Fig 4.

Increased IFN-β secretion and enhanced IRF3 nuclear translocation in Tr/E-pretreated HEC-1A cells after HSV-2 challenge. IFN-β release from HEC-Ad/Tr cells following exposure to HSV-2 at an MOI of 1. (A) HEC-1A cells were left untreated or treated with an MOI of 0 to 50 of Ad/dl or Ad/Tr and incubated for 6 to 24 h in the presence of media or HSV-2 at an MOI of 1. At 6 h posttreatment, total RNA was harvested, and relative expression of IFN-β mRNA by real-time quantitative reverse transcription (RT)-PCR was assessed. Values were normalized to internal control 18S. (B) At 6 and 24 h posttreatment with HSV-2 at an MOI of 1, secreted IFN-β was measured by ELISA in cell-free supernatants and presented as pg/ml. The data are representative of three independent experiments performed in triplicate and are shown as the means ± SD. Statistical analysis was performed using Student's t test, with significance indicated in the graphs. In panels C and D, immunofluorescence analysis of IRF3 nuclear translocation is demonstrated for HEC-1A cells that were left untreated (UT) or treated with an MOI of 0 to 50 of Ad/dl or Ad/Tr (C) or pretreated with Tr or E for 1 h (D) and subsequently exposed to either medium alone or HSV-1 at an MOI of 1 or 5 for 4 h. Representative staining is shown for IRF3 (green), nuclear stain propidium iodide (red), HSV-2 gD (blue), and composite (yellow) at magnifications of ×2520 or ×1260 (D, left column). The data are representative of two independent experiments with similar results.