Journal: European Journal of Medical Research

Article Title: The S1PR1–CCN1 axis drives endothelial-to-mesenchymal transition and vascular instability in brain arteriovenous malformations

doi: 10.1186/s40001-025-03484-5

Figure Lengend Snippet: Endothelial enrichment of S1PR1 in the human brain and its downregulation in ruptured bAVMs. A Normalized transcript levels of S1PR1 across multiple brain regions in the Human Protein Atlas (HPA) dataset, with peak expression observed in the cortex, hippocampus, thalamus, and basal ganglia. B Independent validation of regional S1PR1 expression patterns in the Genotype-Tissue Expression (GTEx) dataset. C Preferential enrichment of S1PR1 in endothelial cells shown by single-cell RNA-seq from the HPA brain atlas. D Immunohistochemistry (IHC) analysis of human bAVM tissues showing strong and continuous endothelial S1PR1 expression in unruptured tissues, and reduced staining in ruptured tissues. Right panel: Quantification of IHC staining intensity for endothelial S1PR1 expression ( n = 3). E Dual immunofluorescence staining of S1PR1 and CD31 (red) shows extensive colocalization in unruptured bAVMs, whereas ruptured bAVMs tissues exhibit reduced colocalization and fragmented endothelial S1PR1 expression

Article Snippet: A Normalized transcript levels of S1PR1 across multiple brain regions in the Human Protein Atlas (HPA) dataset, with peak expression observed in the cortex, hippocampus, thalamus, and basal ganglia.

Techniques: Expressing, Biomarker Discovery, RNA Sequencing, Immunohistochemistry, Staining, Immunofluorescence

Journal: European Journal of Medical Research

Article Title: The S1PR1–CCN1 axis drives endothelial-to-mesenchymal transition and vascular instability in brain arteriovenous malformations

doi: 10.1186/s40001-025-03484-5

Figure Lengend Snippet: S1PR1-mediated regulation of endothelial plasticity and migration. A – D Validation of S1PR1 knockdown (sh-S1PR1#1/#2) and overexpression in HUVECs by qRT-PCR and Western blot; β-actin used as loading control. E , F Expression of EndoMT-associated markers (N-cadherin (CDH2), α-SMA (ACTA2), VE-cadherin (CDH5)) assessed by qRT-PCR and Western blot following S1PR1 knockdown ( n = 3). G , H Representative Transwell migration images and quantification at 24 h and 48 h time points following S1PR1 knockdown and overexpression ( n = 3). I , J Representative scratch wound images and quantification at 0 h and 24 h following S1PR1 knockdown and overexpression ( n = 3). K Schematic overview of CRISPR/Cas9-mediated generation of s1pr1 crispant zebrafish. L Representative light field imaging of control and s1pr1 crispant embryos at 2 days post-fertilization (2 dpf). Yellow arrow indicates cranial hemorrhage observed in the s1pr1 -deficient embryo. The graph on the right summarizes the percentage of embryos exhibiting cranial hemorrhage. n = 100 for Cas9 control group, n = 100 for s1pr1 crispant. M Confocal images of cranial vasculature in Tg (flk1:EGFP); Tg (gata1:dsRed) embryos at 2 dpf. s1pr1 crispants exhibited extravasation of red blood cells (gata1:dsRed) and formation of localized hemorrhagic regions. Scale bar: 100 μm

Article Snippet: A Normalized transcript levels of S1PR1 across multiple brain regions in the Human Protein Atlas (HPA) dataset, with peak expression observed in the cortex, hippocampus, thalamus, and basal ganglia.

Techniques: Migration, Biomarker Discovery, Knockdown, Over Expression, Quantitative RT-PCR, Western Blot, Control, Expressing, CRISPR, Imaging

Journal: European Journal of Medical Research

Article Title: The S1PR1–CCN1 axis drives endothelial-to-mesenchymal transition and vascular instability in brain arteriovenous malformations

doi: 10.1186/s40001-025-03484-5

Figure Lengend Snippet: Global transcriptomic shifts associated with inflammation and ECM remodeling following S1PR1 silencing. A Violin plots showing the distribution of log-transformed FPKM values across biological replicates in S1PR1-silenced (sh-S1PR1) and control (sh-NC) HUVECs, indicating high data quality and consistency. B Scatter plot of log₂FPKM values comparing gene expression between sh-S1PR1 and sh-NC HUVECs. Each dot represents a gene; red and green denote significantly upregulated and downregulated genes, respectively. C Volcano plot of global DEGs between sh-S1PR1 and sh-NC groups. D Heatmap of differentially expressed genes (DEGs), showing hierarchical clustering and clear transcriptomic separation between groups. E Circos plot mapping the chromosomal distribution of upregulated (red) and downregulated (blue) DEGs. F and downregulated G genes across Biological Process, Cellular Component, and Molecular Function categories

Article Snippet: A Normalized transcript levels of S1PR1 across multiple brain regions in the Human Protein Atlas (HPA) dataset, with peak expression observed in the cortex, hippocampus, thalamus, and basal ganglia.

Techniques: Transformation Assay, Control, Gene Expression

Journal: European Journal of Medical Research

Article Title: The S1PR1–CCN1 axis drives endothelial-to-mesenchymal transition and vascular instability in brain arteriovenous malformations

doi: 10.1186/s40001-025-03484-5

Figure Lengend Snippet: ECM remodeling and epigenetic repression as concurrent downstream effects following S1PR1 silencing. A , B KEGG pathway enrichment analysis of DEGs in S1PR1-knockdown HUVECs. C , D Reactome pathway analysis of DEGs following S1PR1 knockdown

Article Snippet: A Normalized transcript levels of S1PR1 across multiple brain regions in the Human Protein Atlas (HPA) dataset, with peak expression observed in the cortex, hippocampus, thalamus, and basal ganglia.

Techniques: Knockdown

Journal: European Journal of Medical Research

Article Title: The S1PR1–CCN1 axis drives endothelial-to-mesenchymal transition and vascular instability in brain arteriovenous malformations

doi: 10.1186/s40001-025-03484-5

Figure Lengend Snippet: Proteomic landscape and pathway alterations in S1PR1-knockdown endothelial cells. A Boxplot of total protein abundance in S1PR1-knockdown (sh_1–3) and control (sh-NC_1–3) HUVECs. B Heatmap and hierarchical clustering of differentially expressed proteins (DEPs) between sh-S1PR1 and sh-NC groups. C Volcano plot of DEPs between sh-S1PR1 and sh-NC groups. D Protein–protein interaction (PPI) network of DEPs. E GO annotation between sh-S1PR1 and sh-NC groups. (F–G) KEGG pathway enrichment of upregulated F and downregulated G proteins

Article Snippet: A Normalized transcript levels of S1PR1 across multiple brain regions in the Human Protein Atlas (HPA) dataset, with peak expression observed in the cortex, hippocampus, thalamus, and basal ganglia.

Techniques: Knockdown, Quantitative Proteomics, Control

Journal: European Journal of Medical Research

Article Title: The S1PR1–CCN1 axis drives endothelial-to-mesenchymal transition and vascular instability in brain arteriovenous malformations

doi: 10.1186/s40001-025-03484-5

Figure Lengend Snippet: CCN1 is upregulated upon S1PR1 silencing and correlates with EndoMT-associated endothelial activation in human bAVMs. A Proteomic profiling identifies CCN1 as significantly upregulated following S1PR1 knockdown in HUVECs. B , C Validation of CCN1 upregulation at the mRNA (qRT-PCR) and protein (Western blot) levels in S1PR1 knockdown endothelial cells ( n = 3). D Quantification of CCN1 expression in ruptured versus unruptured human bAVM tissues ( n = 3). E Representative Immunohistochemistry images of CCN1 in human bAVM tissues, showing elevated expression in ruptured versus unruptured samples. F Immunofluorescence staining of human bAVM tissue shows co-localization of CCN1 with the endothelial marker CD31. G , H Re-analysis of publicly available single-cell RNA sequencing data comparing CCN1 expression across normal and bAVM vascular endothelium. I Schematic illustration of endothelial-specific ccn1 overexpression in zebrafish embryos using Tol2-mediated transgenesis. J Quantification of ccn1 mRNA levels in control and ccn1 -overexpressing embryos at 2 days post-fertilization (2 dpf) by qRT-PCR ( n = 3). K Light field images of zebrafish embryos at 2 dpf. Cranial hemorrhage (yellow arrow) is observed in the ccn1 -overexpressing group but not in controls. Right panel: Quantification of cerebral hemorrhage incidence ( n = 100). L Confocal images of cranial vasculature and red blood cells in Tg (gata1:Ds Red; flk1:EGFP) embryos at 2 dpf. Overexpression of ccn1 led to erythrocyte leakage and blood pooling in the cranial region. M Representative images of trunk vasculature in 5 dpf embryos. Overexpression of ccn1 caused significant dilation of intersomitic vessels (ISVs), as shown by increased vessel diameter. Right panel: Quantification of ISV diameter ( n = 6). N Confocal images of cerebral vasculature at 5 dpf showing increased cerebrovascular density in ccn1-overexpressing embryos. Right panel: Quantification of cerebrovascular fluorescence area ( n = 6). Scale bars: 100 μm

Article Snippet: A Normalized transcript levels of S1PR1 across multiple brain regions in the Human Protein Atlas (HPA) dataset, with peak expression observed in the cortex, hippocampus, thalamus, and basal ganglia.

Techniques: Activation Assay, Knockdown, Biomarker Discovery, Quantitative RT-PCR, Western Blot, Expressing, Immunohistochemistry, Immunofluorescence, Staining, Marker, RNA Sequencing, Over Expression, Control, Fluorescence

Journal: European Journal of Medical Research

Article Title: The S1PR1–CCN1 axis drives endothelial-to-mesenchymal transition and vascular instability in brain arteriovenous malformations

doi: 10.1186/s40001-025-03484-5

Figure Lengend Snippet: Rescue of S1PR1 deficiency-induced EndoMT and migration by CCN1 silencing. A – C mRNA and protein expression of S1PR1 and CCN1 in HUVECs following S1PR1 knockdown with or without co-transfection of sh-CCN1. D – F Rescue of EndoMT markers (N-cadherin, α-SMA, VE-cadherin) assessed by qRT-PCR ( n = 3). G Representative transwell migration images and quantification at 24 h and 48 h post-treatment ( n = 3). H Representative scratch wound images and quantification of wound closure at 0 h and 24 h ( n = 3)

Article Snippet: A Normalized transcript levels of S1PR1 across multiple brain regions in the Human Protein Atlas (HPA) dataset, with peak expression observed in the cortex, hippocampus, thalamus, and basal ganglia.

Techniques: Migration, Expressing, Knockdown, Cotransfection, Quantitative RT-PCR