Journal: Translational Oncology

Article Title: REST-driven upregulation of SFXN3 promotes AML progression via Wnt/β-catenin activation and confers decitabine resistance

doi: 10.1016/j.tranon.2026.102705

Figure Lengend Snippet: Effects of SFXN3 Knockdown on Proliferation, Apoptosis, and Signaling Pathways in AML Cells. (A–C) qRT-PCR and Western blot analyses were used to measure SFXN3 expression levels in various leukemia cell lines (THP-1, KG-1, U937, K562) and in normal bone marrow stromal cells (HS-5). (D) Two independent shRNAs (sh-SFXN3–1 and sh-SFXN3–2) were used to knock down SFXN3 expression in THP-1 and KG-1 cells. Western blot was performed to assess the knockdown efficiency and specificity. (E) Quantification of SFXN3 knockdown efficiency by different shRNAs. (F) CCK-8 cell proliferation assays were conducted to evaluate the effects of SFXN3 knockdown on cell growth dynamics over time. (G) EdU incorporation assays were used to assess DNA synthesis activity, indirectly reflecting cellular proliferation, and to compare differences between knockdown and control groups, (bar=50ųm). (H) Western blot analysis of key cell cycle regulatory proteins (CDK4, CDK6, P27, and P21) to investigate the potential mechanism by which SFXN3 affects cell cycle progression. (I) TUNEL assays were used to evaluate apoptosis levels in the knockdown versus control groups, assessing the role of SFXN3 in apoptosis suppression, (bar=50ųm). (J) Western blot analysis of pro-apoptotic proteins (BAX and BAK) and anti-apoptotic proteins (Bcl-2 and Bcl-xl) in THP-1 and KG-1 cells following SFXN3 knockdown. (K) Correlation analysis between SFXN3 expression and key proteins in the Wnt/β-Catenin signaling pathway. (L) Subcellular fractionation followed by Western blotting was performed to assess β-catenin nuclear translocation. Cytoplasmic (Cyto) and nuclear (Nuc) fractions were probed for β-catenin, with β-actin (cytoplasmic marker) and Histon H3 (nuclear marker) used to confirm fractionation quality. Data are presented as mean ± SD. from three independent experiments ( n = 3). One-way ANOVA was used in (A, B, E), and two-way ANOVA was used in (F). *, p < 0.05.

Article Snippet: SKL2001 was used to activate the Wnt/β-Catenin signaling route (Selleck Chemicals).

Techniques: Knockdown, Protein-Protein interactions, Quantitative RT-PCR, Western Blot, Expressing, CCK-8 Assay, DNA Synthesis, Activity Assay, Control, TUNEL Assay, Fractionation, Translocation Assay, Marker

Journal: Translational Oncology

Article Title: REST-driven upregulation of SFXN3 promotes AML progression via Wnt/β-catenin activation and confers decitabine resistance

doi: 10.1016/j.tranon.2026.102705

Figure Lengend Snippet: The Wnt/β-Catenin Pathway Agonist SKL2001 Reverses the Effects of SFXN3 Knockdown on Leukemia Cell Proliferation and Apoptosis. (A) Western blot analysis of SFXN3 protein expression following SFXN3 knockdown and treatment with SKL2001, to assess whether SKL2001 significantly modulates SFXN3 expression. (B) CCK-8 assays were performed to evaluate whether SKL2001 could reverse the inhibitory effects of SFXN3 knockdown on the proliferation of THP-1 and KG-1 leukemia cells. (C) EdU staining assays were used to assess DNA synthesis activity, analyzing the ability of SKL2001 to restore proliferation suppressed by SFXN3 knockdown, (bar=50 ųm). (D) Quantitative analysis of EdU fluorescence intensity to evaluate DNA replication across different treatment groups. (E) Western blot analysis of cell cycle regulators CDK4, CDK6, Cyclin D1, and Cyclin E1 to determine whether SKL2001 rescues the expression of these proteins in SFXN3-silenced cells. (F) Western blot analysis of pro-apoptotic proteins (BAX, BAK) and anti-apoptotic proteins (Bcl-2, Bcl-xl) to confirm that SKL2001 mitigates the apoptosis-promoting effects of SFXN3 knockdown. (G) TUNEL assays were conducted to assess whether SKL2001 suppresses the enhanced apoptosis induced by SFXN3 knockdown, (bar=50ųm). (H) Quantification of TUNEL fluorescence intensity, reflecting apoptosis levels under different treatment conditions. (I) Subcellular fractionation followed by Western blotting was performed to assess β-catenin nuclear translocation. Cytoplasmic (Cyto) and nuclear (Nuc) fractions were probed for β-catenin, with β-actin (cytoplasmic marker) and Histon H3 (nuclear marker) used to confirm fractionation quality. Data are presented as mean ± SD. from at least three independent experiments. One-way ANOVA was used in (D, H), and two-way ANOVA was used in (B). *, p < 0.05; **, p < 0.01; ***, p < 0.001 vs. control or scramble group.

Article Snippet: SKL2001 was used to activate the Wnt/β-Catenin signaling route (Selleck Chemicals).

Techniques: Knockdown, Western Blot, Expressing, CCK-8 Assay, Staining, DNA Synthesis, Activity Assay, Fluorescence, TUNEL Assay, Fractionation, Translocation Assay, Marker, Control

Journal: Translational Oncology

Article Title: REST-driven upregulation of SFXN3 promotes AML progression via Wnt/β-catenin activation and confers decitabine resistance

doi: 10.1016/j.tranon.2026.102705

Figure Lengend Snippet: The REST–SFXN3 Axis Promotes Malignant Phenotypes in AML Cells via the Wnt/β-Catenin Signaling Pathway. (A) Western blot analysis of the effect of REST knockdown (sh-REST) on SFXN3 expression, and the reversal of this effect by SFXN3 overexpression. (B) CCK-8 assays assess the impact of sh-REST and SFXN3 overexpression on AML cell proliferation. (C) EdU incorporation assays evaluate the effects of sh-REST and SFXN3 overexpression on DNA synthesis activity in AML cells, (bar=50ųm). (D) Quantification of EdU-positive cells to compare DNA synthesis capacity across groups. (E) Western blot analysis of proliferation-related proteins CDK4, CDK6, Cyclin D1, and Cyclin E1 under sh-REST and SFXN3 overexpression conditions. (F) Band intensities were quantified using ImageJ software and normalized to the indicated internal controls. (G) TUNEL assays detect apoptotic cells after REST knockdown and SFXN3 overexpression, (bar=50ųm). (G) Quantitative analysis of apoptotic cells in THP-1 and KG-1 cell lines. (H) Quantification of TUNEL fluorescence intensity, reflecting apoptosis levels under different treatment conditions. (I) Western blot evaluation of pro-apoptotic proteins (BAX, BAK) and anti-apoptotic proteins (Bcl-2, Bcl-xl), demonstrating REST knockdown promotes apoptosis, which is reversed by SFXN3 overexpression. (J) Subcellular fractionation followed by Western blotting was performed to assess β-catenin nuclear translocation. Cytoplasmic (Cyto) and nuclear (Nuc) fractions were probed for β-catenin, with β-actin (cytoplasmic marker) and Histon H3 (nuclear marker) used to confirm fractionation quality. Data are presented as mean ± SD from three independent experiments ( n = 3).One-way ANOVA was used in (D, F,H), and two-way ANOVA was used in (B). **, p < 0.01; ***, p < 0.001.

Article Snippet: SKL2001 was used to activate the Wnt/β-Catenin signaling route (Selleck Chemicals).

Techniques: Western Blot, Knockdown, Expressing, Over Expression, CCK-8 Assay, DNA Synthesis, Activity Assay, Software, TUNEL Assay, Fluorescence, Fractionation, Translocation Assay, Marker

Journal: Translational Oncology

Article Title: REST-driven upregulation of SFXN3 promotes AML progression via Wnt/β-catenin activation and confers decitabine resistance

doi: 10.1016/j.tranon.2026.102705

Figure Lengend Snippet: Decitabine Suppresses AML Cell Proliferation and Promotes Apoptosis via SFXN3 Inhibition. (A) RT-PCR analysis of the effects of Gefitinib, Disulfiram, and Decitabine on SFXN3 mRNA expression. (B) Western blot analysis of SFXN3 protein levels following treatment with Gefitinib, Disulfiram, and Decitabine. (C) CCK-8 assay to calculate the IC50 values of Decitabine in THP-1 and KG-1 cells, identifying appropriate drug concentrations for subsequent experiments (D) CCK-8 assays were performed to evaluate AML cell viability at 6,12,24,48, and 72 h following treatment with 50 nm decitabine, thereby determining the optimal treatment duration. E) EdU incorporation assay evaluating the proliferation capacity of AML cells after Decitabine treatment, (bar=50ųm). (F) Western blot analysis of proliferation-related proteins (P21, P27, CDK4, and CDK6) following Decitabine treatment. (G) TUNEL staining to detect DNA fragmentation at the 3′-OH ends, marking apoptotic cells after Decitabine exposure, (bar=50ųm). (H) Western blot analysis of pro-apoptotic (e.g., BAX, BAK) and anti-apoptotic (e.g., Bcl-2, Bcl-xl) protein expression in response to Decitabine. (I) Western blot analysis of key components of the Wnt/β-Catenin signaling pathway after Decitabine treatment, revealing pathway inhibition. Subcellular fractionation followed by Western blotting was performed to assess β-catenin nuclear translocation. Cytoplasmic (Cyto) and nuclear (Nuc) fractions were probed for β-catenin, with β-actin (cytoplasmic marker) and Histon H3 (nuclear marker) used to confirm fractionation quality. n = 3,Error bars indicate mean ± SD; One-way ANOVA in (D, F); **, p < 0.01, *** p <0.001.

Article Snippet: SKL2001 was used to activate the Wnt/β-Catenin signaling route (Selleck Chemicals).

Techniques: Inhibition, Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot, CCK-8 Assay, TUNEL Assay, Staining, Fractionation, Translocation Assay, Marker

Journal: Journal of Orthopaedic Translation

Article Title: Silencing SFRP1 in bone mesenchymal stem cells alleviates pediatric B-ALL-driven bone loss by activating Wnt/β-catenin signaling

doi: 10.1016/j.jot.2026.101071

Figure Lengend Snippet: SFRP1 suppresses osteogenesis of B-ALL–derived BMSCs through the Wnt/β-catenin pathway. (A) Venn diagram showing overlap between upregulated mRNA and protein datasets. (B) Protein-protein interaction network of DEPs (red: upregulated; green: downregulated). (C) KEGG pathway enrichment analysis of DEGs. (D, E) Quantification of SFRP1 protein expression in hBMSCs from hCTR and hB-ALL patients. (F, G) Quantification of osteogenic proteins (COL1A1, ALP, RUNX2) in hBMSCs from hB-ALL with WAY-316606. (H, I) Quantification of SFRP1 protein expression in mBMSCs from mCTR and mB-ALL mice. (J, M) β-catenin and RUNX2 protein expression in mBMSCs with different treatments. (K, L) Immunofluorescence imaging and quantification of SFRP1 (red) in mouse trabecular bone. (N–P) Immunohistochemical analysis of β-catenin and RUNX2 expression in mouse trabecular bone (n = 3; ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, ∗∗∗∗ P < 0.0001).

Article Snippet: Selected sections were used for immunohistochemistry (IHC) to detect the expression of the osteogenic marker RUNX2 (Servicebio, China) and the Wnt signaling marker β-catenin (Servicebio, China), and immunofluorescence (IF) to detect the expression of the SFRP1 (Proteintech, China).

Techniques: Derivative Assay, Expressing, Immunofluorescence, Imaging, Immunohistochemical staining

Journal: Journal of Orthopaedic Translation

Article Title: Silencing SFRP1 in bone mesenchymal stem cells alleviates pediatric B-ALL-driven bone loss by activating Wnt/β-catenin signaling

doi: 10.1016/j.jot.2026.101071

Figure Lengend Snippet: SFRP1 inhibition enhances β-catenin expression and nuclear translocation (A, C) Immunofluorescence analysis and quantification of SFRP1 expression (red) in femoral sections of mB-ALL mice across treatment groups. (B, D) β-catenin expression and nuclear localization in femoral trabeculae across treatment groups (n = 5; ∗∗ P < 0.01, ∗∗∗ P < 0.001, ∗∗∗∗ P < 0.0001).

Article Snippet: Selected sections were used for immunohistochemistry (IHC) to detect the expression of the osteogenic marker RUNX2 (Servicebio, China) and the Wnt signaling marker β-catenin (Servicebio, China), and immunofluorescence (IF) to detect the expression of the SFRP1 (Proteintech, China).

Techniques: Inhibition, Expressing, Translocation Assay, Immunofluorescence

Journal: Journal of Orthopaedic Translation

Article Title: Silencing SFRP1 in bone mesenchymal stem cells alleviates pediatric B-ALL-driven bone loss by activating Wnt/β-catenin signaling

doi: 10.1016/j.jot.2026.101071

Figure Lengend Snippet: shSFRP1@Lipo-E7 targets BMSCs to regulate the SFRP1/β-catenin signaling pathway, promoting osteogenic differentiation of BMSCs and alleviating bone loss induced by B-ALL. (A) B-ALL cells induce abnormal elevation of SFRP1 in BMSCs, promoting adipogenic differentiation and disrupting the osteogenic-osteoclastic coupling balance, ultimately leading to bone loss. (B) Tail vein delivery of shSFRP1-loaded BMSC-targeted liposomes downregulates SFRP1 to enhance BMSC osteogenic differentiation, suppress adipogenic differentiation, and indirectly inhibit osteoclastogenesis by remodeling the bone microenvironment. (C) SFRP1 inhibit Wnt signaling by promoting the phosphorylation of β-catenin and reducing its nuclear translocation.

Article Snippet: Selected sections were used for immunohistochemistry (IHC) to detect the expression of the osteogenic marker RUNX2 (Servicebio, China) and the Wnt signaling marker β-catenin (Servicebio, China), and immunofluorescence (IF) to detect the expression of the SFRP1 (Proteintech, China).

Techniques: Liposomes, Phospho-proteomics, Translocation Assay

Journal: International Journal of Oncology

Article Title: Ephrin-B2 promotes gastric cancer growth by inhibiting apoptosis and regulating the cell cycle via the Wnt/β-catenin signaling pathway

doi: 10.3892/ijo.2025.5821

Figure Lengend Snippet: Wnt/β-catenin signaling pathway is responsible for EFNB2-mediated biological effects in gastric cancer cells. (A) Gene Expression Profiling Interactive Analysis and (B) Tumor Immune Estimation Resource databases were employed to explore the transcriptional correlation between EFNB2 and CTNNB1, GSK3β and downstream MYC (Spearman). Representative western blot bands and semi-quantification of protein expression levels of p-GSK3β, GSK3β, β-catenin and c-myc detected under the condition of EFNB2 (C) knockdown and (D) overexpression (one-way ANOVA for AGS cells; unpaired Student's test for HGC-27 cells). Representative western blot bands and semi-quantification of protein expression levels in the presence of (E) an agonist (CHIR99021) and (F) an inhibitor (DIF-3) of the Wnt/β-catenin signaling pathway in the EFNB2 knockdown or overexpression groups, respectively. Protein levels of p-GSK3β, GSK3β, β-catenin and c-myc were detected by western blotting (one-way ANOVA). Data are presented as the mean ± SD. * P<0.05, ** P<0.01, *** P<0.001 vs. sh-NC or vector group. ns, not significant; NC, negative control; DIF-3, differentiation-inducing factor-3; p-, phosphorylated; OE, overexpression vector; sh, short hairpin RNA; CTNNB1, catenin β1; TPM, transcript per million; EFNB2, ephrin-B2.

Article Snippet: For the rescue experiments, cells in designated groups were treated at 37°C for 24 h with either CHIR99021 (5 μ M; MedChemExpress) to activate the Wnt/β-catenin signaling cascade or DIF-3 (30 μ M; MedChemExpress) to inhibit the Wnt/β-catenin signaling cascade.

Techniques: Gene Expression, Western Blot, Expressing, Knockdown, Over Expression, Plasmid Preparation, Negative Control, shRNA

Journal: International Journal of Oncology

Article Title: Ephrin-B2 promotes gastric cancer growth by inhibiting apoptosis and regulating the cell cycle via the Wnt/β-catenin signaling pathway

doi: 10.3892/ijo.2025.5821

Figure Lengend Snippet: EFNB2 promotes the tumor growth of gastric cancer via the Wnt/β-catenin signaling pathway in vivo . (A) Tumors from the immunodeficient nude mice in the four experimental groups were excised and imaged. (B) Subcutaneous tumor growth (mm 3 ) was measured every 3-4 days following various treatments (one-way ANOVA). (C) Excised tumor masses (g) were weighed and compared across groups (one-way ANOVA). (D) Histopathological evaluation of xenograft tumors included H&E staining to examine tissue morphology, IHC analysis of EFNB2, β-catenin and Ki67 expression, and TUNEL staining for apoptotic cell evaluation. All images were captured at a magnification of ×200. (E) Quantification of IHC scores for EFNB2 and β-catenin, and the percentage of positive cells for Ki67 and TUNEL staining (Kruskal-Wallis H test for EFNB2 and β-catenin; one-way ANOVA for Ki67 and TUNEL staining). Data are presented as the mean ± SD. * P<0.05, ** P<0.01, *** P<0.001. ns, not significant; EFNB2, ephrin-B2; DIF-3, differentiation-inducing factor-3; OE, overexpression vector; IHC, immunohistochemical.

Article Snippet: For the rescue experiments, cells in designated groups were treated at 37°C for 24 h with either CHIR99021 (5 μ M; MedChemExpress) to activate the Wnt/β-catenin signaling cascade or DIF-3 (30 μ M; MedChemExpress) to inhibit the Wnt/β-catenin signaling cascade.

Techniques: In Vivo, Staining, Expressing, TUNEL Assay, Over Expression, Plasmid Preparation, Immunohistochemical staining