Journal: International Journal of Molecular Sciences

Article Title: Bone Marrow-Suppressive Treatment in Children Is Associated with Diminished IFN-γ Response from T Cells upon Polyclonal and Varicella Zoster Virus Peptide Stimulation

doi: 10.3390/ijms25136960

Figure Lengend Snippet: Simplified view of memory T cell subsets examined in this study, distinguished based on differential expression patterns of CCR7, CD45RO, and CD95. Naïve T cells are CCR7 + CD45RO − CD95 − ; central memory T cells (TCM) are CCR7 + CD45RO + CD95 + ; effector memory T cells (TEM) are CCR7 − CD45RO + CD95 + ; and terminal effector T cells, also named terminally differentiated T cells (TEFF), are CCR7 − CD45RO − CD95 + , in accordance with previous findings by Gattinoni et al. . As shown, the proliferative ability of T cells decreases along with differentiation and gain of effector function.

Article Snippet: Following the removal of the blocking medium, the T-cell mitogen Staphylococcal enterotoxin A (SEA, m Staphylococcus aureus , Sigma Aldrich, St. Louis, MO, USA), serving as polyclonal activator, was transferred to adequate wells at a final concentration of 20 ng/mL.

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: Bone Marrow-Suppressive Treatment in Children Is Associated with Diminished IFN-γ Response from T Cells upon Polyclonal and Varicella Zoster Virus Peptide Stimulation

doi: 10.3390/ijms25136960

Figure Lengend Snippet: Variations in T-cell populations in peripheral venous blood among the groups. ( A ) Gating of naïve (CCR7 + CD95 − ) and memory (CCR7 +/− ) T cells was performed on DCM − CD3 + single lymphocytes, which expressed either CD4 or CD8. Memory T cells were further sub-grouped into TCM (CCR7 + CD95 + ) or TEM/TEFF (CCR7 − CD95 + ) cells. Fluorescence minus one (FMO) controls were used for the gating of naïve and memory T-cell populations. Gating of the control cohort was similar, but included CD45RO, and is described in detail by Nilsson et al. . ( B ) The differences in the proportions of T-cell subpopulations among the three groups (n = 47), and according to age, specifically looking at CD4 + and CD8 + bulk T cells, CD8 + and CD4 + naïve T-cell subsets, as well as memory T-cell subsets for both CD4 + and CD8 + T cells. A Kruskal–Wallis, with Dunn’s multiple comparison test, was used on the grouped scatter plots to determine any significant differences. Statistical significance was determined as p < 0.05 (* p < 0.05, ** p < 0.01, *** p < 0.001), and the median is shown as a red horizontal line.

Article Snippet: Following the removal of the blocking medium, the T-cell mitogen Staphylococcal enterotoxin A (SEA, m Staphylococcus aureus , Sigma Aldrich, St. Louis, MO, USA), serving as polyclonal activator, was transferred to adequate wells at a final concentration of 20 ng/mL.

Techniques: Fluorescence, Control, Comparison

Journal: International Journal of Molecular Sciences

Article Title: Bone Marrow-Suppressive Treatment in Children Is Associated with Diminished IFN-γ Response from T Cells upon Polyclonal and Varicella Zoster Virus Peptide Stimulation

doi: 10.3390/ijms25136960

Figure Lengend Snippet: T-cell function in response to Staphylococcal enterotoxin A (SEA) and varicella zoster virus (VZV) peptides IE63 and gE in children with or without BM suppression. ( A ) Representative FluoroSpot wells at 1× magnification showing cellular expression of IL-22 (blue), IFN-γ (green), IL-10 (yellow), and IL-17A (red) when exposed to a negative control (dimethyl sulfoxide), T-cell mitogen SEA, or specific VZV peptides IE63 and gE. ( B ) Total counts per million of spot-forming cells (SFC) for individuals examined for all four cytokines following exposure to SEA (n = 40) and VZV (n = 38). ( C ) Cytokine profile depicting proportions of cytokine-producing T-cells based on the frequency—measured as SFCs/million cells—of IL-22-, IFN-γ-, IL-10-, and IL17-A-producing cells within each group. Significant differences after VZV exposure were found for the proportion of IL-22-secreting cells between groups I and III ( p < 0.05), for IFN-γ-secreting cells between groups I and III ( p < 0.01), and for IL-17A-secreting cells between groups I and III ( p < 0.01) and between groups II and III ( p < 0.05). ( D ) Frequencies of IL-22 (SEA n = 40, VZV n = 37), IFN-γ (SEA n = 44, VZV n = 41), IL-10 (SEA n = 44, VZV n = 41), and IL17-A (SEA n = 44, VZV n = 41) producing T cells shown at the polyclonal level, upon T-cell mitogen activation with SEA (top row) or at the antigen-specific level following activation with VZV peptides (bottom row). A Kruskal–Wallis test, with Dunn’s multiple comparison, was used to determine significant differences. Statistical significances were determined as p < 0.05 (* p < 0.05, ** p < 0.01, *** p < 0.001), and the median is shown as a red horizontal line.

Article Snippet: Following the removal of the blocking medium, the T-cell mitogen Staphylococcal enterotoxin A (SEA, m Staphylococcus aureus , Sigma Aldrich, St. Louis, MO, USA), serving as polyclonal activator, was transferred to adequate wells at a final concentration of 20 ng/mL.

Techniques: Cell Function Assay, Virus, Expressing, Negative Control, Activation Assay, Comparison

Journal: International Journal of Molecular Sciences

Article Title: Bone Marrow-Suppressive Treatment in Children Is Associated with Diminished IFN-γ Response from T Cells upon Polyclonal and Varicella Zoster Virus Peptide Stimulation

doi: 10.3390/ijms25136960

Figure Lengend Snippet: Residual spot volumes (RSV) upon stimulation with Staphylococcal enterotoxin A (SEA) or varicella zoster virus (VZV) peptides. ( A ) Schematic representation of RSV analysis. Using software algorithms, the RSV of individual spots was measured. These values were then determined as the area under the curve. ( B ) Differences in RSV from IL-22 (SEA n = 40, VZV n = 37), IFN-γ (SEA n = 44, VZV n = 40), IL-10 (SEA n = 44, VZV n = 40), and IL17-A (SEA n = 44, VZV n = 40) spot-forming cells following SEA or VZV stimuli among groups. Statistical analysis was performed using a Kruskal–Wallis test with Dunn’s multiple comparison. Statistical significance was determined as p < 0.05 (* p < 0.05, ** p < 0.01, **** p < 0.0001), and the median is shown as a red horizontal line.

Article Snippet: Following the removal of the blocking medium, the T-cell mitogen Staphylococcal enterotoxin A (SEA, m Staphylococcus aureus , Sigma Aldrich, St. Louis, MO, USA), serving as polyclonal activator, was transferred to adequate wells at a final concentration of 20 ng/mL.

Techniques: Virus, Software, Comparison