recombinant mouse rm cxcl9  (R&D Systems)

Journal: Scientific reports

Article Title: Inhibition of the CXCL9-CXCR3 axis suppresses the progression of experimental apical periodontitis by blocking macrophage migration and activation.

doi: 10.1038/s41598-021-82167-7

Figure Lengend Snippet: Figure 2. Inflammatory markers especially chemokines are highly expressed in periapical lesions. (a) Clustergram analysis of the gene expression patterns for inflammation-associated factors during the development of AP (n = 1 per group). (b) Validation of the inflammation-associated factors by quantitative PCR analysis. Normalization was based on the GAPDH expression in the without (W/O) exposure group. Statistical analysis was performed using the Dunnett test in comparison with the W/O group; *p < 0.05, **p < 0.01 (mean ± SD, n = 4 per group). (c) Clustergram analysis of inflammatory mediator and receptor genes identified during the development of AP (n = 1 per group). (d) Validation of the inflammatory markers by quantitative PCR analysis. Statistical analysis was performed using the Dunnett test in comparison with the W/O group; *p < 0.05, **p < 0.01 (mean ± SD, n = 4 per group). (e) Representative images of immunostained CXCL9- and Mac2-positive macrophages (arrows) at 28 days after pulp exposure. The lower panel shows a higher magnification image of the framed region in the upper panel.

Article Snippet: These cells were later treated with LPS (100 ng/mL), recombinant mouse (rm) CXCL9 (100 ng/mL, 492-MM; R&D Systems) or LPS (100 ng/mL) and rmCXCL9 (100 ng/mL) in the presence of 10% FBS for 24 h. After incubation, the cells were assayed by the gene expression analysis of Cxcl9, Cxcl10, Il1b, Il6, Mmp2, Tnf (primer lists: sTable 1), as described previously44.

Techniques: Gene Expression, Biomarker Discovery, Real-time Polymerase Chain Reaction, Expressing, Comparison

Journal: Scientific reports

Article Title: Inhibition of the CXCL9-CXCR3 axis suppresses the progression of experimental apical periodontitis by blocking macrophage migration and activation.

doi: 10.1038/s41598-021-82167-7

Figure Lengend Snippet: Figure 5. CXCL9 induces macrophage migration and enhances the inflammatory response to LPS. (a) Representative images of peritoneal macrophages that have moved through the membrane. Peritoneal macrophages were stained using a commercial solution in accordance with the manufacturer’s protocol. (b) Cell migration assay for 24 h was analyzed using the colorimetric absorbance at 560 nm (mean ± SD, n = 4 per each group). (c) Validation of the inflammatory markers by quantitative PCR analysis. Statistical analysis was conducted using the Dunnett test to compare with the PBS group; *p < 0.05, **p < 0.01 (mean ± SD, n = 4 per group).

Article Snippet: These cells were later treated with LPS (100 ng/mL), recombinant mouse (rm) CXCL9 (100 ng/mL, 492-MM; R&D Systems) or LPS (100 ng/mL) and rmCXCL9 (100 ng/mL) in the presence of 10% FBS for 24 h. After incubation, the cells were assayed by the gene expression analysis of Cxcl9, Cxcl10, Il1b, Il6, Mmp2, Tnf (primer lists: sTable 1), as described previously44.

Techniques: Migration, Membrane, Staining, Cell Migration Assay, Biomarker Discovery, Real-time Polymerase Chain Reaction

Journal: Scientific reports

Article Title: Inhibition of the CXCL9-CXCR3 axis suppresses the progression of experimental apical periodontitis by blocking macrophage migration and activation.

doi: 10.1038/s41598-021-82167-7

Figure Lengend Snippet: Figure 6. The CXCL9-CXCR3 axis is involved in the regulation of osteoclast activity but does not directly affect RANKL-induced osteoclast formation. (a) Representative immunostaining images of cathepsin K-positive inflammatory cells (arrows) at 28 days after pulp exposure in the AP mouse model. (b) TRAP staining around the periapical area at 28 days after pulp exposure. The arrow denotes TRAP-positive multinuclear cells. (c) Microscopic images of TRAP-positive cells. Osteoclast precursors were treated for 5 days with (i) Macrophage colony stimulating factor (M-CSF) alone; (ii) M-CSF and RANKL; (iii) M-CSF, RANKL and CXCL9; or (iv) M-CSF and CXCL9. (d) Numbers of TRAP-positive cells (cell/section). For quantification of TRAP positive cells, we selected 5 independent areas per one well and quantified using ImageJ software. Statistical analysis was conducted using he Turkey-Kramer test; *p < 0.05, **p < 0.01 (mean ± SD, 20 areas/n = 4 per group).

Article Snippet: These cells were later treated with LPS (100 ng/mL), recombinant mouse (rm) CXCL9 (100 ng/mL, 492-MM; R&D Systems) or LPS (100 ng/mL) and rmCXCL9 (100 ng/mL) in the presence of 10% FBS for 24 h. After incubation, the cells were assayed by the gene expression analysis of Cxcl9, Cxcl10, Il1b, Il6, Mmp2, Tnf (primer lists: sTable 1), as described previously44.

Techniques: Activity Assay, Immunostaining, Staining, Software

Journal: Scientific reports

Article Title: Inhibition of the CXCL9-CXCR3 axis suppresses the progression of experimental apical periodontitis by blocking macrophage migration and activation.

doi: 10.1038/s41598-021-82167-7

Figure Lengend Snippet: Figure 7. Model for the inflammatory mechanism mediated by the CXCL9,10—CXCR3 axis during AP progression. The production of CXCL9 and10 and their receptor CXCR3 is induced by bacterial infection in the root canal. These molecules induce chemotaxis, promote the differentiation of macrophages, and produce osteoclastogenic cytokines to promote bone destruction by activating osteoclasts during the development of AP. A CXCR3 blockade suppresses the tissue destruction caused by an AP lesion through the inhibition of macrophage activation and migration.

Article Snippet: These cells were later treated with LPS (100 ng/mL), recombinant mouse (rm) CXCL9 (100 ng/mL, 492-MM; R&D Systems) or LPS (100 ng/mL) and rmCXCL9 (100 ng/mL) in the presence of 10% FBS for 24 h. After incubation, the cells were assayed by the gene expression analysis of Cxcl9, Cxcl10, Il1b, Il6, Mmp2, Tnf (primer lists: sTable 1), as described previously44.

Techniques: Infection, Chemotaxis Assay, Inhibition, Activation Assay, Migration