Journal: Journal of neuroinflammation

Article Title: Inhibition of neutrophil extracellular traps alleviates blood-brain barrier disruption and cognitive dysfunction via Wnt3/β-catenin/TCF4 signaling in sepsis-associated encephalopathy.

doi: 10.1186/s12974-025-03395-6

Figure Lengend Snippet: Fig. 2 The inhibition of NETs formation improved SAE mice survival, defended neuroinflammation and alleviated cognitive impairment. A 7-day survival rates of SAE mice treated with different concentrations of GSK484 (n = 15). B Quantification of NETs numbers (n = 6). C and D GSK484 inhibited NETs formation in the CA1 region of the hippocampus by IF staining (n = 6). E–H ELISA test showed that GSK484 decreased the levels of TNF-α, IL-6, IL-1β and IL-10 in hippocampus tissue (n = 6). I The trace heat map of OFT. The time (J) and the movement distance (K) in center in OFT. L The trace heat map of MWM test. The movement distance (M) in island, and the time (N) in NW area in the MWM test were increased after GSK484 treatment (n = 6). NETs: Neutrophil extracellular traps; CLP: Cecal Ligation and Puncture; SAE: Sepsis-associated encephalopathy; IF: immunofluorescence; OFT: open field test; MWM: Morris water maze. (ns: no significance, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001)

Article Snippet: GSK484 was administered three days prior to and two days after CLP surgery [30], For β-catenin/ TCF4 inhibition, LF3 (MCE, USA), a selective antagonist, was dissolved in a vehicle solution of 2% DMSO, 40% PEG300, 5% Tween-80, and 53% saline to achieve a final concentration of 250 mg/mL.

Techniques: Inhibition, Staining, Enzyme-linked Immunosorbent Assay, Ligation, Immunofluorescence

Journal: Journal of neuroinflammation

Article Title: Inhibition of neutrophil extracellular traps alleviates blood-brain barrier disruption and cognitive dysfunction via Wnt3/β-catenin/TCF4 signaling in sepsis-associated encephalopathy.

doi: 10.1186/s12974-025-03395-6

Figure Lengend Snippet: Fig. 3 Distinct transcriptional signature between CLP- and GSK484-treated hippocampus. A A vulcano plot was depicted with p < 0.05 of all 705 detected genes from RNA-Seq dataset. Differential expression genes (DEGs) were colored with blue and red, regarding to 163 down-regulated and 542 up-regulated genes, respectively. B Bar plot of GO enrichment analysis; (BP) biological process, (CC) cellular components, (MF) molecular function. C Bubble plot of KEGG pathway analysis. D Cytoscape was used to visualize the PPI network generated by String. E The top 10 hub genes were obtained using the CytoHubba plug-in in Cytoscape. F The top 1 modules are presented based on MCODE scores in descending order. DEGs: Differential expression genes; BP: biological process; CC: cellular components; MF: molecular function; CLP: Cecal Ligation and Puncture; GO: Gene Ontology; KEGG: Kyoto Encyclopedia of Genes and Genomes

Article Snippet: GSK484 was administered three days prior to and two days after CLP surgery [30], For β-catenin/ TCF4 inhibition, LF3 (MCE, USA), a selective antagonist, was dissolved in a vehicle solution of 2% DMSO, 40% PEG300, 5% Tween-80, and 53% saline to achieve a final concentration of 250 mg/mL.

Techniques: RNA Sequencing, Quantitative Proteomics, Generated, Ligation

Journal: Journal of neuroinflammation

Article Title: Inhibition of neutrophil extracellular traps alleviates blood-brain barrier disruption and cognitive dysfunction via Wnt3/β-catenin/TCF4 signaling in sepsis-associated encephalopathy.

doi: 10.1186/s12974-025-03395-6

Figure Lengend Snippet: Fig. 4 The inhibition of NETs formation reduced BBB permeability in SAE mice by alleviating TJs disruption. A The BWC was measured. Quantification of the FS extravasation in the hippocampus (B) and the global brain tissue (C) (n = 6). D Representative images showing the extravasation of FS. E Expression levels of TJs (ZO-1, Occludin, Claudin-5) proteins in the hippocampus were detected by Western blot (n = 6). F Expression levels of TJ proteins were quantified and normalized to GAPDH levels (n = 6). Double labeling shows colocalization of Occludin (G) and ZO-1 (I) with ECs (CD31) in the CA1 region of the hippocampus (40x, n = 6). Quantification of the fluorescence intensity of Occludin (H) and ZO-1 (J) in the hippocampus. NETs: Neutrophil extracellular traps; CLP: Cecal Ligation and Puncture; SAE: Sepsis-associated encephalopathy; ECs: endothelial cells; BBB: Blood–brain barrier; TJs: Tight junctions; BWC: Brain water content; FS: Fluorescein sodium. (ns: no significance, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001)

Article Snippet: GSK484 was administered three days prior to and two days after CLP surgery [30], For β-catenin/ TCF4 inhibition, LF3 (MCE, USA), a selective antagonist, was dissolved in a vehicle solution of 2% DMSO, 40% PEG300, 5% Tween-80, and 53% saline to achieve a final concentration of 250 mg/mL.

Techniques: Inhibition, Permeability, Disruption, Expressing, Western Blot, Labeling, Fluorescence, Ligation

Journal: Journal of neuroinflammation

Article Title: Inhibition of neutrophil extracellular traps alleviates blood-brain barrier disruption and cognitive dysfunction via Wnt3/β-catenin/TCF4 signaling in sepsis-associated encephalopathy.

doi: 10.1186/s12974-025-03395-6

Figure Lengend Snippet: Fig. 7 The inhibition of NETs improved the top 1 modules associated proteins expression in the hippocampus. A Double labeling IF staining of VCL with ECs (CD31) after GSK484 treatment in the hippocampus (n = 6). B Quantification of the fluorescence intensity of VCL in CD31-positive area (n = 6). C Quantification of the mean fluorescence intensity of VCL in all area (n = 6). D Double labeling IF staining of Fermt1 with ECs (CD31) after GSK484 treatment in the hippocampus (n = 6). E Quantification of the fluorescence intensity of Fermt1 in CD31-positive area (n = 6). F Quantification of the mean fluorescence intensity of Fermt1 in all area (n = 6). NETs: Neutrophil extracellular traps; CLP: Cecal Ligation and Puncture; SAE: Sepsis-associated encephalopathy; IF: immunofluorescence. (*p < 0.05, ***p < 0.001, ****p < 0.0001; ###p < 0.001, ####p < 0.0001; *sham vs CLP, #CLP vs CLP + GSK484)

Article Snippet: GSK484 was administered three days prior to and two days after CLP surgery [30], For β-catenin/ TCF4 inhibition, LF3 (MCE, USA), a selective antagonist, was dissolved in a vehicle solution of 2% DMSO, 40% PEG300, 5% Tween-80, and 53% saline to achieve a final concentration of 250 mg/mL.

Techniques: Inhibition, Expressing, Labeling, Staining, Fluorescence, Ligation, Immunofluorescence

Journal: Journal of neuroinflammation

Article Title: Inhibition of neutrophil extracellular traps alleviates blood-brain barrier disruption and cognitive dysfunction via Wnt3/β-catenin/TCF4 signaling in sepsis-associated encephalopathy.

doi: 10.1186/s12974-025-03395-6

Figure Lengend Snippet: Fig. 8 The inhibition of NETs formation alleviated the AJs disruption may by regulating VCL/β-catenin/VE-cadherin functional complex. A Triple labeling IF staining showed colocalization of VCL, β-catenin and VE-cadherin (n = 6); Quantification of the fluorescence intensity of VCL (B), β-catenin (C) and VE-cadherin (D) in the hippocampus (n = 6). NETs: Neutrophil extracellular traps; CLP: Cecal Ligation and Puncture; SAE: Sepsis-associated encephalopathy; IF: immunofluorescence. (ns: no significance, *p < 0.05, **p < 0.01, ***p < 0.001)

Article Snippet: GSK484 was administered three days prior to and two days after CLP surgery [30], For β-catenin/ TCF4 inhibition, LF3 (MCE, USA), a selective antagonist, was dissolved in a vehicle solution of 2% DMSO, 40% PEG300, 5% Tween-80, and 53% saline to achieve a final concentration of 250 mg/mL.

Techniques: Inhibition, Disruption, Functional Assay, Labeling, Staining, Fluorescence, Ligation, Immunofluorescence

Journal: Journal of neuroinflammation

Article Title: Inhibition of neutrophil extracellular traps alleviates blood-brain barrier disruption and cognitive dysfunction via Wnt3/β-catenin/TCF4 signaling in sepsis-associated encephalopathy.

doi: 10.1186/s12974-025-03395-6

Figure Lengend Snippet: Fig. 9 The inhibition of NETs formation alleviated the disruption of TJs and AJs by activating Wnt3/β-catenin/TCF4 signaling pathway. A Expression levels of Wnt/β-catenin/TCF4 signaling proteins in the hippocampus were detected by Western blot (n = 6). B–H The quantification analysis of Wnt/ β-catenin/TCF4 signaling proteins expression levels (n = 6). I Wnt3 is predominantly secreted by neurons in the hippocampal CA1 region of septic mice. J Quantification of the mean fluorescence intensity of Wnt3 in CA1 region (n = 6). NETs: Neutrophil extracellular traps; CLP: Cecal Ligation and Puncture; SAE: Sepsis-associated encephalopathy; BBB: Blood–brain barrier; TJs: Tight junctions; AJs: adherens junction (ns: no significance, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001)

Article Snippet: GSK484 was administered three days prior to and two days after CLP surgery [30], For β-catenin/ TCF4 inhibition, LF3 (MCE, USA), a selective antagonist, was dissolved in a vehicle solution of 2% DMSO, 40% PEG300, 5% Tween-80, and 53% saline to achieve a final concentration of 250 mg/mL.

Techniques: Inhibition, Disruption, Expressing, Western Blot, Fluorescence, Ligation

Journal: Journal of neuroinflammation

Article Title: Inhibition of neutrophil extracellular traps alleviates blood-brain barrier disruption and cognitive dysfunction via Wnt3/β-catenin/TCF4 signaling in sepsis-associated encephalopathy.

doi: 10.1186/s12974-025-03395-6

Figure Lengend Snippet: Fig. 10 The LF3 treatment reversed the expression of Wnt3/β-catenin/TCF4 signaling pathway, TJs and AJs proteins. A Western blot detected the Wnt/β-catenin/TCF4 signaling proteins expression levels in the hippocampus after LF3 treatment (n = 6). B–F The quantification analysis of Wnt/β-catenin/TCF4 signaling proteins expression levels after LF3 treatment (n = 6). G Western blot detected the TJs and AJs proteins expression levels in the hippocampus after LF3 treatment (n = 6). H–M The quantification analysis of TJs and AJs proteins expression levels after LF3 treatment (n = 6). NETs: Neutrophil extracellular traps; CLP: Cecal Ligation and Puncture; SAE: Sepsis-associated encephalopathy; BBB: Blood–brain barrier; TJs: Tight junctions; AJs: adherens junction (ns: no significance, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001)

Article Snippet: GSK484 was administered three days prior to and two days after CLP surgery [30], For β-catenin/ TCF4 inhibition, LF3 (MCE, USA), a selective antagonist, was dissolved in a vehicle solution of 2% DMSO, 40% PEG300, 5% Tween-80, and 53% saline to achieve a final concentration of 250 mg/mL.

Techniques: Expressing, Western Blot, Ligation