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 2

Th9 differentiation from naïve CD4⁺ T cells is inhibited by 7P in vitro. (a) Dendritic cells (DC, CD11c⁺ cells) were isolated from the spleen and matured with LPS overnight. Naïve CD4⁺ T cells were then isolated from spleens of C57BL6J mice, and LPS-treated DCs were treated with anti-CD3 (3 μg/mL), anti-CD28 (3 μg/mL), anti-INF-γ (3 μg/mL), transforming growth factor- (TGF-) β (10 ng/mL), and IL-4 (10 ng/mL) for 4–5 days with 7P, sham-7P, or vehicle. The cells were then restimulated for 4 h with 12-O-tetradecanoylphorbol-13-acetate and ionomycin (500 ng/mL each) in the presence of brefeldin A (1 mg/mL) before real-time PCR analysis. (b–d) Cells treated with TGF-β and IL-4 were collected, and prepared RNA was reversed to cDNA. Il9, Il10, and Irf4 mRNA were amplified by quantitative polymerase chain reaction (qPCR) after treatment by anti-CD3, anti-CD28, anti-INF-γ, TGF-β, and IL-4 with 7P, sham-7P, or vehicle. Data were representative of 7–14 independent experiments (, ). (e) Human naïve CD4⁺ T cells were isolated from peripheral blood and differentiated into Th9 cells with anti-CD3, anti-CD28, TGF-β, and IL-4 plus 7P and vehicle. Il9 mRNA was also assayed by qPCR (; ).

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