human endometrial stromal cells 12z  (Procell Inc)

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: Cellular & Molecular Biology Letters

Article Title: HK2-driven histone H3K18 lactylation promotes stromal cell senescence and decidualization deficiency in URSA via CUX1-mediated SASP factor transcription

doi: 10.1186/s11658-026-00879-y

Figure Lengend Snippet: Glycolysis related lactate could accounts for stromal cell senescence and decidualization deficiency in URSA. A , B Total ion chromatogram of metabolites between stromal cells derived from control and URSA group ( A : positive ion model; B : negative ion model); C , D PCA score plot between stromal cells derived from control and URSA group, showing that samples in the groups were closely clustered to one another ( C : positive ion model; D : negative ion model); E Volcano plot of differential metabolites between stromal cells of control and URSA group. Red dots represent upregulated metabolites in the URSA group, while blue dots represent downregulated metabolites; F Heat map of differential metabolites between stromal cells of control and URSA group. The upregulated metabolites are marked in red, while the downregulated metabolites levels are presented in blue; G Bubble map of the impact values of metabolic pathways, where the horizontal coordinate is the impact value enriched into different metabolic pathways and the vertical coordinate is the enrichment pathway. The size of the dots indicates the corresponding number of metabolites on the pathway. The color of the dot reflects the P value, where the yellower the color, the smaller the P value, and the bluer the color, the larger the P value; H The levels of lactic acid between stromal cells derived from control and URSA group on the basis of global untargeted metabolomics; I The levels of lactic acid detected by lactate content assay kit in stromal cells between control and URSA group ( n = 6 per group); J The correlation of the fluorescence intensity between lactate and p16, p21, respectively; K The experimental plan outline of in vitro decidualization of T-hESCs by using 8-Br-cAMP and MPA with or without NaLa incubation; L The T-hESCs were examined for cell senescence by staining for SA-β-gal activity. Scale bar: 100 µm ( n = 3 per group); M , N Immunofluorescence and western blot to detect the level of p16 (M), p21 (N), with or without NaLa incubation (confocal laser fluorescence microscopy, scale bar: 20 µm) ( n = 3–5 per group); O , P Immunofluorescence to detect the level of γH2AX (O) and F-actin (P), with or without NaLa incubation (confocal laser fluorescence microscopy, scale bar: 20 µm); Q mRNA levels of PRL and IGFBP1 in different groups were verified by qRT-PCR ( n = 6 per group). * P < 0.05, ** P < 0.01, by two-tailed Student’s t -test

Article Snippet: Human: telomerase-immortalized human endometrial stromal cells (T-hESC) , ATCC , Cat. no. CRL-4003.

Techniques: Derivative Assay, Control, Fluorescence, In Vitro, Incubation, Staining, Activity Assay, Immunofluorescence, Western Blot, Microscopy, Quantitative RT-PCR, Two Tailed Test

Journal: Cellular & Molecular Biology Letters

Article Title: HK2-driven histone H3K18 lactylation promotes stromal cell senescence and decidualization deficiency in URSA via CUX1-mediated SASP factor transcription

doi: 10.1186/s11658-026-00879-y

Figure Lengend Snippet: H3K18 lactylation mediated stromal cell senescence and decidualization deficiency in URSA. A – F T-hESCs were randomly divided into Con + NC plasmid, Con + HK2 OE, and Con + HK2 OE + oxamate group and were induced for decidualization. The confocal laser fluorescence microscopy was utilized to assess the abundance of H3K18la of H3K18la ( A ), p16 ( B ), p21 ( C ), γH2AX ( D ), and F-actin ( E ). And the mRNA levels of PRL and IGFBP1 were verified by qRT-PCR in the three mentioned above groups ( F ); G – L T-hESCs were randomly divided into CoCl 2 + scramble siRNA, CoCl 2 + HK2 siRNA, and CoCl 2 + HK2 siRNA + NaLa group, and were then induced for decidualization. The confocal laser fluorescence microscopy was utilized to assess the level of H3K18la ( G ), p16 ( H ), p21 ( I ), γH2AX ( J ), and F-actin ( K ). The mRNA levels of PRL and IGFBP1 in different groups were verified by qRT-PCR ( L ). * P < 0.05, ** P < 0.01, by repeated-measures one-way ANOVA followed by post hoc Dunnett’s multiple-comparisons test ( n = 3–4 per group). Scale bar: 20 µm

Article Snippet: Human: telomerase-immortalized human endometrial stromal cells (T-hESC) , ATCC , Cat. no. CRL-4003.

Techniques: Plasmid Preparation, Fluorescence, Microscopy, Quantitative RT-PCR

Journal: Cellular & Molecular Biology Letters

Article Title: HK2-driven histone H3K18 lactylation promotes stromal cell senescence and decidualization deficiency in URSA via CUX1-mediated SASP factor transcription

doi: 10.1186/s11658-026-00879-y

Figure Lengend Snippet: H3K18la promotes stromal cell senescence by promoting CUX1 transcription. A Schematic of stromal cell isolation from healthy controls and individuals with URSA; B The binding density of H3K18la visualized by deepTools: the heatmap presents the CUT&Tag tag counts on the different H3K18la binding peaks in stromal cells between controls and women with URSA, ordered by signal strength; C Genome-wide distribution of upregulated and downregulated H3K18la-binding peaks in stromal cells between controls and women with URSA; D GO-BP analysis of the H3K18la binding peaks at promoter genes; E The intersection of senescence related genes with upregulated DEGs in URSA marked with H3K18la at their promoter regions; F qRT-PCR assays monitoring the expression of the CUX1, PAM, and RAPGEF4 in stromal cells between controls and women with URSA ( n = 12 per group); G Expression and co-localization of H3K18la and CUX1 in stromal cells of control and URSA groups analyzed by immunofluorescence-based three-dimensional reconstruction technique. Scale bar: 20 µm; H The correlation between of CUX1 and global lactylation (left panel) and H3K18la (right panel) levels in decidual tissue of patients with URSA; I The upper panel is an IGV snapshot of H3K18la tracks across the CUX1 gene locus. The lower panel shows a zoomed-in view of H3K18la ChIP-seq signals at the promoter region of CUX1. The red box in the lower panel indicates the peaks closest to the transcription start site (TSS) region of the CUX1 gene; J H3K18la levels at the CUX1 promoter regions in control and URSA stromal cells analyzed by ChIP-qPCR assay; Normal rabbit immunoglobulin G (IgG) was used as a negative control. Data were normalized to input and are expressed as mean fold enrichment over input relative to an IgG control ( n = 3 per group); K H3K18la levels at the CUX1 promoter regions analyzed by ChIP-qPCR assay in T-hESC versus T-hESC treated with 25 mM NaLa ( n = 3 per group); L qRT-PCR assays monitoring expression of the CUX1 in T-hESC treated with different concentrations of NaLa (0, 5, 10, and 25 mM) for 24 h ( n = 6 per group); M Western blot to detect the level of CUX1, with or without 25 mM NaLa incubation ( n = 5 per group); N Schematic proposing a mechanism of H3K18la promoting stromal cell senescence via targeting CUX1 transcription in URSA.* P < 0.05, ** P < 0.01, by two-tailed Student’s t -test. L * P < 0.05, by repeated-measures one-way ANOVA followed by post hoc Tukey multiple-comparisons test

Article Snippet: Human: telomerase-immortalized human endometrial stromal cells (T-hESC) , ATCC , Cat. no. CRL-4003.

Techniques: Cell Isolation, Binding Assay, Genome Wide, Quantitative RT-PCR, Expressing, Control, Immunofluorescence, ChIP-sequencing, ChIP-qPCR, Negative Control, Western Blot, Incubation, Two Tailed Test

Journal: Cellular & Molecular Biology Letters

Article Title: HK2-driven histone H3K18 lactylation promotes stromal cell senescence and decidualization deficiency in URSA via CUX1-mediated SASP factor transcription

doi: 10.1186/s11658-026-00879-y

Figure Lengend Snippet: CUX1 may promote stromal cell senescence by activating promoter activity of SASP factors. A Heat map of differentially expressed genes in T-hESCs treated with CUX1 OE plasmid versus NC plasmid group during induced decidualization. The upregulated genes are marked in red, and the downregulated genes are presented in blue; B Volcano plot illustrating the differentially expressed genes in T-hESCs following treatment with the CUX1 overexpression (OE) plasmid, in comparison with the negative control (NC) plasmid group, during the process of induced decidualization; C KEGG pathway enrichment analysis on differentially expressed genes between T-hESCs treated with CUX1 OE plasmid versus NC plasmid group during induced decidualization; D – I ChIP-qPCR analysis on the enrichment status of CUX1 at the promoter of EDA ( D ), CXCL1 ( E ), CXCL3 ( F ), PRKCB ( G ), TNF-α ( H ), and TNFSF13B ( I ) in T-hESCs treated with CUX1 OE plasmid versus NC plasmid group during induced decidualization. * P < 0.05, by two-tailed Student’s t test ( n = 3 per group). ns: not significant

Article Snippet: Human: telomerase-immortalized human endometrial stromal cells (T-hESC) , ATCC , Cat. no. CRL-4003.

Techniques: Activity Assay, Plasmid Preparation, Over Expression, Comparison, Negative Control, ChIP-qPCR, Two Tailed Test

Journal: Cellular & Molecular Biology Letters

Article Title: HK2-driven histone H3K18 lactylation promotes stromal cell senescence and decidualization deficiency in URSA via CUX1-mediated SASP factor transcription

doi: 10.1186/s11658-026-00879-y

Figure Lengend Snippet: H3K18la facilitated stromal cell senescence via CUX1-mediated SASP factor transcription in URSA. A – D T-hESCs were randomly divided into CUX1 scramble, NaLa + CUX1 scramble, and NaLa + CUX1 siRNA group, and were induced for decidualization. qRT-PCR was utilized to detect the mRNA levels of CXCL1, CXCL3, and TNF-α ( A ); The confocal laser fluorescence microscopy was utilized to assess the p16, p21, and γH2AX levels ( B ) (scale bar: 20 µm); The stromal cell senescence was assessed by SA-β-gal staining ( C ) (scale bar: 100 µm); The confocal laser fluorescence microscopy was utilized to assess the F-actin level ( D ) (scale bar: 20 µm); Besides, PRL and IGFBP1 in different groups were verified by qRT-PCR ( D ) ( n = 3–6 per group); E – I T-hESCs were randomly divided into CoCl 2 + CUX1 NC plasmid, CoCl2 + oxamate + CUX1 NC plasmid, and CoCl 2 + oxamate + CUX1 OE plasmid group, and were induced for decidualization. The confocal laser fluorescence microscopy was utilized to detect H3K18la ( E ) (scale bar: 20 µm) and CUX1 ( F ) (scale bar: 20 µm) in the three above-mentioned groups; Besides, the mRNA levels of CXCL1, CXCL3, and TNF-α were verified by qRT-PCR ( G ); Confocal laser fluorescence microscopy was utilized to assess the p16, p21, and γH2AX levels ( H ) (scale bar: 20 µm), as well as the F-actin level ( I ) (scale bar: 20 µm); qRT-PCR was utilized to to assess the mRNA level of PRL and IGFBP1 ( I ) ( n = 3–6 per group). * P < 0.05, ** P < 0.01, by two-tailed Student’s t -test or repeated-measures one-way ANOVA followed by post hoc Dunnett’s multiple-comparisons test

Article Snippet: Human: telomerase-immortalized human endometrial stromal cells (T-hESC) , ATCC , Cat. no. CRL-4003.

Techniques: Quantitative RT-PCR, Fluorescence, Microscopy, Staining, Plasmid Preparation, Two Tailed Test