Journal: Journal of Nanobiotechnology

Article Title: Cu-doped TiO 2 nanoparticles improve local antitumor immune activation and optimize dendritic cell vaccine strategies

doi: 10.1186/s12951-023-01844-z

Figure Lengend Snippet: A Histogram displaying the ratio of CD8 + T cells over CD4 + T cells obtained upon in vitro stimulation by OVA-peptide loaded DCs that had either been left untreated (control) or were activated classically, or upon exposure to 33% Cu-doped TiO 2 NPs. B , C Histograms displaying the relative level (control: 100%) of B) granzyme B release, or C) perforin release from T cells obtained from part A that had been exposed to the OVA-derived SIINFEKL peptide. D , E Histograms displaying the level of D) IL12p70 release or E) relative Ccr7 surface expression (control: 100%) on DCs that had either been left untreated (control) or were activated classically, or exposed to 33% Cu-doped TiO 2 NPs. F , G) Histograms displaying the F) number of migrated DCs or G) the level of IL12p70 secreted from migratory DCs obtained from DCs that had either been left untreated (control) or were activated classically, or exposed to 33% Cu-doped TiO 2 NPs and subsequently exposed to the CCR7 ligand 6C-kine. H , I Histograms displaying the in vivo antibody titer for H) OVA-specific IgG1 or I) OVA-specific IgG2α obtained upon intravenous administration of OVA-pulsed DCs that had either been left untreated (control) or were activated classically, or exposed to 33% Cu-doped TiO 2 NPs. J Histogram displaying the level of IFNγ obtained from ex vivo splenocytes obtained from animals receiving the different types of OVA-pulsed DCs and incubated with OVA ex vivo. K Tumor volumes obtained from OVA-expressing E0771 tumors grafted subcutaneously in C57Bl6 mice receiving the different OVA-pulsed DCs that had either been left untreated (control) or were activated classically, or exposed to 33% Cu-doped TiO 2 NPs. L , M Histograms displaying the relative level (control: 100%) of L) granzyme B release, or M) perforin release from CD8 + T cells isolated from OVA-E0771 tumor-bearing animals having received OVA-pulsed DC grafts of the different DC types and then exposed to the OVA-derived SIINFEKL peptide ex vivo. N Histogram displaying the level of OVA-specific CD8 + T cells isolated from the spleen of OVA-E0771 tumor-bearing animals having received OVA-pulsed DC grafts of the different DC types

Article Snippet: Media were then removed and fresh media (5 ml) was given with either no additions (controls), 33% Cu-doped TiO 2 NPs at 40 μg/ml (NP condition) or addition of maturation factors IL1β (25 ng/ml) TNFα (50 ng/ml) and IFNγ (1000 units/ml) (all from PeproTech; this is the “classical” activation scheme) for 24 h. Media were then removed and cells were incubated with soluble recombinant CD40L (BioTechne) at 16 μg/ml for 24 h after which the supernatants was collected and used to determine IL12p70 levels by ELISA (M1270, R&D Systems).

Techniques: In Vitro, Control, Derivative Assay, Expressing, In Vivo, Ex Vivo, Incubation, Isolation

Journal: PLoS ONE

Article Title: TRIF Is a Critical Negative Regulator of TLR Agonist Mediated Activation of Dendritic Cells In Vivo

doi: 10.1371/journal.pone.0022064

Figure Lengend Snippet: ( A ) CD11c + dendritic cells were isolated from C57BL/6 WT (N = 2), MyD88-KO (N = 2), TRIF-KO (N = 2), and MyD88/TRIF-DKO (N = 2) mice, in vitro stimulated with rEA, then used to perform a IL12p70 ELISA as described in . One (of three) representative experiments is shown. The bars represent Mean ± SD. Statistical analysis was completed using a two-way ANOVA with a Bonferroni post-hoc test (genotypes×rEA treatments). *, ** - Indicate values that are statistically different from those in unstimulated DCs (for the same genotype), p<0.05, p<0.001 respectively. #, ## - Indicate values statistically different from those in WT DCs (for the same rEA dose), p<0.05, p<0.01 respectively. No significant differences between mock-injected animals of different genotypes were detected. No significant activation of IL12p70 was observed in MyD88-KO and MyD88/TRIF-DKO DCs. ( B ) DC culture media was collected at 18 hours post-rEA stimulation (0.2 ng/ml) and was analyzed for cytokines/chemokines levels using a multiplexed bead array based quantitative system. Statistical analysis was completed using a one-way ANOVA with a Student-Newman-Keuls post-hoc test. The bars represent Mean ± SD. *, ** - Indicate cytokine values that are statistically different from those in mock injected animals, p<0.05, p<0.001 respectively. No significant differences between mock-injected animals of different genotypes were detected. No significant activation of pro-inflammatory cytokines was observed in MyD88-KO and MyD88/TRIF-DKO animals (only anti-inflammatory IL10 cytokine was induced in MyD88-KO mice).

Article Snippet: Following incubation, culture medium was analyzed for mouse IL12p70 levels using an ELISA kit and following its enclosed instructions (R&D Systems, Minneapolis, MN) or for 23 mouse cytokines/chemokines using multiplex system (Bio-Rad, Hercules, CA).

Techniques: Isolation, In Vitro, Enzyme-linked Immunosorbent Assay, Injection, Activation Assay

Journal: PLoS ONE

Article Title: TRIF Is a Critical Negative Regulator of TLR Agonist Mediated Activation of Dendritic Cells In Vivo

doi: 10.1371/journal.pone.0022064

Figure Lengend Snippet: ( A ) CD11c + dendritic cells were isolated from C57BL/6 WT (N = 3), MyD88-KO (N = 3), TRIF-KO (N = 3), and MyD88/TRIF-DKO (N = 3) mice, in vitro stimulated with various TLR agonists, and used to perform a IL12p70 ELISA as described in . The bars represent Mean ± SEM. Statistical analysis was completed using a one-way ANOVA with Student-Newman-Keuls post-hoc test. ** - Indicate values that are statistically different from those in unstimulated DCs (for the same genotype), p<0.001. No significant differences between mock-injected animals of different genotypes were detected. No significant activation of IL12p70 was observed in MyD88-KO and MyD88/TRIF-DKO DCs. ( B ) DC culture media was collected at 15 hours post stimulation with various TLR agonists (rEA, LPS, ODN2006) and was analyzed for cytokines/chemokines levels using a multiplexed bead array based quantitative system. Statistical analysis was completed using a one-way ANOVA with a Student-Newman-Keuls post-hoc test. The bars represent Mean ± SD. *, ** - Indicate cytokine values that are statistically different from those in mock injected animals, p<0.05, p<0.001 respectively.

Article Snippet: Following incubation, culture medium was analyzed for mouse IL12p70 levels using an ELISA kit and following its enclosed instructions (R&D Systems, Minneapolis, MN) or for 23 mouse cytokines/chemokines using multiplex system (Bio-Rad, Hercules, CA).

Techniques: Isolation, In Vitro, Enzyme-linked Immunosorbent Assay, Injection, Activation Assay

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Critical Role of Zinc Transporter (Zip8) in Myeloid Innate Immune Cell Function and the Host Response against Bacterial Pneumonia

doi: 10.4049/jimmunol.2001395

Figure Lengend Snippet: Analysis of broncho-alveolar lavage fluid for cellularity, cytokine, and chemokine content following S. pneumoniae instillation. WT and Zip8-KO mice were administered 4 x 108 CFU S. pneumoniae intranasally and euthanized 24 or 72 hours later. Lungs were lavaged with PBS, cells collected, counted, immobilized on glass slides, and stained for morphological characterization for (A) total macrophage and (B) neutrophil cell counts. (C-E) Cell-free BAL fluid was also analyzed by ELISA (C) IL-12/23p40 (D) IL-6 and (E) TNF-α 24 hours post infection. Data are presented as the mean ± SEM and represent at least two to three independent studies. (n=18-19 mice per group (24 hours) and 7-8 mice per group (72 hours); **p < 0.01).

Article Snippet: Cytokine levels (TNF-α, IL-2, IL-4, IL-17A/F, IL-6, IL-10, IL-12/23p40, IFN-γ and CXCL-1) were determined using commercially available ELISA kits (BioLegend and R&D, Minneapolis, Mn) according to manufacturer’s instructions.

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Infection

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Critical Role of Zinc Transporter (Zip8) in Myeloid Innate Immune Cell Function and the Host Response against Bacterial Pneumonia

doi: 10.4049/jimmunol.2001395

Figure Lengend Snippet: Cytokine production from BMDCs following stimulation with gram negative and gram-positive bacterial cell wall products in vitro. BMDCs from WT and Zip8-KO mice were stimulated with either LPS (1 μg/ml) or LTA (10 μg/ml) for 6 hours. (A-E) Cytokine production of IL-12/23p40, IL-6, TNF-α, IL-10 and CXCL-1 from LPS/LTA stimulated or unstimulated BMDCs was compared and measured by ELISA. Data are presented as the mean ± SEM and represent at least four independent studies. (n= 16 samples per group; *p < 0.05, **p < 0.01, ****p < 0.0001).

Article Snippet: Cytokine levels (TNF-α, IL-2, IL-4, IL-17A/F, IL-6, IL-10, IL-12/23p40, IFN-γ and CXCL-1) were determined using commercially available ELISA kits (BioLegend and R&D, Minneapolis, Mn) according to manufacturer’s instructions.

Techniques: In Vitro, Enzyme-linked Immunosorbent Assay