Journal: Nature Communications

Article Title: Toll-like receptor signaling in thymic epithelium controls monocyte-derived dendritic cell recruitment and Treg generation

doi: 10.1038/s41467-020-16081-3

Figure Lengend Snippet: a Representative flow cytometry plots (left plot) and their quantification (right plot) comparing the frequencies of main thymic T cell populations between MyD88 fl/fl and MyD88 ΔTECs mice (mean ± SEM, n = 14 mice). b Representative flow cytometry plots comparing the frequencies of CD4 + CD25 + Foxp3 + thymic Tregs between MyD88 fl/fl and MyD88 ΔTECs mice. c Quantification of frequencies from b (mean ± SEM, n = 14 mice). d Representative flow cytometry histograms showing the expression of CD73 by CD4 + CD25 + Foxp3 + thymic Tregs (gated as in b). e Quantification of the total numbers of CD73 – and CD73 + thymic Tregs from d (mean ± SEM, n = 7 mice). f Quantification of the frequencies of thymic Tregs from CpG ODN or PBS intrathymically stimulated (7 days) MyD88 fl/fl or MyD88 ΔTECs mice (mean ± SEM, n = 4 for MyD88 ΔTECs and n = 9 for MyD88 fl/fl mice). g Quantification of the total numbers of CD73 – and CD73 + thymic Tregs from CpG ODN or PBS intrathymically stimulated (7 days) WT (C57Bl/6J) mice (mean ± SEM, n = 6 for ODN + and n = 7 for ODN – mice). h Quantification of frequencies of thymic Tregs from CpG ODN or PBS intrathymically stimulated (7 days) H2-Ab1 fl/fl or H2-Ab1 fl/fl Itgax Cre (H2-Ab1 ΔDCs ) mice (mean ± SEM, n = 6 for H2-Ab1 fl/fl and ODN + H2-Ab1 ΔDCs and n = 7 for ODN – H2-Ab1 ΔDCs mice). i Quantification of the total numbers of CD73 – and CD73 + thymic Tregs from CpG ODN or PBS intrathymically stimulated (7 days) H2-Ab1 ΔDCs mice (mean ± SEM, n = 6 for ODN + and n = 7 for ODN – mice). Statistical analysis in a , c, e – i was performed by unpaired, two-tailed Student’s t -test, p ≤ 0.05 = *, p ≤ 0.01 = **, p ≤ 0.001*** p < 0.0001****, ns not significant.

Article Snippet: CD4-enriched T cells (CD4 + T Cell Isolation Kit, Miltenyi biotec) were stained by anti-CD25 biotin conjugated antibody and CD4 + CD25 + Tregs were isolated using Anti-Biotin MicroBeads (Miltenyi biotec).

Techniques: Flow Cytometry, Expressing, Two Tailed Test

Journal: Nature Communications

Article Title: Toll-like receptor signaling in thymic epithelium controls monocyte-derived dendritic cell recruitment and Treg generation

doi: 10.1038/s41467-020-16081-3

Figure Lengend Snippet: a Experimental design of induced colitis. b Relative quantification of mice weight normalized to its value on day 0 (100% of original weight) after T cell transfer over the time-course of the colitis experiment (mean ± SEM, n = 5–6 mice) Statistical analysis was performed by unpaired, two-tailed Student’s t -test comparing the relative weight of WT CD4 + with MyD88 ΔTECs CD4 + transferred mice (blue) or with WT CD4 + CD45RB high CD25 - transferred mice (red), p ≤ 0.05 = *, ns not significant. c Representative H&E-stained slides of colon sections performed 8 weeks after T cell transfer. Scale bar represents 500 μm ( n = 5 for CD4 + WT and n = 6 for CD4 + MyD88 ΔTECs and CD4 + No Tregs WT mice). d Relative quantification (normalized to average of control mice from each experiment) of colon weight/length ratio of T cell induced colitis experimental mice (mean ± SEM, n = 5–6 mice). e Relative quantification of the frequencies (normalized to average of control mice from each experiment) of CD4 + CD25 + Foxp3 + Tregs isolated from the spleens of experimental mice 8 weeks after T cell transfer (mean ± SEM, n = 5–6 mice). f Quantification of the Means fluorescent intensity (MFI) of CD25 protein expression in CD25 + Foxp3 + Tregs (gated as in Fig. ) in MyD88 fl/fl and MyD88 ΔTECs mice (mean ± SEM, n = 12 mice) Statistical analysis in b , d – f was performed by unpaired, two-tailed Student’s t -test, p ≤ 0.05 = *, p < 0.0001****, ns not significant. g Representative flow cytometry plots showing the frequency of proliferating OT-II T cells, co-cultivated with OVA pulsed BMDC and CD4 + CD25 + Tregs cells (alternatively with CD4 + CD25 – Tconv cells, black) isolated from LNs of MyD88 fl/fl (WT control, red) or MyD88 ΔTECs (blue) for 72 h. h Quantification of frequencies of proliferating OT-II Tcells form h (mean ± SEM, n = 4 wells from two independent experiments).

Article Snippet: CD4-enriched T cells (CD4 + T Cell Isolation Kit, Miltenyi biotec) were stained by anti-CD25 biotin conjugated antibody and CD4 + CD25 + Tregs were isolated using Anti-Biotin MicroBeads (Miltenyi biotec).

Techniques: Quantitative Proteomics, Two Tailed Test, Staining, Control, Isolation, Expressing, Flow Cytometry

Journal: iScience

Article Title: DUSP6 deletion protects mice and reduces disease severity in autoimmune arthritis

doi: 10.1016/j.isci.2024.110158

Figure Lengend Snippet:

Article Snippet: anti-CD25-APC (clone: 4E3) , Miltenyi Biotec, Germany , Cat# 130-113-846.

Techniques: Transgenic Assay, Knock-Out

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Th2 cells lacking T-bet suppress naive and memory T cell responses via IL-10

doi: 10.1073/pnas.2002787118

Figure Lengend Snippet: Tbx21−/− Th2 cells impair CD8+ T cell activation via IL-10 in vitro. (A) Schematic experimental layout to investigate the effect of LCMV-infected DCs previously conditioned by Tbx21−/− Th2 or Il10−/−Tbx21−/− Th2 cells on p14 cell activation in vitro. After 2 d of LCMV infection, CD4+ LCMV-specific T cells were depleted from T cell–DC cocultures. These conditioned DCs were subsequently cocultured with p14 cells in the presence of GP33 for 3 additional days. (B) Expression of surface markers CD69, KLRG-1, CD44, CD25, CD62L, and CD127 as well as (C) IFN-γ and CD107a expression (geometric mean of fluorescence intensity) by p14 cells after activation in vitro with LCMV-infected DCs that were previously conditioned with Tbx21−/− Th2 cells or Il10−/−Tbx21−/− Th2 cells. All experiments were performed at least twice, and each experimental group included n ≥ 3. Data are pooled and expressed as mean ± SEM. Asterisks indicate statistically significant differences as analyzed by one-way ANOVA with Bonferroni’s post test (*P < 0.05).

Article Snippet: For CD4 + T cell transfers, SMARTA1 LCMV-specific T cells of Tbx21 −/− , Il10 −/− Tbx21 −/− , and WT Thy1.1 + tg mice of the respective knockout backgrounds (6 to 8 wk old) were purified by staining with biotinylated antibodies against CD8 (53-6.7), CD11c (N418), CD11b (M1/70), CD19 (1D3), NK1.1 (PK136), Gr-1 (RB6-8C5), and CD25 (7D4), followed by anti-biotin microbeads and magnetic depletion (Miltenyi Biotec).

Techniques: Activation Assay, In Vitro, Infection, Expressing, Fluorescence