Journal: Arthritis Research & Therapy

Article Title: Expansion of myeloid-derived suppressor cells contributes to metabolic osteoarthritis through subchondral bone remodeling

doi: 10.1186/s13075-021-02663-z

Figure Lengend Snippet: High-fat diet increases monocytic-MDSC expansion during PTOA. A Gating strategy used to define the M-MDSC subpopulation (CD11b + Ly6C + Ly6G − cells) in mouse bone marrow. Following the initial FS/SC discrimination, the gate was set on CD11b + cells. After exclusion of doublets (not shown), live CD11b + cells were gated and Ly6C + and Ly6G + populations. B Cytospin images of M-MDSC cells from the bone marrow from LFD and HFD mice (at 16 weeks). Scale bar denotes 10 microns. C HFD increases the BM M-MDSC population in mice ( n = 4 in each group) by total number and percentage with concomitant suppression of CD8 + and CD4 + T-cells compared to LFD mice ( n = 6 in each group, at 16 weeks). D HFD increases the BM M-MDSC population in DMM mice ( n = 3 in each group). * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001 by two-way ANOVA Tukey’s multiple comparison test

Article Snippet: The following antibodies were added in staining buffer (eBioscience) for 30 min: CD11b-APC (clone M1/70, # 130-113-231, Miltenyi), Ly6G-PE (clone 1A8,130-102-392), and Ly6C-FITC (clone REA796, #130-111-915, Miltenyi).

Techniques: Comparison

Journal: Science Advances

Article Title: Elevated type I interferon responses potentiate metabolic dysfunction, inflammation, and accelerated aging in mtDNA mutator mice

doi: 10.1126/sciadv.abe7548

Figure Lengend Snippet: ( A and B ) Twelve-month-old WT ( n = 8) and POLG ( n = 7) mice were challenged with LPS [50 mg/kg by intraperitoneal (i.p.) injection]. Kaplan-Meyer survival analysis was performed (A). Plasma cytokine profiles were determined by multianalyte bead-based immunoassay on n ≥ 6 biological samples per group (B). Log-rank (Mantel-Cox) test was used to compare percent survival between different groups. ( C and D ) CD11b + Ly6C hi inflammatory monocyte population in whole blood from 12-month-old WT and POLG mice was evaluated by flow cytometry. Pseudo-color plots are representative of four independent experiments (C), and quantification of the percentage of CD11b + Ly6C hi cells is shown in (D). ( E and F ) CD11b + Ly6G + blood neutrophil population in 12-month-old WT and POLG mice was determined by flow cytometry. Pseudo-color plots are representative of four independent experiments (E), and quantification of the percentage of CD11b + Ly6G + cells is shown in (F). ( G to I ) CD11b + Ly6C hi TNFα + inflammatory monocyte population in unstimulated (C) or LPS-challenged (L) whole blood from 12-month-old WT and POLG mice was evaluated by flow cytometry. Histograms are representative of four independent experiments (G). Quantification of CD11b + Ly6C hi TNFα + mean fluorescent intensity (MFI) is shown in (H), and the percentage of CD11b + Ly6C hi TNFα + cells is shown in (I). ( J to L ) CD11b + Ly6G + TNFα + neutrophil population in unstimulated or LPS-challenged whole blood from 12-month-old WT and POLG mice was evaluated by flow cytometry. Histograms are representative of four independent experiments (J). Quantification of CD11b + Ly6G + TNFα + MFI is shown in (K), and the percentage of CD11b + Ly6G + TNFα + neutrophils is shown in (L). Unless stated, statistical significance was determined using unpaired Student’s t test after Shapiro-Wilk normality test. * P < 0.05, ** P < 0.01, and *** P < 0.001. Error bars represent SEM.

Article Snippet: Antibodies and reagents used for flow cytometry included the following: phycoerythrin/Cy7 anti-mouse TNFα antibody (506324), PerCP/Cy5.5 anti-mouse Ly6C antibody (128012), fluorescein isothiocyanate anti-mouse Ly6C antibody (128006), and PerCP/Cy5.5 anti-mouse IL-12/IL-23 p40 (505211) (from BioLegend); purified anti-mouse CD16 / CD32 (2.4G2) (70-0161), violetFluor 450 anti-mouse Ly6G (1A8) (75-1276), and allophycocyanin anti-human/mouse CD11b (M1/70) (20-0112) (from Tonbo); IL-6 (11-7061-81) (from Invitrogen); IL-1β (31202) (from Cell Signaling Technology); brefeldin A solution (420601) and monensin solution (420701) (from BioLegend); mouse TNFα (430904) and IL-6 (431304) enzyme-linked immunosorbent assay (ELISA) kits (from BioLegend); and Griess Reagent System (G2930) (from Promega).

Techniques: Injection, Clinical Proteomics, Bead-based Assay, Flow Cytometry

Journal: Science Advances

Article Title: Elevated type I interferon responses potentiate metabolic dysfunction, inflammation, and accelerated aging in mtDNA mutator mice

doi: 10.1126/sciadv.abe7548

Figure Lengend Snippet: ( A and B ) Twelve-month-old WT, POLG, POLG cGAS −/− , POLG Sting −/− , and POLG Ifnar −/− ( n = 5 to 8 per group) mice were intraperitoneally injected with LPS (50 mg/kg). Kaplan-Meyer survival analysis was performed (A). Plasma was collected at indicated time points ( n ≥ 6 at 2 hours and n = 4 at 6 hours) and subjected to multianalyte cytokine analysis (B). Statistical comparisons in (B) were made against LPS-injected POLG mice. Log-rank (Mantel-Cox) test was used to compare percent survival between different groups. ( C and D ) CD11b + Ly6C hi inflammatory monocyte population in whole blood from 12-month-old mice was evaluated by flow cytometry. Pseudo-color plots are representative of four independent experiments (C), and quantification of the percentage of CD11b + Ly6C hi cells is shown in (D). ( E and F ) CD11b + Ly6C hi TNFα + inflammatory monocyte population in unstimulated or LPS challenged whole blood from 12-month-old cohorts was evaluated by flow cytometry. Quantification of CD11b + Ly6C hi TNFα + MFI is shown in (E), and the percentage of CD11b + Ly6C hi TNFα + cells is shown in (F). Unless stated, statistical significance was determined using analysis of variance (ANOVA) and Tukey post hoc test. * P < 0.05, ** P < 0.01, and *** P < 0.001. Error bars represent SEM.

Article Snippet: Antibodies and reagents used for flow cytometry included the following: phycoerythrin/Cy7 anti-mouse TNFα antibody (506324), PerCP/Cy5.5 anti-mouse Ly6C antibody (128012), fluorescein isothiocyanate anti-mouse Ly6C antibody (128006), and PerCP/Cy5.5 anti-mouse IL-12/IL-23 p40 (505211) (from BioLegend); purified anti-mouse CD16 / CD32 (2.4G2) (70-0161), violetFluor 450 anti-mouse Ly6G (1A8) (75-1276), and allophycocyanin anti-human/mouse CD11b (M1/70) (20-0112) (from Tonbo); IL-6 (11-7061-81) (from Invitrogen); IL-1β (31202) (from Cell Signaling Technology); brefeldin A solution (420601) and monensin solution (420701) (from BioLegend); mouse TNFα (430904) and IL-6 (431304) enzyme-linked immunosorbent assay (ELISA) kits (from BioLegend); and Griess Reagent System (G2930) (from Promega).

Techniques: Injection, Clinical Proteomics, Flow Cytometry