Journal: International journal of immunopathology and pharmacology

Article Title: Administration of anti-GFAP antibodies increases CGRP expression and increases pain hypersensitivity in spinal cord injured animals.

doi: 10.1177/03946320251320754

Figure Lengend Snippet: Figure 2. Plasma levels of immunoreactivity to GFAP were significantly higher in the anti-GFAP treated group at day 14 and day 28 compared to IgG-treated and sham saline-treated rats. Those receiving the anti-GFAP showed a mean difference on day 14: 2.97, t = 4.58, p < 0.001; and mean difference on day 28: 1.81, t = 2.8, p = 0.007 compared to SCI rats receiving normal rat IgG(*). A standard curve was generated by serial dilution of purified recombinant GFAP protein for between- assay comparison, measurements are expressed as ng/ul of GFAP. GFAP: glial fibrillary acidic protein; IgG: immunoglobulin G.

Article Snippet: The specificity and sensitivity of antibodies was verified using immunohistochemistry compared to commercial controls, and using Western blot against both purified recombinant GFAP protein (OriGene, Rockville, MD) and brain tissue.

Techniques: Clinical Proteomics, Saline, Generated, Serial Dilution, Purification, Recombinant, Comparison

Journal: International journal of immunopathology and pharmacology

Article Title: Administration of anti-GFAP antibodies increases CGRP expression and increases pain hypersensitivity in spinal cord injured animals.

doi: 10.1177/03946320251320754

Figure Lengend Snippet: Figure 3. There was a significant difference in mean CGRP expression among the treatment groups (F(2,12) = 6.519, p = 0.012)(*). Anti-GFAP treated mean CGRP expression (M = 20.09, SD = 2.42) was significantly increased compared to both laminectomy-only (M = 13.71, SD = 2.555), p = 0.007(*), and IgG treated groups (M = 15.62, SD = 3.19), p = 0.017(*). No significant difference in CGRP expression was determined between the IgG-treated rats and laminectomy-only rats (p = 0.351). These results indicate that the anti-GFAP treatment increased expression of CGRP. CGRP: calcitonin gene-related peptide.

Article Snippet: The specificity and sensitivity of antibodies was verified using immunohistochemistry compared to commercial controls, and using Western blot against both purified recombinant GFAP protein (OriGene, Rockville, MD) and brain tissue.

Techniques: Expressing

Journal: International journal of immunopathology and pharmacology

Article Title: Administration of anti-GFAP antibodies increases CGRP expression and increases pain hypersensitivity in spinal cord injured animals.

doi: 10.1177/03946320251320754

Figure Lengend Snippet: Figure 4. (a) Von Frey assay testing indicated a lower paw withdrawal threshold in the injured groups compared to laminectomy control rats at 1- and 2-month time points. (b) MCAP revealed significantly reduced number of crosses in the anti-GFAP treated rats compared to laminectomy controls at 2 months post-injury (*) indicating avoidance of the crossing, suggesting hyperalgesia. This was not observed in the IgG- or saline-treated animals. GFAP: glial fibrillary acidic protein; IgG: immunoglobulin G; MCAP: mechanical conflict-avoidance paradigm.

Article Snippet: The specificity and sensitivity of antibodies was verified using immunohistochemistry compared to commercial controls, and using Western blot against both purified recombinant GFAP protein (OriGene, Rockville, MD) and brain tissue.

Techniques: Control, Saline

Journal: Cell reports

Article Title: Astrocytes Amplify Neuronal Dendritic Volume Transmission Stimulated by Norepinephrine

doi: 10.1016/j.celrep.2019.11.092

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: rabbit anti GFAP , Phosphosolution , Cat# 620-GFAP; RRID:AB_2492124.

Techniques: Virus, Plasmid Preparation, Recombinant, Software

Journal: Biomolecules & therapeutics

Article Title: PEGylated Erythropoietin Protects against Brain Injury in the MCAO-Induced Stroke Model by Blocking NF-κB Activation.

doi: 10.4062/biomolther.2019.147

Figure Lengend Snippet: Fig. 2. Effect of P-EPO on MDA and GSH/GSSG levels and expression of GFAP and Iba-1. Vehicle, 5000 U/kg of P-EPO, was adminis- tered via i.v. before MCAO. Ischemic brain tissues were processed for immunohistochemistry 24 h after reperfusion following MCAO. (A) MDA levels in the brain infarction site was measured using a TBARS assay kit. (B) GSH/GSSG levels in the brain infarction site were mea- sured using a GSH/GSSG ratio detection assay kit. Representative photomicrographs of (C) GFAP-positive astrocytes, (D) Iba-1-positive microglia/macrophages. Each value is presented as mean ± SD from five mice. **p<0.01 significant difference from MCAO mice. ##p<0.01, ###p<0.0001 significant difference from Sham mice (Sham n=6-7, MCAO n=8-9, M-EPO n=10-12).

Article Snippet: Membranes were incubated at room temperature for 2 h with the following specific antibodies: anti-COX-2, anti-IκB, anti-p-IκB, anti- STAT1, anti-p-STAT1, anti- STAT3, anti-pSTAT3, anti- STAT5, anti-p-STAT5 (Cell Signaling Technology, Inc., Beverly, MA, USA), anti-inducible nitric oxide synthase (iNOS) and anti-Glial fibrillary acidic protein (GFAP) (1:1,000 Novus Biologicals, Inc., Littleton, CO, USA), anti-p50, antip65, anti-BAX, anti-cleaved caspase-3, histone H1 (1:1,000; Santa Cruz Biotechnologies, Inc., Santa Cruz, CA, USA), antihEPO (1:1,000, R&D systems, Inc., Minneapolis, MN), antiPEG-2-128, anti-PEG-B47 (1:1,000; Abcam, Cambridge, UK) and anti-β-actin (1:2,500; Sigma, St Louis, MO).

Techniques: Expressing, Immunohistochemistry, TBARS Assay, Detection Assay

Journal: Biomolecules & therapeutics

Article Title: PEGylated Erythropoietin Protects against Brain Injury in the MCAO-Induced Stroke Model by Blocking NF-κB Activation.

doi: 10.4062/biomolther.2019.147

Figure Lengend Snippet: Fig. 3. Effects of P-EPO on inflammatory protein and NF-κB, STAT5 activity. 10 h after reperfusion following 2-h MCAO, vehicle (Sham), 5,000 U/kg of P-EPO was administered via i.v. The ischemic cortex in the striatum was collected for Western blot or immunohistochemistry 24 h after reperfusion following MCAO. (A) Representative Western blots of iNOS, COX-2, GFAP and Iba-1 in the ischemic brain. (B) Rep- resentative Western blots of transcription factors STAT1, STAT3, and STAT5 and phosphorylated form and NF-κB factor p50 and p65. Equal loading was confirmed by monitoring the β-actin and histone-H1 protein levels. (C) The expression levels were quantified by stereological analysis using the ImageJ program. (D) Serum levels of cytokines after MCAO measured by ELISA. Anti-inflammatory cytokines (TNF-α, IL-6, IL-1β) were examined. The data are the mean ± SD each sample was probed in triplicate. n=5/group; *p<0.05, **p<0.01, ***p<0.0001 significant difference from MCAO mice. ##p<0.01, ###p<0.0001 significant difference from Sham mice (Sham n=8-9, MCAO n=8-9, M-EPO n=10-12).

Article Snippet: Membranes were incubated at room temperature for 2 h with the following specific antibodies: anti-COX-2, anti-IκB, anti-p-IκB, anti- STAT1, anti-p-STAT1, anti- STAT3, anti-pSTAT3, anti- STAT5, anti-p-STAT5 (Cell Signaling Technology, Inc., Beverly, MA, USA), anti-inducible nitric oxide synthase (iNOS) and anti-Glial fibrillary acidic protein (GFAP) (1:1,000 Novus Biologicals, Inc., Littleton, CO, USA), anti-p50, antip65, anti-BAX, anti-cleaved caspase-3, histone H1 (1:1,000; Santa Cruz Biotechnologies, Inc., Santa Cruz, CA, USA), antihEPO (1:1,000, R&D systems, Inc., Minneapolis, MN), antiPEG-2-128, anti-PEG-B47 (1:1,000; Abcam, Cambridge, UK) and anti-β-actin (1:2,500; Sigma, St Louis, MO).

Techniques: Activity Assay, Western Blot, Immunohistochemistry, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Cell reports

Article Title: Angiopoietin-2 aggravates Alzheimer’s disease by promoting blood-brain barrier dysfunction and neuroinflammation

doi: 10.1016/j.celrep.2025.116621

Figure Lengend Snippet: (A and B) Increased CD74 + activated microglia co-labeled with Iba1 in the brain cortex of 6-month-old 5xFAD mice treated with ANGPT2 AAV compared to control-AAV-treated mice (A), with corresponding quantification (B). Arrowheads indicate CD74 + activated microglia. Scale bars: 50 μm. (C and D) Representative images (C) and quantification (D) showing increased cell body diameter and decreased dendritic length of Iba1 + microglia in ANGPT2-AAV-treated 5xFAD mice compared with controls. Asterisks mark the centers of Iba1 + microglial nuclei. Scale bars: 10 μm. (E and F) Elevated GFAP + reactive astrocytes co-labeled with S100β in the cortex of ANGPT2-AAV-treated 5xFAD mice compared with controls (E), with corresponding quantification (F). Arrowheads indicate GFAP + reactive astrocytes. Scale bars: 50 μm. (G and H) Morphological analysis of GFAP + reactive astrocytes showing increased branching and area coverage in ANGPT2-AAV-treated 5xFAD mice compared to control-AAV-treated mice (G), with corresponding quantification (H). Scale bars: 50 μm (white) and 20 μm (yellow). (I) Cytokine profile analysis showing increased levels of pro-inflammatory cytokines in ANGPT2-AAV-treated 5xFAD mice compared to control-AAV-treated mice. Each dot indicates an individual mouse for (B), (F), (H), and (I). Data are presented as the mean ± SEM.

Article Snippet: Next, sections were incubated overnight at 4°C with the following primary antibodies: For mouse tissue sections: CD31 (Thermo Fisher Scientific, MA3105), β-amyloid (Abcam, ab2539, ab126649), TIE2 (Regeneron Pharmaceuticals Inc., human monoclonal, REGN1376), pTIE2 (R&D systems, AF2720), ANGPT2 (Regeneron Pharmaceuticals Inc., human monoclonal, REGN910), PDGFR β (Thermo Fisher Scientific, 14-1402-82), fibrin(ogen) (Dako, A0080), claudin-5 (Thermo Fisher Scientific, 34-1600), GFAP (Aves Lab, GFAP), S100 β (Abcam, ab52642), Iba1 (Fujifilm Wako, 019-19741), CD74 (BioLegend, 151002), occludin (Thermo Fisher Scientific, 71-1500), PLVAP (BD Biosciences, 550563), and caveolin-1 (Santa Cruz Biotechnology, sc53564).

Techniques: Labeling, Control