Journal: Stroke

Article Title: Role of SCH79797 in Maintaining Vascular Integrity in Rat Model of Subarachnoid Hemorrhage

doi: 10.1161/strokeaha.113.678474

Figure Lengend Snippet: Figure 1. Experimental designs. A, Experiment 1 was designed to investigate the effect of the protease-activated receptor-1 (PAR-1) agonist, SFLLRN. B, Experiment 2 was designed to evalu- ate the dose-dependent effect of SCH79797. C, Experiment 3 was designed to show the effects of SCH79797 treatment. D, Experiment 4 was designed to explore the mechanism of SCH79797 treatment. DMSO indicates dimethyl sulfoxide; IF, immunofluorescence staining; IP, intraperitoneally injection; IV, intravenously injection; SAH, sub- arachnoid hemorrhage; and WB, Western blot.

Article Snippet: The roles of a PAR-1–specific agonist, SFLLRN (Tocris, Ellisville, MO; 1 mg/kg as reported by the others),15,16 and a thrombin-specific inhibitor, argatroban (Sigma-Aldrich, St. Louis, MO), were evaluated (Figure 1A).

Techniques: Immunofluorescence, Staining, Injection, Western Blot

Journal: Stroke

Article Title: Role of SCH79797 in Maintaining Vascular Integrity in Rat Model of Subarachnoid Hemorrhage

doi: 10.1161/strokeaha.113.678474

Figure Lengend Snippet: Figure 2. The effects of the protease-activated receptor-1 (PAR-1) agonist, SFLLRN. There were 2 peak levels of PAR-1 expression at 2 to 6 hours and 24 hours after subarachnoid hemorrhage (SAH; A, n=6 each group). SFLLRN neutralized the protective effects of the thrombin inhibitor, argatroban, on attenuating brain edema (B) and neurological deficits (C and D) at 24 hours after SAH. B to D, n=5, 7, 7, 8 in sham, SAH+normal saline (NS), SAH+argatroban (ARG), and SAH+ ARG+SFLLRN groups. *P<0.05, **P<0.01 compared with the sham group; #P<0.05, ##P<0.01 compared with the SAH+NS group; §P<0.05, §§P<0.01 compared with the SAH+ARG group. The West- ern blot results were representative of 3 independent experiments. BS indicates brain stem; C, cerebellum; LH, left hemisphere; and RH, right hemisphere.

Article Snippet: The roles of a PAR-1–specific agonist, SFLLRN (Tocris, Ellisville, MO; 1 mg/kg as reported by the others),15,16 and a thrombin-specific inhibitor, argatroban (Sigma-Aldrich, St. Louis, MO), were evaluated (Figure 1A).

Techniques: Expressing, Saline

Journal: Scientific reports

Article Title: Disordered metabolism in mice lacking irisin.

doi: 10.1038/s41598-020-74588-7

Figure Lengend Snippet: Figure 6. Increased bone resorption in irisin lacking mice. (A, B) Sections of the distal shaft of the femur were stained with osteoprotegerin (OPG) (1:60) and receptor activator of nuclear factor-kB ligand (RANKL) (1:80) antibodies. Positive (black arrows) osteoblasts are shown, 200×. (C) Representative images of the distal metaphyseal region of the femur, together with cell counts (osteoclasts) per bone perimeter (Bpm). Red arrows, tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts, 100×, (NIS-Elements Viewer, v4.2.0; https:// www.downza.cn/soft/2751-21.html); (D) serum levels of osteocalcin TRAP (with three replicates). (Graphpad Prism, v7.0, https://www.xue51.com/soft/3932.html). Data are presented as the mean ± SEM (n = 15 per group). *P < 0.05, **P < 0.01 compared to the WT group.

Article Snippet: The enzyme-linked immunosorbent assay (ELISA) kit for osteocalcin (OCN) (E06917m), Alkaline Phosphatase (ALP) (E0200m), and Tartrate-resistant acid phosphatase (TRAP) (E08492m) were purchased from Elabscience (Wuhan, China).

Techniques: Staining

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Tereticornate A suppresses RANKL-induced osteoclastogenesis via the downregulation of c-Src and TRAF6 and the inhibition of RANK signaling pathways.

doi: 10.1016/j.biopha.2022.113140

Figure Lengend Snippet: Fig. 3. TA inhibits osteoclast-specific protein and gene expression during osteoclastogenesis. (A,B) Representative immunoblots for the effects of TA on RANKL- induced expression of TRAP (A) and cathepsin K (B) proteins. The gray densities of the bands for TRAP and cathepsin K were quantified using the AlphaView software. (C–F) TA inhibited RANKL-induced mRNA expression of β-Integrin (C), MMP-9 (D), ATP6V0D2 (E), and DC-STAMP (F). Total proteins or RNA from RAW 264.7 cells pretreated with or without TA for 15 min, then stimulated with RANKL (50 ng/mL) for 24 h, were subjected to western blotting or qRT-PCR analysis. Data are presented as the mean ± SEM from three independent biological experiments. ***p < 0.001 compared to the control group; #p < 0.05, ##p < 0.01, and ###p <

Article Snippet: Primary antibodies against NFATc1, c-Fos, TRAP, cathepsin K, TRAF6, and c-Src were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Gene Expression, Western Blot, Expressing, Software, Quantitative RT-PCR, Control

Journal: Immunology

Article Title: Anti‐CD52 antibody treatment in murine experimental autoimmune encephalomyelitis induces dynamic and differential modulation of innate immune cells in peripheral immune and central nervous systems

doi: 10.1111/imm.13437

Figure Lengend Snippet: Effect of anti‐CD52‐Ab treatment on the phenotype of periphery innate immune cells in EAE mice at first day post‐injection. Single‐cell suspensions were obtained from blood and spleen tissue of anti‐CD52‐Ab or vehicle‐treated EAE mice at first day after the final dose injection. Cells were stained for various surface markers and analysed by flow cytometry. (A) Flow cytometry gating strategy. (B) Blood and spleen CD11c + cells were analysed for expression of MHC‐II ( n = 9), CD40 ( n = 5), CD80 ( n = 5) and CD86 ( n = 9 blood; n = 5 spleen). (C) Blood and spleen CD11b + cells were analysed for expression of MHC‐II ( n = 8), CD40 ( n = 5), CD80 ( n = 5), CD86 ( n = 5), CD38 ( n = 5) and CD206 ( n = 9 blood; n = 7 spleen). Unpaired two‐tailed Student's t test. * p < 0·05, ** p < 0·01, *** p < 0·001

Article Snippet: To block non‐specific Fc receptors, cells were resuspended in anti‐mouse CD16/CD32 Fc block (eBioscience) for 10 min before incubation with the appropriate antibodies against: CD11b (#130‐113‐236), CD11c (#130‐102‐545), CD38 (#130‐109‐257), CD40 (#130‐105‐376 or #130‐102‐533), CD80 (#130‐116‐465), CD86 (#130‐105‐137), MHC‐II (#130‐107‐942), all were purchased from MACS Miltenyi Biotec; and CD45 (#11‐0451‐82) and CD206 (#12‐2061‐82), all were purchased from Thermo Fisher Scientific.

Techniques: Injection, Staining, Flow Cytometry, Expressing, Two Tailed Test

Journal: Immunology

Article Title: Anti‐CD52 antibody treatment in murine experimental autoimmune encephalomyelitis induces dynamic and differential modulation of innate immune cells in peripheral immune and central nervous systems

doi: 10.1111/imm.13437

Figure Lengend Snippet: Effect of anti‐CD52‐Ab treatment on the phenotype and function of CNS innate immune cells in EAE mice at first day post‐injection. Single‐cell suspensions were obtained from CNS (brain and spinal cord) tissue of anti‐CD52‐Ab or vehicle‐treated EAE mice at first day after the final treatment dose. Cells were stained for various surface markers and analysed by flow cytometry. (A) Flow cytometry gating strategy. (B) CD11b + CD45 lo/neg resident microglia and (C) CNS CD11b + CD45 hi infiltrating macrophages and were analysed for expression of MHC‐II ( n = 10), CD40 ( n = 5), CD80 ( n = 5), CD86 ( n = 5 microglia; n = 9 macrophages) and CD206 ( n = 7 microglia; n = 5 macrophages). Unpaired two‐tailed Student's t test. CD11b + cells were isolated from CNS tissue of anti‐CD52‐Ab or vehicle‐treated mice at first day after the final treatment dose. (D) Respiratory burst activity of CD11b + ‐isolated cells. (E) Phagocytosis of FITC‐coated beads by CD11b + isolated cells. Unpaired two‐tailed Student's t test. (F) Cells were left unstimulated or were stimulated with LPS or Poly(I:C)(PIC) for 24 h and cytokine production determined by ELISA. Two‐way ANOVA with Sidak multiple comparisons test, * p < 0·05, ** p < 0·01, *** p < 0·001

Article Snippet: To block non‐specific Fc receptors, cells were resuspended in anti‐mouse CD16/CD32 Fc block (eBioscience) for 10 min before incubation with the appropriate antibodies against: CD11b (#130‐113‐236), CD11c (#130‐102‐545), CD38 (#130‐109‐257), CD40 (#130‐105‐376 or #130‐102‐533), CD80 (#130‐116‐465), CD86 (#130‐105‐137), MHC‐II (#130‐107‐942), all were purchased from MACS Miltenyi Biotec; and CD45 (#11‐0451‐82) and CD206 (#12‐2061‐82), all were purchased from Thermo Fisher Scientific.

Techniques: Injection, Staining, Flow Cytometry, Expressing, Two Tailed Test, Isolation, Activity Assay, Enzyme-linked Immunosorbent Assay

Journal: Immunology

Article Title: Anti‐CD52 antibody treatment in murine experimental autoimmune encephalomyelitis induces dynamic and differential modulation of innate immune cells in peripheral immune and central nervous systems

doi: 10.1111/imm.13437

Figure Lengend Snippet: Effect of anti‐CD52‐Ab treatment on the phenotype of periphery and CNS innate immune cells in EAE mice at three weeks post‐injection. Single‐cell suspensions were obtained from blood, spleen and CNS (brain and spinal cord) tissue of anti‐CD52‐Ab or vehicle‐treated mice three weeks after the final treatment dose. Cells were stained for various surface markers and analysed by flow cytometry. (A) Blood and spleen CD11c + cells were analysed for expression of MHC‐II ( n = 9), CD40 ( n = 5), CD80 ( n = 5) and CD86 ( n = 9 blood; n = 5 spleen). (B) Blood and spleen CD11b + cells were analysed for expression of MHC‐II ( n = 8), CD40 ( n = 5), CD80 ( n = 5), CD86 ( n = 5), CD38 ( n = 5) and CD206 ( n = 9 blood; n = 7 spleen). (C) CNS CD11b + CD45 hi ‐infiltrating macrophages and CD11b + CD45 lo/neg resident microglia were analysed for expression of MHC‐II ( n = 10), CD40 ( n = 5), CD80 ( n = 5), CD86 ( n = 5 microglia; n = 9 macrophages) and CD206 ( n = 7 microglia; n = 5 macrophages). Unpaired two‐tailed Student's t test. * p < 0·05, ** p < 0·01, *** p < 0·001

Article Snippet: To block non‐specific Fc receptors, cells were resuspended in anti‐mouse CD16/CD32 Fc block (eBioscience) for 10 min before incubation with the appropriate antibodies against: CD11b (#130‐113‐236), CD11c (#130‐102‐545), CD38 (#130‐109‐257), CD40 (#130‐105‐376 or #130‐102‐533), CD80 (#130‐116‐465), CD86 (#130‐105‐137), MHC‐II (#130‐107‐942), all were purchased from MACS Miltenyi Biotec; and CD45 (#11‐0451‐82) and CD206 (#12‐2061‐82), all were purchased from Thermo Fisher Scientific.

Techniques: Injection, Staining, Flow Cytometry, Expressing, Two Tailed Test

Journal: Immunology

Article Title: Anti‐CD52 antibody treatment in murine experimental autoimmune encephalomyelitis induces dynamic and differential modulation of innate immune cells in peripheral immune and central nervous systems

doi: 10.1111/imm.13437

Figure Lengend Snippet: Effect of anti‐CD52‐Ab treatment on the function of periphery and CNS innate immune cells in SCID mice at first day post‐injection. SCID mice were treated with either vehicle or 10 mg/kg murine anti‐CD52‐Ab‐Ab for five consecutive days. Single‐cell suspensions were then obtained from blood and spleen tissues first day after the final treatment dose. Cells were stained for various surface markers and analysed by flow cytometry. (A) Blood and spleen CD11c + cells were analysed for expression of MHC‐II, CD40, CD80 and CD86 (all n = 7). (B) Blood and spleen CD11b + cells were analysed for expression of MHC‐II, CD40, CD80, CD86, and CD206 (all n = 7). Unpaired two‐tailed Student's t test. * p < 0·05, ** p < 0·01, *** p < 0·001

Article Snippet: To block non‐specific Fc receptors, cells were resuspended in anti‐mouse CD16/CD32 Fc block (eBioscience) for 10 min before incubation with the appropriate antibodies against: CD11b (#130‐113‐236), CD11c (#130‐102‐545), CD38 (#130‐109‐257), CD40 (#130‐105‐376 or #130‐102‐533), CD80 (#130‐116‐465), CD86 (#130‐105‐137), MHC‐II (#130‐107‐942), all were purchased from MACS Miltenyi Biotec; and CD45 (#11‐0451‐82) and CD206 (#12‐2061‐82), all were purchased from Thermo Fisher Scientific.

Techniques: Injection, Staining, Flow Cytometry, Expressing, Two Tailed Test