Figure 2. HAV induces the activation of NKT cells.

(A) NKT cells constitutively express TIM-1. NKT cells were identified by flow cytometry by gating on 6B11⁺CD3⁺ NKT cells in total PBMCs (upper panels) or in an NKT cell line (lower panels). TIM-1 expression was then analyzed on gated NKT cells (right panels). The numbers in the dot plot represent the percentage of NKT cells after gating on lymphocytes. Shaded histogram represents background staining with isotype control (mIgG1). (B) NKT cells are cytotoxic for HAV-infected hepatoma cells. Naive or HAV-infected hepatoma cells were cocultured with NKT cell lines from different donors (black: homozygous short form of TIM-1 [n = 6], light gray: heterozygous form of TIM-1 [n = 10], and white: homozygous long form of TIM-1 [n = 2]). AST levels of noninfected and HAV-infected hepatoma cells cultured alone are shown. After 48 hours, culture supernatants were analyzed for AST level. Greater AST indicates greater hepatoma cell lysis. Data are given as mean ± SEM; *P < 0.05, ***P < 0.001 (ANOVA). (C) WT HAV is found at the cell surface of HAV-infected hepatoma cells. Human hepatoma Huh7-A-I cells were infected with WT HAV (HAV-infected) or mock infected (Naive) and examined with a cell surface ELISA using an anti-HAV mAb, detected with peroxidase-labeled anti-IgG. Absorbance at 450 nm was determined in an ELISA plate reader, and the mean absorbance of duplicate wells ± SD was plotted versus the anti-HAV mAb concentration (***P < 0.001 [ANOVA]). The assay was performed twice, and the data represent 1 experiment.