Characterization of the human CD8⁺ T cell response following infection with 2009 pandemic influenza H1N1 virus

Abstract

The 2009 H1N1 influenza pandemic provided an opportunity to study human virus-specific T cell responses after infection with a novel influenza virus against which limited humoral immunity existed in the population. Here we describe the magnitude, kinetics, and nature of the virus-specific T cell response using intracellular gamma interferon (IFN-γ) staining and fluorochrome-labeled major histocompatibility complex (MHC) class I-peptide complexes. We demonstrate that influenza virus-infected patients develop recall T cell responses that peak within 1 week postinfection and that contract rapidly. In particular, effector cell frequencies declined rapidly postinfection in favor of relatively larger proportions of central memory cells.

Fig. 1.

Example of the analysis of virus-specific T cells. (A) The frequency of IFN-γ⁺ CD8⁺ T cells was determined by intracellular IFN-γ staining after stimulation with influenza virus A/Netherlands/602/09 (pH1N1) virus (left panel) and subtracting the background values of nonstimulated control cells (right panel). (B) The subsets of Tm⁺ cells were quantified on the basis of expression of CCR7, CD45RA, CD28, and CD27 using flow cytometry. Phycoerythrin (PE), allophycocyanin (APC), peridinin chlorophyll protein (PerCP), FITC, and Cy7 fluorophore were used to label CD27, CD28, CCR7, etc.

Fig. 2.

Analysis of the magnitude of the T cell response upon infection with 2009 H1N1 virus. (A to D) The frequencies of virus-specific CD8⁺ (A and C) and CD4⁺ (B and D) T cells were determined by intracellular IFN-γ staining in individual patients at various time points after the onset of clinical symptoms as indicated. The data are averages of duplicate values. In panels C and D, trend lines and their R² values were added using Excel software.

Fig. 3.

Subset analysis of Tm⁺ T cells. PBMC were obtained from individual patients at various time points after the onset of clinical symptoms, and Tm⁺ cells were tested for the expression of CCR7, CD45RA, CD28, and CD27 to determine the proportion of effector T cells, naïve T cells, T_EM, T_CM, T_EMRA and CD45RA⁺ CCR7⁺ CD28⁻ cells. Cells not belonging to any of these subsets are shown in white. The data are averages of duplicate values.