Allostimulatory ability of DCs stimulated with G-β₂GPI and T-cell priming. (A) Irradiated DCs prestimulated with G-β₂GPI, M-β₂GPI, or LPS increased the proliferative ability (mean counts per minute, cpm) of resting allogenic T cells compared with unstimulated DCs. At a DC:T-cell ratio of 1:16, unstimulated DCs vs G-β₂GPI, P = .004, and unstimulated DCs vs M-β₂GPI, P = .04. (B) G-β₂GPI–treated DCs stimulated allogeneic naive human T cells to produce IFN-γ and IL-4. Five-day human DCs were stimulated with G-β₂GPI, M-β₂GPI, or LPS, or were left unstimulated for 18 hours. A total of 5 × 10⁴ DCs were used to stimulate 1 × 10⁶ allogeneic naive negatively selected CD4⁺CD45RA⁺ T cells. Activated T cells were expanded with recombinant human IL-2 (30 U/mL). On day 10, T-cell lines were stimulated with PMA and ionomycin for 4 hours in the presence of brefeldin A. Cells were stained with anti-hu-CD3PerCP and processed for intracellular labeling with anti-human IFN-γ–FITC and anti-human IL-4–PE. The numbers show the percentage of activated CD3⁺ cells producing the cytokine. Samples were analyzed on a FACSCanto cytofluorometer using FACSDiva software (BD Biosciences). The results of 1 representative experiment of 3 are shown. (C) Dose-response production of IL-6 and IL-10 in β₂GPI–stimulated DC culture supernatants. Five-day human DCs were stimulated with LPS (100 ng/mL), G-β₂GPI (0-30 μg/mL), or M-β₂GPI (0-30 μg/mL), or were left unstimulated. Supernatants were collected after 18 hours to measure IL-6 and IL-10 by specific ELISA experiments. Results are expressed as means ± SD; n = 3.