Journal: Frontiers in Immunology

Article Title: Decreased CD57 expression of natural killer cells enhanced cytotoxicity in patients with primary sclerosing cholangitis

doi: 10.3389/fimmu.2022.912961

Figure Lengend Snippet: Characteristics of peripheral NK cells in patients with PSC. PBMCs from PSC and HCs were isolated and stained with fluorochrome-conjugated antibodies for cell subsets. (A) Strategy of FCM of NK cell subsets. CD3 and CD56 expression in PBMCs was analyzed after application of a lymphocyte gate. CD3-negative cells were further gated for analysis of CD3 − CD56 bright CD16 − (CD56 bright ) and CD3 − CD56 dim CD16 − (CD56 dim ) NK cell subsets. (B) The frequency (%) of CD56 bright and CD56 dim cells in total lymphocytes and (C) MFI of natural cytotoxicity receptors in each NK subset were compared between PSC (n = 34) and HCs (n = 29). *, p < 0.05; **, p < 0.01 (two-tailed Mann–Whitney test). NK, natural killer; PSC, primary sclerosing cholangitis; PBMCs, peripheral blood mononuclear cells; HCs, healthy controls; FCM, flow cytometry; MFI, mean fluorescence intensity.

Article Snippet: NK cells were purified from the PBMCs using a human CD56 + CD16 + NK cell isolation kit, following the manufacturer’s recommendations (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Isolation, Staining, Expressing, Two Tailed Test, MANN-WHITNEY, Flow Cytometry, Fluorescence

Journal: Frontiers in Immunology

Article Title: Decreased CD57 expression of natural killer cells enhanced cytotoxicity in patients with primary sclerosing cholangitis

doi: 10.3389/fimmu.2022.912961

Figure Lengend Snippet: The expression of surface molecules on CD56 dim NK cells from patients with PSC. PBMCs were stained with fluorochrome-conjugated antibodies for different markers and analyzed by FCM. Percentage (%) of positive cells for each molecule marker in CD56 dim NK cells or MFI of each molecule were compared between PSC (n = 34) and HCs (n = 29). *, p < 0.05 (two-tailed unpaired t-test or non-parametric Mann–Whitney test). NK, natural killer; PSC, primary sclerosing cholangitis; PBMCs, peripheral blood mononuclear cells; FCM, flow cytometry; MFI, mean fluorescence intensity; HCs, healthy controls.

Article Snippet: NK cells were purified from the PBMCs using a human CD56 + CD16 + NK cell isolation kit, following the manufacturer’s recommendations (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Expressing, Staining, Marker, Two Tailed Test, MANN-WHITNEY, Flow Cytometry, Fluorescence

Journal: Frontiers in Immunology

Article Title: Decreased CD57 expression of natural killer cells enhanced cytotoxicity in patients with primary sclerosing cholangitis

doi: 10.3389/fimmu.2022.912961

Figure Lengend Snippet: Phenotypic changes related to CD57 expression in PSC CD56 dim NK cells. (A) Pattern of NKp46 versus CD57 expression in CD56 bright and CD56 dim NK cell subpopulations. (B) Linear regression analysis for the correlation between the frequency of CD56 dim and CD57 + NK cells and between the MFI level of NKp46 and percentage of CD57 + cells in CD56 dim NK cells from PSC (n = 34) and HCs (n = 29). The p and r 2 values are indicated in each graph. (C) Expression of different markers on CD57 − and CD57 + CD56 dim NK cells. NK cells from PSC (n = 17) were compared with HCs (n = 15). *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001 (two-tailed unpaired t-test or non-parametric test for comparisons between PSC and HCs; two-tailed paired t-test or non-parametric test for CD57 − versus CD57 + NK cells in either patients or HCs). PSC, primary sclerosing cholangitis; NK, natural killer; MFI, mean fluorescence intensity; HCs, healthy controls.

Article Snippet: NK cells were purified from the PBMCs using a human CD56 + CD16 + NK cell isolation kit, following the manufacturer’s recommendations (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Expressing, Two Tailed Test, Fluorescence

Journal: Frontiers in Immunology

Article Title: Decreased CD57 expression of natural killer cells enhanced cytotoxicity in patients with primary sclerosing cholangitis

doi: 10.3389/fimmu.2022.912961

Figure Lengend Snippet: Correlation of CD57 + CD56 dim NK cells with disease severity of PSC. (A) Scatter diagram showing the relationship between the frequency (%) of CD57 + cells in CD56 dim NK cells and the levels of biochemistry markers in patients with PSC (n = 25). The p and r 2 values are indicated in each graph. (B) The expression of NKp46 (MFI) and CD57 (%) was analyzed in CD56 dim NK cells from PSC patients without IBD (PSC w/o IBD, n = 16) and those with IBD (PSC w/IBD, n = 17). NK, natural killer; PSC, primary sclerosing cholangitis; MFI, mean fluorescence intensity; IBD, inflammatory bowel disease.

Article Snippet: NK cells were purified from the PBMCs using a human CD56 + CD16 + NK cell isolation kit, following the manufacturer’s recommendations (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Expressing, Fluorescence

Journal: Frontiers in Immunology

Article Title: Decreased CD57 expression of natural killer cells enhanced cytotoxicity in patients with primary sclerosing cholangitis

doi: 10.3389/fimmu.2022.912961

Figure Lengend Snippet: Cytotoxicity of CD56 dim NK cells. (A) CD107a degranulation in CD56 dim NK cells were compared between PSC (n = 12) and HCs (n = 13). PBMCs were cultured with or without IL-2 for 16 h. After being washed with culture medium, the cells were re-incubated with K562 cells at serial E:T (PBMC effector to K562 target) ratios in the presence of anti-CD107a antibody for 4 h. The percentages of CD107a + cells in CD56 dim NK cells were analyzed by FCM. (B) NKp46-mediated NK cells redirected killing of P815 target cells by purified CD16 + CD56 dim NK cells from PBMCs. Percentage of apoptotic Annexin V + P815 cells was compared between PSC and HCs. *, p < 0.05 (two-tailed non-parametric Mann–Whitney test). NK, natural killer; PSC, primary sclerosing cholangitis; PBMCs, peripheral blood mononuclear cells; FCM, flow cytometry.

Article Snippet: NK cells were purified from the PBMCs using a human CD56 + CD16 + NK cell isolation kit, following the manufacturer’s recommendations (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Cell Culture, Incubation, Purification, Two Tailed Test, MANN-WHITNEY, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Decreased CD57 expression of natural killer cells enhanced cytotoxicity in patients with primary sclerosing cholangitis

doi: 10.3389/fimmu.2022.912961

Figure Lengend Snippet: Cytotoxicity of CD57 + versus CD57 − CD56 dim NK cells in PSC. (A) PBMCs from PSC (n = 9) or HCs (n = 8) were co-incubated with K562 target cells at an E:T ratio of 1:1 in the presence of anti-CD107a antibody for 4 h. The percentages of CD107a + CD57 + and CD107a + CD57 − cells in the CD56 dim NK cell subset were compared. (B, C) PBMCs from PSC patients (n = 15) and HCs (n = 9) were cultured with the K562 target cells at an E:T ratio of 1:1 in an anti-NKp46 Ab-coated plate for 4 h. CD107a + cells (B) and intracellular IFN-γ + cells (C) in NK cells were analyzed by FCM. *p < 0.05; **p < 0.01 (two-tailed unpaired t-test or non-parametric test for comparisons between PSC and HCs; two-tailed paired t-test or non-parametric test for CD57 − versus CD57 + NK cells in either patients or HCs). NK, natural killer; PSC, primary sclerosing cholangitis; PBMCs, peripheral blood mononuclear cells; HCs, healthy controls.

Article Snippet: NK cells were purified from the PBMCs using a human CD56 + CD16 + NK cell isolation kit, following the manufacturer’s recommendations (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Incubation, Cell Culture, Two Tailed Test

Journal: BMC Biotechnology

Article Title: Quantitative whole-cell MALDI-TOF MS fingerprints distinguishes human monocyte sub-populations activated by distinct microbial ligands

doi: 10.1186/s12896-015-0140-1

Figure Lengend Snippet: Phenotypic characterization of isolated monocyte populations. Representative FACS-plot and scatter-plot frequencies of A) Pan-monocytes and B) CD16 + monocytes based on the expression of CD14 versus CD16 (left plots) and CD16 versus P-selectin glycoprotein ligand-1 (MDC8 antibody specificity , right plots). Scatter-plots summarizing frequencies of each sub-population based on the combinations between the three monocytic markers are shown across all monocytes preparations (n = 8 healthy donors).

Article Snippet: Pan- and CD16+ monocyte purification were performed using Pan Monocyte Isolation Kit and CD16+ Monocyte Isolation Kit respectively according to manufacturer’s instructions (Miltenyi Biotec GmbH).

Techniques: Isolation, Expressing

Journal: BMC Biotechnology

Article Title: Quantitative whole-cell MALDI-TOF MS fingerprints distinguishes human monocyte sub-populations activated by distinct microbial ligands

doi: 10.1186/s12896-015-0140-1

Figure Lengend Snippet: Quantitative whole-cell MALDI-TOF MS discriminates resting and stimulated human monocyte subpopulations. Principal component analysis of all samples combined (A) ; stimulated and resting pan-monocytes (n = 8) (B) and CD16+ monocytes (n = 7) (C) , respectively, based on the peak intensity matrix.

Article Snippet: Pan- and CD16+ monocyte purification were performed using Pan Monocyte Isolation Kit and CD16+ Monocyte Isolation Kit respectively according to manufacturer’s instructions (Miltenyi Biotec GmbH).

Techniques:

Journal: BMC Biotechnology

Article Title: Quantitative whole-cell MALDI-TOF MS fingerprints distinguishes human monocyte sub-populations activated by distinct microbial ligands

doi: 10.1186/s12896-015-0140-1

Figure Lengend Snippet: Whole-cell MALDI-TOF MS based identification of biomarkers specific for pan-monocyte and CD16+ monocytes. A) Smoothed mass spectra quadruplicates obtained from resting pan-monocytes (n = 8, 32 red spectra) overlaid with those obtained from CD16+ monocytes (n = 7, 28 green spectra). m/z peaks significantly over-represented in pan-monocytes (left overlay plot) or CD16+ monocytes (right overlay plot) are indicated. B) Scatter-plot of quadruplicate’s averaged mass peak intensities from pan-monocytes and CD16+ monocytes from individual donor are shown for the peak at 9296.1 m/z (left) and 9720.3 m/z (right). The two-way ANOVA with Bonferroni post-tests correction was used with a significance at p < 0.001.

Article Snippet: Pan- and CD16+ monocyte purification were performed using Pan Monocyte Isolation Kit and CD16+ Monocyte Isolation Kit respectively according to manufacturer’s instructions (Miltenyi Biotec GmbH).

Techniques:

Journal: BMC Biotechnology

Article Title: Quantitative whole-cell MALDI-TOF MS fingerprints distinguishes human monocyte sub-populations activated by distinct microbial ligands

doi: 10.1186/s12896-015-0140-1

Figure Lengend Snippet: Whole-cell MALDI-TOF can monitor immune activation events. Vent diagrams depicting for A) Pan-monocytes or B) CD16+ monocytes, m/z peak intensities significantly different between the detailed culture conditions (two-way ANOVA statistical analysis, p < 0.001). C) Smoothed mass spectra quadruplicates and respective scatter-plot of quadruplicate’s averaged mass peak intensities from resting CD16 + monocytes (green overlay) and LPS-treated CD16 + monocytes (red overlay) highlighting m/z areas containing putative phosphorylation of m/z 11316.24 into m/z 11396.28 (two-way ANOVA with Bonferroni post-tests correction, *** p < 0.001).

Article Snippet: Pan- and CD16+ monocyte purification were performed using Pan Monocyte Isolation Kit and CD16+ Monocyte Isolation Kit respectively according to manufacturer’s instructions (Miltenyi Biotec GmbH).

Techniques: Activation Assay, Phospho-proteomics