Journal: Signal Transduction and Targeted Therapy

Article Title: Visinin-like protein 1 disrupts calcium homeostasis and promotes atrial fibrillation in human and rodent models

doi: 10.1038/s41392-026-02615-6

Figure Lengend Snippet: Multi-omics analysis reveals that Vsnl1 is associated with Ca 2+ homeostasis dysregulation in atrial fibrillation (AF). a Schematic workflow depicting the integration of bulk RNA sequencing (RNA-seq) and full-length single-cell RNA sequencing (scRNA-seq) data from human AF patients and rat AF models, alongside electrophysiological recordings. b UMAP analysis illustrating the transcriptional clustering of atrial cardiomyocytes, highlighting the distinct cellular subpopulations associated with AF pathophysiology. c Heatmap of differentially expressed genes in atrial cardiomyocytes during AF progression, derived from pseudotime trajectory analysis, demonstrating dynamic shifts in gene expression patterns over disease progression. The pseudotime trajectory is mapped onto the UMAP projection to visualize cellular state transitions. d Volcano plot displaying differentially expressed genes (DEGs) between AF and sinus rhythm (SR) patient samples, with Gene Ontology (GO) analysis indicating a significant enrichment of Ca 2+ signaling pathway-related genes in AF. e Venn diagram illustrating the intersection of DEGs from human AF samples ( P .adj < 0.05), genes whose expression levels correlated with abnormal Ca 2+ dynamics ( | Pearson’s ( r ) correlation coefficient | > 0.2, P < 0.01), Cluster 3 marker genes ( P .adj < 0.05), and genes from the pseudotime C4 module ( q _value < 0.05 and morans_I > 0.25), identifying Vsnl1 (encoding VILIP-1) and Kcnip2 as a key AF-associated genes

Article Snippet: The enzymatic digestion was performed at 37 °C in Ca 2+ -free PBS (Gibco, USA) containing an enzyme cocktail of 0.125 mg/mL trypsin (Gibco, USA), 0.1 mg/mL collagenase type IV (Sigma-Aldrich, USA) and 10 μg/mL DNase II (Sigma-Aldrich, USA).

Techniques: Biomarker Discovery, RNA Sequencing, Single Cell, Derivative Assay, Gene Expression, Expressing, Marker

Journal: Signal Transduction and Targeted Therapy

Article Title: Visinin-like protein 1 disrupts calcium homeostasis and promotes atrial fibrillation in human and rodent models

doi: 10.1038/s41392-026-02615-6

Figure Lengend Snippet: Atrial cardiomyocyte-specific overexpression of VILIP-1 promotes atrial fibrillation (AF) by increasing delayed afterdepolarizations (DADs), action potential duration (APD) alternans, and Ca 2+ leakage. a Electrophysiological recordings of surface electrocardiogram (ECG) and intracardiac electrograms. Left: representative traces from Ctrl and VILIP-1-OE mice, indicating sinus rhythm (SR) in Ctrl and AF episodes in VILIP-1-OE mice following burst pacing. Right: incidence and duration of pacing-induced AF. * P < 0.05, Mann-Whitney test or Fisher’s exact test, n = 7–9 per group. b APD alternans analysis in isolated Langendorff-perfused hearts. Left: representative action potential traces of Ctrl and VILIP-1-OE mice. Right: quantification of APD alternans incidence. * P < 0.05, Fisher’s exact test, n = 11–12 per group. c Delayed afterdepolarizations (DADs) recordings in atrial cardiomyocytes isolated from Ctrl and VILIP-1-OE mice. Left: representative traces from Ctrl and VILIP-1-OE mice. Right: incidence of DADs. * P < 0.05, Fisher’s exact test, n = 19–21 per group. d Confocal Ca 2+ imaging in isolated atrial cardiomyocytes. Left: representative Ca 2+ wave recordings in Ctrl and VILIP-1-OE cells. Right: quantification of the frequency of spontaneous Ca 2+ waves, indicating that VILIP-1-OE increased Ca 2+ leakage in atrial cardiomyocytes. * P < 0.05, Mann-Whitney test, n = 16-19 per group. e Representative line-scan images of caffeine (10 mM)-induced Ca 2+ transients in atrial cardiomyocytes from Ctrl and VILIP-1-OE mice. Quantification of sarcoplasmic reticulum Ca 2+ content, measured by peak caffeine-induced Ca 2+ release amplitude, showing significantly reduced sarcoplasmic reticulum Ca 2+ stores in the VILIP-1-OE group (* P < 0.05, Student’s t test, n = 7 cells per group)

Article Snippet: The enzymatic digestion was performed at 37 °C in Ca 2+ -free PBS (Gibco, USA) containing an enzyme cocktail of 0.125 mg/mL trypsin (Gibco, USA), 0.1 mg/mL collagenase type IV (Sigma-Aldrich, USA) and 10 μg/mL DNase II (Sigma-Aldrich, USA).

Techniques: Over Expression, MANN-WHITNEY, Isolation, Imaging

Journal: Signal Transduction and Targeted Therapy

Article Title: Visinin-like protein 1 disrupts calcium homeostasis and promotes atrial fibrillation in human and rodent models

doi: 10.1038/s41392-026-02615-6

Figure Lengend Snippet: Sodium-calcium exchanger (NCX-1) is identified as a key downstream target for VILIP-1. a Schematic workflow illustrating immunoprecipitation-mass spectrometry (left panel) and proximity labeling experiments using VILIP-1-V5-miniTurbo under varying Ca 2+ concentrations (right panel) to identify VILIP-1 interactors. b MaSigPro (R package) identified four dynamic VILIP-1-proximity labeling patterns in response to increased Ca 2+ concentrations. Proteins in Cluster 1 were identified as VILIP-1’s candidate interactors based on their progressive enrichment in VILIP-1-V5-miniTurbo-mediated biotinylation profiles with increasing Ca 2+ concentrations. c Venn diagram illustrating the overlap between interactors identified by co-immunoprecipitation and proximity labeling, confirming high-confidence candidates. d Western blot analysis of biotinylated proteins from VILIP-1-V5-miniTurbo proximity labeling, showing Ca 2+ -dependent enrichment of NCX-1. e Confocal microscopy images confirming the localization of biotinylated VILIP-1-interacting proteins (within a 10 nm radius), NCX-1, in neonatal rat cardiomyocytes (NRCMs) expressing VILIP-1-V5-miniTurbo. Cells were treated with 500 μM biotin for 13 min at 37 °C, and biotinylated proteins were visualized using Neutravidin–Alexa Fluor 488 staining. f Representative immunoblots showing co-immunoprecipitation of VILIP-1 and NCX-1 in rat atrial lysate, demonstrating their interaction. g Western blot analysis of in vitro GST pull-down experiments from VILIP-1-GST and HA-NCX-1, confirming the direct interaction between VILIP-1 and NCX-1. h Confocal microscopy images displaying colocalization of VILIP-1 and NCX-1 in rat atrial cardiomyocytes

Article Snippet: The enzymatic digestion was performed at 37 °C in Ca 2+ -free PBS (Gibco, USA) containing an enzyme cocktail of 0.125 mg/mL trypsin (Gibco, USA), 0.1 mg/mL collagenase type IV (Sigma-Aldrich, USA) and 10 μg/mL DNase II (Sigma-Aldrich, USA).

Techniques: Immunoprecipitation, Mass Spectrometry, Labeling, Western Blot, Confocal Microscopy, Expressing, Staining, In Vitro

Journal: Signal Transduction and Targeted Therapy

Article Title: Visinin-like protein 1 disrupts calcium homeostasis and promotes atrial fibrillation in human and rodent models

doi: 10.1038/s41392-026-02615-6

Figure Lengend Snippet: Atrial cardiomyocyte-specific overexpression of VILIP-1 increases atrial fibrillation (AF) susceptibility through enhancing the membrane expression and function of NCX-1. a Western blot analysis of VILIP-1 and NCX-1 protein levels in the atria of VILIP-1-OE and Ctrl mice. Left: representative immunoblot images. Right: quantification of protein levels normalized to GAPDH. * P < 0.05, Student’s t test, n = 3 per group. b Capillary western immunoassay results showed elevated NCX-1 and VILIP-1 protein levels in membrane fractions of atria from Ctrl and VILIP-1-OE mice. Quantification was performed using the Total Protein Detection Module (DM-TP01, ProteinSimple, USA). * P < 0.05, Student’s t test, n = 4 per group. c Whole-cell patch-clamp recordings from isolated atrial cardiomyocytes illustrating a significant increase in the I NCX current density in VILIP-1-OE mice compared to Ctrl. * P < 0.05, Student’s t test, n = 14 per group. d Confocal Ca 2+ imaging of atrial cardiomyocytes from VILIP-1-OE mice pretreated with vehicle or SEA0400. Left: representative Ca 2+ wave recordings. Right: quantification of the frequency of spontaneous Ca 2+ waves, showing reduced spontaneous Ca 2+ waves upon SEA0400 treatment. * P < 0.05, Mann-Whitney test, n = 13-18 per group. e Electrophysiological recordings of surface ECG and intracardiac electrograms in VILIP-1-OE mice. Left: representative traces of pacing-induced AF episodes in VILIP-1-OE mice with or without NCX-1 inhibition using SEA0400. Right: quantification of AF incidence and duration, showing a significant reduction in AF susceptibility upon SEA0400 treatment. * P < 0.05, Mann-Whitney test or Fisher’s exact test, n = 7–8 per group

Article Snippet: The enzymatic digestion was performed at 37 °C in Ca 2+ -free PBS (Gibco, USA) containing an enzyme cocktail of 0.125 mg/mL trypsin (Gibco, USA), 0.1 mg/mL collagenase type IV (Sigma-Aldrich, USA) and 10 μg/mL DNase II (Sigma-Aldrich, USA).

Techniques: Over Expression, Membrane, Expressing, Western Blot, Patch Clamp, Isolation, Imaging, MANN-WHITNEY, Inhibition

Journal: Signal Transduction and Targeted Therapy

Article Title: Visinin-like protein 1 disrupts calcium homeostasis and promotes atrial fibrillation in human and rodent models

doi: 10.1038/s41392-026-02615-6

Figure Lengend Snippet: Inhibition of VILIP-1 myristoylation rescues atrial fibrillation (AF) susceptibility in rat AF models. a Capillary western immunoassay analysis and quantification of NCX-1 expression in atrial membrane fractions from AF rat models treated with vehicle or Desloratadine. Quantification was performed using the Total Protein Detection Module (DM-TP01, ProteinSimple, USA). * P < 0.05, Student’s t test, n = 4 per group. b Whole-cell patch-clamp recordings demonstrate that AF rats treated with vehicle exhibit significantly increased I NCX current density compared to the control, while desloratadine treatment restores the I NCX current density to control levels. * P < 0.05, one-way ANOVA, n = 14–18 per group. c Confocal Ca 2+ imaging of atrial cardiomyocytes from rat AF models pretreated with vehicle or desloratadine. Left: representative Ca 2+ wave recordings. Right: quantification of the frequency of spontaneous Ca 2+ waves, showing reduced Ca 2+ waves upon desloratadine treatment. * P < 0.05, Mann-Whitney test, n = 17 per group. d Electrophysiological recordings of surface ECG and intracardiac electrograms in pacing-induced AF rats. Left: representative traces showing AF episodes in vehicle-pretreated versus 5 mg/kg desloratadine-pretreated rats. Right: quantification of AF incidence and duration, demonstrating a significant reduction in AF susceptibility following desloratadine administration. * P < 0.05, Mann-Whitney test or Fisher’s exact test, n = 6–8 per group

Article Snippet: The enzymatic digestion was performed at 37 °C in Ca 2+ -free PBS (Gibco, USA) containing an enzyme cocktail of 0.125 mg/mL trypsin (Gibco, USA), 0.1 mg/mL collagenase type IV (Sigma-Aldrich, USA) and 10 μg/mL DNase II (Sigma-Aldrich, USA).

Techniques: Inhibition, Western Blot, Expressing, Membrane, Patch Clamp, Control, Imaging, MANN-WHITNEY

Journal: Signal Transduction and Targeted Therapy

Article Title: Visinin-like protein 1 disrupts calcium homeostasis and promotes atrial fibrillation in human and rodent models

doi: 10.1038/s41392-026-02615-6

Figure Lengend Snippet: VILIP-1 inhibition alleviates Ca 2+ leakage and prevents atrial fibrillation (AF) inducibility in both human patients and rat AF models. a Molecular docking simulation demonstrating the interaction between repaglinide and VILIP-1 (docking score = -318.2), supporting its potential as a VILIP-1 inhibitor. b Thermal shift assay (TSA) analysis showing VILIP-1 protein stabilization following 10 μM repaglinide treatment at varying temperatures. c Binding affinities of repaglinide to VILIP-1 measured by surface plasmon resonance (SPR) assays. d Capillary western immunoassay demonstrating a reduction in NCX-1 expression in membrane fractions of atria from pacing-induced AF rat models treated with repaglinide. Quantification was performed using the Total Protein Detection Module (DM-TP01, ProteinSimple, USA). * P < 0.05, Mann-Whitney test, n = 4 per group. e Immunofluorescence staining of VILIP-1 and NCX-1 in atria from AF rat models pretreated with vehicle or repaglinide, showing reduced NCX-1 surface expression upon repaglinide treatment. f Confocal Ca 2+ imaging of atrial cardiomyocytes from AF rat models pretreated with vehicle or repaglinide. Top: representative Ca 2+ wave recordings. Bottom: quantification of the frequency of spontaneous Ca 2+ waves, demonstrating a significant reduction in spontaneous Ca 2+ waves following repaglinide treatment. * P < 0.05, Mann-Whitney test, n = 15–16 per group. g Electrophysiological recordings of surface ECG and intracardiac electrograms in pacing-induced AF rats pretreated with vehicle or repaglinide. Top: representative traces showing a decrease in pacing-induced AF episodes following repaglinide administration. Bottom: quantification of AF incidence and duration. * P < 0.05, Mann-Whitney test or Fisher’s exact test, n = 6 – 7 per group. h Capillary western immunoassay analysis of NCX-1 expression in membrane fractions of human atria, showing decreased NCX-1 surface expression following repaglinide treatment. Quantification was performed using the Total Protein Detection Module (DM-TP01, ProteinSimple, USA). * P < 0.05, Mann-Whitney test, n = 6-8 per group. i Confocal Ca 2+ imaging of atrial cardiomyocytes from human AF patients pretreated with vehicle or 10 μM repaglinide. Left: representative Ca 2+ wave recordings. Right: quantification of the frequency of spontaneous Ca 2+ waves, demonstrating a significant reduction in spontaneous Ca 2+ waves following repaglinide treatment. * P < 0.05, Mann-Whitney test, n = 17–19 per group

Article Snippet: The enzymatic digestion was performed at 37 °C in Ca 2+ -free PBS (Gibco, USA) containing an enzyme cocktail of 0.125 mg/mL trypsin (Gibco, USA), 0.1 mg/mL collagenase type IV (Sigma-Aldrich, USA) and 10 μg/mL DNase II (Sigma-Aldrich, USA).

Techniques: Inhibition, Thermal Shift Assay, Binding Assay, SPR Assay, Western Blot, Expressing, Membrane, MANN-WHITNEY, Immunofluorescence, Staining, Imaging

Journal: Signal Transduction and Targeted Therapy

Article Title: Visinin-like protein 1 disrupts calcium homeostasis and promotes atrial fibrillation in human and rodent models

doi: 10.1038/s41392-026-02615-6

Figure Lengend Snippet: Upregulated VILIP-1 promotes atrial fibrillation (AF) through a myristoylation-dependent pathway. In pathological conditions, upregulated VILIP-1 binds Ca 2+ , triggering myristoylation-dependent membrane anchoring. At the membrane, VILIP-1 directly increases the surface membrane expression of NCX-1 and enhances its function. The sustained Ca 2+ overload combined with intracellular Na + accumulation drives NCX-1 reverse-mode activation, exacerbating both Ca 2+ influx and cytoplasmic Ca 2+ accumulation. The resulting Ca 2+ overload reinforces VILIP-1 membrane targeting by promoting its myristoylation, creating a self-perpetuating cycle driving NCX-1 hyperactivity and AF progression. The schematic illustration was created in Adobe Illustrator 2024

Article Snippet: The enzymatic digestion was performed at 37 °C in Ca 2+ -free PBS (Gibco, USA) containing an enzyme cocktail of 0.125 mg/mL trypsin (Gibco, USA), 0.1 mg/mL collagenase type IV (Sigma-Aldrich, USA) and 10 μg/mL DNase II (Sigma-Aldrich, USA).

Techniques: Membrane, Expressing, Activation Assay

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Evaluation of a human 3D multicellular hepatic spheroid model as a platform for studying hepatic transporters

doi: 10.1007/s00018-025-05996-z

Figure Lengend Snippet: Tight junction modulation in human multicellular hepatic spheroids (MHS) on culture day 14. Donor WID MHS were incubated in standard HBSS containing 5 µM CDFDA for 30 min at 37 °C on culture day 14. MHS were washed and maintained in standard HBSS (“cells + bile”) or Ca 2+ -free HBSS (“cells only”) for up to 20 min and imaged over time using a CellInsight CX7 High Content Scanner (10 µm step size, 12 steps, 120 µm extended depth of field, 10X objective). Transmitted light (grey) was used to visualize MHS structure. White arrows depict CDF accumulation in bile pockets in standard HBSS (top panel) or CDF dispersing from bile pockets in Ca 2+ -free HBSS (bottom panel). Scale bar = 100 µm. CDFDA, 5(6)-Carboxy-2′,7′-dichlorofluorescein diacetate; HBSS, Hank’s balanced salt solution

Article Snippet: Acetonitrile (Cat. No. A995), methanol (Cat. No. A456), water (Cat. No. W6), formic acid (Cat. No. A117), phosphate-buffered saline (PBS; Cat. No. 14190), standard Hank’s balanced salt solution (HBSS; Cat. No. 14025), Ca 2+ -free HBSS (Cat. No. 14175), 5(6)-carboxy-2′,7′-dichlorofluorescein diacetate (CDFDA; Cat. No. C369), and qPCR TaqMan probes were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Incubation