A short protocol using dexamethasone and monophosphoryl lipid A generates tolerogenic dendritic cells that display a pot
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RESEARCH
Open Access
A short protocol using dexamethasone and monophosphoryl lipid A generates tolerogenic dendritic cells that display a potent migratory capacity to lymphoid chemokines Paulina García-González1,2†, Rodrigo Morales1,2†, Lorena Hoyos1,2, Jaxaira Maggi1,2, Javier Campos1,2, Bárbara Pesce1,2, David Gárate1, Milton Larrondo3, Rodrigo González3, Lilian Soto1,4, Verónica Ramos1,2, Pía Tobar1, María Carmen Molina5, Karina Pino-Lagos1,2, Diego Catalán1,2* and Juan Carlos Aguillón1,2*
Abstract Background: Generation of tolerogenic dendritic cells (TolDCs) for therapy is challenging due to its implications for the design of protocols suitable for clinical applications, which means not only using safe products, but also working at defining specific biomarkers for TolDCs identification, developing shorter DCs differentiation methods and obtaining TolDCs with a stable phenotype. We describe here, a short-term protocol for TolDCs generation, which are characterized in terms of phenotypic markers, cytokines secretion profile, CD4+ T cell-stimulatory ability and migratory capacity. Methods: TolDCs from healthy donors were generated by modulation with dexamethasone plus monophosphoryl lipid A (MPLA-tDCs). We performed an analysis of MPLA-tDCs in terms of yield, viability, morphology, phenotypic markers, cytokines secretion profile, stability, allogeneic and antigen-specific CD4+ T-cell stimulatory ability and migration capacity. Results: After a 5-day culture, MPLA-tDCs displayed reduced expression of costimulatory and maturation molecules together to an anti-inflammatory cytokines secretion profile, being able to maintain these tolerogenic features even after the engagement of CD40 by its cognate ligand. In addition, MPLA-tDCs exhibited reduced capabilities to stimulate allogeneic and antigen-specific CD4+ T cell proliferation, and induced an anti-inflammatory cytokine secretion pattern. Among potential tolerogenic markers studied, only TLR-2 was highly expressed in MPLA-tDCs when compared to mature and immature DCs. Remarkable, like mature DCs, MPLA-tDCs displayed a high CCR7 and CXCR4 expression, both chemokine receptors involved in migration to secondary lymphoid organs, and even more, in an in vitro assay they exhibited a high migration response towards CCL19 and CXCL12. Conclusion: We describe a short-term protocol for TolDC generation, which confers them a stable phenotype and migratory capacity to lymphoid chemokines, essential features for TolDCs to be used as therapeutics for autoimmunity and prevention of graft rejection. Keywords: Tolerance, Monocyte-derived dendritic cells, Chemokine receptors, CCR7, CXCR4, Cell-based therapy
* Correspondence: [email protected]; [email protected] † Equal contributors 1 Immune Regulation and Tolerance Research Group, Programa Disciplinario de Inmunología, ICBM, Facultad de Medicina, Universidad de Chile, Santiago, Chile 2 Millennium Institute on Immunology and Immunotherapy, Santiago, Chile Full list of author information is available at the end of the arti
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