Journal:

Article Title: Role of early- or late-phase activation of p38 mitogen-activated protein kinase induced by tumour necrosis factor-? or 2,4-dinitrochlorobenzene during maturation of murine dendritic cells

doi: 10.1046/j.1365-2567.2003.01746.x

Figure Lengend Snippet: Effect of SB203580 on tumour necrosis factor-α (TNF-α)- or 2,4-dinitrochlorobenzene (DNCB)-induced phenotypic maturation of BC1 cells. BC1 cells were pretreated with or without SB203580 (20 µm) for 1 hr and then treated with TNF-α (40 ng/ml) or DNCB (2·0 µm) in the presence or absence of SB203580 for 24 hr. The cell-surface expression of CD86, CD80 or major histocompatibility complex (MHC) class II was analysed by flow cytometry. Each column represents the mean ± standard error (SE) of three or four independent experiments. Statistical significance was calculated by the Student's t-test (*P < 0·05; **P < 0·01).

Article Snippet: SB203580 was obtained from Calbiochem (La Jolla, CA) and used at 20 µ m . Fluorescein isothiocyanate (FITC)-conjugated anti-mouse CD11c monoclonal antibody (mAb) (HL3), phycoerythrin (PE)-conjugated anti-mouse CD80 mAb (16-10A1), PE-conjugated anti-mouse CD86 mAb (GL1), biotin-conjugated anti-mouse H-2K d mAb (SF1-1.1), biotin-conjugated anti-mouse I-A d mAb (AMS-32.1), and streptavidin Cy-Chrome™, were obtained from PharMingen (La Jolla, CA).

Techniques: Expressing, Flow Cytometry

Journal: Drug Design, Development and Therapy

Article Title: Osteopontin promotes dendritic cell maturation and function in response to HBV antigens

doi: 10.2147/DDDT.S81656

Figure Lengend Snippet: Blockage of OPN suppressed DC maturation and cytokine production after hepatitis B virus (HBV) antigenic stimulations in human PBMC. Notes: After administering OPN neutralizing antibody in vitro, the histograms ( A ) and expressions ( B ) of CD80, CD86, and HLA-DR in BDCA-1 + cells after HBV antigenic stimulations in human PBMCs are presented. Overlay plots are representative and show isotype control (shadow) and the expressions of the indicated molecules on DCs in the absence (green line) or presence (red line) of neutralizing antibody in each group. Levels of IL-12 ( C ), TNF-α, IFN-γ, and IL-4 ( D ) in human PBMC supernatant were measured after administering the OPN-neutralizing antibody in vitro as well as HBcAg and HBsAg stimulations. * P <0.05. Abbreviations: OPN, osteopontin; DCs, dendritic cells; HBV, hepatitis B virus; HBcAg, hepatitis B core antigen; HBsAg, hepatitis B surface antigen; PBMC, peripheral blood mononuclear cells; IL, interleukin; TNF, tumor necrosis factor; IFN, interferon; Ab, antibody; IgG, Immunoglobulin gamma; HLA-DR, human leukocyte antigen-DR; BDCA-1, blood dendritic cell antigen 1.

Article Snippet: Expressions of surface molecules on DC were detected by flow cytometry using the following antibodies: PE-anti-mouse I-Ab/I-Ed (M5/114), PE-anti-mouse B7-1 (1G10), PE-anti-mouse B7-2 (Gl-1), FITC-anti-mouse CD11c (HL3), APC-anti-mouse TCRβ (H57-597), FITC-anti-human CD80 (L307.4), FITC-anti-human CD86 (2331 (FUN-1)), FITC-anti-human human leukocyte antigen DR (HLA-DR) (G46-6), PE-anti-human BDCA-1 (F10/21A3) and APC-anti-human CD3 (HIT3A) (all from BD Biosciences, San Diego, CA, USA).

Techniques: In Vitro

Journal: Drug Design, Development and Therapy

Article Title: Osteopontin promotes dendritic cell maturation and function in response to HBV antigens

doi: 10.2147/DDDT.S81656

Figure Lengend Snippet: OPN deficiency impaired DCs maturation and function after HBV antigenic stimulation in vitro. Notes: DCs derived from BM of the WT and OPN −/− mice were stimulated by HBV antigens. The histograms ( A ) and percentages ( B–D ) of CD80, CD86, and MHC-II molecules on the DC surface were detected by flow cytometry. Overlay plots are representative and show isotype control (shadow) and the expressions of the indicated molecules on DCs from WT mice (green line) or OPN −/− mice (red line) in each group. ( E ) Levels of IL-12 in the BMDC culture supernatants following different stimulations were measured by ELISA. * P <0.05; ** P <0.01. Abbreviations: OPN, osteopontin; DCs, dendritic cells; HBV, hepatitis B virus; BM, bone marrow; WT, wild-type; HBcAg, hepatitis B core antigen; HBsAg, surface antigen of the hepatitis B virus; MHC, major histocompatibility complex; IL, interleukin; BMDC, bone marrow-derived dendritic cell; ELISA, enzyme-linked immunosorbent assay.

Article Snippet: Expressions of surface molecules on DC were detected by flow cytometry using the following antibodies: PE-anti-mouse I-Ab/I-Ed (M5/114), PE-anti-mouse B7-1 (1G10), PE-anti-mouse B7-2 (Gl-1), FITC-anti-mouse CD11c (HL3), APC-anti-mouse TCRβ (H57-597), FITC-anti-human CD80 (L307.4), FITC-anti-human CD86 (2331 (FUN-1)), FITC-anti-human human leukocyte antigen DR (HLA-DR) (G46-6), PE-anti-human BDCA-1 (F10/21A3) and APC-anti-human CD3 (HIT3A) (all from BD Biosciences, San Diego, CA, USA).

Techniques: In Vitro, Derivative Assay, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Drug Design, Development and Therapy

Article Title: Osteopontin promotes dendritic cell maturation and function in response to HBV antigens

doi: 10.2147/DDDT.S81656

Figure Lengend Snippet: OPN produced by DCs plays an important role in response to HBV antigenic stimulation in vivo. Notes: At 24 hours after HepG2.2.15 cells supernatant direct injection into liver from the WT mice and OPN −/− mice ( A ) liver histology was assessed. Original magnification, 400×. ( B ) Leukocyte number and ( C ) expressions of CD80, CD86, and MHC-II molecules on DCs in liver from the WT mice and OPN −/− mice were measured. ( D ) Number of TCR + cells per gram of liver tissue were detected by an automated cell counter and flow cytometry. ( E ) Number of IFN-γ-producing T-cells and IL-4-producing T-cells were detected. HepG2 supernatant injection was used as control. n=5 per group. Adoptive transfer of CD11c + cells from the WT and OPN −/− mice as well as HepG2.2.15 supernatant direct injection into mice liver were performed. Then ( F ) the number of leukocyte per gram of liver tissue was counted in the OPN −/− mice at 0 and 24 hours; n=5 per group. ( G ) IFN-γ-producing cells in T-cells in liver from the OPN −/− mice were analyzed. n=5 per group. * P <0.05; ** P <0.01; *** P <0.001. Abbreviations: OPN, osteopontin; DCs, dendritic cells; HBV, hepatitis B virus; WT, wild-type; MHC, major histocompatibility complex; IL, interleukin; IFN, interferon; TCR, T-cell receptor; h, hour.

Article Snippet: Expressions of surface molecules on DC were detected by flow cytometry using the following antibodies: PE-anti-mouse I-Ab/I-Ed (M5/114), PE-anti-mouse B7-1 (1G10), PE-anti-mouse B7-2 (Gl-1), FITC-anti-mouse CD11c (HL3), APC-anti-mouse TCRβ (H57-597), FITC-anti-human CD80 (L307.4), FITC-anti-human CD86 (2331 (FUN-1)), FITC-anti-human human leukocyte antigen DR (HLA-DR) (G46-6), PE-anti-human BDCA-1 (F10/21A3) and APC-anti-human CD3 (HIT3A) (all from BD Biosciences, San Diego, CA, USA).

Techniques: Produced, In Vivo, Injection, Flow Cytometry, Adoptive Transfer Assay

Journal: Drug Design, Development and Therapy

Article Title: Osteopontin promotes dendritic cell maturation and function in response to HBV antigens

doi: 10.2147/DDDT.S81656

Figure Lengend Snippet: OPN administration significantly promoted the maturation of DCs from CHB patients after HBcAg stimulation. Notes: The expressions of CD80 ( A ), CD86 ( B ), and HLA-DR ( C ) in BDCA-1 + cells from the peripheral blood of CHB patients following HBcAg stimulation with or without exogenous OPN administration were measured by flow cytometry. CHB patients, n=20. ** P <0.01, *** P <0.001. ( D ) The percentages of IFN-γ-producing cell in CD3 + T-cells from the peripheral blood of CHB patients following HBcAg stimulation with or without exogenous OPN administration were measured by flow cytometry. Abbreviations: OPN, osteopontin; DCs, dendritic cells; CHB, chronic hepatitis B; HBV, hepatitis B virus; HBcAg, hepatitis B core antigen; IFN, interferon; HLA-DR, human leukocyte antigen-DR; BDCA-1, Blood Dendritic Cell Antigen 1.

Article Snippet: Expressions of surface molecules on DC were detected by flow cytometry using the following antibodies: PE-anti-mouse I-Ab/I-Ed (M5/114), PE-anti-mouse B7-1 (1G10), PE-anti-mouse B7-2 (Gl-1), FITC-anti-mouse CD11c (HL3), APC-anti-mouse TCRβ (H57-597), FITC-anti-human CD80 (L307.4), FITC-anti-human CD86 (2331 (FUN-1)), FITC-anti-human human leukocyte antigen DR (HLA-DR) (G46-6), PE-anti-human BDCA-1 (F10/21A3) and APC-anti-human CD3 (HIT3A) (all from BD Biosciences, San Diego, CA, USA).

Techniques: Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Targeting immunosuppressive Ly6C+ classical monocytes reverses anti-PD-1/CTLA-4 immunotherapy resistance

doi: 10.3389/fimmu.2023.1161869

Figure Lengend Snippet: Anti-PD-1/CTLA-4/Ly6C treatment reduces tumor growth and increases CD8 T cells/dendritic cells. (A) 344SQ tumor bearing 129/Sv mice were weekly treated with anti-PD-1_CTLA-4 (200 µg of anti-PD-1 plus 200 µg of anti-CTLA-4 per mouse), anti-Ly6C alone (200 µg of anti-Ly6C per mouse), anti-PD-1_CTLA-4_Ly6C (200 µg of anti-PD-1 plus 200 µg of anti-CTLA-4 plus 200 µg of anti-Ly6C per mouse), or their IgG control mixture (IgG control) beginning on week 2 after a subcutaneous cancer cell injection (1 x 10 6 cells per mouse; n = 5) for 4 weeks. Mice received total of 4 treatments starting at week 2 post tumor cell implantation. (B) tSNE CD45 plots from 344SQ tumors treated with IgG control, anti-PD-1_CTLA-4, or anti-PD-1_CTLA-4_Ly6C from (A) . Tumors from (A) were harvested to prepare single cell suspensions for FACS analysis. (C) Percentages of CD8 T cells (left), Effector/memory CD8 T cells (middle), and dendritic cells (right). (D) Percentage of Ly6C- (left) and Ly6C+ (right) CD14+CD115+ monocytes. Kras LSL-G12D /p53 fl/fl mice generated through intratracheal administration of adenovirus expressing Cre recombinase were treated with either anti-PD-1/CTLA-4 or anti-PD-1/CTLA-4/Ly6C for 8 weeks (E) Micro-CT images shown at week 0 (baseline) and week 8 (endpoint) for anti-PD-1/CTLA-4 or anti-PD-1/CTLA-4/Ly6C. Dashed yellow circles indicate lung tumors. H indicates of heart. (F) Percentage change of tumor area was calculated taking into account prior time point and normalized to the baseline measurement. (G) H&E stained lung sections at week 8 from anti-PD-1/CTLA-4 or anti-PD-1/CTLA-4/Ly6C treated mice from (E) . bar= 5 mm. (H) Number of lung tumors on H&E sections from (G) treated with anti-PD-1/CTLA-4 or anti-PD-1/CTLA-4/Ly6C weekly for 8 weeks. (I) IHC of CD8 stained lung tumors from (F) week 8. Percentage of CD8 T cells found in the lung tumors from Kras LSL-G12D /p53 fl/fl mice treated for 8 weeks with anti-PD-1/CTLA-4 or anti-PD-1/CTLA-4/Ly6C. (J) The dendritic cells from CD11c-DTR mice were transferred into C57BL/6 mice to generate the chimerical mice, after 8-week stabilization, the mice were treated with diphtheria toxin twice a week to maintain the depletion of dendritic cells. The B16 melanoma-bearing mice were treated weekly with anti-PD-1_CTLA-4_Ly6C for 3 weeks staring on week 1 after tumor cells inoculation. (K) 344SQ tumor bearing 129/Sv mice were treated weekly with blockade of PD-1, CTLA-4, and Ly6C (200 µg of anti-PD-1 plus 200 µg of anti-CTLA-4 plus 200 µg of anti-Ly6C per mouse) or their IgG control mixture (IgG control) beginning on day 7 after a subcutaneous cancer cell injection (0.1 x 10 6 cells per mouse; n = 5) for 4 weeks. For blocking B7 signal, antibodies (anti-B7: 300 μg of anti-CD80 and 300 μg of anti-CD86 per mouse) were intraperitoneally administered 1 day before the first dose of therapy, and then once a week to maintain the blockade. ANOVA test was used to analyze the data. n.s., not significance; *p < 0.05; **p < 0.01; ***p< 0.001; ****p < 0.0001.

Article Snippet: Anti-mouse PD-1 (clone RMP1-14), anti-mouse CTLA-4 (clone 9D9), anti-mouse CD80 (clone 16-10A1), anti-mouse CD86 (clone GL-1), anti-mouse Ly6C (clone Monts 1), anti-mouse CCL2 (clone 2H5), and the isotype-matched IgG controls were purchased from BioXCell.

Techniques: Control, Injection, Generated, Expressing, Micro-CT, Staining, Blocking Assay

Journal: Science Advances

Article Title: Biofunctional lipid nanoparticles for precision treatment and prophylaxis of bacterial infections

doi: 10.1126/sciadv.adk9754

Figure Lengend Snippet: ( A ) Expression levels of CD80 + and CD86 + in BMDCs (CD11c + ) and ( B ) production of IL-6 and TNF-α of the cell supernatant measured by ELISA after different treatments (concentration, 2 μg ml −1 NPs) for 24 hours ( n = 3). ( C ) Schematics of experimental design to evaluate the in vivo short-term immune responses activated by a single dose of various LNPs (20 μg/20 g) via subcutaneous injection. ( D ) Percentage of CD80 + and ( E ) CD86 + cells (gated on CD11c + cells) in the LNs at day 3 after immunization ( n = 6). ( F ) Percentage of CD3 + T cells, ( G ) CD8 + T cells (gated on CD3 + T cells), and ( H ) CD4 + T cells (gated on CD3 + T cells) in the spleen at day 3 after immunization ( n = 6). Naive mice without immunization were used as control. * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: Fluorescein isothiocyanate (FITC)–conjugated anti-mouse CD11c, phycoerythrin (PE)–conjugated anti-mouse CD80, PE-conjugated anti-mouse CD86, allophycocyanin (APC)–conjugated anti-mouse CD3, PE-conjugated anti-mouse CD8a, and FITC-conjugated anti-mouse CD4 were purchased from Tonbo Biosciences, USA.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Concentration Assay, In Vivo, Injection, Control