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: CUX1 overexpression plasmid (OE: pSLenti-CMV-CUX1-3xFLAG-PGK-Puro-WPRE) , Obio Technology , N/A.

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: CUX1 overexpression plasmid (OE: pSLenti-CMV-CUX1-3xFLAG-PGK-Puro-WPRE) , Obio Technology , N/A.

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: CUX1 overexpression plasmid (OE: pSLenti-CMV-CUX1-3xFLAG-PGK-Puro-WPRE) , Obio Technology , N/A.

Techniques: Quantitative RT-PCR, Fluorescence, Microscopy, Staining, Plasmid Preparation, 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 knockdown results in cell senescence, decidualization deficiency, and embryo absorption in URSA murine model. A Schematic representation of the CUX1 or scramble AAV injection for in vivo CUX1 manipulation in URSA murine model; B ELISA assay detected the CXCL1, CXCL3, and TNF-α levels in stromal cells of URSA murine model treated with CUX1 or scramble shRNA ( n = 8 per group); C Double-immunofluorescence staining for the p16 and p21 levels in stromal cells derived from URSA murine model treated with CUX1 or scramble shRNA. Scale bar: 50 µm (left), 10 µm (right); D p16 and p21 expression levels verified by Western blot in URSA murine model treated with CUX1 or scramble shRNA ( n = 4 per group); E PRL and IGFBP1 mRNA expression verified by qRT-PCR in URSA murine model treated with CUX1 or scramble shRNA ( n = 6 per group); F Mice embryo absorption in URSA murine model treated with CUX1 or scramble shRNA ( n = 6 per group); G Positron emission tomography (PET)–computed tomography (CT) imaging to identify successful pregnancy in URSA murine model treated with CUX1 or scramble shRNA. * P < 0.05, ** P < 0.01, by repeated-measures one-way ANOVA followed by post hoc Tukey multiple-comparisons test. ns: not significant

Article Snippet: CUX1 overexpression plasmid (OE: pSLenti-CMV-CUX1-3xFLAG-PGK-Puro-WPRE) , Obio Technology , N/A.

Techniques: Knockdown, Injection, In Vivo, Enzyme-linked Immunosorbent Assay, shRNA, Double Immunofluorescence Staining, Derivative Assay, Expressing, Western Blot, Quantitative RT-PCR, Positron Emission Tomography, Computed Tomography, Imaging