mouse mammary tumor cell line 4t1  (ATCC)

Journal: iScience

Article Title: Downregulation of IRF8 in alveolar macrophages by G-CSF promotes metastatic tumor progression

doi: 10.1016/j.isci.2024.109187

Figure Lengend Snippet: IRF8 expression declines among alveolar macrophages (AMs), recruited macrophages and monocytes prior to 4T1 micro-metastasis in lung tissue (A) Mammary tumor, lung and spleen weights of BALB/c orthotopic 4T1-bearing WT mice compared to non-tumor-bearing (NTB) control tissue. (B) 4T1 micro-metastasis detection in lung tissue by staining of colony formation (CF). Scale bar: 370 μm. (C) Myeloid cell percentages within lung tissue or blood during 4T1 growth. (D) Myeloid intracellular IRF8 levels within lung tissue of NTB or 4T1-bearing mice. (E) Flow-sorted AMs have reduced Irf8 and target gene expression during 4T1 growth. (F) IRF8 levels as in (D) within blood. Data are represented as mean ± SEM. Significance was determined by Wilcoxon rank-sum tests with Holm-Bonferroni correction for pre-planned comparisons (A and B), Dunnett’s test for correction of comparisons to NTB control (C; D and F, left), Spearman correlation (D and F, right; line indicates simple linear regression), or Mann-Whitney (E; n ≥ 5 mice/group). ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. See also Figures S1–S3 .

Article Snippet: Mouse: 4T1 mammary tumor cell line , ATCC , Cat#CRL-2539.

Techniques: Expressing, Control, Staining, Targeted Gene Expression, MANN-WHITNEY

Journal: iScience

Article Title: Downregulation of IRF8 in alveolar macrophages by G-CSF promotes metastatic tumor progression

doi: 10.1016/j.isci.2024.109187

Figure Lengend Snippet: IRF8 expression in AMs promotes anti-metastatic activity (A) Greater spontaneous 4T1 metastatic tumor CF occurs in BALB/c IRF8 cKO ( Lyz2CreIRF8 fl/fl ) lung tissue than IRF8 fl/fl lung tissue. (B) IRF8 cKO hosts have greater 4T1 experimental lung metastasis and larger nodule sizes than IRF8 fl/fl controls. (C) Schematic of control or clodronate-encapsulated liposome treatment before and during 4T1 experimental metastasis. (D and E) AM depletion via intranasal (i.n.) clodronate treatment reduces 4T1 experimental metastatic burden in IRF8 cKO hosts (D). Depletion of systemic macrophages, but not AMs, by intraperitoneal (i.p.) clodronate treatment increases 4T1 experimental metastasis in IRF8 cKO hosts (E). Representative H&E staining and images of lung metastasis displayed on the left. Scale bar: 370 μm. All data are represented as mean ± SEM. Significance was determined by Mann-Whitney (A and B) or Wilcoxon rank-sum tests with Holm-Bonferroni correction for pre-planned comparisons (D and E; n ≥ 5 mice/group). ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001. See also Figures S1–S6 .

Article Snippet: Mouse: 4T1 mammary tumor cell line , ATCC , Cat#CRL-2539.

Techniques: Expressing, Activity Assay, Control, Staining, MANN-WHITNEY

Journal: iScience

Article Title: Downregulation of IRF8 in alveolar macrophages by G-CSF promotes metastatic tumor progression

doi: 10.1016/j.isci.2024.109187

Figure Lengend Snippet: IRF8-deficient neonate liver-derived alveolar macrophages (NLDAMs) directly facilitate tumor growth (A) IRF8 cKO NLDAMs have reduced Irf8 and target gene expression (top) while increased inducible pro-tumor factor gene expression (bottom) compared to IRF8 fl/fl NLDAMs. (B) Schematic of 4T1 indirect co-culture with NLDAMs or BMDMs. (C and D) IRF8 cKO NLDAMs stimulate the greatest CF compared to IRF8 fl/fl controls or BMDMs (C). IRF8-transfected WT NLDAMs stimulate lower CF than IRF8-transfected IRF8 cKO NLDAMs (D). Scale bar: 370 μm. (E) Schematic of i.n. adoptive cell transfers of NLDAMs during 4T1 experimental metastasis. (F and G) IRF8 cKO NLDAM transfers augment 4T1 experimental metastasis in IRF8 cKO hosts (F). Reduced 4T1 experimental metastatic burden in IRF8 cKO hosts following transfers of IRF8-transfected WT NLDAMs than IRF8-transfected IRF8 cKO NLDAMs (G). Representative images are displayed. All data are represented as mean ± SEM. Significance was determined by Wilcoxon rank-sum tests with Holm-Bonferroni correction for pre-planned comparisons ($ indicates significantly different from all groups; n ≥ 6 mice/group). ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. See also Figures S7 and .

Article Snippet: Mouse: 4T1 mammary tumor cell line , ATCC , Cat#CRL-2539.

Techniques: Derivative Assay, Targeted Gene Expression, Gene Expression, Co-Culture Assay, Transfection

Journal: iScience

Article Title: Downregulation of IRF8 in alveolar macrophages by G-CSF promotes metastatic tumor progression

doi: 10.1016/j.isci.2024.109187

Figure Lengend Snippet: Mammary tumor expression of G-CSF reduces IRF8 expression in AMs and promotes lung metastasis (A) G-CSF levels within in vitro tumor-conditioned media (left) or in vivo orthotopic mammary tumors (right). (B) Triple-negative breast cancer (TNBC) cells express more G-CSF than non-TNBC cells. (C) Lung and spleen weights of NTB WT mice following local recombinant G-CSF (rG-CSF) exposure. (D and E) Myeloid cell percentages (D) and IRF8 levels (E) within lung tissue of control or rG-CSF-treated mice. (F) Schematic of local G-CSF blockade during orthotopic 4T1 growth. Tumor, lung and spleen weights from treated 4T1-bearing WT mice compared to isotype controls and baseline NTB tissue. (G) I.n. anti-G-CSF reduces lung micro-metastasis compared to isotype control treatment of 4T1-bearing mice. (H) Myeloid cell percentages within lung tissue following anti-G-CSF treatment. (I) Schematic of local G-CSF blockade during orthotopic 231/LM2-4 LUC+ growth and weights as in (F). (J) I.n. blockade of human or murine G-CSF reduces lung micro-metastasis of 231/LM2-4 LUC+ -bearing SCID mice compared to isotype controls. (K) Myeloid cell percentages within lung tissue following human or murine G-CSF blockade. All data are represented as mean ± SEM. Significance was determined by Wilcoxon rank-sum tests with Holm-Bonferroni correction for pre-planned comparisons (A, F, I, and J), unpaired t test with Welch’s correction (B and E), Mann-Whitney (C, D and G), or Dunnett’s test for correction of comparisons to NTB control (H and K; n ≥ 5 mice/group). ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. See also Figure S9 and Table S1 .

Article Snippet: Mouse: 4T1 mammary tumor cell line , ATCC , Cat#CRL-2539.

Techniques: Expressing, In Vitro, In Vivo, Recombinant, Control, MANN-WHITNEY

Journal: iScience

Article Title: Downregulation of IRF8 in alveolar macrophages by G-CSF promotes metastatic tumor progression

doi: 10.1016/j.isci.2024.109187

Figure Lengend Snippet: Local G-CSF targeting recovers anti-metastatic activity via AM- and IRF8-dependent mechanisms (A) Schematic of local neoadjuvant G-CSF blockade during orthotopic 4T1 growth compared to neoadjuvant isotype treatment. Tumor weights measured upon surgical removal. Post-surgery endpoint weights of lung or spleen. (B and C) Local neoadjuvant G-CSF blockade extends survival (B) and reduces overall metastasis (C) of WT but not IRF8 cKO mice. (D) Schematic of local neoadjuvant G-CSF blockade followed by single i.n. control or clodronate treatment during orthotopic 4T1 growth. Weights as in (A). (E and F) AM depletion via i.n. clodronate treatment does not alter post-surgery survival (E) of WT mice but negates anti-metastatic effects of local neoadjuvant G-CSF blockade (F). Representative images at post-surgery endpoint. All data are represented as mean ± SEM. Significance was determined by Wilcoxon rank-sum tests with Holm-Bonferroni correction for pre-planned comparisons (A and C), Log rank (B and E) or Mann-Whitney (D and F; n ≥ 8 mice/group). ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001.

Article Snippet: Mouse: 4T1 mammary tumor cell line , ATCC , Cat#CRL-2539.

Techniques: Activity Assay, Control, MANN-WHITNEY

Journal: iScience

Article Title: Downregulation of IRF8 in alveolar macrophages by G-CSF promotes metastatic tumor progression

doi: 10.1016/j.isci.2024.109187

Figure Lengend Snippet:

Article Snippet: Mouse: 4T1 mammary tumor cell line , ATCC , Cat#CRL-2539.

Techniques: Control, Recombinant, Virus, Derivative Assay, Liposomes, Lysis, Transfection, Staining, Enzyme-linked Immunosorbent Assay, cDNA Synthesis, Reverse Transcription, Variant Assay, Transgenic Assay, Plasmid Preparation, Expressing, Software, Pore Size, Cell Culture, Microscopy

Journal: iScience

Article Title: Controllable self-replicating RNA vaccine delivered intradermally elicits predominantly cellular immunity

doi: 10.1016/j.isci.2023.106335

Figure Lengend Snippet:

Article Snippet: Mouse mammary tumor cell line 4T1-Luc2 , ATCC , CRL-2539-Luc2.

Techniques: Virus, Recombinant, Modification, Derivative Assay, Variant Assay, Saline, Plasmid Preparation, Enzyme-linked Immunospot, Double-Color Enzymatic ELISPOT, Enzyme-linked Immunosorbent Assay, Plaque Reduction Neutralization Test, Software, Imaging

Journal: iScience

Article Title: Controllable self-replicating RNA vaccine delivered intradermally elicits predominantly cellular immunity

doi: 10.1016/j.isci.2023.106335

Figure Lengend Snippet: EXG-5006, encoding a fusion protein of SARS-CoV-2 and MERS-CoV nucleoproteins, can eliminate cells expressing those antigens in mouse (A) A schematic drawing of EXG-5006, encoding a fusion protein of CD5 signal peptide, nucleoprotein of SARS-CoV-2, and a nucleoprotein of MERS-CoV. (B–E) Cellular immunity by ELISpot assays, showing the frequency of IFN-γ (B, D) spot-forming cells or IL-4 (C and E) spot-forming cells in 1 × 10ˆ6 splenocytes obtained from BALB/c mice on Day 14 after vaccinating by a single intradermal injection of 100 μL solution containing either 5 μg (n = 1) or 25 μg (n = 4) of EXG-5006, or a placebo (PBO: buffer only: n = 5). The splenocytes were restimulated by culturing in the presence and absence of a pool of 15mer peptides for nucleoprotein of SARS-CoV-2 (original strain) (B, C) or a pool of 15mer peptides for nucleoprotein of MERS-CoV (D, E). The frequency obtained in the presence of peptides is plotted in the graph after subtracting the frequency obtained in the absence of peptides (background). Data are presented as mean (SD). (F) shows the survival (%) of female BALB/c mice vaccinated with EXG-5006, followed by injection of 4T1 tumor cells expressing the same antigen (A fusion protein of nucleoproteins of SARS-CoV-2 and MERS-CoV, without CD5 signal peptides).

Article Snippet: Mouse mammary tumor cell lines, 4T1 (ATCC, CRL-2539, female) and 4T1-Luc2 (ATCC, CRL-2539-Luc2, female), were purchased from ATCC.

Techniques: Expressing, Enzyme-linked Immunospot, Injection

Journal: iScience

Article Title: Controllable self-replicating RNA vaccine delivered intradermally elicits predominantly cellular immunity

doi: 10.1016/j.isci.2023.106335

Figure Lengend Snippet:

Article Snippet: Mouse mammary tumor cell lines, 4T1 (ATCC, CRL-2539, female) and 4T1-Luc2 (ATCC, CRL-2539-Luc2, female), were purchased from ATCC.

Techniques: Virus, Recombinant, Modification, Derivative Assay, Variant Assay, Saline, Plasmid Preparation, Enzyme-linked Immunospot, Double-Color Enzymatic ELISPOT, Enzyme-linked Immunosorbent Assay, Plaque Reduction Neutralization Test, Software, Imaging