Journal: Redox Biology

Article Title: LDB1 represses fetal hemoglobin expression by enhancing BCL11A transcription

doi: 10.1016/j.redox.2026.104070

Figure Lengend Snippet: The LDB1 complex containing LMO2 and GATA1 directly enhances the transcription of the Bcl11a gene. ( A-E ) ChIP-qPCR assays were performed to assess the occupancy of putative LDB1 complex component proteins, including LDB1, LMO2, GATA1, TAL1, and E2A, at the Bcl11a locus in E14.5 WT and KO FLCs. n = 3. Control, isotype control Ab; LDB1, anti-LDB1 Ab; LMO2, anti-LMO2 Ab; GATA1, anti-GATA1 Ab; TAL1, anti-TAL1 Ab, E2A, anti-E2A Ab; WT, E14.5 FLCs; KO, E14.5 KO FLCs. Statistical significance was assessed by one-way ANOVA with Tukey HSD analysis. Mean values not sharing the same superscript letter ( a, b ) differ significantly at P < 0.05. ( F and G ) Dual-luciferase reporter assays were performed using the 210 bp region covering primer #3 ( Bcl11a intron #3) or the 225 bp region covering primer #4 ( Bcl11a intron #4) at Bcl11a locus. The relative luciferase activity of each construct containing either Bcl11a intron #3 or Bcl11a intron #4 was measured in HEK293T cells transfected with the indicated combination of each expression vector encoding Ldb1 , Lmo2 , or Gata1 cDNA. The schematic diagrams of each cloned enhancer region ( F ) and the results of the luciferase reporter assays ( G ) are presented. Ldb1 , expression vector encoding Ldb1 cDNA; Lmo2 , expression vector encoding Lmo2 cDNA; Gata1 , expression vector encoding Gata1 cDNA; ◯, transfected cells; ☓, untransfected cells. n = 5. Statistical significance was assessed by one-way ANOVA with Tukey HSD analysis. Mean values not sharing the same superscript letter ( a, b, c ) differ significantly at P < 0.05. ( H and I ) ChIP-qPCR assays were performed to assess LDB1 occupancy at the human BCL11A locus in K562 cells. The locations of each indicated primer set within the BCL11A locus are shown in the schematic diagram ( H ) along with the corresponding ChIP-qPCR results ( I ). n = 3. Mock K562, empty vector transfected K562 cells; KO K562, human LDB1 knockout K562 cells; Control, isotype control Ab; LDB1, anti-LDB1 Ab. Statistical significance was assessed by two-tailed Student's t -test. ∗ P < 0.05. All data are presented as the mean ± SEM.

Article Snippet: The lysates of E14.5 FLCs or K562 cells (ATCC CCL-243) were prepared for the ChIP-qPCR assay following established protocols [ ].

Techniques: ChIP-qPCR, Control, Luciferase, Activity Assay, Construct, Transfection, Expressing, Plasmid Preparation, Clone Assay, Knock-Out, Two Tailed Test

Journal: Journal of Bone Oncology

Article Title: Doxorubicin enhances adipogenesis in an FGF2-dependent manner and induces a tumour-promoting secretory phenotype

doi: 10.1016/j.jbo.2026.100754

Figure Lengend Snippet: Tumour cell proliferation is increased by culture with or media transfer from doxorubicin-treated adipogenic differentiated MSC and is dependent in part on FGF2. A) Proliferation of PC3 cells was assessed at 24 h, 48 h and 72 h following I) treatment with conditioned media from doxorubicin-treated BMA or II) direct seeding on top of doxorubicin-treated BMA. B) Proliferation of PC3 following direct co-culture with day 12 adipogenic differentiated MSC that were treated at day 9 of adipogenic differentiation with vehicle control or 25 nM or 50 nM doxorubicin. PC3 cells were enumerated at 24 h, 48 h and 72 h following addition to adipogenic differentiated MSC. Proliferation of PC3 treated with 30% conditioned media isolated at C) day 10 or D) day 12 from differentiated MSC treated at day 9 of adipogenic differentiation with vehicle control or 25 nM doxorubicin. E) Proliferation of PC3 in response to treatment with 30% conditioned media isolated at day 10 from adipogenic differentiating MSC following treatment at day 9 with 25 nM doxorubicin and siRNA targeting FGF2 or non-targeting control siRNA. F) Conditioned media was collected at day 10 from differentiating MSC/BMA cells treated with vehicle or 25 nM doxorubicin at day 9 of the differentiation process. K562 or 4T1 tumour cells were treated with 30% adipocyte conditioned media and were imaged and counted 48 h after addition of conditioned media. Mean cell count normalized to vehicle control is presented. In all graphs, mean cell count is normalized to cell count at 0 h prior to addition of conditioned media. Graphs show the mean ± SEM, using the statistical test two-way ANOVA in Figure B) and one-way ANOVA in Figure C), D) and E), (*p < 0.05, ***p < 0.001; ****p < 0.0001), n = 3 biological replicates each with 3 technical replicates.

Article Snippet: PC3 prostate tumour cells (ATCC CRL-1435), 4T1 breast tumour cells (CRL-2539) and K562 leukemia tumour cells (CCL-243) were obtained from the American Type Culture Collection (ATCC, Manassas, VA).

Techniques: Co-Culture Assay, Control, Isolation, Cell Characterization

Journal: Molecular Therapy Oncology

Article Title: Survivin/BIRC5-derived peptide disrupts survivin dimerization and cell division and induces multifaceted anti-cancer effects

doi: 10.1016/j.omton.2025.201123

Figure Lengend Snippet: Cell death induction by 1H13-survivin/BIRC5-derived peptides targeted to the cytoplasm, mitochondria, or nucleus (A) The sequences added to the 1H13-BIRC5 peptide to target it to the: (1) cytoplasm, (2) mitochondria, or (3) nucleus. The underlined sequence represents the 1H13 peptide; in red, amino acids indicate D-amino acid substitutions. (B) A549 cells were incubated for 90 min with 5 μM FITC-labeled, nucleus-targeted peptide IH13-Nuc, and were also IF-stained with anti--SMAC, anti-IP3R or anti-GM130 antibodies, and stained with DAPI to visualize the mitochondria, ER, Golgi, and nucleus, respectively. Confocal microscope images are shown, with white arrows indicating peptide co-localization with the mitochondria (SMAC). Orange and yellow arrows indicate peptide presence in the nucleus and cytosol, respectively. (C) A549 cells were incubated with the mitochondria- or nucleus-targeted 1H13-BIRC5-derived peptide for 24 h in serum-free medium, followed by a cell proliferation assay using the SRB method. (D and E) Apoptotic cell death as induced in A549 cells following incubation for 24 h with the nucleus-targeted peptide (2/3D-1H13-Nuc) in the presence or absence of the indicated concentrations of the peptides in serum-free medium and subjected to FITC–annexin V/PI staining, followed by a flow cytometry analysis. Representative histograms for control and selected peptide concentration (D) and analysis of early and late apoptotic stages are shown (E). (F and G) Cell death as induced by 2/3D-1H13-Nuc in different cell lines, A549, SH-SY5Y, U-87MG, PC-3, and HUV-EC-C (F) or Jurkat, K562, and KMH2-LC (G) were treated with the indicated concentrations of the peptide for 24 h, then subjected to cell death analysis using propidium iodide (PI) staining and flow cytometry. (H and I) A549 cells were seeded at a density of 2 × 10 5 cells per well in a 12-well plate. After 24 h, the cells were transfected with 2 μg of a pCMV3-survivin expression plasmid (HG10356-UT, Sino Biological, China) or with an empty pCMV3 plasmid (control) using JetPrime transfection reagent (Polyplus, France), following the manufacturer’s instructions. Twenty-four hours post-transfection, the cells were re-seeded at 1 × 10 5 cells per well in a 12-well plate. After another 24 h, the culture medium was replaced with serum-free medium, and the cells were treated with the indicated concentrations of the 2/3D-1H13-Nuc peptide. Survivin overexpression levels were assessed by immunoblotting (H), and cell death was analyzed by propidium iodide (PI) staining followed by FACS analysis (I). Results represent the means ± SEM ( n = 3).

Article Snippet: A549 (human lung adenocarcinoma epithelial), MDA-MB-231 (human breast cancer), PC-3 (human prostate adenocarcinoma epithelial), U-87MG (human glioblastoma), SH-SY5Y (human neuroblastoma), HeLa (human cervix adenocarcinoma), Jurkat (human acute T cell leukemia), K562 (human chronic myelogenous leukemia), and KMH2-LC (human anaplastic thyroid carcinoma) cell lines were obtained from the American type culture collection (ATCC, Manassas, VA) and maintained according to ATCC instructions.

Techniques: Derivative Assay, Sequencing, Incubation, Labeling, Staining, Microscopy, Proliferation Assay, Flow Cytometry, Control, Concentration Assay, Transfection, Expressing, Plasmid Preparation, Over Expression, Western Blot