| Precise regulation of effector function is critical for mounting a potent, yet specific
immune response to a given antigenic challenge. It has been hypothesized that the
cytokine content of secondary lymphoid organs and at the actual site(s) of inflammation
exert localized control over immune cell populations. Distinct cytokine combinations
present within a given microenvironment can induce changes in the phenotype and
effector functions of immune cells that differ dramatically from the effects exerted by
each cytokine alone. Using a multiparametric analysis approach, we investigated the
effects in culture of all possible combinations of as many as 6 cytokines (IL-2, IL-4, IL-7,
IL-12, TNF-α, and TGF-β) on the phenotype and functional capacity of CD4+ and CD8+
isolates derived from human peripheral blood. Following stimulation, cultures were
assessed for changes in maturation/activation state (as measured by surface marker
expression and phospho-protein analysis), as well as determining their individual proliferative
capacity and viability (ViaCount®). Stimulated CD4+ and CD8+ cultures were
also subject to cytokine production profiling through intracellular staining. The cytolytic
capacity of CD8+ cultures was measured both indirectly, through intracellular Granzyme
B staining, and directly through killing of autologous target B cells (CellToxicity).
All samples were acquired and analyzed using the Guava EasyCyte platform, a microcapillary-
based cell cytometry system. Briefly, we found that unique combinations of
cytokines had profound and highly varied effects on lymphocyte fate and function in
vitro. Demonstrations of synergy, antagonism, dominance, and substitution amongst
multiple cytokines in our experimental system may reflect actual paradigms underlying
such processes as the differentiation of T cell subsets into effector and resting
memory phenotypes in vivo. |
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