Ex vivo generation of mature equine monocyte-derived dendritic cells

Cavatorta, Derek J.; Erb, Hollis N.; Flaminio, M. Julia B.F.

doi:10.1016/j.vetimm.2009.04.019

Cited by 15 publications

(18 citation statements)

References 36 publications

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“…CD206 was also able to discriminate MoDC and MΦ, this time with expression restricted to the MΦ. However, BMDC also expressed CD206 which is in agreement with observations done in other species where this c-type lectin can also be found on both MΦ and DC [10, 34, 41]. Based on our data demonstrating CD40 expression even on unstimulated MoDC and BMDC, but not on MΦ and monocytes, we propose that this surface molecule is a potential discriminatory marker for DC.…”

Section: Discussionsupporting

confidence: 91%

Comparative analysis of canine monocyte- and bone-marrow-derived dendritic cells

Gutzwiller

Moulin²,

Zurbriggen

et al. 2010

Vet. Res.

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Dendritic cells (DC) represent a heterogeneous cell family of major importance for innate immune responses against pathogens and antigen presentation during infection, cancer, allergy and autoimmunity. The aim of the present study was to characterize canine DC generated in vitro with respect to their phenotype, responsiveness to toll-like receptor (TLR) ligands and T-cell stimulatory capacity. DC were derived from monocytes (MoDC) and from bone marrow hematopoietic cells cultured with either Flt3-ligand (FL-BMDC) or with GM-CSF (GM-BMDC). All three methods generated cells with typical DC morphology that expressed CD1c, CD11c and CD14, similar to macrophages. However, CD40 was only found on DC, CD206 on MΦ and BMDC, but not on monocytes and MoDC. CD1c was not found on monocytes but on all in vitro differentiated cells. FL-BMDC and GM-BMDC were partially positive for CD4 and CD8. CD45RA was expressed on a subset of FL-BMDC but not on MoDC and GM-BMDC. MoDC and FL-DC responded well to TLR ligands including poly-IC (TLR2), Pam3Cys (TLR3), LPS (TLR4) and imiquimod (TLR7) by up-regulating MHC II and CD86. The generated DC and MΦ showed a stimulatory capacity for lymphocytes, which increased upon maturation with LPS. Taken together, our results are the basis for further characterization of canine DC subsets with respect to their role in inflammation and immune responses.

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Section: Discussionsupporting

confidence: 91%

Comparative analysis of canine monocyte- and bone-marrow-derived dendritic cells

Gutzwiller

Moulin²,

Zurbriggen

et al. 2010

Vet. Res.

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“…In horses, previous studies to generate MoDC under the influence of GM-CSF and IL-4 used FBS [14, 16, 17] and pilot studies have also shown the potential use of such ex vivo generated MoDC for treatment of tumors [40]. However, previous experience where FBS-specific IgE was induced through MoDC application [36], eliminated such equine MoDC for the purpose of allergen immunotherapy.…”

Section: Discussionmentioning

confidence: 99%

Equine dendritic cells generated with horse serum have enhanced functionality in comparison to dendritic cells generated with fetal bovine serum

et al. 2016

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BackgroundDendritic cells are professional antigen-presenting cells that play an essential role in the initiation and modulation of T cell responses. They have been studied widely for their potential clinical applications, but for clinical use to be successful, alternatives to xenogeneic substances like fetal bovine serum (FBS) in cell culture need to be found. Protocols for the generation of dendritic cells ex vivo from monocytes are well established for several species, including horses. Currently, the gold standard protocol for generating dendritic cells from monocytes across various species relies upon a combination of GM-CSF and IL-4 added to cell culture medium which is supplemented with FBS. The aim of this study was to substitute FBS with heterologous horse serum. For this purpose, equine monocyte-derived dendritic cells (eqMoDC) were generated in the presence of horse serum or FBS and analysed for the effect on morphology, phenotype and immunological properties. Changes in the expression of phenotypic markers (CD14, CD86, CD206) were assessed during dendritic cell maturation by flow cytometry. To obtain a more complete picture of the eqMoDC differentiation and assess possible differences between FBS- and horse serum-driven cultures, a transcriptomic microarray analysis was performed. Lastly, immature eqMoDC were primed with a primary antigen (ovalbumin) or a recall antigen (tetanus toxoid) and, after maturation, were co-cultured with freshly isolated autologous CD5+ T lymphocytes to assess their T cell stimulatory capacity.ResultsThe microarray analysis demonstrated that eqMoDC generated with horse serum were indistinguishable from those generated with FBS. However, eqMoDC incubated with horse serum-supplemented medium exhibited a more characteristic dendritic cell morphology during differentiation from monocytes. A significant increase in cell viability was also observed in eqMoDC cultured with horse serum. Furthermore, eqMoDC generated in the presence of horse serum were found to be superior in their functional T lymphocyte priming capacity and to elicit significantly less non-specific proliferation.ConclusionsEqMoDC generated with horse serum-supplemented medium showed improved morphological characteristics, higher cell viability and exhibited a more robust performance in the functional T cell assays. Therefore, horse serum was found to be superior to FBS for generating equine monocyte-derived dendritic cells.

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“…As in humans and mice, the equine AMLR was contact and MHC class II dependent and primarily involved CD3 ϩ CD4 ϩ CD8 Ϫ T cells (24,27). Equine DCs express high levels of MHC class II, contain a subpopulation of cells with a mature flow cytometric phenotype, and more potently induce the AMLR compared to macrophages (9). This confirms the superior antigen-presenting and costimulatory capacity of equine DCs, because the AMLR requires DC expression of CD86 and MHC class II (44).…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, the AMLR displays characteristics of a normal immune response (including specificity and memory) and is reduced in a variety of human disease states (23, 53). Furthermore, the self-reactive T cells in the AMLR demonstrate a capacity for immunosuppression and increased transcription of the Treg transcription factor FOXP3, suggesting that such DC-stimulated T cells are involved in immune regulation during the normal immune response in vivo (25,45,51).A protocol for generating monocyte-derived DCs has been described in the horse, and the phenotype of these cells has been characterized by several groups (9,12,19). However, relatively little is known about the ability of equine DCs to induce proliferation and differentiation of autologous T cells.…”

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confidence: 99%

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Activation-Induced FoxP3 Expression Regulates Cytokine Production in Conventional T Cells Stimulated with Autologous Dendritic Cells

Cavatorta

Erb

Felippe

2012

Clin. Vaccine Immunol.

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A defining feature of dendritic cells (DCs) is their ability to induce the proliferation of autologous T cells in the absence of foreign antigen-a process termed the "autologous mixed leukocyte reaction" (AMLR). We report that equine monocytederived DCs, but not macrophages, are potent inducers of the AMLR. The response is contact dependent and major histocompatibility complex class II dependent and primarily involves CD3؉ CD4 ؉ CD8 ؊ T cells. Upon stimulation with DCs or the mitogen concanavalin A, a subset of the proliferating T cells expresses the regulatory T-cell (Treg) transcription factor FoxP3. Although many of these FoxP3 ؉ T cells are capable of producing the effector cytokines interleukin-4 (IL-4) and gamma interferon (IFN-␥), they are more likely to produce IL-10 and less likely to produce IFN-␥ than equivalent FoxP3 DCs take up antigen from the peripheral tissues, process it, and present it to naive T cells in the draining lymph node. Depending on the local cytokine environment during DC-mediated T cell activation, the antigen-specific T cell can proliferate and differentiate into one of a variety of effector T cell phenotypes, including (i) gamma interferon (IFN-␥)-producing Th1 cells that are important for immunity to intracellular pathogens and tumors; (ii) interleukin-4 (IL-4)-, IL-5-, and IL-13-producing Th2 cells that promote antibody production, help prevent parasitic diseases, and are involved in allergic responses; and (iii) IL-17-producing Th17 cells that protect against extracellular infections (58). DCs also play a critical role in immune regulation because they are capable of both expanding thymus-derived FoxP3 ϩ (designated FOXP3 in humans) natural regulatory T cells (nTregs) and inducing naive T cells to develop into induced regulatory T cells (iTregs) in the periphery (55, 56). In turn, Tregs use a variety of mechanisms (such as the production of the anti-inflammatory cytokines IL-10 and transforming growth factor ␤ [TGF-␤]) to keep effector responses in check and to prevent immune-mediated disease (7).DCs also possess the unique ability to efficiently induce proliferation of autologous T cells in the absence of exogenous antigen. This process was first demonstrated with murine DCs several decades ago and was termed the "autologous mixed leukocyte reaction" (AMLR) (39). The AMLR has since been described with human cells and is thought to represent polyclonal activation of autoreactive T cells specific for self-antigens presented by DCs (2,3,10,38,44). Interestingly, the AMLR displays characteristics of a normal immune response (including specificity and memory) and is reduced in a variety of human disease states (23, 53). Furthermore, the self-reactive T cells in the AMLR demonstrate a capacity for immunosuppression and increased transcription of the Treg transcription factor FOXP3, suggesting that such DC-stimulated T cells are involved in immune regulation during the normal immune response in vivo (25,45,51).A protocol for generating monocyte-derived DCs has been described in the h...

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Ex vivo generation of mature equine monocyte-derived dendritic cells

Cited by 15 publications

References 36 publications

Comparative analysis of canine monocyte- and bone-marrow-derived dendritic cells

Comparative analysis of canine monocyte- and bone-marrow-derived dendritic cells

Equine dendritic cells generated with horse serum have enhanced functionality in comparison to dendritic cells generated with fetal bovine serum

Activation-Induced FoxP3 Expression Regulates Cytokine Production in Conventional T Cells Stimulated with Autologous Dendritic Cells

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