RESUMO
BACKGROUND AIMS: Dendritic cell (DC)-tumor cell hybrids have been used clinically in cancer immunotherapy, but their advantage over the simple mixture of tumor cells and DCs is still a matter of controversy. In this study, we compared DC-tumor cell hybrids with the non-fused mixture of DC and tumor cells directly in their ability to induce a specific immune response. METHODS: Hybrids were obtained by electrofusion of tumor cells and monocyte-derived DCs. Cell phenotype was evaluated by flow cytometry and antigen-presenting ability by co-culture with syngeneic T cells followed by tetramer analysis and interferon (IFN)-γ ELISPOT. RESULTS: Less than half the cells in the mixture expressed DC co-stimulatory molecules. Furthermore, DCs in the mixture had significantly lower expression of MHC class I molecules than DCs in the fusion. Conversely, nearly all CD11c(+)Her2/neu(+) hybrids expressed CD80, CD86, CD83, HLA-DR and MHC class I from both tumor cells and DCs. Using tumor cells constitutively expressing a cytomegalovirus (CMV) antigen, we show that expansion of CMV-specific cytotoxic T lymphocytes (CTLs) restricted by DCs' MHC class I molecules was higher when DC-tumor hybrids were the stimulators. Furthermore, only hybrids stimulated CTLs to produce IFN-γ in response to CMV-positive target cells. CONCLUSIONS: These data show the superiority of DC-tumor cell hybrids over their simple mixture as T-cell stimulators. Hybrids expressed more co-stimulatory and MHC molecules, induced higher antigen-specific T-cell expansion and were the only cells able to induce IFN-γ-producing antigen-specific T cells. Thus, these data offer further support for cancer immunotherapeutic approaches using DC-tumor cell hybrids.
Assuntos
Células Dendríticas/imunologia , Células Híbridas/imunologia , Imunidade Celular , Imunoterapia/métodos , Neoplasias/imunologia , Neoplasias/terapia , Apresentação de Antígeno , Vacinas Anticâncer/imunologia , Fusão Celular , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/patologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Células Híbridas/patologia , Neoplasias/patologia , Linfócitos T Citotóxicos/imunologiaRESUMO
Mast cells (MCs) are tissue resident cells, rich in inflammatory mediators, involved in allergic reactions, and with an increasingly recognized role in immunomodulation. Dendritic cells (DCs), on the other hand, are central to the determination of immune response patterns, being highly efficient antigen-presenting cells that respond promptly to changes in their microenvironment. Here, we show that direct cell contact between immature monocyte-derived DCs (iDCs) and MC bends DCs toward tolerance induction. DCs that had direct contact with MC (MC-iDC) decreased HLA-DR but increased PD-L1 expression and stimulated regulatory T lymphocytes, which expresses FoxP3(+), secrete TGF-ß and IL-10, and suppress the proliferation of mitogen-stimulated naïve T lymphocytes. Furthermore, MC-iDC expressed higher levels of indoleamine-2,3-deoxigenase (IDO), a phenomenon that was blocked by treatment of MC with anti-PD-1 or by the treatment of DCs with anti-PD-L1 or anti-PD-L2, but not by blocking of H1 and H2 histamine receptors on DCs. Contact with MC also increased phosphorylated STAT-3 levels in iDCs. When a STAT-3 inhibitor, JSI-124, was added to the DCs before contact with MC, the MC-iDC recovered their ability to induce allogeneic T cell proliferation and did not increase their IDO expression.
Assuntos
Imunoterapia , Neoplasias/imunologia , Anticorpos Monoclonais , Imunização , Imunoterapia Ativa , VacinasRESUMO
DCs orchestrate immune responses contributing to the pattern of response developed. In cancer, DCs may play a dysfunctional role in the induction of CD4(+)CD25(+)Foxp3(+) Tregs, contributing to immune evasion. We show here that Mo-DCs from breast cancer patients show an altered phenotype and induce preferentially Tregs, a phenomenon that occurred regardless of DC maturation stimulus (sCD40L, cytokine cocktail, TNF-α, and LPS). The Mo-DCs of patients induced low proliferation of allogeneic CD3(+)CD25(neg)Foxp3(neg) cells, which after becoming CD25(+), suppressed mitogen-stimulated T cells. Contrastingly, Mo-DCs from healthy donors induced a stronger proliferative response, a low frequency of CD4(+)CD25(+)Foxp3(+) with no suppressive activity. Furthermore, healthy Mo-DCs induced higher levels of IFN-γ, whereas the Mo-DCs of patients induced higher levels of bioactive TGF-ß1 and IL-10 in cocultures with allogeneic T cells. Interestingly, TGF-ß1 blocking with mAb in cocultures was not enough to completely revert the Mo-DCs of patients' bias toward Treg induction. Altogether, these findings should be considered in immunotherapeutic approaches for cancer based on Mo-DCs.
Assuntos
Neoplasias da Mama/imunologia , Células Dendríticas/imunologia , Ativação Linfocitária/imunologia , Linfócitos T Reguladores/imunologia , Evasão Tumoral/imunologia , Técnicas de Cocultura , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Fatores de Transcrição Forkhead/imunologia , Humanos , Subunidade alfa de Receptor de Interleucina-2/imunologia , Monócitos/imunologiaRESUMO
BACKGROUND: Patients with X-linked hyper-IgM syndrome (X-HIGM) due to CD40 ligand (CD40L) mutations are susceptible to fungal pathogens; however, the underlying susceptibility mechanisms remain poorly understood. OBJECTIVE: To determine whether monocyte-derived dendritic cells (DCs) from patients with X-HIGM exhibit normal responses to fungal pathogens. METHODS: DCs from patients and controls were evaluated for the expression of costimulatory (CD80 and CD86) and MHC class II molecules and for their ability to produce IL-12 and IL-10 in response to Candida albicans and Paracoccidioides brasiliensis. We also evaluated the ability of C albicans- and P brasiliensis-pulsed mature DCs to induce autologous T-cell proliferation, generation of T helper (T(H)) 17 cells, and production of IFN-γ, TGF-ß, IL-4, IL-5, and IL-17. RESULTS: Immature DCs from patients with X-HIGM showed reduced expression of CD80, CD86, and HLA-DR, which could be reversed by exogenous trimeric soluble CD40L. Most important, mature DCs from patients with X-HIGM differentiated by coculturing DCs with fungi secreted minimal amounts of IL-12 but substantial amounts of IL-10 compared with mature DCs from normal individuals. Coculture of mature DCs from X-HIGM patients with autologous T cells led to low IFN-γ production, whereas IL-4 and IL-5 production was increased. T-cell proliferation and IL-17 secretion were normal. Finally, in vitro incubation with soluble CD40L reversed the decreased IL-12 production and the skewed T(H)2 pattern response. CONCLUSION: Absence of CD40L during monocyte/DC differentiation leads to functional DC abnormalities, which may contribute to the susceptibility to fungal infections in patients with X-HIGM.