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Differential capacity of human interleukin-4 and interferon-α monocyte-derived dendritic cells for cross-presentation of free versus cell-associated antigen.

Ruben JM, Bontkes HJ, Westers TM, Hooijberg E, Ossenkoppele GJ, de Gruijl TD, van de Loosdrecht AA - Cancer Immunol. Immunother. (2015)

Bottom Line: One of the particular interest is the induction of an immune response targeting multiple (unknown) tumor-associated antigens (TAA), which requires a polyvalent source of TAA.Recent reports suggest that MoDC cultured in the presence of GM-CSF supplemented with IFNα (IFNα MoDC), as compared to IL-4 (IL-4 MoDC), have an increased capacity to cross-present antigen to CD8(+) T cells.In conclusion, our data indicate the use of IFNα MoDC over IL-4 MoDC in the context of DC vaccination with SLP, whereas IL-4 MoDC are preferred for vaccination with bleb-derived antigens.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology, Cancer Center Amsterdam, VU University Medical Center, De Boelelaan 1117, 1081HV, Amsterdam, The Netherlands.

ABSTRACT
Dendritic cells (DC) vaccination is a potent therapeutic approach for inducing tumor-directed immunity, but challenges remain. One of the particular interest is the induction of an immune response targeting multiple (unknown) tumor-associated antigens (TAA), which requires a polyvalent source of TAA. Previously, we described the preferred use of apoptotic cell-derived blebs over the larger apoptotic cell remnants, as a source of TAA for both in situ loading of skin-resident DC and in vitro loading of monocyte-derived DC (MoDC). Recent reports suggest that MoDC cultured in the presence of GM-CSF supplemented with IFNα (IFNα MoDC), as compared to IL-4 (IL-4 MoDC), have an increased capacity to cross-present antigen to CD8(+) T cells. As culture conditions, maturation methods and antigen sources differ between the conducted studies, we analyzed the functional differences between IL-4 MoDC and IFNα MoDC, loaded with blebs, in a head-to-head comparison using commonly used protocols. Our data show that both MoDC types are potent (cross-) primers of CD8(+) T cells. Whereas IFNα MoDC were more potent in their capacity to cross-present a 25-mer MART-1 synthetic long peptide (SLP) to a MART-1aa26-35 recognizing CD8(+) T cell line, IL-4 MoDC proved more potent cross-primers of antigen-specific CD8(+) T cells when loaded with blebs. The latter is likely due to the observed greater capacity of IL-4 MoDC to ingest apoptotic blebs. In conclusion, our data indicate the use of IFNα MoDC over IL-4 MoDC in the context of DC vaccination with SLP, whereas IL-4 MoDC are preferred for vaccination with bleb-derived antigens.

No MeSH data available.


Related in: MedlinePlus

Analysis of MoDC immunophenotype and endocytic capacity. a Immunophenotype of monocyte-derived dendritic cells (MoDC) generated in the presence of either GM-CSF and IL-4 (black bars), or GM-CSF and IFNα (white bars), for, respectively, 5 or 3 days, (n = 4). *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001. b Immunophenotype of MoDC 48 h after loading with blebs and subsequent maturation induction by IL-1β, IL-6, TNFα, and PGE-2 (n = 4). *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001. c Fluorescently labeled MoDC were co-cultured overnight with fluorescently labeled blebs, after which the percentage of double-positive IL-4 (black bars), or IFNα MoDC (white bars), were quantified (apoptotic blebs). Alternatively, MoDC were cultured in the presence of Lucifer Yellow (pinocytosis), or dextran-FITC (receptor-mediated endocytosis). (n = 4). d Expression levels of scavenger receptors on immature IL-4 (black bars), or IFNα MoDC (white bars), using flow cytometry (n = 4). Statistical significance was for all figures determined using a Student’s t test
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Fig1: Analysis of MoDC immunophenotype and endocytic capacity. a Immunophenotype of monocyte-derived dendritic cells (MoDC) generated in the presence of either GM-CSF and IL-4 (black bars), or GM-CSF and IFNα (white bars), for, respectively, 5 or 3 days, (n = 4). *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001. b Immunophenotype of MoDC 48 h after loading with blebs and subsequent maturation induction by IL-1β, IL-6, TNFα, and PGE-2 (n = 4). *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001. c Fluorescently labeled MoDC were co-cultured overnight with fluorescently labeled blebs, after which the percentage of double-positive IL-4 (black bars), or IFNα MoDC (white bars), were quantified (apoptotic blebs). Alternatively, MoDC were cultured in the presence of Lucifer Yellow (pinocytosis), or dextran-FITC (receptor-mediated endocytosis). (n = 4). d Expression levels of scavenger receptors on immature IL-4 (black bars), or IFNα MoDC (white bars), using flow cytometry (n = 4). Statistical significance was for all figures determined using a Student’s t test

Mentions: We first assessed MoDC phenotype after a 3- or 5-day differentiation period for IFNα or IL-4 MoDC, respectively (Fig. 1a). IFNα MoDC had significantly higher expression levels of HLA class II (p = 0.04), and CD14 (p = 0.0005), as compared to IL-4 MoDC. Moreover, IFNα MoDC showed a higher expression of CD80 (p = 0.01) and CD86 (p = 0.03). Next, we assessed the MoDC phenotype upon maturation induction in the presence of apoptotic blebs. In contrast to the immature state, IL-4 MoDC displayed a significantly higher expression of both HLA class I (p = 0.0006) and HLA class II (p = 0.01) and CD1a (p = 0.009) following loading with blebs and subsequent maturation (Fig. 1b). Moreover, IL-4 MoDC had a significantly higher expression of CD40 (p = 0.003), CD80 (p = 0.002), CD83 (p = 0.001), and CD86 (p = 0.002), whereas IFNα MoDC retained a higher CD14 expression after maturation (p = 0.009). After inducing maturation, IFNα MoDC down-regulated CD1a, compared to their immature expression levels (p = 0.02), at levels lower than those on mature IL-4 MoDC (p = 0.002).Fig. 1


Differential capacity of human interleukin-4 and interferon-α monocyte-derived dendritic cells for cross-presentation of free versus cell-associated antigen.

Ruben JM, Bontkes HJ, Westers TM, Hooijberg E, Ossenkoppele GJ, de Gruijl TD, van de Loosdrecht AA - Cancer Immunol. Immunother. (2015)

Analysis of MoDC immunophenotype and endocytic capacity. a Immunophenotype of monocyte-derived dendritic cells (MoDC) generated in the presence of either GM-CSF and IL-4 (black bars), or GM-CSF and IFNα (white bars), for, respectively, 5 or 3 days, (n = 4). *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001. b Immunophenotype of MoDC 48 h after loading with blebs and subsequent maturation induction by IL-1β, IL-6, TNFα, and PGE-2 (n = 4). *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001. c Fluorescently labeled MoDC were co-cultured overnight with fluorescently labeled blebs, after which the percentage of double-positive IL-4 (black bars), or IFNα MoDC (white bars), were quantified (apoptotic blebs). Alternatively, MoDC were cultured in the presence of Lucifer Yellow (pinocytosis), or dextran-FITC (receptor-mediated endocytosis). (n = 4). d Expression levels of scavenger receptors on immature IL-4 (black bars), or IFNα MoDC (white bars), using flow cytometry (n = 4). Statistical significance was for all figures determined using a Student’s t test
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Related In: Results  -  Collection

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Fig1: Analysis of MoDC immunophenotype and endocytic capacity. a Immunophenotype of monocyte-derived dendritic cells (MoDC) generated in the presence of either GM-CSF and IL-4 (black bars), or GM-CSF and IFNα (white bars), for, respectively, 5 or 3 days, (n = 4). *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001. b Immunophenotype of MoDC 48 h after loading with blebs and subsequent maturation induction by IL-1β, IL-6, TNFα, and PGE-2 (n = 4). *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001. c Fluorescently labeled MoDC were co-cultured overnight with fluorescently labeled blebs, after which the percentage of double-positive IL-4 (black bars), or IFNα MoDC (white bars), were quantified (apoptotic blebs). Alternatively, MoDC were cultured in the presence of Lucifer Yellow (pinocytosis), or dextran-FITC (receptor-mediated endocytosis). (n = 4). d Expression levels of scavenger receptors on immature IL-4 (black bars), or IFNα MoDC (white bars), using flow cytometry (n = 4). Statistical significance was for all figures determined using a Student’s t test
Mentions: We first assessed MoDC phenotype after a 3- or 5-day differentiation period for IFNα or IL-4 MoDC, respectively (Fig. 1a). IFNα MoDC had significantly higher expression levels of HLA class II (p = 0.04), and CD14 (p = 0.0005), as compared to IL-4 MoDC. Moreover, IFNα MoDC showed a higher expression of CD80 (p = 0.01) and CD86 (p = 0.03). Next, we assessed the MoDC phenotype upon maturation induction in the presence of apoptotic blebs. In contrast to the immature state, IL-4 MoDC displayed a significantly higher expression of both HLA class I (p = 0.0006) and HLA class II (p = 0.01) and CD1a (p = 0.009) following loading with blebs and subsequent maturation (Fig. 1b). Moreover, IL-4 MoDC had a significantly higher expression of CD40 (p = 0.003), CD80 (p = 0.002), CD83 (p = 0.001), and CD86 (p = 0.002), whereas IFNα MoDC retained a higher CD14 expression after maturation (p = 0.009). After inducing maturation, IFNα MoDC down-regulated CD1a, compared to their immature expression levels (p = 0.02), at levels lower than those on mature IL-4 MoDC (p = 0.002).Fig. 1

Bottom Line: One of the particular interest is the induction of an immune response targeting multiple (unknown) tumor-associated antigens (TAA), which requires a polyvalent source of TAA.Recent reports suggest that MoDC cultured in the presence of GM-CSF supplemented with IFNα (IFNα MoDC), as compared to IL-4 (IL-4 MoDC), have an increased capacity to cross-present antigen to CD8(+) T cells.In conclusion, our data indicate the use of IFNα MoDC over IL-4 MoDC in the context of DC vaccination with SLP, whereas IL-4 MoDC are preferred for vaccination with bleb-derived antigens.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology, Cancer Center Amsterdam, VU University Medical Center, De Boelelaan 1117, 1081HV, Amsterdam, The Netherlands.

ABSTRACT
Dendritic cells (DC) vaccination is a potent therapeutic approach for inducing tumor-directed immunity, but challenges remain. One of the particular interest is the induction of an immune response targeting multiple (unknown) tumor-associated antigens (TAA), which requires a polyvalent source of TAA. Previously, we described the preferred use of apoptotic cell-derived blebs over the larger apoptotic cell remnants, as a source of TAA for both in situ loading of skin-resident DC and in vitro loading of monocyte-derived DC (MoDC). Recent reports suggest that MoDC cultured in the presence of GM-CSF supplemented with IFNα (IFNα MoDC), as compared to IL-4 (IL-4 MoDC), have an increased capacity to cross-present antigen to CD8(+) T cells. As culture conditions, maturation methods and antigen sources differ between the conducted studies, we analyzed the functional differences between IL-4 MoDC and IFNα MoDC, loaded with blebs, in a head-to-head comparison using commonly used protocols. Our data show that both MoDC types are potent (cross-) primers of CD8(+) T cells. Whereas IFNα MoDC were more potent in their capacity to cross-present a 25-mer MART-1 synthetic long peptide (SLP) to a MART-1aa26-35 recognizing CD8(+) T cell line, IL-4 MoDC proved more potent cross-primers of antigen-specific CD8(+) T cells when loaded with blebs. The latter is likely due to the observed greater capacity of IL-4 MoDC to ingest apoptotic blebs. In conclusion, our data indicate the use of IFNα MoDC over IL-4 MoDC in the context of DC vaccination with SLP, whereas IL-4 MoDC are preferred for vaccination with bleb-derived antigens.

No MeSH data available.


Related in: MedlinePlus