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

Article Title: N-?-Benzoyl-N5-(2-Chloro-1-Iminoethyl)-L-Ornithine Amide, a Protein Arginine Deiminase Inhibitor, Reduces the Severity of Murine Collagen-Induced Arthritis

doi: 10.4049/jimmunol.1001620

Figure Lengend Snippet: Cl-amidine treatment reduces clinical disease activity and joint destruction in CIA. Arthritis was induced by two injections of bovine CII in CFA at days 0 and 21. Mice were treated as described in Materials and Methods and assessed three times weekly for clinical disease activity. A, All treatment groups showed significantly reduced clinical disease activity scores compared with the PBS vehicle control beginning on day 27 and continuing through day 35. *p < 0.05 at day 24 and p < 0.03 at day 27 for 10-mg group, #p < 0.01 for 50-mg group, ***p < 0.03 for 10- and 50-mg groups, and p < 0.05 for 1-mg group. (n = 8 for all groups, except n = 7 for 1-mg group.) B, Histologic sections were scored for changes in synovial inflammation, pannus, cartilage damage, and bone damage, all on a scale of 0–5. C, Sections were scored for C3 deposition in the synovium and cartilage. Data are expressed as the mean ± SEM based upon a set of five joints per animal. Treatment with Cl-amidine resulted in decreased histopathology scores for all parameters in the 10- and 50-mg/kg/d treatment groups compared with the PBS vehicle group. (n = 8 for all groups, except n = 7 for 1-mg group.) *p ≤ 0.03, **p < 0.02, ***p < 0.01.

Article Snippet: IgG2a and IgG1 Abs to murine and bovine CII were measured in triplicate by specific ELISA by coating 96-well plates with 5 μg/ml murine or bovine CII (Chondrex).

Techniques: Activity Assay, Histopathology

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

Article Title: N-?-Benzoyl-N5-(2-Chloro-1-Iminoethyl)-L-Ornithine Amide, a Protein Arginine Deiminase Inhibitor, Reduces the Severity of Murine Collagen-Induced Arthritis

doi: 10.4049/jimmunol.1001620

Figure Lengend Snippet: Cl-amidine treatment reduces serum and synovial citrulline content in vivo. A, Synovial citrulline content. Arthritis was induced in control and Cl-amidine–treated mice by a single injection of bovine CII in CFA at day 0. Mice were treated daily, beginning on day 0, for 21 d with Cl-amidine (1, 10, or 50 mg/kg) in PBS or PBS alone or were left untreated. Results are displayed by dose (left panel) and grouped (right panel) as unimmunized (WT), control (PBS and untreated), and Cl-amidine (1, 10, or 50 mg/kg)-treated mice. B, Serum citrulline content was measured in mice at day 35. Results are displayed by dose (left panel) and grouped (right panel) as control (untreated and PBS) and Cl-amidine (1, 10, or 50 mg/kg)-treated mice.

Article Snippet: IgG2a and IgG1 Abs to murine and bovine CII were measured in triplicate by specific ELISA by coating 96-well plates with 5 μg/ml murine or bovine CII (Chondrex).

Techniques: In Vivo, Injection

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

Article Title: N-?-Benzoyl-N5-(2-Chloro-1-Iminoethyl)-L-Ornithine Amide, a Protein Arginine Deiminase Inhibitor, Reduces the Severity of Murine Collagen-Induced Arthritis

doi: 10.4049/jimmunol.1001620

Figure Lengend Snippet: Cl-amidine treatment reduces IgG autoantibody response to mouse but not bovine CII. Sera from individual mice were incubated on a 96-well plate coated with bovine (A) or mouse (B) CII. The presence of anti-mouse IgG1 or IgG2a Ab was detected by ELISA. The 50-mg/kg/d group showed significantly lower IgG1 anti-mouse CII Ab titers, whereas the 10-mg/kg/d group had significantly lower IgG2a anti-mouse CII Ab titers compared with the PBS vehicle control group at day 35. C, IgG1 and IgG2a Ab titers at day 35. *p ≤ 0.05. (n = 8 for all group, except n = 7 for 1-mg group.)

Article Snippet: IgG2a and IgG1 Abs to murine and bovine CII were measured in triplicate by specific ELISA by coating 96-well plates with 5 μg/ml murine or bovine CII (Chondrex).

Techniques: Incubation, Enzyme-linked Immunosorbent Assay

Journal: Nature Communications

Article Title: IFN-β is a macrophage-derived effector cytokine facilitating the resolution of bacterial inflammation

doi: 10.1038/s41467-019-10903-9

Figure Lengend Snippet: Non-phagocytic resolution phase macrophages express IFN-β. a , b Male mice were injected intraperitoneally with zymosan A (1 mg/mouse) followed by an injection of PKH2-PCL at 62 h. After 4 h, the peritoneal cells were recovered and immuno-stained for F4/80 and CD11b. Then, F4/80 + macrophages were sorted based on the extent of PKH2-PCL acquisition (PKH2 + /− populations; >98% purity) using the FACSAria III sorter (illustrated in ( a ). The collected cells were immediately used for RNA extraction (with RNA integrity value above 7.5), and a gene expression microarray analysis was performed using Illumina hiSeq 2500. Differential gene expression analysis and gene ontology (GO) enrichment were performed for genes that were significantly upregulated ( b , left panel) or downregulated ( b , right panel) in non-phagocytic/satiated (PKH2-PCL lo ) macrophages in comparison to phagocytic (PKH2-PCL hi ) ones. The results indicate the statistical significance of the GO term and the percentage of enrichment is presented. c – e Expression of IFN-β and ISG15 in sorted satiated and phagocytic macrophages. Representative results ( c ) and mean ± SEM ( d , e ) for three independent experiments. * P < 0.05 (Student’s t test). g , h Peritoneal exudates were collected from unchallenged mice (0 h) or following peritonitis for 4–96 h. IFN-β content in cell-free fluids was determined by ELISA ( f ). Results are mean ± SEM from three (24, 72, 96 h) or four (0, 4, 48 h) mice. * P < 0.05, ** P < 0.01, *** P < 0.005 (Tukey’s HSD). Alternatively, resolution phase macrophages were recovered 66 h post peritonitis initiation (PPI) and incubated with TGF-β (5 ng/ml), poly (I:C) (4 μg/ml) or apoptotic cells (AC, at a ratio of 1:5) for 24 h. Culture supernatants were then collected and IFN-β content was measured ( g ). Culture media from apoptotic cells served as control. Results are representative from three independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.005 (Tukey’s HSD). Source data are provided as a Source Data file

Article Snippet: Peritoneal exudates and bronchoalveolar lavage fluid were collected at the indicated times and the levels of IFN-β in cell-free fluids were determined by VeriKine-HS Mouse Interferon Beta Serum ELISA Kit (Pestka Biomedical Laboratories, Inc).

Techniques: Injection, Staining, RNA Extraction, Gene Expression, Microarray, Comparison, Expressing, Enzyme-linked Immunosorbent Assay, Incubation, Control

Journal: Nature Communications

Article Title: IFN-β is a macrophage-derived effector cytokine facilitating the resolution of bacterial inflammation

doi: 10.1038/s41467-019-10903-9

Figure Lengend Snippet: IFN-β favors macrophage reprogramming during the resolution of inflammation. a – d Macrophages were recovered from peritoneal exudates of male Ifnb +/+ or Ifnb − / − mice at 48–66 h PPI and cultured with LPS (1 μg/ml) for 24 h. Culture supernatants were then collected and levels of IL-10 ( a ), IL-6 ( b ), IL-12 ( c ) and CCL3 ( d ) were determined by selective ELISAs. Results are means ± SEM from four independent experiments. * P < 0.05, *** P < 0.005 (Tukey’s HSD). e – f Macrophages were recovered from peritoneal exudates of Ifnb +/+ mice at 48–66 h PPI and cultured with mouse IFN-β or IFN-α (20 ng/ml each) for 48 h. Then, culture supernatants were collected and levels of IL-10 ( e ) and IL-12 ( f ) were determined by ELISA. Results are means ± SEM (n = 4). *** P < 0.005 (Tukey’s HSD). g Macrophages were recovered from peritoneal exudates of Ifnb +/+ mice 48–66 h PPI and incubated with IFN-β (20 ng/ml) for 48 h. The cells were then immunostained for F4/80 and CD11b and the percentage of CD11b low macrophages was determined by flow cytometry. Results are means ± SEM from three independent experiments. *** P < 0.005 (Tukey’s HSD). h – i Mice undergoing peritonitis were treated with IFN-β (20 ng/mouse, i.p.) or vehicle at 24 h PPI. Peritoneal macrophages were collected at 48 h PPI, lysed and immunoblotted for 12/15-LO, arginase 1, ISG15 and GAPDH. Representative blots ( h ) and densitometry analysis (means ± SEM) ( i ) for three independent experiments. * P < 0.05 (Student’s t test). Source data are provided as a Source Data file

Article Snippet: Peritoneal exudates and bronchoalveolar lavage fluid were collected at the indicated times and the levels of IFN-β in cell-free fluids were determined by VeriKine-HS Mouse Interferon Beta Serum ELISA Kit (Pestka Biomedical Laboratories, Inc).

Techniques: Cell Culture, Enzyme-linked Immunosorbent Assay, Incubation, Flow Cytometry