Journal: International Journal of Molecular Sciences

Article Title: Colonic Mucosal Immune Activation in Mice with Ovalbumin-Induced Allergic Airway Disease: Association between Allergic Airway Disease and Irritable Bowel Syndrome

doi: 10.3390/ijms23010181

Figure Lengend Snippet: The results of cytokine ELISA assay. T-helper 1 (Th1) (IL-1β, interferon [IFN]-γ, and tumor necrosis factor [TNF]-α), Th2 (IL-4, IL-5 and IL-10), and proinflammatory cytokine (IL-6) levels were measured using ELISA with a multiplex protein assay kit. * p < 0.05, ** p = 0.064.

Article Snippet: A multiplex protein assay kit (Bio-Rad, Hercules, CA, USA) was used to measure the mucosal levels of Th1 (IFN-γ, IL-1β and TNF-α), Th2 (IL-4, IL-5 and IL-10), and proinflammatory cytokines (IL-6).

Techniques: Enzyme-linked Immunosorbent Assay, Multiplex Assay

Journal: PLoS Pathogens

Article Title: A20 Deficiency in Lung Epithelial Cells Protects against Influenza A Virus Infection

doi: 10.1371/journal.ppat.1005410

Figure Lengend Snippet: Intratracheal administration of 0.5μg TNF (A and B) or 50μg poly(I:C) (C and D) to A20 AEC-KO or wild type littermates (A20 WT ). Absolute numbers of neutrophils or monocytes in bronchoalveolar lavages (BAL) as determined by flow cytometry at 6h and 24h post-treatment for TNF (A) or 24h post-treatment for poly(I:C) (C). IL-6, CXCL1 (KC), CCL2 (MCP-1) and TNF [only for poly(I:C)] protein levels in BAL fluid detected by Multiplex immunoassay (B and D). Data represent mean ± SEM of at least 4 mice per group (*p < 0.05; Student’s t -test).

Article Snippet: Recombinant mouse CCL2 (R&D Systems, endotoxin levels <0.01 EU per μg of protein as measured by the LAL method) was administered intranasally at a dose of 50 μg/kg at day 6 post infection.

Techniques: Flow Cytometry, Multiplex Assay

Journal: PLoS Pathogens

Article Title: A20 Deficiency in Lung Epithelial Cells Protects against Influenza A Virus Infection

doi: 10.1371/journal.ppat.1005410

Figure Lengend Snippet: (A) Absolute numbers of monocytes, neutrophils and alveolar macrophages in bronchoalveolar lavages (BAL) of A20 AEC-KO or A20 WT mice at 2, 5, 8 and 12 days post-infection (days p.i.) with 0.05 X LD 50 X-47. (B) Absolute numbers of resident CD11b - or recruited CD11b + macrophages in the lungs of A20 WT and A20 AEC-KO mice. (C) CCL2 (MCP-1) protein levels in BAL fluid measured by Multiplex immunoassay at indicated time points post-infection. (D) Weight loss of A20 AEC-KO and A20 WT mice infected with 0.05 X LD 50 X-47. At day 6 p.i. (indicated by an arrow) mice received intranasal treatment with 50 μg/kg recombinant CCL2 (rCCL2) or PBS. Data were analysed using Student’s t -test (A, B and C *p < 0.05) and 2-way ANOVA (D, *p < 0.05 for A20 AEC-KO PBS vs. A20 WT PBS and # p < 0.05 for A20 AEC-KO PBS vs A20 AEC-Cre rCCL2). Data represent mean ± SEM of at least 3 mice per group. Data are representative of at least 2 independent experiments.

Article Snippet: Recombinant mouse CCL2 (R&D Systems, endotoxin levels <0.01 EU per μg of protein as measured by the LAL method) was administered intranasally at a dose of 50 μg/kg at day 6 post infection.

Techniques: Infection, Multiplex Assay, Recombinant

Journal: Infection and Immunity

Article Title: Group A Streptococcus Infection of the Nasopharynx Requires Proinflammatory Signaling through the Interleukin-1 Receptor

doi: 10.1128/IAI.00356-20

Figure Lengend Snippet: Anakinra antagonizes GAS colonization of the nasopharynx. Induction of inflammatory cytokines and their contribution to GAS infection. C57BL/6 mice were treated with anakinra (50 μg/kg) or PBS control and inoculated intranasally with 108 CFU of GAS M1T1 5448. Mice were euthanized after 24 or 72 h, and the nasopharynxes were subjected to lavage to quantify GAS CFU by dilution plating (A), quantify cytokines with a Meso Scale Diagnostics multiplex ELISA (means are displayed; comparisons indicated reach significance only for IL-1β and IL-6) (B), and measure levels of IL-1 signaling using a transgenic IL-1 receptor reporter specific for active IL-1α and IL-1β (C). Data are means ± standard deviations (SD) (n = 10 each) and are representative of at least 3 experiments. ND, none detected; *, P < 0.05; **, P < 0.005; ***, P < 0.0005; ns, not significant.

Article Snippet: Mice were euthanized after 24 or 72 h, and the nasopharynxes were subjected to lavage to quantify GAS CFU by dilution plating (A), quantify cytokines with a Meso Scale Diagnostics multiplex ELISA (means are displayed; comparisons indicated reach significance only for IL-1β and IL-6) (B), and measure levels of IL-1 signaling using a transgenic IL-1 receptor reporter specific for active IL-1α and IL-1β (C).

Techniques: Infection, Control, Multiplex Assay, Enzyme-linked Immunosorbent Assay, Transgenic Assay

Journal: Infection and Immunity

Article Title: Group A Streptococcus Infection of the Nasopharynx Requires Proinflammatory Signaling through the Interleukin-1 Receptor

doi: 10.1128/IAI.00356-20

Figure Lengend Snippet: SpeB and caspase-1 contribute to IL-1β generation in the nasopharynx. Effects of IL-1 and inflammasome signaling on GAS survival. (A) C57BL/6, casp-1/11−/−, or IL1R1−/− mice were inoculated intranasally with 108 CFU of wild-type GAS M1T1 5448. Mice were euthanized after 24 h, nasopharyngeal lavage fluids were plated to enumerate CFU, and cytokine levels were quantified by ELISA and IL-1R reporter assay. (B and C) Role of SpeB and CovRS in GAS survival. C57BL/6 mice were given anakinra (50 μg/kg) or PBS and infected as described above with wild-type, ΔspeB, or AP covS-frameshift GAS, and CFU and cytokines were examined as described above. (D) Role of host pathways in selection for covS-frameshift SpeB− clones. Isolated colonies from the experiments for panels A, B, and C were screened for SpeB hydrolysis of azocasein, and the fraction where SpeB activity is lost is indicated. ND, none detected (for experiments where no GAS was recoverable from the mouse). Data are means ± SD (n = 5 each) and are representative of at least 3 experiments. *, P < 0.05; **, P < 0.005; ***, P < 0.0005; ns, not significant.

Article Snippet: Mice were euthanized after 24 or 72 h, and the nasopharynxes were subjected to lavage to quantify GAS CFU by dilution plating (A), quantify cytokines with a Meso Scale Diagnostics multiplex ELISA (means are displayed; comparisons indicated reach significance only for IL-1β and IL-6) (B), and measure levels of IL-1 signaling using a transgenic IL-1 receptor reporter specific for active IL-1α and IL-1β (C).

Techniques: Enzyme-linked Immunosorbent Assay, Reporter Assay, Infection, Selection, Clone Assay, Isolation, Activity Assay

Journal: Infection and Immunity

Article Title: Group A Streptococcus Infection of the Nasopharynx Requires Proinflammatory Signaling through the Interleukin-1 Receptor

doi: 10.1128/IAI.00356-20

Figure Lengend Snippet: IL-1 mediates microbial interference in the nasopharynx. (A) SpeB-dependent effects on IL-1β and growth. C57BL/6 mice were inoculated intranasally with 108 CFU of GAS, L. lactis, or SpeB-expressing L. lactis. IL-1β was quantified by ELISA and CFU by dilution plating from nasopharyngeal lavage samples collected after 24 h infection. (B) IL-1β-dependent effects on L. lactis growth. C57BL/6 mice treated with anakinra (50 μg/kg) or PBS control were infected and CFU enumerated after 24 h as described above. (C) In vivo competition experiment with GAS and L. lactis. C57BL/6 mice were inoculated with 108 CFU GAS, L. lactis, or a mix of both (5 × 107 each; 1:1), and CFU were enumerated after 24 h as described above. (D) In vitro competition experiment with GAS and L. lactis. GAS (104 CFU), L. lactis (104 CFU), or a mix of both (5 × 103 CFU each; 1:1) were grown 18 h in 3 ml Todd-Hewitt broth at 37°C in 5% CO2; then, the CFU of each were enumerated by dilution and differential plating. Data are means ± SD (n = 5 each) and are representative of at least 3 experiments. ND, none detected; *, P < 0.05; **, P < 0.005; ***, P < 0.0005; ns, not significant.

Article Snippet: Mice were euthanized after 24 or 72 h, and the nasopharynxes were subjected to lavage to quantify GAS CFU by dilution plating (A), quantify cytokines with a Meso Scale Diagnostics multiplex ELISA (means are displayed; comparisons indicated reach significance only for IL-1β and IL-6) (B), and measure levels of IL-1 signaling using a transgenic IL-1 receptor reporter specific for active IL-1α and IL-1β (C).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Infection, Control, In Vivo, In Vitro

Journal: Frontiers in Immunology

Article Title: Simian Immunodeficiency Virus Infection Mediated Changes in Jejunum and Peripheral SARS-CoV-2 Receptor ACE2 and Associated Proteins or Genes in Rhesus Macaques

doi: 10.3389/fimmu.2022.835686

Figure Lengend Snippet: Dynamics of different renin angiotensin system proteins, lactate dehydrogenase, and cytokine production in plasma/serum. (A) Plasma concentration of ACE2 (ng/mL) at different time points of infection. The plasma ACE2 gradually decreased from acute to chronic infection with the lowest concentration at 90 dpi, then recovered during the late chronic stage of infection. (B) Plasma concentration of Angiotensin II (Ang II, pg/mL) at different time points. Though not significant, post infection showed a slight increase of Ang II compared to the pre infection time point, except at 112 and 145 dpi. (C) Plasma concentration of angiotensin II receptor 1 (AGTR1) (ng/mL) during infection. A gradual decrease of plasma AGTR1 was detected from 14 dpi onward. (D) Serum LDH activity (U/L) in infected RMs and uninfected controls were evaluated using a Beckman Coulter AU 480 analyzer. No significant changes in serum LDH activity were detected across the different time points. Plasma IL-6 (E) , IL-1β (F) , and TNF-α (G) concentrations (pg/mL) at pre and post infection time points were evaluated by U-plex biomarker NHP multiplex assay. No significant changes were detected at any time points for any of the proinflammatory cytokines tested. The error bars represent the mean ± SE for each time point (n=10). Each symbol represents individual macaque in each plot. Asterisks indicate statistical differences between time points, as calculated by Bonferroni for ACE2 and Ang II and Tukey-Kramer for AGTR1 (* p < 0.05; ** p < 0.01; *** p < 0.001 and **** p < 0.0001).

Article Snippet: Inflammatory cytokines (IL-1β, IL-6, and TNF-α) and MCP-1 chemokine (monocyte chemoattractant protein-1) in plasma were quantified using a U-plex biomarker NHP multiplex assay (Meso Scale Diagnostics, USA) following manufacturer instruction with minor modification.

Techniques: Clinical Proteomics, Concentration Assay, Infection, Activity Assay, Biomarker Discovery, Multiplex Assay