Journal: Veterinary Research

Article Title: The local immune response of mice after Helicobacter suis infection: strain differences and distinction with Helicobacter pylori

doi: 10.1186/1297-9716-43-75

Figure Lengend Snippet: Number of colonizing H. suis bacteria. Shown is the average number of H. suis bacteria/milligram tissue in the stomach of BALB/c and C57BL/6 mice 59 days after experimental infection.

Article Snippet: Seven-week-old, female specific-pathogen-free BALB/c and C57BL/6 mice, free of Helicobacter spp., were purchased from Harlan NL (Horst, The Netherlands).

Techniques: Bacteria, Infection

Journal: Veterinary Research

Article Title: The local immune response of mice after Helicobacter suis infection: strain differences and distinction with Helicobacter pylori

doi: 10.1186/1297-9716-43-75

Figure Lengend Snippet: General cytokine expression profile after experimental H. suis infection in BALB/c and C57BL/6 mice. Shown are the mean fold changes in mRNA expression of indicated cytokines in H. suis -infected BALB/c and C57BL/6 mice. The mean fold change in the relevant uninfected control groups is equal to 1. An * indicates a statistically significant difference compared to uninfected control mice ( p < 0.05).

Article Snippet: Seven-week-old, female specific-pathogen-free BALB/c and C57BL/6 mice, free of Helicobacter spp., were purchased from Harlan NL (Horst, The Netherlands).

Techniques: Expressing, Infection, Control

Journal: Veterinary Research

Article Title: The local immune response of mice after Helicobacter suis infection: strain differences and distinction with Helicobacter pylori

doi: 10.1186/1297-9716-43-75

Figure Lengend Snippet: Cytokine expression profiles for each group of animals infected with H. suis strains HS1-9 and H. pylori strains SS1 or pMSS1. Shown are the mean fold changes of mRNA expression in HS1-HS9- and SS1- or pMSS1-infected BALB/c and C57BL/6 mice for IL-1β ( A ), IL-4 ( B ), IL-6 ( C ), IL-10 ( D ) and IL-17 ( E ). The mean fold change in the relevant uninfected control groups is equal to 1. An * indicates a significant upregulation of mRNA expression compared to uninfected control mice ( p < 0.05). An ** indicates a significant upregulation of mRNA expression compared to uninfected control mice ( p < 0.0056, i.e. the new cut-off p value obtained after Bonferroni correction for multiple comparisons).

Article Snippet: Seven-week-old, female specific-pathogen-free BALB/c and C57BL/6 mice, free of Helicobacter spp., were purchased from Harlan NL (Horst, The Netherlands).

Techniques: Expressing, Infection, Control

Journal: Veterinary Research

Article Title: The local immune response of mice after Helicobacter suis infection: strain differences and distinction with Helicobacter pylori

doi: 10.1186/1297-9716-43-75

Figure Lengend Snippet: Correlation between cytokine expression and H. suis colonization. Shown are the correlation analyses between IL-6 ( A ), IL-17 ( B ), MIP-2 ( C ), IL-1β ( D ) and IL-10 ( E ) mRNA expression levels and the number of colonizing H. suis bacteria in stomachs of indicated mouse strains (BALB/c and C57BL/6). Correlation was measured by Spearman’s Rho (ρ).

Article Snippet: Seven-week-old, female specific-pathogen-free BALB/c and C57BL/6 mice, free of Helicobacter spp., were purchased from Harlan NL (Horst, The Netherlands).

Techniques: Expressing, Bacteria

Journal: Science Advances

Article Title: Mitochondrial elongation impairs breast cancer metastasis

doi: 10.1126/sciadv.adm8212

Figure Lengend Snippet: ( A and B ) Representative images (A, scale bars, 5 μm) and quantification of mitochondrial length (B) in Tom 20–stained 67NR, 4T07, 66cl4, and 4T1 breast cancer cells. In (B), each data point represents an individual cell ( n = 34 to 81 per group). **** P < 0.0001 by Welch’s analysis of variance (ANOVA) and Dunnett’s T3 post-hoc test. ( C ) Schematic representation of the 4T1 KO cell lines generated for this study. Two independent clones (#1 and #2) were derived from each individual KO cell population (Drp1, Fis1, and Mff). ( D ) Schematic representation of mitochondrial fission highlighting the mitochondrial fission proteins Drp1, Fis1, and Mff. ( E ) Validation of KO cell lines by immunoblot. Representative images of three independent experiments. Actin or Vinculin served as loading controls. ( F and G ) Representative images (F, scale bars, 5 μm) and quantification of mitochondrial length (G) in Tom 20–stained parental, Drp1 KO, Fis1 KO, and Mff KO 4T1 breast cancer cells. In (G), each data point represents an individual cell ( n = 33 to 42 per group). **** P < 0.0001 by Welch’s ANOVA and Dunnett’s T3 post-hoc test. ( H ) Live cell counts of parental versus Drp1 KO, Fis1 KO, and Mff KO 4T1 breast cancer cells over the course of 5 days. Bar graphs depict live cell counts on day 5. Results represented as mean ± SEM of three independent experiments. **** P < 0.0001, *** P < 0.001, ** P < 0.01 by one-way ANOVA and Dunnett’s post-hoc test. ( I and J ) Number of colonies (I) and representative images of colonies (J) observed in 4T1 parental versus Drp1 KO, Fis1 KO, and Mff KO clones after culture in ultralow attachment plates for 7 (I) and 14 (I and J) days. In (I), results are represented as mean ± SEM of five (7 days) or three (14 days) independent experiments. **** P < 0.0001 by two-way ANOVA and Dunnett’s post-hoc test. ns, nonsignificant.

Article Snippet: For spontaneous metastasis assays, 10,000 cells (4T1) or 250,000 cells (MDA-MB-231), were injected into the mammary fad pads of female Balb/c mice (Charles River, 4T1 model) or NOD-scid IL2Rgamma null mice (The Jackson Laboratory, MDA-MB-231 model) in a 1:1 mixture of PBS:Matrigel Growth Factor Reduced (Corning).

Techniques: Staining, Generated, Clone Assay, Derivative Assay, Western Blot

Journal: Science Advances

Article Title: Mitochondrial elongation impairs breast cancer metastasis

doi: 10.1126/sciadv.adm8212

Figure Lengend Snippet: ( A ) Venn diagrams depicting common down-regulated and up-regulated transcripts in all KO cell lines versus parental 4T1 breast cancer cells that were identified by DESeq analysis of RNA-seq data ( n = 3), |FC| > 1.5 and P adjusted value <0.05. ( B ) Dot plots of common down-regulated and up-regulated pathways in all KO cell lines versus parental 4T1 breast cancer cells, according to GSEA. GeneRatio is calculated as follows: count of core enrichment genes/count of genes present in the indicated pathway. ( C ) Heatmaps showing common affected genes in all KO cell lines versus parental 4T1 breast cancer cells from selected pathways in (B). ( D ) PLS-DA of metabolite profile data between parental and fission KO 4T1 cell lines. All KO cell lines were grouped into one category (fKO), contained in the light shaded orange circle. Each data point represents one sample ( n = 6 for parental 4T1, n = 3 for each KO clone). ( E ) VIP scores of top 15 metabolites. All fission KO cell lines were grouped into one category (fKO) and compared to parental 4T1 breast cancer cells. ( F ) Dot plots of common altered metabolic pathways in all KO cell lines versus parental 4T1 breast cancer cells, according to MSEA.

Article Snippet: For spontaneous metastasis assays, 10,000 cells (4T1) or 250,000 cells (MDA-MB-231), were injected into the mammary fad pads of female Balb/c mice (Charles River, 4T1 model) or NOD-scid IL2Rgamma null mice (The Jackson Laboratory, MDA-MB-231 model) in a 1:1 mixture of PBS:Matrigel Growth Factor Reduced (Corning).

Techniques: RNA Sequencing Assay

Journal: Science Advances

Article Title: Mitochondrial elongation impairs breast cancer metastasis

doi: 10.1126/sciadv.adm8212

Figure Lengend Snippet: ( A ) Left: Oxygen consumption rate (OCR) after injection of oligomycin (O), FCCP (F), rotenone/antimycin A (R/A) and monensin (M). Right: Rate of ATP production by oxidative phosphorylation (J ATP OXPHOS) in Drp1 KO, Fis1 KO, and Mff KO clones versus parental 4T1 breast cancer cells. **** P < 0.0001, *** P < 0.001, ** P < 0.01, * P < 0.05 by one-way ANOVA and Dunnett’s post-hoc test. ( B ) Left: Extracellular acidification rate (ECAR) after injection of oligomycin (O), FCCP (F), rotenone/antimycin A (R/A), and monensin (M). Right: rate of ATP production by glycolysis (J ATP glycolysis) in Drp1 KO, Fis1 KO, and Mff KO clones versus parental 4T1 breast cancer cells. **** P < 0.0001, *** P < 0.001, ** P < 0.01, by one-way ANOVA and Dunnett’s post-hoc test. ( C ) Left: Bioenergetic space plots of Drp1 KO, Fis1 KO, and Mff KO clones versus parental 4T1 breast cancer cells. Data points with dashed lines depict maximal ATP production rates, whereas data points with solid lines depict ATP production rates under basal conditions. The areas delimited by the dashed and solid lines represent the maximal and basal bioenergetic scope, respectively. Right: Calculated bioenergetic scope values under basal and maximal conditions. *** P < 0.001, * P < 0.05 by two-way ANOVA and Dunnett’s post-hoc test. All results shown in this figure are depicted as mean ± SEM of four independent experiments. ns, nonsignificant.

Article Snippet: For spontaneous metastasis assays, 10,000 cells (4T1) or 250,000 cells (MDA-MB-231), were injected into the mammary fad pads of female Balb/c mice (Charles River, 4T1 model) or NOD-scid IL2Rgamma null mice (The Jackson Laboratory, MDA-MB-231 model) in a 1:1 mixture of PBS:Matrigel Growth Factor Reduced (Corning).

Techniques: Injection, Clone Assay

Journal: Science Advances

Article Title: Mitochondrial elongation impairs breast cancer metastasis

doi: 10.1126/sciadv.adm8212

Figure Lengend Snippet: ( A ) Schematic depicting the experimental design of the spontaneous metastasis assays conducted to assess metastatic potential of parental versus Drp1 KO, Fis1 KO, and Mff KO 4T1 breast cancer cells in vivo. Cells were injected into the mammary fat pads (MFPs) of mice and tumor growth was assessed for 20 to 38 days. Tumors were resected when they reached 500 mm 3 . Twenty-one days after resection, lung metastatic burden was evaluated. ( B ) Diagram showing the median time required for each cell line to reach resection size. ( C ) Primary tumor growth of parental versus Drp1 KO, Fis1 KO, and Mff KO 4T1 breast cancer cells. Graph shows mean tumor volume over time ± SEM ( n = 9 per group). **** P < 0.0001, ** P < 0.01 by mixed-effects model and Dunnett’s posttest. ( D and E ) Number of visible lesions (D) and total metastatic lesion area (E) quantified in the lungs of mice bearing parental versus Drp1 KO, Fis1 KO, and Mff KO 4T1 tumors. Results are shown as individual data points, where each data point represents an individual mouse, and mean is depicted as a line ( n = 8 for parental 4T1, n = 9 for all KOs). **** P < 0.0001, by one-way ANOVA and Dunnett’s post-hoc test. ( F ) Representative images of H&E-stained lungs from mice injected with parental versus Drp1 KO, Fis1 KO, and Mff KO 4T1 breast cancer cells. Scale bar, 4 mm. ( G and H ) Kaplan-Meier overall survival (G) and time to distant recurrence (H) analysis of patients from the METABRIC study. A gene expression signature associated with impaired mitochondrial fission was constructed using the genes identified in . Patients were divided in two groups according to their gene signature score.

Article Snippet: For spontaneous metastasis assays, 10,000 cells (4T1) or 250,000 cells (MDA-MB-231), were injected into the mammary fad pads of female Balb/c mice (Charles River, 4T1 model) or NOD-scid IL2Rgamma null mice (The Jackson Laboratory, MDA-MB-231 model) in a 1:1 mixture of PBS:Matrigel Growth Factor Reduced (Corning).

Techniques: In Vivo, Injection, Staining, Expressing, Construct

Journal: Science Advances

Article Title: Mitochondrial elongation impairs breast cancer metastasis

doi: 10.1126/sciadv.adm8212

Figure Lengend Snippet: ( A and B ) Representative images (A, scale bars, 5 μm) and quantification of mitochondrial length (B) in Tom 20–stained 4T1 cells treated with Leflu or DMSO. In (B), each data point represents an individual cell ( n = 29 per group). **** P < 0.0001 by Welch’s t test. ( C ) Left: Proliferation curve of Leflu- and DMSO-treated 4T1 cells. Right: Live cell counts on day 5 ( N = 4). *** P < 0.001 by unpaired t test. ( D ) Venn diagrams (top) and heatmap (bottom) depicting common altered genes in fission KO (fKO) and Leflu-treated 4T1 cells. ( E ) Common altered metabolic pathways in Leflu-treated and fission KO 4T1 cells. ( F ) Representative immunoblot and densitometric analysis of Ddr1 signal relative to actin signal. * P < 0.05 by unpaired t test ( N = 4). ( G ) Tumor growth curve of Leflu- versus DMSO-treated mice ( n = 6 per group). P = 0.0627 by mixed-effects model and Dunnett’s post-hoc test. ( H and I ) Number of visible lesions (H) and total metastatic area (I) in lungs of Leflu- versus DMSO-treated mice. Each data point represents an individual mouse, and mean is depicted as a line ( n = 6 per group). ** P < 0.01, * P < 0.05, by one-way ANOVA and Dunnett’s post-hoc test. ( J ) Representative images of H&E-stained lungs from Leflu- versus DMSO-treated mice. Scale bar, 2 mm. ( K ) Kaplan-Meier overall survival analysis of patients from the METABRIC study. A gene expression signature associated with enhanced mitochondrial elongation was constructed using the genes identified in (D). Patients were divided into two groups according to their signature score. ( L and M ) Overall (L) and progression-free (M) survival of patients belonging to an aggressive breast cancer cohort. Patients were categorized according to their signature scores and separated by the median score into two groups (low and high) to conduct survival analyses. Unless indicated otherwise, all data are presented as mean ± SEM.

Article Snippet: For spontaneous metastasis assays, 10,000 cells (4T1) or 250,000 cells (MDA-MB-231), were injected into the mammary fad pads of female Balb/c mice (Charles River, 4T1 model) or NOD-scid IL2Rgamma null mice (The Jackson Laboratory, MDA-MB-231 model) in a 1:1 mixture of PBS:Matrigel Growth Factor Reduced (Corning).

Techniques: Staining, Western Blot, Expressing, Construct

Journal: Oncoimmunology

Article Title: Anticancer DNA vaccine based on human telomerase reverse transcriptase generates a strong and specific T cell immune response

doi: 10.1080/2162402X.2015.1083670

Figure Lengend Snippet: INVAC‑1 induces hTERT specific T‑cell responses in mice. (A) C57BL/6 mice (6 mice per group), BALB/c mice (6‑7 mice per group), HLA‑B7 mice (4‑6 mice per group) and HLA‑A2/DR1 mice (5-6 mice per group) were immunized once. Fourteen days later, an IFNγ ELISpot assay was performed with splenocytes stimulated with a pool of hTERT restricted peptides according to mouse MHC. IFNγ hTERT specific CD8+ or CD4+ T‑cells/200,000 splenocytes are represented as mean ± SD. Mann–Whitney non-parametric test against mice control, **p < 0.01. (B) HLA‑A2/DR1 mice (3‑5 mice per group) were immunized twice (prime-boost) with INVAC‑1 (D0 and D21). At D31, splenocytes were Ficoll purified and stimulated with a pool of 3 hTERT specific peptides restricted to HLA‑DRB1. Supernatants from stimulated cells were recovered and tested in a cytokine binding assay in order to evaluate the concentration of Th1, Th2 and Th17 cytokines secreted by hTERT specific CD4+ T‑cells. Cytokine concentrations in pg/mL are represented as mean ± SD. Mann–Whitney non-parametric test against mice control, *p < 0.05.

Article Snippet: Female 6-week-old C57BL/6JRj and BALB/cJRj mice were purchased from Janvier laboratories (Saint-Berthevin, France).

Techniques: Enzyme-linked Immunospot, MANN-WHITNEY, Purification, Binding Assay, Concentration Assay

Journal: Oncoimmunology

Article Title: Anticancer DNA vaccine based on human telomerase reverse transcriptase generates a strong and specific T cell immune response

doi: 10.1080/2162402X.2015.1083670

Figure Lengend Snippet: Prime‑boost vaccination enhanced, shortened and broadened hTERT‑specific T‑cell responses. (A) Ten C57BL/6 mice were immunized four times with INVAC-1 (D0, D21, D123 and D144). Peripheral blood was collected before the first immunization D0, and at D7, D14, D21, D33, D40, D118, D132, D139, D154, D159 and D166. PBMCs were Ficoll purified and stimulated with a pool of 4 hTERT specific peptides restricted to the H2‑Kb/Db and analyzed by an IFNγ ELISPOT assay. Black arrows represent days of vaccination. IFNγ hTERT specific CD8+ T‑cells/200,000 PBMCs are represented as mean ± SD. Mann–Whitney non-parametric test against D0, *p < 0.05. (B) Seven to 13‑weeks‑old C57BL/6 mice (6 mice per group) were immunized twice (D0 and D21). At D31, splenocytes were Ficoll purified and stimulated with 13 pools of hTERT 15‑mer overlapping peptides (10 peptides/pool) and analyzed by an IFNγ ELISPOT assay. IFNγ hTERT specific T‑cells /200,000 splenocytes are represented as mean ± SD.

Article Snippet: Female 6-week-old C57BL/6JRj and BALB/cJRj mice were purchased from Janvier laboratories (Saint-Berthevin, France).

Techniques: Purification, Enzyme-linked Immunospot, MANN-WHITNEY

Journal: Oncoimmunology

Article Title: Anticancer DNA vaccine based on human telomerase reverse transcriptase generates a strong and specific T cell immune response

doi: 10.1080/2162402X.2015.1083670

Figure Lengend Snippet: INVAC‑1 induces hTERT specific cytotoxic T‑cells. (A) HLA‑B7 mice (4‑6 mice per group), were immunized twice (D0 and D21). At D31, an ELISpot granzyme B (GrB) assay was performed with splenocytes stimulated with a pool of hTERT HLA‑B7 restricted peptides. GrB hTERT specific CD8+ T‑cells/200,000 splenocytes are represented as mean ± SD. Mann–Whitney non-parametric test against mice control, **p < 0.01. (B) C57BL/6 mice (8 mice per group) were immunized twice (D0 and D21). At D31, syngeneic splenocytes, pulsed with individual hTERT peptides restricted to the H2‑Kb/Db (either p660 or p1034) were labeled with CFSE and injected IV to immunized mice. After 15‑18 h, the disappearance of peptide‑pulsed cells in spleens was analyzed by flow cytometry. (C) Percent of killing was presented as mean ± SD. (D) C57BL/6 mice (6 mice per group) were injected s.c. in the right flank with 2 × 105 TC‑1 cells. On day 5 (when tumors are palpable), mice were immunized with INVAC‑1 or left non-treated (NT). Fourteen days later, mice were sacrificed and splenocytes, tumor draining lymph nodes and tumor infiltrated lymphocytes were isolated. An ELISpot IFNγ was performed with H2‑Kb/Db restricted peptides (p429, p660, p1021 and p1034). IFNγ hTERT specific CD8+ T-cells/200,000 cells are represented as mean ± SD. Mann–Whitney non-parametric test **p < 0.01.

Article Snippet: Female 6-week-old C57BL/6JRj and BALB/cJRj mice were purchased from Janvier laboratories (Saint-Berthevin, France).

Techniques: Enzyme-linked Immunospot, MANN-WHITNEY, Labeling, Injection, Flow Cytometry, Isolation

Journal: Oncoimmunology

Article Title: Anticancer DNA vaccine based on human telomerase reverse transcriptase generates a strong and specific T cell immune response

doi: 10.1080/2162402X.2015.1083670

Figure Lengend Snippet: hTERT and INVAC‑1 peptides.

Article Snippet: Female 6-week-old C57BL/6JRj and BALB/cJRj mice were purchased from Janvier laboratories (Saint-Berthevin, France).

Techniques: Sequencing