cat af791  (R&D Systems)

Journal: Acta Neuropathologica Communications

Article Title: CX3CL1/CX3CR1 axis dysregulation contributes to epileptogenic mechanisms in focal cortical dysplasia

doi: 10.1186/s40478-026-02274-2

Figure Lengend Snippet: Characterization of CX3CL1 expression and its impact on immune and pathway modulation in interneurons. a t-SNE plot showing clustering of interneurons. b t-SNE plot of interneurons grouped by CX3CL1 expression level. c Violin plots of CX3CL1 expression across clusters. d Heatmap of pathway enrichment analysis between high and low expression groups, revealing significant differences in multiple pathways including adenosine receptor signaling, GABA degradation. e Estimated immune cell subpopulation proportions from CIBERSORTx deconvolution combining RNA-seq and scRNA-seq datasets. The x-axis represents CX3CL1/CX3CR1 high-expression subpopulations, and the y-axis represents the estimated proportion. Orange indicates disease group, green indicates normal group, and statistical significance is annotated above (ns, not significant; * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, Kruskal–Wallis test). f Ligand–receptor interaction analysis. The bubble plot shows ligand activity (AUPR), proportion of cells expressing each ligand, and average expression levels. The bar plot indicates LFC of ligands in sender cells. The heatmap displays the regulatory potential of candidate ligands across different receptors and target genes.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h at room temperature and incubated overnight at 4 °C with primary antibodies against CX3CL1 (1:1000; #29282, Signalway Antibody), CX3CR1 (1:1000; #38481, Signalway Antibody), ARG1 (1:1000; #sc-271430, Santa Cruz), iNOS (1:1000; #sc-7271, Santa Cruz), TNF-α (1:1000; #17590-1-AP, Proteintech), CD206 (1:1000; #18704-1-AP, Proteintech), β-actin (1:1000; #3700, CST), and GAPDH (1:1000; #5174, CST).

Techniques: Expressing, RNA Sequencing, Activity Assay

Journal: Acta Neuropathologica Communications

Article Title: CX3CL1/CX3CR1 axis dysregulation contributes to epileptogenic mechanisms in focal cortical dysplasia

doi: 10.1186/s40478-026-02274-2

Figure Lengend Snippet: CX3CL1/CX3CR1 dysregulation in human FCD epileptogenic foci. a H&E staining of normal cerebral cortex and FCD epileptogenic foci. Boxed area: Disorganized microcolumnar architecture; arrows: DNs; triangles: BCs. Scale bars: 200 μm (top), 20 μm (middle), 10 μm (bottom). b IHC analysis of CX3CL1/CX3CR1 localization and expression in normal cerebral cortex and FCD epileptogenic foci. Representative images shown ( n = 3 per group). Scale bars: 200 μm (left), 20 μm (right). c CX3CL1/CX3CR1 mRNA expression quantified by RT-qPCR in normal cortex and FCD epileptogenic foci. ( n = 4 per group). d CX3CL1/CX3CR1 protein expression in normal cortex and FCD epileptogenic foci. Control ( n = 6), FCDIa ( n = 5), FCDIIa and FCDIIb ( n = 4). Each data point represents an individual sample; horizontal lines indicate the mean. (Comparisons between control and each FCD subgroup were performed using unpaired two-tailed Student’s t-tests with Welch’s correction. * P < 0.05, *** P < 0.001; ns, not significant.)

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h at room temperature and incubated overnight at 4 °C with primary antibodies against CX3CL1 (1:1000; #29282, Signalway Antibody), CX3CR1 (1:1000; #38481, Signalway Antibody), ARG1 (1:1000; #sc-271430, Santa Cruz), iNOS (1:1000; #sc-7271, Santa Cruz), TNF-α (1:1000; #17590-1-AP, Proteintech), CD206 (1:1000; #18704-1-AP, Proteintech), β-actin (1:1000; #3700, CST), and GAPDH (1:1000; #5174, CST).

Techniques: Staining, Expressing, Quantitative RT-PCR, Control, Two Tailed Test

Journal: Acta Neuropathologica Communications

Article Title: CX3CL1/CX3CR1 axis dysregulation contributes to epileptogenic mechanisms in focal cortical dysplasia

doi: 10.1186/s40478-026-02274-2

Figure Lengend Snippet: Dynamic changes of CX3CR1 correlate with alterations in microglial polarization. a CX3CL1/CX3CR1 protein expression: Protein expression and statistical analysis in lesioned brain tissues of Sham and FL-CD groups at different time points ( n = 4). b Effects of PTZ on CX3CL1/CX3CR1: Protein expression and statistical analysis in Sham-PTZ vs. FL-CD-PTZ groups and FL-CD vs. FL-CD-PTZ groups ( n = 4). c mRNA levels of CX3CR1 and M1/M2-related cytokines (M1: IL-6, IL-1β; M2: ARG1, IL-10) in lesioned brain tissues of Sham vs. FL-CD and Sham-PTZ vs. FL-CD-PTZ groups ( n = 4). d Protein expression of CX3CR1 and M1/M2 markers (M1: iNOS, TNF-α; M2: CD206, ARG1) in lesioned brain tissues of Sham vs. FL-CD and Sham-PTZ vs. FL-CD-PTZ groups ( n = 4). e Double immunofluorescence staining demonstrated co-localization of CX3CR1(green) with Iba1(red)(Top). Double immunofluorescence staining of (1) iNOS (red) + Iba1 (green) and (2) ARG1 (red) + Iba1 (green) in lesioned brain tissues of Sham vs. FL-CD groups. Scale bars: 20 μm. f Double immunofluorescence staining of (1) iNOS (red) + Iba1 (green) and (2) ARG1 (red) + Iba1 (green) in lesioned brain tissues of Sham-PTZ vs. FL-CD-PTZ groups. Scale bars: 20 μm. g Quantitative analysis of the M1/M2 microglia (M1: iNOS, M2:ARG1) per mm2 at different groups ( n = 9). (Comparisons between groups were performed using unpaired two-tailed Student’s t-tests for normally distributed continuous variables. * P < 0.05, ** P < 0.01, *** P < 0.001; ns, not significant.)

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h at room temperature and incubated overnight at 4 °C with primary antibodies against CX3CL1 (1:1000; #29282, Signalway Antibody), CX3CR1 (1:1000; #38481, Signalway Antibody), ARG1 (1:1000; #sc-271430, Santa Cruz), iNOS (1:1000; #sc-7271, Santa Cruz), TNF-α (1:1000; #17590-1-AP, Proteintech), CD206 (1:1000; #18704-1-AP, Proteintech), β-actin (1:1000; #3700, CST), and GAPDH (1:1000; #5174, CST).

Techniques: Expressing, Double Immunofluorescence Staining, Two Tailed Test

Journal: Acta Neuropathologica Communications

Article Title: CX3CL1/CX3CR1 axis dysregulation contributes to epileptogenic mechanisms in focal cortical dysplasia

doi: 10.1186/s40478-026-02274-2

Figure Lengend Snippet: Summary of the dysregulation CX3CL1/CX3CR1 axis in FCD-related epileptogenesis: A Protective vs. Pathogenic Model. This diagram illustrates the dual role of the CX3CL1/CX3CR1 axis in the context of FL-CD and PTZ-induced seizures. It highlights how the balance between M2 polarization (protective) and M1 polarization (pathogenic) of microglia influences neuronal excitability, seizure threshold, and the pro-inflammatory microenvironment. The mechanistic hypothesis proposes that overexpression of sCX3CL1 disrupts neurotransmitter function, shifts the excitatory/inhibitory balance, and engages in a vicious cycle leading to neuronal damage and increased seizure frequency.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h at room temperature and incubated overnight at 4 °C with primary antibodies against CX3CL1 (1:1000; #29282, Signalway Antibody), CX3CR1 (1:1000; #38481, Signalway Antibody), ARG1 (1:1000; #sc-271430, Santa Cruz), iNOS (1:1000; #sc-7271, Santa Cruz), TNF-α (1:1000; #17590-1-AP, Proteintech), CD206 (1:1000; #18704-1-AP, Proteintech), β-actin (1:1000; #3700, CST), and GAPDH (1:1000; #5174, CST).

Techniques: Over Expression