Abstract
Because dendritic cells (DCs) play a critical role in the regulation of adaptive immune responses, they have been ideal candidates for cell-based immunotherapy of cancers and infections in humans. Generally, monocyte-derived DCs (MDDCs) were generated from purified monocytes by multiple steps of time-consuming physical manipulations for an extended period cultivation. In this study, we developed a novel, simple and rapid method for the generation of type-1 helper T cell (Th1)-stimulating human DCs directly from bulk peripheral blood mononuclear cells (PBMCs). PBMCs were cultivated in the presence of 20 ng/ml of granulocyte-macrophage colony-stimulating factor, 20 ng/ml of interleukin-4 (IL-4) and 1,000 U/ml of interferon-β for 24 h followed by 24 h maturation with a cytokine cocktail containing 10 ng/ml of tumor necrosis factor-α (TNF-α), 10 ng/ml of IL-1β and 1 μg/ml of prostaglandin E2. The phenotype and biological activity of these new DCs for induction of allogeneic T cell proliferation and cytokine production were comparable to those of the MDDCs. Importantly, these new DCs pulsed with inactivated HIV-1 could generated HIV-1-reactive CD4+ T cell responses in humanized mice reconstituted with autologous PBMCs from HIV-1-negative donors. This simple and quick method for generation of functional DCs will be useful for future studies on DC-mediated immunotherapies.
Highlights
Dendritic cells (DCs) are professional antigen-presenting cells (APCs) which play a critical role in the regulation of the adaptive immune response through activation and polarization of naive T cells (Banchereau et al, 2000)
One possible concern on using whole peripheral blood mononuclear cells (PBMCs) was that the nonmonocyte cells, such as T, B or natural killer (NK) cells, in the PBMCs might interfere with differentiation and function of DCs
IL-6 that is included in the maturation cytokine cocktail tumor necrosis factor-α (TNF-α) and IL-1β and prostaglandin E2 (PGE2) to mature DCs was not necessary in the present culture conditions
Summary
Dendritic cells (DCs) are professional antigen-presenting cells (APCs) which play a critical role in the regulation of the adaptive immune response through activation and polarization of naive T cells (Banchereau et al, 2000). Since different ex vivo DC generation methods affect the DC phenotype and function (Kalantari et al, 2011), it is critical to choose appropriate method for generating functional DCs. In general, the DC precursor monocytes are purified from PBMCs by adherence (Jonuleit et al, 2001), elutriation (Berger et al, 2005) or positive or negative selection using immunomagnetic beads (Babatz et al, 2003). The DC precursor monocytes are purified from PBMCs by adherence (Jonuleit et al, 2001), elutriation (Berger et al, 2005) or positive or negative selection using immunomagnetic beads (Babatz et al, 2003) These enriched monocytes are induced to differentiate into DCs by 5 days-in vitro cultivation in medium supplemented with granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-4 followed by a 2-daysmaturation procedure (Sallusto and Lanzavecchia, 1994; Gilboa, 2007; Dauer et al, 2008). A lines of evidence are increasing that mature monocyte-derived DCs can be generated even after short-term cell culture for 2–3 days (Dauer et al, 2003a,b; Jarnjak-Jankovic et al, 2007; Zhang et al, 2008; Tawab et al, 2009)
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