Role of IL-12 in T cell activation in old mice
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Publisher:The Ohio State University
Series/Report no.:The Ohio State University. College of Biological Sciences Honors Theses; 2008
The cytokine interleukin-12 (IL-12) has potent biological activities and is particularly important for the synergistic activation of CD4+ and CD8+ T cells to secrete IFN-γ during an antigen specific exposure. Additionally, IL-12 can stimulate cells of the innate immune system (natural killer cells) directly to secrete IFN-γ. In old mice it has been shown that IL-12 can stimulate CD8+ T cells to secrete IFN-γ in an antigen-independent manner, demonstrating an alternative T cell activation pathway in old age. This project seeks to dissect the mechanism of CD8+ T cell activation in more detail. Biologically active IL-12 exists as a heterodimer comprised of two peptide chains, p40 and p35. In this form IL-12 (p70) can interact with the IL-12 receptor β1 and β2 chains and signal the receptive cells to produce IFN-γ. The single peptide p40 can also exist as a monomer (p40) or a homodimer (p40/p40, designated p80). It has been shown under various conditions that p80 can stimulate or inhibit T cells. This project determined whether the stimulatory effect of IL-12 on CD8+ T cells from old mice to produce IFN-γ is mediated by IL-12p70, p80 homodimer, or p40 monomer. Additionally, we determined whether IL-23, a close relative of IL-12 consisting of p40 and p19, can stimulate CD8+ T cells directly. Using combinations of p40 monomer, p40 homodimer, IL-12, or IL-23 we stimulated T cells to secrete IFN-γ and measured cytokine production by enzyme linked immuno-sorbent assay (ELISA). Dose response curves and time course were performed to optimize the assays. Once the optimum assay conditions were ascertained, the stimulatory component of IL-12 was identified. Additional studies determined whether p40 or p80 can inhibit the biological action of IL-12. Furthermore, an additional synergistic cytokine, IL-18 was added into the assay conditions to determine whether the stimulatory capacity of IL-12 and its family members can be boosted. Finally, to conclusively demonstrate that CD8+ T cells from old mice are directly stimulated to produce IFN-γ by an IL-12 family member, cell purification techniques were implemented.