A 7-Amino Acid Peptide Mimic from Hepatitis C Virus Hypervariable Region 1 Inhibits Mouse Lung Th9 Cell Differentiation by Blocking CD81 Signaling during Allergic Lung Inflammation

Figure 4

CD81 expression during mouse and human Th9 cell differentiation. Naïve CD4⁺ T cells and 7P bind to CD81. (a–d) Mouse and human naïve CD4⁺ T cells isolated from mouse spleen and human peripheral blood were stimulated with anti-CD3 (3 μg/mL), anti-CD28 (3 μg/mL), and anti-INF-γ (3 μg/mL) with TGF-β (10 ng/mL) and IL-4 (10 or 20 ng/mL) for 5 days to induce (a) mouse and (c) human Th9 cell differentiation. Cells were analyzed for CD81 expression by qPCR (, ). (b) Mouse and (d) human CD81 were examined by flow cytometry. (e, f) Naïve CD4⁺ T cells transfected with pcDNA3.1-CD81 and purified by FACS. Cells were stimulated with anti-CD3, anti-CD28, anti–INF-γ, TGF-β, and IL-4 (10 and 20 ng/mL) for 5 days, then stimulated for 4 h with 12-O-tetradecanoylphorbol-13-acetate and ionomycin (500 ng/mL each) in the presence of brefeldin A (1 μg/mL). Intracellular staining was performed with anti-CD4, IL-9, and CD81. CD81 expression was analyzed by flow cytometry with 7P-FITC in the gated CD4⁺IL-9⁺ cell population. The percentage of 7P-positive Th9 cells was examined by FACS. Results were representative of three independent experiments (, ).

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