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

Journal: Scientific Reports

Article Title: In vivo neutralization of the protagonist role of macrophages during the chronic inflammatory stage of Huntington’s disease

doi: 10.1038/s41598-018-29792-x

Figure Lengend Snippet: Immune activation in late-stage HD mice as indicated by cytokine levels. ( a ) Cytokine upregulation in the blood of 14 week old R6/2 mice. Plasma levels for R6/2 mice compared to WT are shown ± SEM, ( n = 20/group IL1β; 16/group IL2; 19/group IL6; 26/group IL10 and 28/group TNFα). ( b ) Increases in cytokine gene expression was limited to the striatum of late-stage R6/2 mice. Il1β , Il2 , Il4 , Il6 , Il10 , Il12 , Il17 and Tnfα gene expression levels in cerebellum, striatum and cortex of R6/2 mice ( n = 5/group) compared to WT ( n = 5/group) ± SEM as measured by qPCR. ( c ) Increased cytokine gene expression in R6/2 splenic and peritoneal macrophage enriched cell populations. R6/2 mouse splenic ( n = 10/group) and peritoneal macrophage ( n = 7/group) gene expression for cytokines relative to WT ± SEM quantified by qPCR. * p < 0.05 and ** p < 0.01 vs WT by Student’s t test. WT = wild type, pM0 = peritoneal macrophages.

Article Snippet: Cytokine (IL1β, IL2, IL6, IL10, IL12 and TNFα) levels in plasma were quantified using Meso Scale Discovery (MSD) assays as per the manufacturer’s protocol and analysed on a SECTOR 2400 instrument (MSD).

Techniques: Activation Assay, Clinical Proteomics, Gene Expression

Journal: Scientific Reports

Article Title: In vivo neutralization of the protagonist role of macrophages during the chronic inflammatory stage of Huntington’s disease

doi: 10.1038/s41598-018-29792-x

Figure Lengend Snippet: Immune activity in the periphery of early stage R6/2 mice. Upregulated cytokine and activated macrophage cell marker gene expression levels in 8-week-old R6/2 mice. Gene expression of ( a ) Il6 in peritoneal macrophages ( n = 10/group), ( b ) Il12 ( n = 5/group) and ( c ) Cd40 in splenic cells ( n = 5/group), ( d ) Cd40 in peritoneal macrophages ( n = 10/group) in R6/2 mice compared to WT. ( e ) Increased frequencies of activated macrophages and DC in early-symptomatic stage R6/2 mice. Mean % ± SEM of peritoneal macrophages (gated as; CD11b + , CD8 − , CD11c − , Ly6G − ) and splenic DC (gated as; CD11b + , CD8 − , CD11c + , Ly6G − ) highly expressing cell surface CD40 in 8 week old WT ( n = 5/group) and R6/2 mice ( n = 6/group) as assessed by FACS. ( f ) Increased frequencies of activated T cells in early-symptomatic stage R6/2 mice. Mean percentage ± SEM of splenic T cells (gated as; CD3 + , CD4 + or CD3 + , CD8 + ) highly expressing cell surface OX40 in 8 week old WT ( n = 5/group) and R6/2 mice ( n = 8/group) as assessed by FACS. ( g ) Presence of exon 1 mutant huntingtin ( mHTT ) in splenic mononuclear cells and peritoneal macrophages in 8 week old R6/2 ( n = 6) but not in WT ( n = 5) mice as assessed by qPCR, mean CT values ± SEM are shown for both mHTT and the geometric mean of ATP5B and B2M housekeeping genes (HK). Agarose gel electrophoresis of the corresponding mHTT qPCR products are shown below each graph. # p = 0.06, * p < 0.05 vs WT by Student’s t test. WT = wild type, pM0 = peritoneal macrophages.

Article Snippet: Cytokine (IL1β, IL2, IL6, IL10, IL12 and TNFα) levels in plasma were quantified using Meso Scale Discovery (MSD) assays as per the manufacturer’s protocol and analysed on a SECTOR 2400 instrument (MSD).

Techniques: Activity Assay, Marker, Gene Expression, Expressing, Mutagenesis, Agarose Gel Electrophoresis

Journal: Scientific Reports

Article Title: In vivo neutralization of the protagonist role of macrophages during the chronic inflammatory stage of Huntington’s disease

doi: 10.1038/s41598-018-29792-x

Figure Lengend Snippet: Immune activation in zQ175 mice. ( a ) Increased gene expression in late-stage zQ175 mice for several cytokines in splenic ( n = 5–6/group) and peritoneal ( n = 5–6/group) macrophage populations. 22-month-old zQ175 mice ± SEM splenic and peritoneal macrophage gene expression for cytokines relative to WT as quantified by qPCR. ( b ) Increased activated T cell numbers in 22-month-old Q175 mice. Mean % ± SEM of splenic T cells (gated as; CD3 + , CD4 + or CD8 + ) expressing OX40 highly in 22 month old WT ( n = 5/group) and zQ175 mice ( n = 6/group) as assessed by FACS. ( c ) Increased activated peritoneal macrophage and DC frequencies in 22-month-old zQ175 mice. Mean percentage ± SEM of peritoneal macrophage and splenic DC expressing OX40L highly in 22 month old WT ( n = 5/group) and zQ175 mice ( n = 5/group) as assessed by FACS. ( d ) Increases in cytokine gene expression was limited to the striatum of 12 month old zQ175 mice. Mean Il1β , Il6 , Il10 , Il12 and Tnfα gene expression levels in cerebellum, striatum and cortex of zQ175 mice ( n = 5/group) compared to WT ( n = 5/group) ± SEM as measured by qPCR. ( e ) There were no increases in cytokine gene expression in 21 month old zQ175 mice. Mean Il1β , Il6 , Il10 , Il12 and Tnfα gene expression levels in cerebellum, striatum and cortex of zQ175 mice ( n = 5/group) compared to WT ( n = 5/group) ± SEM as measured by qPCR. * p < 0.05 and ** p < 0.01 vs WT by Student’s t test. WT = wild type, pM0 = peritoneal macrophages, DC = dendritic cells.

Article Snippet: Cytokine (IL1β, IL2, IL6, IL10, IL12 and TNFα) levels in plasma were quantified using Meso Scale Discovery (MSD) assays as per the manufacturer’s protocol and analysed on a SECTOR 2400 instrument (MSD).

Techniques: Activation Assay, Gene Expression, Expressing

Journal: Scientific Reports

Article Title: In vivo neutralization of the protagonist role of macrophages during the chronic inflammatory stage of Huntington’s disease

doi: 10.1038/s41598-018-29792-x

Figure Lengend Snippet: Clodronate liposome normalisation of cytokine levels in late-stage R6/2 mice. ( a ) Clodronate liposome treatment reduced IL1β, IL2 and IL6 levels in late-stage R6/2 mice post IV treatment ( n = 8–12/group). Mean concentration of IL10, IL1β, IL2, IL6 and TNFα ± SEM following clodronate liposome IV injection, at the day of greatest splenic M0 depletion and one day later as assessed by MSD. ( b ) Clodronate liposome treatment normalised TNFα, IL2 and IL6 levels in late-stage R6/2 mice post IP treatment ( n = 9–12/group). Mean concentration of TNFα, IL1β, IL2, IL6 and IL10 ± SEM following clodronate liposome IP injection, at day of highest splenic M0 depletion and one day later as assessed by MSD. All comparisons to R6/2 levels at day 0, ANOVA with Bonferroni correction. * p < 0.05, # p = 0.06. WT = wild type, IV = intravenous, IP = intraperitoneal, M0 = splenic macrophages.

Article Snippet: Cytokine (IL1β, IL2, IL6, IL10, IL12 and TNFα) levels in plasma were quantified using Meso Scale Discovery (MSD) assays as per the manufacturer’s protocol and analysed on a SECTOR 2400 instrument (MSD).

Techniques: Concentration Assay, IV Injection, Injection

Journal: Viruses

Article Title: MAVS Genetic Variation Is Associated with Decreased HIV-1 Replication In Vitro and Reduced CD4 + T Cell Infection in HIV-1-Infected Individuals

doi: 10.3390/v12070764

Figure Lengend Snippet: MAVS genetic variation does not affect intracellular cytokine levels of HIV-1-specific T cells. ( a ) PBMCs of individuals carrying a MAVS minor or MAVS major genotype obtained 2.5–3.5 years p.SC were left unstimulated (-), or were treated with HIV-1 consensus B Gag peptide pool (Gag) or staphylococcal enterotoxin B (SEB). After 6 h, the percentages of surface molecules expressing CD137 + and CD107a + and intracellular expression of IFN-γ + , IL-2 + , Mip-1β + , and TNF-α + CD4 + T cells and ( b ) CD8 + T cells were analyzed using flow cytometry. Each square or dot represents a different study participant (median ± IQR). No significant differences between HIV-1-infected individuals with a MAVS minor or MAVS major genotype were observed.

Article Snippet: Then, cells were washed, permeabilized using a Cytofix/Cytoperm kit (BD Biosciences, San Jose, CA, USA) and stained for 30 min at 4 °C to determine the expression levels of intracellular cytokines IL-2 (PE, 340450, BD Biosciences, San Jose, CA, USA), TNF-α (Alexa Fluor 700, 557996, BD Biosciences, San Jose, CA, USA), Mip-1β (PE-Cy7, 560687, BD Biosciences, San Jose, CA, USA), and IFN-γ (APC-Alexa Fluor 750, MHCIFG27, Thermo Fisher, Waltham, MA, USA).

Techniques: Expressing, Flow Cytometry, Infection

Journal: Transboundary and Emerging Diseases

Article Title: Construction and Evaluation of Quadruple-Gene-Deleted Pseudorabies Virus Platforms for ASFV Antigen Delivery

doi: 10.1155/tbed/3628600

Figure Lengend Snippet: Effects of p54, p72, CD2v, and pp62 antigen immunization on immune responses in mice. (A) Detection of IL-2, IL-4, and IFN-γ levels secreted by splenocytes from mice immunized with rPRV-p54+p72 and rPRV-CD2v+pp62 using ELISA. (B) Effects of p54, p72, CD2v, and pp62 antigen stimulation on mouse splenocyte proliferation. (C, D) Flow cytometric analysis of CD3+, CD4+, and CD8+ T cell percentages in spleens of mice immunized with different antigens. ⁣ ∗ p < 0.05, ⁣ ∗∗ p < 0.01, ⁣ ∗∗∗ p < 0.001. ns : not significant.

Article Snippet: Serum cytokine levels in mice (IL-12, IL-4, IL-6, IL-10, IFN-γ, and TNF-α) were measured using commercial ELISA test kits (catalog numbers KE10004, KE10010, KE10007, KE10103, KE10094, and KE10002, Proteintech).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Transboundary and Emerging Diseases

Article Title: Construction and Evaluation of Quadruple-Gene-Deleted Pseudorabies Virus Platforms for ASFV Antigen Delivery

doi: 10.1155/tbed/3628600

Figure Lengend Snippet: rPRV-p54+p72 and rPRV-CD2v+pp62 protected mice against challenge with PRV SX-10 infection. (A) Survival rates of mice after challenge with PRV SX-10 strain. (B) Histopathological lesions in various organs of mice following PRV challenge. Tissue samples were collected as indicated for histopathological examination. (C) Cytokine expression levels of TNF-α, IL-6, and IL-10 in mouse sera were detected at 3 dpc using ELISA. ⁣ ∗ p < 0.05, ⁣ ∗∗ p < 0.01, ⁣ ∗∗∗ p < 0.001, ⁣ ∗∗∗∗ p < 0.0001, and ns as not significant.

Article Snippet: Serum cytokine levels in mice (IL-12, IL-4, IL-6, IL-10, IFN-γ, and TNF-α) were measured using commercial ELISA test kits (catalog numbers KE10004, KE10010, KE10007, KE10103, KE10094, and KE10002, Proteintech).

Techniques: Infection, Expressing, Enzyme-linked Immunosorbent Assay