Journal: Apoptosis

Article Title: Single-cell profiling and machine learning identify cuproptosis-related fibroblast subpopulations and fibrogenesis modulator AEBP1 in endometriosis

doi: 10.1007/s10495-026-02342-x

Figure Lengend Snippet: FDX1 knockdown attenuates copper ionophore-associated mitochondrial alterations and profibrotic responses in endometrial stromal cells. ThESCs were transfected with negative-control siRNA (siNC) or FDX1 siRNA (siFDX1), followed by treatment with CuCl 2 plus elesclomol (Cu + ele) or vehicle, as indicated. A Representative western blots and densitometric quantification of FDX1, LIAS, and Lip-DLAT. Tubulin was used as the loading control. B Representative JC-1 flow cytometry plots and quantification of JC-1 green fluorescence. C Representative MitoSOX staining images and quantification of mitochondrial ROS levels. Nuclei were counterstained with DAPI. D Representative immunofluorescence staining of AEBP1 and FDX1, with quantification of relative fluorescence intensity. E Representative western blots and densitometric quantification of AEBP1, α-SMA, and CTGF. F Representative western blots and densitometric quantification of β-catenin and c-Myc. Western blot quantification was derived from three independent experiments. MitoSOX fluorescence was quantified from three independent microscopic fields per group. Data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA followed by Tukey’s multiple-comparisons test. ** P < 0.01; **** P < 0.0001. Scale bars = 50 μm. si NC, negative-control small interfering RNA; si FDX1, FDX1-specific small interfering RNA; Cu + ele, CuCl 2 plus elesclomol; α-SMA, alpha-smooth muscle actin; CTGF, connective tissue growth factor; ROS, reactive oxygen species

Article Snippet: Human endometrial stromal cells (ThESCs) were obtained from the American Type Culture Collection (ATCC; catalog no. CRL-4003) and cultured in Dulbecco’s Modified Eagle Medium (DMEM) enriched with 10% fetal bovine serum (FBS; Gibco, Carlsbad, CA, USA).

Techniques: Knockdown, Transfection, Negative Control, Western Blot, Control, Flow Cytometry, Fluorescence, Staining, Immunofluorescence, Derivative Assay, Small Interfering RNA

Journal: Apoptosis

Article Title: Single-cell profiling and machine learning identify cuproptosis-related fibroblast subpopulations and fibrogenesis modulator AEBP1 in endometriosis

doi: 10.1007/s10495-026-02342-x

Figure Lengend Snippet: AEBP1 modulates profibrotic marker expression and β-catenin pathway-related proteins in endometrial stromal cells. ThESCs were transfected with AEBP1-specific siRNA (siAEBP1) or AEBP1 overexpression plasmid (ovAEBP1), followed by treatment with CuCl 2 plus elesclomol (Cu + ele) or vehicle, as indicated. A Representative immunofluorescence staining of AEBP1 and α-SMA in siNC, siAEBP1, Cu + ele, and siAEBP1 + Cu + ele groups, with quantification of relative fluorescence intensity. B Representative western blots and densitometric quantification of AEBP1, α-SMA, and CTGF in siNC, siAEBP1, Cu + ele, and siAEBP1 + Cu + ele groups. C Representative western blots and densitometric quantification of AEBP1, α-SMA, and CTGF in ovNC, Cu + ele, and ovAEBP1 groups. D Representative immunofluorescence staining of AEBP1 and β-catenin in siNC, siAEBP1, Cu + ele, and siAEBP1 + Cu + ele groups, with quantification of relative fluorescence intensity. E Representative western blots and densitometric quantification of β-catenin and c-Myc in siNC, siAEBP1, Cu + ele, and siAEBP1 + Cu + ele groups. F Representative western blots and densitometric quantification of β-catenin and c-Myc in ovNC, Cu + ele, and ovAEBP1 groups. Tubulin was used as the loading control for all western blot analyses. Western blot quantification was derived from three independent experiments. Data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA followed by Tukey’s multiple-comparisons test. ns, not significant; ** P < 0.01; *** P < 0.001; **** P < 0.0001. Scale bars = 50 μm. si NC, negative-control small interfering RNA; si AEBP1, AEBP1-specific small interfering RNA; ov NC, empty vector control; ovAEBP1, AEBP1 overexpression plasmid; α-SMA, alpha-smooth muscle actin; CTGF, connective tissue growth factor

Article Snippet: Human endometrial stromal cells (ThESCs) were obtained from the American Type Culture Collection (ATCC; catalog no. CRL-4003) and cultured in Dulbecco’s Modified Eagle Medium (DMEM) enriched with 10% fetal bovine serum (FBS; Gibco, Carlsbad, CA, USA).

Techniques: Marker, Expressing, Transfection, Over Expression, Plasmid Preparation, Immunofluorescence, Staining, Fluorescence, Western Blot, Control, Derivative Assay, Negative Control, Small Interfering RNA

Journal: bioRxiv

Article Title: Increased chromatin accessibility following 1α,25-dihydroxyvitamin D 3 treatment in human endometrial stromal cells

doi: 10.64898/2026.05.06.723064

Figure Lengend Snippet: The analysis shows that 1,25(OH) 2 D 3 treatment is associated with increased accessibility signal without widespread reorganization of the global accessible chromatin landscape. (A) Overall profiles and heatmaps of ATAC-seq signal centered on called accessible regions (center ± 2 kb) and (B) surrounding transcription start sites (TSS ± 2 kb) in vehicle- and 1,25(OH) 2 D 3 -treated T-HESCs. (C) Overlap of open chromatin peaks identified by ATAC-seq in vehicle- and 1,25(OH) 2 D 3 -treated cells. (D) Representative genome browser loci of ATAC-seq signal at VDR, CYP24A1, PRL, and EFL1 regions from vehicle- and 1,25(OH) 2 D 3 -treated cells. RefSeq gene models are shown below.

Article Snippet: hTERT-immortalized endometrial stromal cells (T-HESCs) were purchased from the American Type Culture Collection (ATCC; Cat. CRL-4003, VA, USA).

Techniques:

Journal: bioRxiv

Article Title: Increased chromatin accessibility following 1α,25-dihydroxyvitamin D 3 treatment in human endometrial stromal cells

doi: 10.64898/2026.05.06.723064

Figure Lengend Snippet: (A) Clustered heatmaps and average signal profiles comparing ATAC-seq intensity patterns between vehicle- and 1,25(OH) 2 D 3 -treated cells across DARs. (B) Volcano plot showing differential chromatin accessibility between 1,25(OH) 2 D 3 -treated and vehicle-treated T-HESCs. Peaks were classified as increased or decreased accessible regions based on adjusted p -value and fold-change thresholds. (C) Comparison of genomic features of regions between loss and gain of accessibility. (D) Metaplot and heatmap of ATAC-seq signal across regions with increased accessibility in response to 1,25(OH) 2 D 3 , showing elevated signal intensity in ligand-treated cells compared with vehicle-treated cells. (E) Metaplot and heatmap of ATAC-seq signal across regions with decreased accessibility in response to 1,25(OH) 2 D 3 . The result shows that 1,25(OH) 2 D 3 induces a distinct set of accessibility changes, with pronounced signal enhancement at selected ligand-responsive regions.

Article Snippet: hTERT-immortalized endometrial stromal cells (T-HESCs) were purchased from the American Type Culture Collection (ATCC; Cat. CRL-4003, VA, USA).

Techniques: Comparison

Journal: bioRxiv

Article Title: Increased chromatin accessibility following 1α,25-dihydroxyvitamin D 3 treatment in human endometrial stromal cells

doi: 10.64898/2026.05.06.723064

Figure Lengend Snippet: (A) The number of overlapped peaks between ATAC-seq and publicly available CUT&RUN. (B) Enriched binding motifs from cistromic modification peaks within the open chromatin areas in T-HESCs by HOMER de novo analysis. (C) Venn diagram showing overlap of genes annotated to ATAC-seq peaks in vehicle- and 1,25(OH) 2 D 3 -treated T-HESCs. Peaks were annotated to the nearest gene within 100 kb of the transcription start site. A total of 19,789 genes were associated with open chromatin regions in vehicle-treated cells, whereas 19,114 genes were associated with open chromatin regions in 1,25(OH) 2 D 3 -treated cells. Most annotated genes were shared between conditions, with 18,322 genes common to both datasets. (D) Overlap between 1,25(OH) 2 D 3 -responsive DEGs and genes associated with open chromatin regions in vehicle- or 1,25(OH) 2 D 3 -treated cells. Among 626 DEGs, 540 overlapped with vehicle-associated open chromatin genes, whereas 530 overlapped with 1,25(OH) 2 D 3 -associated open chromatin genes. The high proportion of overlap indicates that most ligand-responsive transcripts are associated with nearby accessible chromatin regions present in either condition. (E) Ingenuity Pathway Analysis (IPA) of the 530 differentially expressed genes associated with open chromatin regions in 1,25(OH) 2 D 3 -treated T-HESCs. Causal network analysis identified VDR as the most significant predicted activated master regulator. (F) IPA canonical pathway analysis of the same gene set revealed enrichment of pathways related to transcriptional regulation, immune-associated signaling, and vitamin D receptor activity.

Article Snippet: hTERT-immortalized endometrial stromal cells (T-HESCs) were purchased from the American Type Culture Collection (ATCC; Cat. CRL-4003, VA, USA).

Techniques: Binding Assay, Modification, Activity Assay

Journal: bioRxiv

Article Title: Increased chromatin accessibility following 1α,25-dihydroxyvitamin D 3 treatment in human endometrial stromal cells

doi: 10.64898/2026.05.06.723064

Figure Lengend Snippet: (A) Summarized transcript information on the overlap between DARs. (B) Upregulated DARs overlap with 1,25(OH) 2 D 3 DEGs, and downregulated DARs overlap with 1,25(OH) 2 D 3 DEGs. (C) Representative ligand-responsive accessible regions selected for locus-specific quantitative validation. Corresponding validation of the enhancer peak region of GPAT3 and MAMDC2. (D) Proposed working model of 1,25(OH) 2 D 3 -dependent chromatin-transcription coupling in T-HESCs. 1,25(OH) 2 D 3 reinforces a permissive chromatin landscape in human endometrial stromal cells.

Article Snippet: hTERT-immortalized endometrial stromal cells (T-HESCs) were purchased from the American Type Culture Collection (ATCC; Cat. CRL-4003, VA, USA).

Techniques: Biomarker Discovery