Monocyte-derived Dendritic Cells (moDCs) have been traditionally used as vaccines capable of eliciting specific T cell responses against tumors and infections. Another innate immune effector cell, Natural Killer (NK) cells, endowed with antitumor and anti-infection capacities, have been adoptively transferred to treat against malignancy and pathogens. In our study, we investigate the potential of Aspergillus fumigatus derived beta(1,3)glucan (BG) pulsed moDCs as NK cell manipulators, which might give new perspectives in optimization of DC based vaccines. DCs were generated from positive selection of CD14+ cells from peripheral blood mononuclear cells (PBMCs) and cultured for 5-6 days in serum-free GMP grade medium supplemented with GM-CSF and IL-4. On day 2, immature DCs (iDCs) were maturated (mDCs) with BG or Tumor Necrosis Factor- alpha (TNF-a). DC profile was characterized by quantifying the expression of CD1a, CD14, CD40, CD80, CD83, CD86, HLA-I, HLA-DR, CCR7, Dectin-1, IL12p35 and IL15. Autologous NK cells, obtained by negative selection of PBMCs were cocultured with DCs for 24h at a ratio of 1:1. After coculture, NK cells were studied for the expression of defined markers, among them were CD69, HLA DR, CD25, CD83, CCR7, cytokines (interferon gamma, IFN-g), transcription factors (T-bet). NK cell cytotoxicity was evaluated against the leukemic cell line K562 via the degranulation marker CD107a. Results: DC-NK cell interaction further increased the expression of costimulatory molecules and HLA antigens as well as the release of IL12p35 onmDCs. In parallel, mDCs stimulated NK cells stronger than iDCs and they potentiated the cytotoxic capacity of NK cells towards K562. In addition, mDCs induced the expression of HLA DR, CD25 and CD83 on NK cells. Interestingly, this effect was significantly higher when NK cells were stimulated by BG mDCs than with TNF-a mDCs. Using neutralizing antibodies against Dectin-1, which is the receptor of BG on DCs, we observed a decrease of the stimulatory capacity of BG mDCs. Our study proposes the NK cells as a new target for DC vaccination. By comparing the classical DC maturation adjuvants, BG showed to escalate the stimulatory effect of DCs towards NK cells. Furthermore, BG mDCs induce the expression of a specific NK cell profile which could promote the interaction of NK cells with T cells. These data might be relevant for the design of DC vaccines and future immunotherapeutic strategies against cancer and infections.
Full conference title:
- EBMT 39th (2013)