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A T cell costimulatory molecule, OX40, contributes to T cell expansion, survival, and cytokine production. Although several roles for OX40 in CD8+ T cell responses to tumors and viral infection have been shown, the precise function of these signals in the generation of memory CD8+ T cells remains to be elucidated. To address this, we examined the generation and maintenance of memory CD8+ T cells during infection with Listeria monocytogenes in the presence and absence of OX40 signaling. We used the expression of killer cell lectin-like receptor G1 (KLRG1), a recently reported marker, to distinguish between short-lived effector and memory precursor effector T cells (MPECs). Although OX40 was dispensable for the generation of effector T cells in general, the lack of OX40 signals significantly reduced the number and proportion of KLRG1low MPECs, and, subsequently, markedly impaired the generation of memory CD8+ T cells. Moreover, memory T cells that were generated in the absence of OX40 signals in a host animal did not show self-renewal in a second host, suggesting that OX40 is important for the maintenance of memory T cells. Additional experiments making use of an inhibitory mAb against the OX40 ligand demonstrated that OX40 signals are essential during priming, not only for the survival of KLRG1low MPECs, but also for their self-renewing ability, both of which contribute to the homeostasis of memory CD8+ T cells.

Background The pathogenicity of pneumococcus with high morbidity, mortality, and multi-drug resistance patterns has been increasing. The limited coverage of the licensed polysaccharide-based vaccines and the replacement of the non-vaccine serotypes are the main reasons for producing a successful serotype-independent vaccine. Pneumococcal surface protein A (PspA) is an extremely important virulence factor and an interesting candidate for conserved protein-based pneumococcal vaccine classified into two prominent families containing five clades. PspA family-elicited immunity is clade-dependent, and the level of the PspA cross-reactivity is restricted to the same family. Methods To cover and overcome the clade-dependent immunity of the PspAs in this study, we designed and tested a PspA1-5c+p vaccine candidate composed of the highest immunodominant coverage of B- and T-cell epitope truncated domain of each clade focusing on two cross-reactive B and C regions of the PspAs. The antigenicity, toxicity, physicochemical properties, 3D structure prediction, stability and flexibility of the designed protein using molecular dynamic (MD) simulation, molecular docking of the construct withHLADRB1*(01:01) and human lactoferrin N-lop, and immune simulation were assessed using immunoinformatics tools. In the experimental section, after intraperitoneal immunization of the mice with Alum adjuvanted recombinant PspA1-5c+p, we evaluated the immune response, cross-reactivity, and functionality of the Anti-PspA1-5c+p antibody using ELISA, Opsonophagocytic killing activity, and serum bactericidal assay. Results For the first time, this work suggested a novel PspA-based vaccine candidate using immunoinformatics tools. The designed PspA1-5c+p protein is predicted to be highly antigenic, non-toxic, soluble, stable with low flexibility in MD simulation, and able to stimulate both humoral and cellular immune responses. The designed protein also could interact strongly with HLADRB1*(01:01) and human lactoferrin N-lop in the docking study. Our immunoinformatics predictions were validated using experimental data. Results showed that the anti-PspA1-5c+p IgG not only had a high titer with strong and same cross-reactivity coverage against all pneumococcal serotypes used but also had high and effective bioactivity for pneumococcal clearance using complement system and phagocytic cells. Conclusion Our findings elucidated the potential application of the PspA1-5c+p vaccine candidate as a serotype-independent pneumococcal vaccine with a strong cross-reactivity feature. Further in-vitro and in-vivo investigations against other PspA clades should be performed to confirm the full protection of the PspA1-5c+p vaccine candidate.

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