Quantification of ex vivo generated dendritic cells (DC) and leukemia-derived DC contributes to estimate the quality of DC, to detect optimal DC-generating methods or to optimize DC-mediated T-cell-activation-procedures ex vivo or in vivo

Hm, Schmetzer; Kremser, Andreas; Loibl, Julia; Kroell, Tanja; Hj, Kolb

doi:10.1038/sj.leu.2404639

Cited by 40 publications

(57 citation statements)

References 11 publications

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“…34 We defined DC generation as successful if at least >10% DC and >5% leukemia-derived DC (DC leu ) could be generated in the MNC fraction. 32 DC leu were defined by the coexpression of blast-with DC-markers.…”

Section: Generationmentioning

confidence: 99%

“…Quantification and characterization of DC and DC leu were performed by flow cytometry according to our gating strategy already described by former groups. [30][31][32][33][34] To quantify and characterize the several T-cell subpopulations, we defined a "lympho-gate" surrounding lymphocytes and monocytes. T-cell subgroups were defined by antigen coexpression before and after 'MNC' or 'DC' stimulation as shown in Table 3.…”

Section: Mlcmentioning

confidence: 99%

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Antileukemic T-cell Responses Can Be Predicted by the Composition of Specific Regulatory T-cell Subpopulations

Schick¹,

Vogt²,

Zerwes

et al. 2013

Journal of Immunotherapy

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Summary: Regulatory T cells (T reg ) are important regulators of immune responses. In acute myeloid leukemia (AML) patients before/after immunotherapy (stem cell transplantation or donor lymphocyte infusion), their suppressive role can contribute to suppress severe graft-versus-host reactions, but also to impair antileukemic reactions. As leukemia-derived dendritic cells (DC leu ) are known to improve the antileukemic functionality of T cells, we evaluated the composition and development of distinct T reg subtypes in AML patients (n = 12) compared with healthy probands (n = 5) under unstimulated conditions and during stimulation with DC leu -containing DC (DC) or blast-containing mononuclear cells (MNC) in 0-to 7-day mixed lymphocyte cultures by flow cytometry. T-cell subgroups in AML patients were correlated with antileukemic functionality before and after DC or MNC stimulation by functional fluorolysis assays. (1) AML patients' T cells presented with significantly higher frequencies of T reg subgroups in unstimulated T cells compared with healthy probands. (2) After 7 days of DC or MNC stimulation, all T reg subtypes generally increased; significantly higher frequencies of T reg subtypes were still found in AML patients. (3) Antileukemic cytotoxicity was achieved in 36% of T cells after MNC compared with 64% after DC stimulation. Antileukemic activity after DC but not after MNC stimulation correlated with significantly lower frequencies of T reg subtypes (CD8 + T reg /T eff/em reg ). Furthermore, cut-off values for T reg subpopulations could be defined, allowing a prediction of antileukemic response. We demonstrate a crucial role of special T reg subtypes in the mediation of antileukemic functionality. High CD8 + T reg , T eff/em reg , and CD39 + T cells correlated clearly with a reduced antileukemic activity of T cells. DC stimulation of T cells contributes to overcome impaired antileukemic T-cell reactivity. Refined analyses in the context of clinical responses to immunotherapies and graft versus host reactions are required.

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Section: Generationmentioning

confidence: 99%

Section: Mlcmentioning

confidence: 99%

Antileukemic T-cell Responses Can Be Predicted by the Composition of Specific Regulatory T-cell Subpopulations

Schick¹,

Vogt²,

Zerwes

et al. 2013

Journal of Immunotherapy

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“…Proportions of positive events in defined gates compared with the isotype controls were calculated using Cell Quest software (Becton Dickinson, BD, Heidelberg, Germany). Quantification and characterization of DC and DC leu were performed by Flow cytometry according to our gating strategy already described by former groups (Schmetzer et al 2007;Kremser et al 2010;Liepert et al 2010;Grabrucker et al 2010;Dreyßig et al 2011). Flow cytometric analyses were carried out on day 0 and 10 followed by a functional cytotoxic fluorolysis assay on day 10 as described below.…”

Section: Mixed Lymphocyte Culture (Mlc)mentioning

confidence: 99%

Cytokine Release Patterns in Mixed Lymphocyte Culture (MLC) of T-Cells with Dendritic Cells (DC) Generated from AML Blasts Contribute to Predict anti-Leukaemic T-Cell Reactions and Patients’ Response to Immunotherapy

Fischbacher¹,

Merle²,

Liepert³

et al. 2015

Cell Communication & Adhesion

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To enlighten interactions between autologous, allogeneic or T-cells from patients after stem cell transplantation with leukaemia-derived-dendritic-cells containing dendritic cells or blast containing mononuclear cells (n ¼ 21, respectively), we determined cytokine-concentrations (interleukin 2, 4, 6, 10, tumor-necrosis-factor-a, interferon-c) in supernatants of mixed-lymphocyte-culture and in serum (n ¼ 16) of 20 patients with acute myeloid leukaemia and three patients with myelodysplastic syndromes by cytometric-bead-assay. We correlated our data with lytic capabilities of stimulated T-cells in a fluorolysis-assay and clinical data: Dendritic-cell-/mononuclear-cell-stimulation of T-cells resulted in increased cytokine-levels in culture-medium compared to serum. There were no significant differences between cytokine-patterns of cases with/without lytic T-cell-activity, response to immunotherapy (stem cell transplantation/donor-lymphocyte-infusion) or graft-versus-host-disease. However, some predictive cytokine-cut-off-values for antileukaemic T-cell-activity, patients' response to immunotherapy and graft-versus-host-disease could be defined. Cytokine-profiles alone, without functional assays, are no useful tool to predict antileukaemic T-cell-function, although they can indicate lytic T-cell-activity, patients' response to immunotherapy and graft-versus-host-disease. ARTICLE HISTORY

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“…For analysis and quantification of the lymphocytes, monocytes and leukemic cells before culture the total MNC fractions were gated. An AML sample was considered as 'positive' for a surface marker, if the percentage of positive events in a gate surrounding the blasts, lymphocytes and monocytes was more than 20 %, as described [21][22][23].…”

Section: Flow Cytometrymentioning

confidence: 99%

“…In most cases, stimulation with DC leu containing DC fractions (DC/DC leu ) resulted in a very effective cytotoxic T-cell response. But there are a few cases, where opposite T-cell response patterns have been observed, in terms of T cells mediating anergy or even supporting blast proliferation [20][21][22][23][24]. So far, these different T-cell response patterns are not predictable, but it might be expected that different 'qualities' of stimulators may result into different clonal compositions and functional diversities of T-cell subsets.…”

Section: Introductionmentioning

confidence: 99%

In vitro-induced response patterns of antileukemic T cells: characterization by spectratyping and immunophenotyping

et al. 2012

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Quantification of ex vivo generated dendritic cells (DC) and leukemia-derived DC contributes to estimate the quality of DC, to detect optimal DC-generating methods or to optimize DC-mediated T-cell-activation-procedures ex vivo or in vivo

Cited by 40 publications

References 11 publications

Antileukemic T-cell Responses Can Be Predicted by the Composition of Specific Regulatory T-cell Subpopulations

Antileukemic T-cell Responses Can Be Predicted by the Composition of Specific Regulatory T-cell Subpopulations

Cytokine Release Patterns in Mixed Lymphocyte Culture (MLC) of T-Cells with Dendritic Cells (DC) Generated from AML Blasts Contribute to Predict anti-Leukaemic T-Cell Reactions and Patients’ Response to Immunotherapy

In vitro-induced response patterns of antileukemic T cells: characterization by spectratyping and immunophenotyping

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