Early gamma interferon and interleukin-2 responses to vaccination predict the late resting memory in malaria-naïve and malaria-exposed individuals

Abstract

Two different cell populations respond to potent T-cell-inducing vaccinations. The induction and loss of effector cells can be seen using an ex vivo enzyme-linked immunospot (ELISPOT) assay, but the more durable resting memory response is demonstrable by a cultured ELISPOT assay. The relationship of the early effector response to durable resting memory is incompletely understood. Effector phenotype is usually identified by gamma interferon (IFN-gamma) production, but interleukin-2 (IL-2) has been specifically linked to the differentiation of memory cells. Here, IFN-gamma- and IL-2-secreting effector cells were identified by an ex vivo ELISPOT assay 1 week after vaccination and compared with the resting memory responses detected by a cultured ELISPOT assay 3 months later. The different kinetics and induction of IL-2 by different vaccines and natural exposure are described. Furthermore, both early IFN-gamma and IL-2 production independently predicted subsequent memory responses at 3 months in malaria-naïve volunteers, but only IFN-gamma predicted memory in malaria-exposed volunteers. However, dual ELISPOT assays were also performed on malaria-exposed volunteers to identify cells producing both cytokines simultaneously. This demonstrated that double-cytokine-producing cells were highly predictive of memory. This assay may be useful in predicting vaccinations most likely to generate stable, long-term memory responses.

FIG. 1.

Antigen-specific effector cell IFN-γ (IFN g) and IL-2 (IL 2) production by PBMCs isolated from immunized and unimmunized volunteers. Ex vivo IL-2 responses (white boxes) and ex vivo IFN-γ (hatched boxes) are shown. Results are shown for malaria-naïve and malaria-exposed subjects. When only IL-2 production is shown, “ns” is shown in place of the box and whisker plot that would indicate IFN-γ production. Vaccination regimens are abbreviated as shown in Table 1, and the numbers displayed on the category axis (x axis) indicate the assay was conducted at that number of days after the vaccination regimen shown. For instance, DDM + 7 for refers to 7 days after two sequential DNA priming immunizations followed by MVA boosting (encoding either CS or ME-TRAP).

FIG. 2.

Correlations between IFN-γ (IFN g) and IL-2 (IL2) effector responses measured 1 week postvaccination and subsequent resting memory responses at 3 months by vaccination group (for malaria-naïve and malaria-exposed subjects). For the vaccination regimens, the correlations between IFN-γ and IL-2 production 1 week after vaccination and resting memory 3 months after vaccination are shown. Resting memory (RM) is abbreviated as “RM.” The plots labeled “Natural immunity” show the responses seen before vaccination in malaria-exposed volunteers. The plots labeled “FMF/MFM (ME-TRAP), RM at 9 months” compare IFN-γ and IL-2 production 1 week after vaccination with resting memory seen 9 months after vaccination. r and P values are shown underneath each plot.

FIG. 3.

Correlations between single-cytokine- and double-cytokine-producing cells and subsequent resting and effector memory responses. Dual-cytokine ELISPOT assays were used to test malaria-exposed volunteers. Five volunteers had received alternating-vector immunizations, while six volunteers had received prime-boost regimens encoding CS. PBMCs for further study were not available from RRM/MRR vaccinees. The top row of three scatter plots shows the correlation between ex vivo IFN-γ (IFN g) production (left), ex vivo IL-2 production (middle), and ex vivo double-cytokine-positive cells (right) (x axis) with resting memory IFN-γ production at 3 months (y axis). The corresponding r and P values are shown above each plot. The bottom row of three scatter plots shows the same sequence of x-axis labels with persistent ex vivo memory cells at 3 months (y axis). The double-cytokine-producing cells are most predictive of subsequent resting memory.