ARN was observed in the majority of uninoculated eyes of wt mice. Further study revealed that an increase in certain cytokine responses at day 3 and 7 p.i. in the HSV-1 injected eye of mu compared with wt mice. FC showed more cells were infected by HSV-1 in mu compared with wt mice in virus inoculated ocular cells at day 4 p.i. One step growth curve in retinal pigment epithelial cells isolated from either mu or wt mice showed that the replication kinetics of HSV-1 were similar in both. To further investigate the role of TLR4 in virus propagation in neuronal cells within the visual pathway, HSV-1 was injected intravitreally. IHC demonstrated that HSV-1 had infected the retinal ganglion cells, then the inner nuclear cell layer cells in wt mice, whereas in mu mice, fewer cells of the subjacent retina were infected at day 2 p.i. Most HSV-1 + cells showed distinctive beta III-tubulin staining. The optic nerve was HSV-1 + at day 2 p.i. in wt mice whereas in mu mice, at day 4 p.i. The arrival of virus in the optic chiasm, tract, SC, and LGN was similarly delayed in mu compared with wt mice. IHC revealed that TLR4 was in close association with beta III-tubulin in HSV-1 infected rat retinal ganglion cells.
Conclusions: Although absence of TLR4 leads more cells to be infected by HSV-1 in virus inoculated AC, lack/delay of neuronal spread after AC or intravitreal injection of HSV-1 in mu mice suggests that TLR4 may play a role in facilitating HSV-1 transport along microtubules within the visual pathway.
Wednesday, March 24, 2010 1:00 PM
Poster Session VI # 216, Wednesday Afternoon