Journal: Nature communications

Article Title: Conserved regulatory logic at accessible and inaccessible chromatin during the acute inflammatory response in mammals.

doi: 10.1038/s41467-020-20765-1

Figure Lengend Snippet: Fig. 4 Conserved NF-κB binding is shared between cell types near common immune response genes. a Representative genomic regions showing the pan-cell (four-cell-type-shared: between human aortic ECs [HAECs], human umbilical vein ECs [HUVECs], lymphoblastoid cell lines [LCLs], and adipocytes), EC-specific (shared only between HAECs and HUVECs), LCL-specific, and adipocyte-specific RELA peaks. b Stacked bar charts showing the fractions of four-, three-, and two-cell-type-shared and HAEC-specific RELA peaks (one-cell type) that are three-species-conserved, two-species- conserved, or human-specific. c Table showing the top-scoring de novo motifs (MEME-ChIP e values, Supplementary Dataset 4), fold enrichments of RELA binding modes (p value: *<1.0 × 10−6, Chi-squared test with Yates’s correction for continuity of independence and Bonferroni correction for multiple testing), fold enrichments near TNFα target genes (HAEC RNA-seq, ±10 kb of transcriptional start sites, p value: * <1.0 × 10−5, two-sided Fisher’s exact test with Bonferroni correction), and the top GO terms (GREAT FDR q values, Biological Process Ontology, Supplementary Dataset 3) for the pan-cell (four-cell- type-shared), EC-specific, and HAEC-specific RELA peaks. ↑= upregulated, ↓= downregulated, NC no change (i.e., constitutively expressed). d Profile plots comparing ERG binding signal at pan-cell, EC-specific, and HAEC-specific RELA peaks in HAECs. ERG ChIP-seq signal was generated using published raw data from ref. 15. The plot lines indicate mean RPM ± SEM. Arrows and percentages indicate change in signal after TNFα stimulation. The p values were calculated using two-sided Welch Two Sample t-test (p value: *<0.05). e, Model of NF-κB-ERG dynamics and cofactor squelching showing redistribution of cofactors from EC-specific Mode O regions to pan-cell Mode P regions. Source data are provided in the Source Data file for Fig. 4c, d.

Article Snippet: To induce the acute pro-inflammatory response, HAECs (#1487 and #2139) were treated with 10-ng/mL recombinant human TNFα (Cell Applications, cat# RP1111-50), MAECs (#A092913T2MP and #B092913T2MP) were treated with 10-ng/mL recombinant mouse TNFα (Cell Applications, cat# RP2031-20), and BAECs (#1165 and #1190) were treated with 10-ng/mL recombinant bovine TNFα (R&D Systems, cat# 2279-BT-025) for 45 min in basal Endothelial Cell Growth Media MV2 (PromoCell) without supplements.

Techniques: Binding Assay, RNA Sequencing, ChIP-sequencing, Generated

Journal: JCI Insight

Article Title: Evaluating immunotherapeutic outcomes in triple-negative breast cancer with a cholesterol radiotracer in mice

doi: 10.1172/jci.insight.175320

Figure Lengend Snippet: Three days after orthotopically injecting E0771 or AT-3 breast cancer cells plus mouse mammary fibroblasts into syngeneic C57BL/6J mice, we randomly assigned animals to treatment with anti–PD-1 antibody or PBS vehicle every 3 days for 4 doses total. Graphs show mean values ± SEM (symbols) and calculated logistic regression (smooth line) for E0771 ( A ) or AT-3 ( D ) tumors ( n = 6 control; n = 8 anti–PD-1) treated with anti–PD-1 antibody or PBS. ( B and E ) Growth of E0771 and AT3 tumor growth, respectively, for individual mice over time. Three tumors from the anti–PD-1 group failed to grow tumors and are overlapped on the x axis of panel B . We analyzed differences in tumor growth data by logistic regression. Survival curves demonstrate that anti–PD-1 treatment significantly prolonged survival for mice with E0771 tumors ( C ) but not with AT-3 ( F ), as analyzed by the Mantel-Cox test.

Article Snippet: We implanted 5 × 10 5 EO771 or AT-3 mouse mammary tumor cell lines and 1 × 10 5 mouse mammary fibroblasts orthotopically into fourth inguinal mammary fat pads of 6- to 8-week-old female C57BL/6J female mice (The Jackson Laboratory) ( ).

Techniques: Control

Journal: JCI Insight

Article Title: Evaluating immunotherapeutic outcomes in triple-negative breast cancer with a cholesterol radiotracer in mice

doi: 10.1172/jci.insight.175320

Figure Lengend Snippet: We injected C57BL/6J mice intraperitoneally with cholesterol labeled with BODIPY and euthanized animals 24 hours later to collect and dissociate tumors for flow cytometry ( n = 5 each for EO771 and AT-3). Plots for ( A ) CD8 + and ( B ) CD4 + T cells show accumulation of BODIPY-cholesterol in cells from individual tumors from EO771 and AT-3 tumors relative to vehicle only or isotype antibody control. ( C ) CD8 + and ( D ) CD4 + T cells in EO771 tumors showed significantly higher fold change accumulation of fluorescent cholesterol relative to FMO control. ( E ) CD8 + , but not ( F ) CD4 + , T cells in EO771 tumors also expressed higher levels of PD-1. ** P < 0.01,*** P < 0.001, **** P < 0.0001 for differences between means using nonparametric Mann-Whitney tests ( C and D ; n = 5 mice per group), while differences between T cell population percentages were assessed using 2-tailed Student’s t test ( E and F ).

Article Snippet: We implanted 5 × 10 5 EO771 or AT-3 mouse mammary tumor cell lines and 1 × 10 5 mouse mammary fibroblasts orthotopically into fourth inguinal mammary fat pads of 6- to 8-week-old female C57BL/6J female mice (The Jackson Laboratory) ( ).

Techniques: Injection, Labeling, Flow Cytometry, Control, MANN-WHITNEY

Journal: JCI Insight

Article Title: Evaluating immunotherapeutic outcomes in triple-negative breast cancer with a cholesterol radiotracer in mice

doi: 10.1172/jci.insight.175320

Figure Lengend Snippet: When tumors reached approximately 70 mm 2 , we treated mice with anti–PD-1 antibody or control for 4 days. Mice were then injected with 100 μCi of eFNP-59 followed by T cell isolation protocols. ( A ) Graph shows uptake of eFNP-59 per microgram of spleen tissue measured by scintigraphy. Symbols show individual mice with annotations for mean values and standard deviations. ( B ) We isolated CD4 + and CD8 + tumor-infiltrating lymphocytes (TILs) by positive selection with immunomagnetic beads and determined accumulation of eFNP-59 normalized to total cell protein with a BCA assay. Representative data from 2 experimental replicates, with statistical comparisons by 1-way ANOVA with Dunn’s multiple-comparison test. * P < 0.05; ** P < 0.01.

Article Snippet: We implanted 5 × 10 5 EO771 or AT-3 mouse mammary tumor cell lines and 1 × 10 5 mouse mammary fibroblasts orthotopically into fourth inguinal mammary fat pads of 6- to 8-week-old female C57BL/6J female mice (The Jackson Laboratory) ( ).

Techniques: Control, Injection, Cell Isolation, Isolation, Selection, BIA-KA, Comparison