Abstract
Purpose: :
To determine the contribution of resident versus hematopoietic-derived cells to lymphatic vessel integrity in response to herpes simplex virus type 1 (HSV-1) infection.
Methods: :
Male C57BL/6 (wild type, WT), WT mice deficient in the type I interferon receptor A chain (CD118-/-) and WT/CD118-/- mouse chimeras were infected with HSV-1 (1,000 plaque forming units/scarified cornea). Mice were exsanguinated at times post infection (PI), and the corneas were evaluated for lymphangiogenesis (LYVE-1+), hemangiogenesis (CD31+), and VEGFR2 expression by confocal microscopy, VEGF A production by ELISA, and viral titer by plaque assay. In another group of mice, an HSV-1 vesicle-induced model was developed to determine the relationship between vesicle genesis and lymphatic/blood vessel continuity. Tissue was analyzed at times before and after vesicle development. Images were quantified by Metavue imaging software. Data was analyzed by ANOVA and Tukey’s post hoc T test for significance (p<.05).
Results: :
As previously reported, lymphatic and blood vessels grew into the cornea proper in a time-specific manner in WT mice in response to HSV-1 infection. In contrast, there was an initial robust growth of lymphatic vessels into the cornea of HSV-1-infected CD118-/- mice but such vessels disappeared by day 5 PI. VEGF-A levels and VEGFR2 expression were similar between WT and CD118-/- mice. The loss was selective for lymphatic vessels as blood vessel integrity remained intact in the infected CD118-/- mice. Further, the loss of lymphatics corresponded with CD118 deficiency in the resident cell population as opposed to bone marrow-derived cells as determined using WT and CD118-/- chimeras. HSV-1 titers were significantly elevated in CD118-/- mice compared to WT mice. To explore the significance of these findings further, a mouse skin HSV-1-vesicle-inducible model was developed. After the application of a high dose of GFP-expressing HSV-1 (1x106), a vesicle could be observed proximal to the site of injection. Confocal microscopic analysis of the lesion found a loss of lymphatic but not blood vessels whereas surrounding tissue displayed intact blood and lymphatic vessels.
Conclusions: :
Lymphatic vessel integrity following HSV-1 corneal infection requires an intact type I interferon response by resident cell populations. The specific targeting of vessels is likely due to the absence of an intact basement membrane with access to lymphatic endothelium via portals. The loss of lymphatics would allow the virus to collect in high titer within the vesicular fluid increasing the ease of spreading to a naive host.
Keywords: herpes simplex virus • cytokines/chemokines • vascular endothelial growth factor