Figure 5: Low cytokine production by memory-like HCV-specific CD8⁺ T cells.

Virus-specific CD8⁺ T cells were stimulated with epitope-specific peptides (10 μg ml⁻¹) for 5 h at 37 °C and analysed for cytokine production (a,b). (a) Flow cytometric dot plots demonstrate stability of peptide/MHCI tetramer staining (upper row) and IFNγ and TNF production (lower row) of HCV epitope-specific CD8⁺ T cells (red gate). (b) Cytokine production by HCV epitope-specific CD8⁺ T cells of five patients with HCV infection (HCV; six HCV epitope-specific CD8⁺ T-cell responses), seven spontaneous resolvers (SpR; ten HCV-epitope-specific CD8⁺ T-cell responses) and five FLU epitope-specific CD8+ T-cell responses (FLU, n=5). (c–e) In vitro peptide-expanded HCV epitope-specific CD8⁺ T cells from HCV-infected patients (n=9; 12 HCV epitope-specific CD8⁺ T-cell responses) or spontaneous resolvers (n=8; 13 HCV epitope-specific CD8⁺ T-cell responses) were analysed for cytokine production after 5 h of peptide-specific re-stimulation. (c) Representative dot plots showing cytokine production by in vitro HCV peptide-expanded CD8⁺ T cells after HCV peptide-specific re-stimulation (gated on bulk CD8⁺ T cells). Percentages of the respective quadrant are depicted. (d) Frequencies of IFNγ- (left) and TNF-producing (right) in vitro peptide-expanded HCV epitope-specific CD8⁺ T cells are shown in the statistical graphs. (e) TNF production by IFNγ⁺ in vitro peptide-expanded CD8⁺ T cells in per cent. Bar charts show the median value with interquartile range. Statistical significance was assessed by Friedman test (HCV/DAA) and by unpaired Kruskal−Wallis (HCV, SpR, FLU). Multiple comparisons were performed with Dunn’s test. (*P<0.05.)