Human antigen presenting cells stimulated with Salmonella delivered influenza antigens induce cytokine production and proliferation of human CD4⁺ T cells in vitro

This study aimed to investigate whether the human antigen presenting cells (APCs)can process and present Salmonella expressing H7N9 hemagglutinin (Sal-HA), neuraminidase (Sal-NA)or M2 ectodomain (Sal-M2e)to T cells and subsequently activate CD4⁺ T cell responses in vitro. In this study, APCs generated from human peripheral blood mononuclear cells (PBMCs)were first treated with mitomycin-C, followed by stimulation with Sal-HA, Sal-M2e, Sal-NA or Salmonella alone for 24 h. Subsequently, stimulated APCs were coincubated with untreated PBMCs (1:10)of the same individual for 24 or 72 h and then analysed for cytokine induction and T cell proliferations by qRT-PCR assay and flow cytometry, respectively. Our results demonstrated that APCs stimulated with Sal-HA, Sal-M2e or Sal-NA induced significantly (p < .05)higher CD3⁺CD4⁺ T cell proliferations compared to the APCs treated with Salmonella alone. Our data further revealved that APCs treated with Sal-HA induced significantly (p < .05)higher CD3⁺CD4⁺ T cell responses compared to the APCs treated with either Sal-M2e or Sal-NA, which both induced almost comparable levels. The T cell proliferation responses were further measured by lymphocyte proliferation assay and the results showed that Sal-HA and Sal-M2e stimulated APCs induced significantly (p < .05)higher proliferations in T cells compared to the APCs stimulated with either Sal-NA or Salmonella alone. With respect to cytokine inductions, APCs treated with either Sal-HA or Sal-M2e induced significantly (p < .05)higher mRNA transcription levels of proinflammatory (IL-1β, IL-6, IL-12 and IL-23), Th1 (IFN-γ), Th17 (IL-17 and IL-21)and Th2 (IL-10 and TGF-β)cytokines in T cells compared to Sal-NA or Salmonella alone treated APCs. In conclusion, we show that Salmonella system can efficiently deliver vaccine antigens to APCs and is, thus, capable to elicit heterologous antigen-specific adaptive immunity.