Journal: Cell reports

Article Title: Chronic stress physically spares but functionally impairs innate-like invariant T cells

doi: 10.1016/j.celrep.2021.108979

Figure Lengend Snippet:

Article Snippet: Mouse: B16-F10-Red-FLuc (B16-FLuc) melanoma cells , PerkinElmer , Cat # BW124734.

Techniques: Control, Recombinant, Western Blot, Enzyme-linked Immunosorbent Assay, Staining, In Situ, Selection, cDNA Synthesis, Software

Journal: Journal of controlled release : official journal of the Controlled Release Society

Article Title: Surface Engineering Tumor Cells with Adjuvant-loaded Particles for Use as Cancer Vaccines

doi: 10.1016/j.jconrel.2016.12.036

Figure Lengend Snippet: (A1) B16.F10 cells and (A2) RM11 cells treated or not treated (control) with biotinylated anti-CD29 antibodies (anti-CD29-biotin) followed by strepavidin-PE. (B–F) Validation of cell-particle hybrid assembly from B16.F10 or RM11 cells subsequent to surface biotinylation using anti-CD29-biotin and mixed with streptavidin-coated particles loaded with rhodamine B and washed to remove unbound particles (hybrid). Control involved the same conditions except cells were not treated with anti-CD29-biotin. Validation was performed using: (B1–2 and E1–2): flow cytometry where (B1 and E1) representive (n = 1) and (B2 and E2) mean (n = 3) results from measuring relative mean rhodamine fluorescence intensity (RMFI(Rh)) of (B1 and B2) B16.F10 and (E1 and E2) RM11 cells; (C1–2 and F1–2): laser scanning confocal microscopy showing (C1 and F1) hybrid and (C2 and F2) control cell-particle mixtures for (C1 and C2) B16.F10 and (F1 and F2) RM11 cells (blue = DAPI stained cell nuclei, red = rhodamine-labeled PLGA particles); (D): scanning electron microscopy showing (D1) B16.F10 hybrid and (D2) control cell-particle mixtures (arrows in C1, D1, and F1 indicate particles bound to cell surface). Scale bar= 20 micron for C1–2 and F1–2, 10 microns for D1–2. When applicable, error bars = SD. ** p < 0.01, *** p < 0.001, n = 3.

Article Snippet: For vaccinations, a B16.F10 GM-CSF clone expressing 220 ng GM-CSF/10 6 cells/day was derived by transducing B16.F10 cells with a lentiviral vector encoding murine GM-CSF (AMSBIO, Cambridge, MA).

Techniques: Flow Cytometry, Fluorescence, Confocal Microscopy, Staining, Labeling, Electron Microscopy

Journal: Journal of controlled release : official journal of the Controlled Release Society

Article Title: Surface Engineering Tumor Cells with Adjuvant-loaded Particles for Use as Cancer Vaccines

doi: 10.1016/j.jconrel.2016.12.036

Figure Lengend Snippet: Laser scanning confocal microscopy imaging of cell-particle hybrid uptake by BMDC. Cell-particle hybrids were incubated with BMDC Yellow arrows indicating colocalization of B16.F10 cells (green) and particles (red) inside BMDC (magenta). Magenta: Alexa flour®700 (CD11c) stained BMDC, green: CFSE labeled B16.F10 melanoma cells, red: rhodamine B-labeled PLGA particles, gray: DAPI stained cell nuclei. Scale bar: 20 micron.

Article Snippet: For vaccinations, a B16.F10 GM-CSF clone expressing 220 ng GM-CSF/10 6 cells/day was derived by transducing B16.F10 cells with a lentiviral vector encoding murine GM-CSF (AMSBIO, Cambridge, MA).

Techniques: Confocal Microscopy, Imaging, Incubation, Staining, Labeling

Journal: Research in Pharmaceutical Sciences

Article Title: Antiangiogenic and antiproliferative effects of black pomegranate peel extract on melanoma cell line

doi:

Figure Lengend Snippet: The effects of PPE on cell viability. As detailed in Materials and methods, A; C554 HUVECs, B; B16F10 melanoma cells were treated with PPE and the viability of cells was determined by the MTT assay. The data are expressed as the percentage of cell viability compare to control.

Article Snippet: The human umbilical vein endothelial cells (HUVECs) (C554) and melanoma cell line (B16F10) (National Cell bank of Iran affiliated to Pasteur Institute, Tehran, Iran) were cultured in dulbecco's modified eagle's medium (DMEM) supplemented with 1% antibiotics (100 units/ml penicillin and 100 μg/ml streptomycin) and 10% fetal calf serum until the third passage before performing the experiments.

Techniques: MTT Assay, Control

Journal: Frontiers in Immunology

Article Title: Bcl6 Preserves the Suppressive Function of Regulatory T Cells During Tumorigenesis

doi: 10.3389/fimmu.2020.00806

Figure Lengend Snippet: The deficiency of Bcl6 in Treg cells leads to delayed tumor progression. (A) RT-qPCR of Bcl6 expression in tumor infiltrating Treg (CD4 + CD25 + GITR + CXCR5 – , noted as TIL Treg) cells, Tfh (gated in CD4 + CD44 + Foxp3 – CXCR5 + ) and Th1 (gated in CD4 + CD44 + Foxp3 – CXCR5 – ) cells sorted from draining lymph node of WT tumor bearing mice. Naive CD4 (gated in CD4 + CD44 – ) sorted from naive WT mice are included as control. Mice were inoculated with 5 × 10 5 B16-F10 cells per mice intravenously, and were sacrificed (indicated as “Sac” in figure) at day 16 for analysis. The data presented are representative of two technical replicates pooled from at least three mice per group. (B) Bcl6 expression in tumor infiltrating Treg, dLNs Treg and spleen Treg (CD4 + CD25 + ) cells. Bcl6-RFP reporter mice were inoculated with 1 × 10 6 MC-38 subcutaneously, and were sacrificed at day 10 for flow cytometry analysis. Statistical analysis shown on the right. The data presented are representative of two independent experiments with at least three mice per group. Error bars indicate the mean ± SEM. * p < 0.05, *** p < 0.001. (C,D) Pictures of MC-38 tumor samples harvested at day 17 after tumor implantation [1 × 10 6 MC-38 cells per mice subcutaneously, panel (C) ], with tumor growth curve and weight (D) . (E) Tumor progression screened by bioluminescent imaging at D11. Mice were inoculated with 5 × 10 5 B16-F10 with luciferase sequence intravenously per mice. (F) Pictures of lung samples harvested from metastasis model, in which mice were inoculated with 1 × 10 6 B16-F10 cells per mice intravenously and sacrificed at day 16, with foci numbers calculated. (G) Survival curve of Bcl6 fl/fl Foxp3 Cre and WT mice inoculated with 5 × 10 5 B16-F10 per mice intravenously. Data were collected from 8 to 14 mice per group with two independent experiments. Statistical differences were calculated by unpaired t -test (tumor weight), Log-rank test (survival curve) or two-way ANOVA with a post hoc Turkey test (tumor growth curve). *** p < 0.001, **** p < 0.0001. Data are presented as mean ± SEM. See also .

Article Snippet: Tumor cell lines, B16-F10, B-Luciferase B16-F10 (BIOCYTOGEN Cat. B-MCL-002) and MC-38 were cultured for cell injection into C57BL/6J mice.

Techniques: Quantitative RT-PCR, Expressing, Flow Cytometry, Tumor Implantation, Imaging, Luciferase, Sequencing

Journal: Frontiers in Immunology

Article Title: Bcl6 Preserves the Suppressive Function of Regulatory T Cells During Tumorigenesis

doi: 10.3389/fimmu.2020.00806

Figure Lengend Snippet: Tfr cells are dispensable for tumor control. (A) Set up of Tfr deficient Bone Marrow Chimera mice. Bone marrow cells were isolated from Foxp3 DTR and Cxcr5 –/– mice were mixed 1:1 and then transferred 8 × 10 6 per mice to irradiated CD4 –/– mice. After reconstruction, mice were inoculated with 5 × 10 5 B16-F10 cells intravenously and were treated with DT or vehicle, then sacrificed at D16 post tumor inoculation. (B) Pictures of lung samples harvested from metastasis model of Tfr deficient mice. (C–E) Flow cytometry analyzing proportion and total number of Treg, Tfr, Tfh, Th1, CD8 + CXCR5 + subpopulation from dLNs (C) and Treg, CD4 + CD44 – (D) , CD8 + T cells (E) from tumor tissue. The data presented are representative of two independent experiments with at least 4 mice per group. # represents the absolute number and % represents the proportion of indicating population. Statistical differences were calculated by unpaired t -test. Center values indicate mean. See also .

Article Snippet: Tumor cell lines, B16-F10, B-Luciferase B16-F10 (BIOCYTOGEN Cat. B-MCL-002) and MC-38 were cultured for cell injection into C57BL/6J mice.

Techniques: Isolation, Irradiation, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Bcl6 Preserves the Suppressive Function of Regulatory T Cells During Tumorigenesis

doi: 10.3389/fimmu.2020.00806

Figure Lengend Snippet: Bcl6 is essential in maintaining the lineage stability of Treg cells in TME. (A) RT-qPCR analysis of Foxp3 expression, normalized to its expression in Bcl6-deficient tumor infiltrating Treg cells (CD4 + CD25 + GITR + CXCR5 – ), assessed at day 16 after B16-F10 inoculation intravenously. (B) RT-qPCR analysis of Foxp3 expression, normalized to its expression in Treg cells (CD4 + CD25 + GITR + CXCR5 – ) from spleen, mesenteric lymph nodes(mLN) or peripheral non-draining lymph nodes (pLN) of naive Bcl6 fl/fl Foxp3 Cre and WT mice. (C,D) RT-qPCR analysis of Gata3 and Il4 (C) , Il17 and Rorc (D) normalized to their expression in Bcl6-deficient and WT tumor infiltrating Treg cells (CD4 + CD25 + GITR + CXCR5 – ). Naive CD4 (gated in CD4 + CD44 – ) sorted from naive WT mice, Tfh (gated in CD4 + CD44 + Foxp3 – CXCR5 + ) and Th1 (gated in CD4 + CD44 + Foxp3 – CXCR5 – ) cells sorted from WT mice infected with Armstrong (Arm) at day 8 are included as control. (E) RT-qPCR analysis of Il17 , Rorc and Gata3 normalized to their expression in Treg cells (CD4 + CD25 + GITR + CXCR5 – ) from spleen, mesenteric lymph nodes(mLN) or peripheral non-draining lymph nodes (pLN) of naive Bcl6 fl/fl Foxp3 Cre and WT mice. The data (A-E) presented are representative of two technical replicates pooled from at least three mice per group. Error bars indicate the mean ± SEM. (F) Flow cytometry analysis of tumor infiltrating Treg cells in splenic chimera mice assay, assessed at day 18 after B16-F10 inoculation intravenously. Quantification of Tbet and GATA3 in Bcl6-deficient and WT Treg cells. The data presented are representative of two independent experiments with at least 4 mice per group. Statistical differences were calculated by paired t -test. ** p < 0.01, **** p < 0.0001.

Article Snippet: Tumor cell lines, B16-F10, B-Luciferase B16-F10 (BIOCYTOGEN Cat. B-MCL-002) and MC-38 were cultured for cell injection into C57BL/6J mice.

Techniques: Quantitative RT-PCR, Expressing, Infection, Flow Cytometry, Mouse Assay