Journal: NPJ Breast Cancer

Article Title: Retinoid X receptor agonist LG100268 modulates the immune microenvironment in preclinical breast cancer models

doi: 10.1038/s41523-019-0135-5

Figure Lengend Snippet: LG268 treatment modifies T-cell populations in established tumors. a T-cell populations in mammary gland tumors from MMTV-Neu mice treated with rexinoids (100 mg/kg diet) were analyzed by flow cytometry ( n = 6–8 mice/group). Total T cells: CD45 + , CD3 + ; CD4 T cells: CD45 + , CD3 + , CD4 + ; activated CD4 T cells: CD45 + , CD3 + , CD4 + , CD25 + ( t = 3.341; df = 7); CD8 T cells: CD45 + , CD3 + , CD8 + . (ratio %CD8/%CD4, CD25: t = 2.543; df = 6). b Immunostaining of FOXP3 in tumor sections was used to confirm the downregulation of activated CD4 T cells observed by flow cytometry. Scale bar represents 60 μm. n = 7–8, bars represent average and error bars the standard error of the mean (SEM)

Article Snippet: Single cells were resuspended in a solution of PBS/0.5% BSA/0.1% azide and stained for 30 min at 4 °C with the following antibodies: CD45-VioGreen (30F11, Miltenyi, 10:100), Gr1-PE (RB6-8C5, Miltenyi, 10:100), CD11b-FITC (M1/70.15.11.5, Miltenyi, 10:100), CD19-APC (1D3/CD19, Biolegend, 1:100), B220-PerCP-Cy5.5 (RA3-6B2, Biolegend, 1:100), CD3-PE (145–2C11, Biolegend, 1:100), CD4-FITC (Gk1.5, Miltenyi, 10:100), CD8-APC (53–6.7, Biolegend, 1:100), CD25-PE.Cy7 (EBiosciences, 1:100) and 5 μg/ml anti-mouse CD16/CD32 antibody (Biolegend) to reduce antibody binding to Fc receptors.

Techniques: Flow Cytometry, Immunostaining

Journal: NPJ Breast Cancer

Article Title: Retinoid X receptor agonist LG100268 modulates the immune microenvironment in preclinical breast cancer models

doi: 10.1038/s41523-019-0135-5

Figure Lengend Snippet: The combination of LG268 and anti-PD-L1 antibodies increases the infiltration of CD8 T cells and caspase 3 activation. a Experimental design. PyMT mice with tumor(s) 32–64 mm 3 in size were started on control diet or LG268 diet (100 mg/kg) 48 h before the first anti-PD-L1 treatment. Anti-PD-L1 antibodies were injected i.p. (three injections each of 40 µg per mouse). Mice were killed on day 15. ( n = 7 mice/group). b Immune cell infiltration in tumors was analyzed by flow cytometry. Total T cells: CD45 + , CD3 + ; CD4 T cells: CD45 + , CD3 + , CD4 + ; activated CD4 T cells: CD45 + , CD3 + , CD4 + , CD25 + ; CD8 T cells: CD45 + , CD3 + , CD8 + ( t = 2.219; df = 12) (ratio %CD8/%CD4, CD25: t = 2.268; df = 12). c Immunohistochemistry for PD-L1, CD8 and cleaved caspase 3 in tumors. Scale bar represents 60 μm. n = 5–7, bars represent average and error bars the standard error of the mean (SEM)

Article Snippet: Single cells were resuspended in a solution of PBS/0.5% BSA/0.1% azide and stained for 30 min at 4 °C with the following antibodies: CD45-VioGreen (30F11, Miltenyi, 10:100), Gr1-PE (RB6-8C5, Miltenyi, 10:100), CD11b-FITC (M1/70.15.11.5, Miltenyi, 10:100), CD19-APC (1D3/CD19, Biolegend, 1:100), B220-PerCP-Cy5.5 (RA3-6B2, Biolegend, 1:100), CD3-PE (145–2C11, Biolegend, 1:100), CD4-FITC (Gk1.5, Miltenyi, 10:100), CD8-APC (53–6.7, Biolegend, 1:100), CD25-PE.Cy7 (EBiosciences, 1:100) and 5 μg/ml anti-mouse CD16/CD32 antibody (Biolegend) to reduce antibody binding to Fc receptors.

Techniques: Activation Assay, Injection, Flow Cytometry, Immunohistochemistry

Journal: Immunity

Article Title: The E3 ligase VHL promotes group 2 innate lymphoid cell maturation and function via inhibition of glycolysis and induction of interleukin 33 receptor

doi: 10.1016/j.immuni.2017.12.013

Figure Lengend Snippet: (A) Flow cytometric profiles of cells derived from sorted common helper innate lymphoid progenitors (CHILPs; Lin−CD127+Flt3−CD25−α4β7+) of WT and Vhlfl/fl Zbtb16-cre mice cultured on OP9-DL4 stromal cells in the presence of IL-7 and IL-33, and analyzed for the percentage of ILC2s on day 15 (D15) or 18 (D18) of culture by the specific markers, including Sca1+ICOS+, CD44+CD25+, and CD127+ST2+, pre-gated with Lin− cells. Numbers indicate the frequency of flow cytometric events.

Article Snippet: Anti-mouse CD25 PE-Cy7 (clone PC61.5) , Thermo Fisher Scientific , Cat# 25-0251-82;.

Techniques: Derivative Assay, Cell Culture

Journal: Immunity

Article Title: The E3 ligase VHL promotes group 2 innate lymphoid cell maturation and function via inhibition of glycolysis and induction of interleukin 33 receptor

doi: 10.1016/j.immuni.2017.12.013

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Anti-mouse CD25 PE-Cy7 (clone PC61.5) , Thermo Fisher Scientific , Cat# 25-0251-82;.

Techniques: Control, Recombinant, SYBR Green Assay, Staining, Enzyme-linked Immunosorbent Assay, cDNA Synthesis, Software