Abstract
Monocyte-derived dendritic cells (moDC) are an important scientific and clinical source of functional dendritic cells (DC). However, the optimization of the generation process has to date mainly been limited to the variation of soluble factors. In this study, we investigated the impact of the cell culture dish surface on phenotype and cytokine profile. We compared a standard cell culture dish to a non-adherent culture dish for two immunogenic maturation conditions, two tolerogenic conditions, and an unstimulated control. Phenotype, cytokine profile and T cell stimulatory capacity were determined after a 3-day culture. Light microscopy revealed an increase in homotypic cluster formation correlated with the use of non-adherent surfaces, which could be reduced by using blocking antibodies against CD18. All surface markers analyzed showed moderate to strong differences depending on the culture dish surface, including significantly decreased expression of key maturation markers such as CD80, CD86, and CCR7 as well as PD-L1 on cells stimulated with the Jonuleit cytokine cocktail cultured on a non-adherent surface. Significant differences in the secretion of many cytokines were observed, especially for cells stimulated with LPS, with over 10-fold decreased secretion of IL-10, IL12-p40, and TNF-α from the cells cultured on the non-adherent surface. All immunogenic moDC populations showed similar capacity to induce antigen-specific T cells. These results provide evidence that the DC phenotype depends on the surface used during moDC generation. This has important implications for the optimization of DC-based immunotherapy development and underlines that the local surrounding can interfere with the final DC population beyond the soluble factors.
Highlights
Dendritic cells (DC), positioned between the innate and adaptive immune system, play a central role in a great variety of immunological settings
We observed an increase in cell cluster formation on the non-adherent culture dish in comparison to the standard cell culture dish for all Monocyte-derived dendritic cells (moDC) populations (Figure 1, Figure S1)
Smaller clusters were observed on the standard cell culture dish, especially for the immunogenic stimulation conditions with LPS or the Jonuleit cocktail (Figure S1)
Summary
Dendritic cells (DC), positioned between the innate and adaptive immune system, play a central role in a great variety of immunological settings. They play an important role for the pathogenesis of many diseases, and are increasingly under investigation as a clinical tool to treat a great diversity of different challenging conditions, ranging from cancer to autoimmunity [1]. Alone the in vivo regulation of DC adhesion upon maturation [8, 9] gives an indication that adhesional signaling might be of importance in a potentially more diverse way than can be expected from an unspecific surface of a plastic cell culture dish. The choice of the culture dish can potentially have a significant impact on the DC phenotype and function by either supporting the early, integrinmediated adhesion followed by low homotypic clustering, or by reducing culture dish interactions leading to an increase in clustering and cluster-mediated cell-cell interactions
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