Journal: medRxiv

Article Title: Anomalous epithelial variations and ectopic inflammatory response in chronic obstructive pulmonary disease

doi: 10.1101/2020.12.03.20242412

Figure Lengend Snippet: A. Representative images immunostained with CCL2, TNC, and CXCL8 with AT2 cell markers (SFTPC or ABCA3) in COPD and never-smoker lung sections. B . Violin plots for COL4A1, COL4A2, and COL4A3 in AT1 clusters. C. Violin plots for FKBP5 in selected cell types. a) present dataset; b) combined publicly-available datasets.

Article Snippet: Antibodies and reagents were as follows: anti-PD-L1 (rabbit, 1:1000, #ab205921, Abcam, Cambridge, UK); anti-TNC (rabbit, 1:100, #HPA004823, Sigma-Aldrich); anti-CCL2 (rabbit, 1:100, #HPA019163, Sigma-Aldrich); anti-RAGE (rabbit, 1:1000, #ab216329, Abcam); anti-SFTPC (rabbit, 1:1000, #HPA010928, Sigma-Aldrich); anti-ABCA3 (mouse, 1:1000, #WMAB-ABCA3-17, Seven Hills Bioreagents, OH, USA); anti- macrophage inflammatory protein 3 alpha (rabbit, 1:1000, #ab224188, Abcam); anti- CXCL1 (mouse, 1:100, #MAB275, R&D Systems, MN, USA); anti-CXCL8 (mouse, 1:100, #MAB208, R&D Systems); and Hoechst 33242 (1:1000, #H342, Sigma-Aldrich).

Techniques:

Journal: medRxiv

Article Title: Anomalous epithelial variations and ectopic inflammatory response in chronic obstructive pulmonary disease

doi: 10.1101/2020.12.03.20242412

Figure Lengend Snippet: A. 100% stacked bar charts of the percentage of cell populations for AT1 subtype clusters, AT2 subtype clusters, and basal lineage clusters. B. Bar charts displaying the percentages of subpopulations such as AT1-B, AT2-A, AT2-B, club, goblet, and basal clusters across patient states. C. a UMAP plot of epithelial cells focusing on the AT2-C cluster (left). The cell population of the AT2-C cluster (right). The y-axis represents the ratio of cells in the AT2-C cluster across patient states. Brackets () represent the percentage of cells of the AT2-C cluster based on epithelial cells in each of the patient states. D. Violin plots of representative inflammatory-related genes displaying the AT2-C (iAT2) cluster. E. Violin plots for CXCL1 and CXCL8 in AT2 cells in combined publicly-available datasets. F. Representative images immunostained with CD274 (PD-L1), CXCL1 and CXCL20 with AT2 cell markers (SFTPC or ABCA3) in COPD and never-smoker lung sections. Scale bars, 25 μm (left), 12.5 μm (right).

Article Snippet: Antibodies and reagents were as follows: anti-PD-L1 (rabbit, 1:1000, #ab205921, Abcam, Cambridge, UK); anti-TNC (rabbit, 1:100, #HPA004823, Sigma-Aldrich); anti-CCL2 (rabbit, 1:100, #HPA019163, Sigma-Aldrich); anti-RAGE (rabbit, 1:1000, #ab216329, Abcam); anti-SFTPC (rabbit, 1:1000, #HPA010928, Sigma-Aldrich); anti-ABCA3 (mouse, 1:1000, #WMAB-ABCA3-17, Seven Hills Bioreagents, OH, USA); anti- macrophage inflammatory protein 3 alpha (rabbit, 1:1000, #ab224188, Abcam); anti- CXCL1 (mouse, 1:100, #MAB275, R&D Systems, MN, USA); anti-CXCL8 (mouse, 1:100, #MAB208, R&D Systems); and Hoechst 33242 (1:1000, #H342, Sigma-Aldrich).

Techniques:

Journal: Frontiers in Immunology

Article Title: Serum Amyloid A1 (SAA1) Revisited: Restricted Leukocyte-Activating Properties of Homogeneous SAA1

doi: 10.3389/fimmu.2020.00843

Figure Lengend Snippet: hrSAA1 does not induce chemokine expression in CD14 + monocytes and treatment of rSAA1 with LPL diminishes activity. (A,C) Freshly isolated monocytes were induced with homogenous rSAA1 (hrSAA1) (1–12000 ng/ml, 0.08–1000 nM) in parallel with rSAA1 (1–12000 ng/ml, 0.08–1000 nM). (B,D) rSAA1 (100 ng/ml, 8 nM) was pre-incubated with LPL (2000 ng/ml) for 4 h prior to stimulation of monocytes. Following an incubation period of 24 h, cell supernatants were collected and CXCL8 and CCL3 expression was determined via specific ELISAs. Expression in unstimulated cells is indicated by a dashed line (—). Results are represented as the mean chemokine concentration + SEM and are derived from four to six independent experiments. Significant upregulation in comparison to control is indicated by asterisks (Mann-Whitney U -test; * p -value < 0.05, ** p -value < 0.01). Significant reduction in inductive capacity following LPL treatment is indicated by a dagger (Mann-Whitney U -test; † p -value < 0.05).

Article Snippet: The human CXCL8 ELISA was developed in our laboratory using monoclonal mouse anti-human CXCL8 (MAB208) and polyclonal goat anti-human CXCL8 (BAF208) antibodies from R&D Systems ( ).

Techniques: Expressing, Activity Assay, Isolation, Incubation, Concentration Assay, Derivative Assay, Comparison, Control, MANN-WHITNEY

Journal: Frontiers in Immunology

Article Title: Serum Amyloid A1 (SAA1) Revisited: Restricted Leukocyte-Activating Properties of Homogeneous SAA1

doi: 10.3389/fimmu.2020.00843

Figure Lengend Snippet: hrSAA1 retains the capacity to synergize with CXCL8 in the in vitro attraction and activation of neutrophils. (A) Homogenous rSAA1 (hrSAA1, 300 or 3000 ng/ml, 25 or 250 nM) and CXCL8 (1–10 ng/ml, 0.12–1.2 nM) were added to the lower compartment of a Boyden microchamber either alone or in combination. Freshly isolated neutrophils were added to the upper compartment and allowed to migrate for 45 min. The chemotactic potencies are expressed as mean chemotactic index (+SEM) derived from six independent experiments. Control recruitment is indicated by a dashed line (—). (B) MMK-1 (16 μg/ml, 9930 nM), hrSAA1 (3000 ng/ml, 250 nM), CXCL8 (3 or 9 ng/ml, 0.36 or 1.07 nM) or a combination of hrSAA1 and CXCL8 were added to the lower compartment of a Boyden microchamber. Freshly isolated neutrophils in the presence or absence of the FPR2 antagonist WRW 4 (20 μg/ml, 18110 nM) were added to the upper compartment and allowed to migrate for 45 min. The chemotactic potencies are expressed as mean chemotactic index + SEM and are derived from three independent experiments. Control recruitment is indicated by a dashed line (—). (C) Neutrophils were stimulated (3 min) with homogenous hrSAA1 (300 or 3000 ng/ml, 25 or 250 nM), CXCL8 (1 or 3 ng/ml, 0.12 or 0.36 nM) or a combination of hrSAA1 and CXCL8. Data from six independent experiments are expressed as the net percentage of activated cells (blebbed and elongated cells) (+SEM). (D) Freshly isolated neutrophils were stimulated for 30 s with different concentrations of hrSAA1, CXCL8 or a combination of hrSAA1 and CXCL8. Following stimulation, cells were stained with Alexa Fluor 555 Phalloidin. Control actin polymerization is indicated by a dashed line (—). Results are presented as the mean percent florescence intensity relative to buffer-stimulated cells (+SEM) and are derived from six independent experiments. Significant neutrophil recruitment/activation in comparison to control is indicated by asterisks (Mann-Whitney U -test; * p -value < 0.05, ** p -value < 0.01). Significant synergy between hrSAA1 and CXCL8 in neutrophil activation/recruitment is indicated by daggers (Mann-Whitney U -test; † p -value < 0.05, †† p -value < 0.01).

Article Snippet: The human CXCL8 ELISA was developed in our laboratory using monoclonal mouse anti-human CXCL8 (MAB208) and polyclonal goat anti-human CXCL8 (BAF208) antibodies from R&D Systems ( ).

Techniques: In Vitro, Activation Assay, Isolation, Derivative Assay, Control, Staining, Comparison, MANN-WHITNEY, Significance Assay

Journal: Frontiers in Immunology

Article Title: Serum Amyloid A1 (SAA1) Revisited: Restricted Leukocyte-Activating Properties of Homogeneous SAA1

doi: 10.3389/fimmu.2020.00843

Figure Lengend Snippet: Comparison of the biological activities of E. coli -expressed SAA1 before (SAA1) and after (hrSAA1) purification to homogeneity using RP-HPLC.

Article Snippet: The human CXCL8 ELISA was developed in our laboratory using monoclonal mouse anti-human CXCL8 (MAB208) and polyclonal goat anti-human CXCL8 (BAF208) antibodies from R&D Systems ( ).

Techniques: Comparison, Purification, In Vivo, Migration, Expressing

Journal: BMC Microbiology

Article Title: Induction of a chemoattractant transcriptional response by a Campylobacter jejuni boiled cell extract in colonocytes

doi: 10.1186/1471-2180-9-28

Figure Lengend Snippet: Up-regulated genes. Functional classes of genes shown are ordered by the S score of the most highly regulated examples in the class with S score ≥ 5.

Article Snippet: IL8 and CCL20 (MIP-3α) were specifically measured using a sandwich ELISA, by capture with a murine anti-human IL8 or CCL20 and detected using biotinylated goat anti-human IL8 using streptavidin-coupled horseradish-peroxidase, according to the manufacturer's instructions (R&D Systems, Minneapolis, MN, USA).

Techniques: Functional Assay, Ubiquitin Proteomics, Binding Assay, Immunopeptidomics

Journal: BMC Microbiology

Article Title: Induction of a chemoattractant transcriptional response by a Campylobacter jejuni boiled cell extract in colonocytes

doi: 10.1186/1471-2180-9-28

Figure Lengend Snippet: Comparison of results for selected up-regulated genes determined by Affymetrix/S score and RQ-PCR.

Article Snippet: IL8 and CCL20 (MIP-3α) were specifically measured using a sandwich ELISA, by capture with a murine anti-human IL8 or CCL20 and detected using biotinylated goat anti-human IL8 using streptavidin-coupled horseradish-peroxidase, according to the manufacturer's instructions (R&D Systems, Minneapolis, MN, USA).

Techniques: Comparison, Clinical Proteomics, Membrane

Journal: BMC Microbiology

Article Title: Induction of a chemoattractant transcriptional response by a Campylobacter jejuni boiled cell extract in colonocytes

doi: 10.1186/1471-2180-9-28

Figure Lengend Snippet: Cytokine and chemokine levels (pg/ml) pre- and post-induction of HCA-7 cells with C. jejuni BCE for 6 h.

Article Snippet: IL8 and CCL20 (MIP-3α) were specifically measured using a sandwich ELISA, by capture with a murine anti-human IL8 or CCL20 and detected using biotinylated goat anti-human IL8 using streptavidin-coupled horseradish-peroxidase, according to the manufacturer's instructions (R&D Systems, Minneapolis, MN, USA).

Techniques:

Journal: Journal of Clinical Investigation

Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

doi: 10.1172/jci124694

Figure Lengend Snippet: Figure 2. Anti-Nfasc155 autoantibodies target surface Schwann cell antigens. (A) Sciatic nerve fibers were incubated in vitro with purified anti-Nfasc155 IgG4 from patient CIDP1 for 3 hours, and immunolabeled for IgG4 (green) and CNTN1 (red). (B–E) Sciatic nerves were fixed 1 day (B and D) or 3 days (C and E) after intraneural injections of anti-Nfasc155 IgG4, and immunolabeled for IgG4 (green) and β-catenin (red; B and C) or CNTN1 (red; D and E) and Nav channels (blue; D and E). Note that anti-Nfasc155 IgG4 bound to the surface of the Schwann cells and deposited at the vicinity of the node of Ranvier (dou- ble arrowheads) and at adherens junctions along the internode stained here with β-catenin (arrows). However, no penetration across the paranodal region was observed (images are representative of n = 3 independent experiments). Scale bars: 10 μm.

Article Snippet: Teased fibers were permeabilized by immersion in –20°C acetone for 10 minutes, blocked at room temperature for 1 hour with PBS containing 5% fish skin gelatin and 0.1% Triton X-100, then incubated overnight at 4°C with primary antibodies: rabbit antisera against Nfasc186 (1:500; gift of Alex Gow, Wayne State University, Detroit, Michigan, USA) (51), ankyrin-G (1:500; gift of Gisèle Alcaraz, GMGF, INSERM, UMR-S910, Marseille, France) (52), or CASPR1 (1:2000); mouse monoclonal antibodies against Nav channels (1:500; K58/35, Sigma-Aldrich) or β-catenin (MAB2081, Merck Millipore); and goat antibody against CNTN1 (1:2000; R&D Systems).

Techniques: Incubation, In Vitro, Purification, Immunolabeling, Staining

Journal: PLoS ONE

Article Title: ω-3 PUFA Rich Camelina Oil By-Products Improve the Systemic Metabolism and Spleen Cell Functions in Fattening Pigs

doi: 10.1371/journal.pone.0110186

Figure Lengend Snippet: Cytokine concentrations * in plasma and spleen of pigs fed T1 diet (sunflower meal) or T2 diet (camelina oil cakes).

Article Snippet: Purified fractions of anti-swine cytokines IL-8 (MAB5351), TNF-α (MAB6902), IL-1β (MAB6811), IL-6 (MAB686), IL-4 (ASC0944) and IFN-γ (ASC4934) (R&D Systems, Minneapolis, USA and Biosource International, Inc., Camarillo, USA) were used as capture antibody, in conjunction with biotinylated anti-swine cytokines IL-8 (BAF535), TNF-α (BAF690), IL-1β (BAF681), IL-6 (BAF686), IL-4 (ASC0849), IFN-γ (ASC4839).

Techniques: Clinical Proteomics