FIGURE 4.

Pathophysiology of enhanced DENV disease in vaccinated AG129 mice. (A) Enhanced DENV disease associated vascular leak pathology of pVD1-WT and -CRR vaccinated mice following sub-lethal heterologous DENV-2 infection. No pathology was visible in naїve or vaccinated mice 3 DPC, yet by 4 and 5 DPC mice succumbing to enhanced disease exhibited severe vascular leakage associated pathology. The same mouse is shown in the bottom two photos to highlight the severe intestinal hemorrhage observed in pVD1-WT but not in pVD1-CRR vaccinated mice (arrows). (B) Histology of uninfected, pVD1-WT, and pVD1-CRR vaccinated mouse livers 3 DPC. The top row is hematoxylin and eosin stained liver sections and the bottom row is immunohistochemistry for DENV-2 NS1 protein of the same individual animals. NS1+ mononuclear inflammatory cells and sinusoidal endothelial cells stain red in their cytoplasm and are visible (arrows) in liver tissue from vaccinated mice but not uninfected mice. Multiple sections from multiple animals were examined and single representatives are shown. All photos were taken at 400x magnification. (C) In vitro DENV-2 enhancement by pVD1-WT and pVD1-CRR vaccinated serum 12 weeks following vaccinations and one day prior to DENV-2 challenge. Consistent with the In vivo results, pVD1-WT vaccinated serum significantly enhanced DENV-2 infection whereas pVD1-CRR vaccinated serum did not (p = 0.0012). The data are representative of two independent experiments of 4 pools of 6–7 individual serum specimens for each vaccine treatment and were analyzed with two-way ANOVA and Bonferroni post hoc test significance at individual dilutions is depicted with asterisks.