Journal: bioRxiv

Article Title: BCLAF1 links RNA splicing to ATF4-dependent metabolic adaptation in acute myeloid leukemia

doi: 10.64898/2026.01.19.700325

Figure Lengend Snippet: A. BCLAF1 expression levels across cancer types from the TCGA dataset. B. BCLAF1 expression in LAML patient samples (n = 173) and normal hematopoietic tissues (n = 70), shown as log 2 (TPM + 1). C. Growth competition experiments showing the percentage of GFP-positive cells between day 2 and day 15 following infection with LentiCRISPRv2GFP constructs expressing individual sgRNAs targeting the indicated genes. Data represent mean ± SEM of three biological replicates (*p < 0.05, ***p < 0.001; two-way ANOVA). D. Immunoblot analysis of BCLAF1 protein levels in shCONTROL and shBCLAF1 cells treated or not with DOX for 96 hours. β-tubulin was used as a loading control. E. Flow cytometry analysis showing the percentage of CD11b-positive cells in shCONTROL and shBCLAF1 cells treated with DOX for 96h. Data represent mean ± SEM of three biological replicates (*p < 0.05, ***p < 0.001; unpaired t-test). F. Dot plot of GO terms enriched among genes up-regulated in BCLAF1-low versus BCLAF1-high patient samples from the BEAT-AML cohort. GeneRatio indicates the proportion of significantly upregulated genes associated with each GO term. Dot size reflects the number of genes per term; color denotes adjusted p-value. G. Kaplan-Meier curve shows survival of mice transplanted with MLL-AF9 expressing Bclaf1 f/f (black) or Vav-Cre:Bclaf1 f/f (blue) HSPCs. Briefly, HSPCs from CD45.2 + Bclaf1 f/f or Vav-Cre:Bclaf1 f/f mice were transduced with retrovirus co-expressing MLL-AF9 and GFP then transplanted with 300,000 CD45.1 + recipient bone marrow cells into lethally-irradiated CD45.1 + recipient mice (schematic in Figure S1G). Leukemic burden was assessed by percentage of GFP + cells in peripheral blood. Mice were euthanized once peripheral blood had ≥ 80% leukemia. Inset shows median survival. Bclaf1 f/f , n=23; Vav-Cre:Bclaf1 f/f , n=24. H. Primary Bclaf1 f/f AMLs isolated from mice in G were transduced with retroviral vector expressing Thy1.1 (empty control, black) or Thy1.1 and Cre recombinase (blue) then expanded as mixed co-cultures with nontransduced cells. Bar graphs show percentage of transduced cells (Thy1.1 + ) at day 7 of mixed co-culture relative to day 0 transduction efficiency. Data repesent mean ± SD of 3 independent experiments (**p ≤ 0.01; unpaired t-test). I. Primary Bclaf1 f/f AMLs isolated from mice in G were transduced with retroviral vectors as in D. Thy1.1 + cells were enriched by flow cytometric sorting and cultured in complete methocult media. Total colony forming units were quantitated at day 7 of culture. Data repesent mean ± SD of 3 independent experiments (*p ≤ 0.05; unpaired t-test).

Article Snippet: These samples were stained using CD11b surface marker and Annexin V kit (Annexin V-FITC Kit, Miltenyi Biotec).

Techniques: Expressing, Infection, Construct, Western Blot, Control, Flow Cytometry, Transduction, Irradiation, Isolation, Retroviral, Plasmid Preparation, Co-Culture Assay, Cell Culture

Journal: bioRxiv

Article Title: BCLAF1 links RNA splicing to ATF4-dependent metabolic adaptation in acute myeloid leukemia

doi: 10.64898/2026.01.19.700325

Figure Lengend Snippet: A. Immunoblot analysis of BCLAF1 and ATF4 protein levels in shCONTROL and shBCLAF1 cells treated with DOX for 96 hours. β-tubulin was used as a loading control. B. Immunoblot analysis of BCLAF1 and ATF4 protein levels in MLL-AF9-expressing AML cells of indicated genotypes. GAPDH was used as a loading control. C. Immunoblot analysis of BCLAF1 and ATF4 protein levels in shCONTROL and shATF4 cells treated with DOX for 96 hours. β-tubulin was used as a loading control. D. Cell proliferation assay (CellTiter-Blue-based) in shCONTROL and shATF4 cells treated or not with DOX for 96 hours, followed by 96 additional hours in culture. Data represent mean ± SEM of three biological replicates (*p < 0.01; paired t-test). E. Flow cytometry analysis showing the percentage of CD11b-positive cells in shCONTROL and shATF4 cells treated with DOX for 96h. Data represent mean ± SEM of four biological replicates (***p < 0.001; unpaired t-test). F. Annexin V/PI flow cytometry analysis showing the distribution of live, early apoptotic, late apoptotic, and necrotic cells following 96h of DOX treatment. Data represent mean ± SEM of three biological replicates (two-way ANOVA). G. RT-qPCR experiments showing relative mRNA levels of the indicated transcripts in shCONTROL and shATF4 cells treated with DOX for 96 hours. Data were normalized to GAPDH and represent mean ± SEM of three biological replicates (**p < 0.01; unpaired t-test). H. Immunoblot analysis of ATF4, SLC7A5, ASS1, PHGDH and PYCR1 protein levels in shCONTROL and shATF4 cells treated or not with DOX for 96 hours. β-tubulin was used as a loading control.

Article Snippet: These samples were stained using CD11b surface marker and Annexin V kit (Annexin V-FITC Kit, Miltenyi Biotec).

Techniques: Western Blot, Control, Expressing, Proliferation Assay, Flow Cytometry, Quantitative RT-PCR

Journal: Biology

Article Title: Force-Dependent Presence of Senescent Cells Expressing Vascular Endothelial Growth Factor During Orthodontic Tooth Movement

doi: 10.3390/biology15020187

Figure Lengend Snippet: Dynamic changes in angiogenesis induced by OTM. ( A ) Representative CD31 immunofluorescence staining images. CD31 staining without DAPI ( left ) and merged images with DAPI ( right ) are shown. ( B ) Quantitative analysis of CD31 + area in PDL. ( C ) Representative CD31/vascular endothelial growth factor (VEGF) double immunofluorescence staining images. CD31/VEGF staining without DAPI ( left ) and merged images with DAPI ( right ) are shown. ( D ) Quantitative analysis of VEGF + area in PDL. Scale bars: 100 µm (low magnification), 10 µm (high magnification). Data are mean ± SD (n = 4). ns: not significant, * p < 0.05; ** p < 0.01; *** p < 0.001; Welch’s ANOVA with Games–Howell post hoc test. PDL: periodontal ligament, AB: alveolar bone.

Article Snippet: CD31 , Endothelial cell marker , Bioss Antibodies (Shanghai, China) , bs-0195R , BF647 , 1:100.

Techniques: Immunofluorescence, Staining, Double Immunofluorescence Staining

Journal: Biology

Article Title: Force-Dependent Presence of Senescent Cells Expressing Vascular Endothelial Growth Factor During Orthodontic Tooth Movement

doi: 10.3390/biology15020187

Figure Lengend Snippet: Vascular endothelial growth factor (VEGF) expression of senescent cells during OTM. ( A ) Representative p16 and VEGF double immunofluorescence staining images. p16/VEGF staining without DAPI ( left ) and merged images with DAPI ( right ) are shown. ( B ) Quantitative analysis of p16 + VEGF + area in PDL. ( C ) Schematic illustration of the quantitative analysis for p16 + and VEGF + . ( D ) Quantitative analysis of p16 + VEGF + /VEGF + area in PDL. ( E ) Quantitative analysis of p16 + VEGF + /p16 + area in PDL. Scale bars: 100 µm (low magnification), 10 µm (high magnification). Data are mean ± SD (n = 4). ns; not significant, ** p < 0.01; *** p < 0.001; **** p < 0.0001; one-way ANOVA with Tukey’s test for ( B , D ), and Welch’s ANOVA with Games–Howell post hoc test for ( E ). VEGF: vascular endothelial growth factor, PDL: periodontal ligament, AB: alveolar bone.

Article Snippet: CD31 , Endothelial cell marker , Bioss Antibodies (Shanghai, China) , bs-0195R , BF647 , 1:100.

Techniques: Expressing, Double Immunofluorescence Staining, Staining

Journal: Biology

Article Title: Force-Dependent Presence of Senescent Cells Expressing Vascular Endothelial Growth Factor During Orthodontic Tooth Movement

doi: 10.3390/biology15020187

Figure Lengend Snippet: Cellular senescence of vascular endothelial cells (ECs) induced by OTM. ( A ) Representative p21 and CD31 immunofluorescence staining images. p21/CD31 staining without DAPI ( left ) and merged images with DAPI ( right ) are shown. ( B ) Quantitative analysis of p21 + CD31 + area in PDL. ( C ) Representative p16 and CD31 immunofluorescence staining images. p16/CD31 staining without DAPI ( left ) and merged images with DAPI ( right ) are shown. ( D ) Quantitative analysis of p16 + CD31 + area in PDL. Scale bars: 100 µm (low magnification), 10 µm (high magnification). Data are mean ± SD (n = 4). ns: not significant, * p < 0.05; ** p < 0.01; Kruskal–Wallis with Dunn’s post hoc test. PDL: periodontal ligament, AB: alveolar bone.

Article Snippet: CD31 , Endothelial cell marker , Bioss Antibodies (Shanghai, China) , bs-0195R , BF647 , 1:100.

Techniques: Immunofluorescence, Staining

Journal: Biology

Article Title: Force-Dependent Presence of Senescent Cells Expressing Vascular Endothelial Growth Factor During Orthodontic Tooth Movement

doi: 10.3390/biology15020187

Figure Lengend Snippet: Overlapping analysis of senescent and endothelial markers during OTM. ( A ) Schematic illustration of the overlapping analysis for p21 and CD31 in PDL. ( B ) Quantitative analysis of p21 + CD31 + area/p21 + area. ( C ) Quantitative analysis of p21 + CD31 + area/CD31 + area. ( D ) Schematic illustration of the overlapping analysis for p16 and CD31 in PDL. ( E ) Quantitative analysis of p16 + CD31 + area/p16 + area. ( F ) Quantitative analysis of p16 + CD31 + area/CD31 + area. Data are mean ± SD (n = 4). ns: not significant, * p < 0.05; ** p < 0.01; *** p < 0.001; Welch’s ANOVA with Games–Howell post hoc test for ( B ), one-way ANOVA with Tukey’s test for ( C , F ), and Kruskal–Wallis with Dunn’s post hoc test for ( E ).

Article Snippet: CD31 , Endothelial cell marker , Bioss Antibodies (Shanghai, China) , bs-0195R , BF647 , 1:100.

Techniques:

Journal: Biology

Article Title: Force-Dependent Presence of Senescent Cells Expressing Vascular Endothelial Growth Factor During Orthodontic Tooth Movement

doi: 10.3390/biology15020187

Figure Lengend Snippet: VEGF-expressing senescent ECs. ( A ) Representative p16, CD31, and VEGF triple immunofluorescence staining images. p16/CD31/VEGF staining without DAPI ( left ) and merged images with DAPI ( right ) are shown. ( B ) Quantitative analysis of p16 + CD31 + VEGF + area in PDL. ( C ) Schematic illustration of the quantitative analysis for p16, CD31, and VEGF in PDL. ( D ) Quantitative analysis of p16 + CD31 + VEGF + area/p16 + CD31 + area. ( E ) Quantitative analysis of p16 + CD31 + VEGF + area/p16 + VEGF + area. Scale bars: 100 µm (low magnification), 10 µm (high magnification). Data are mean ± SD (n = 4). ns: not significant, ** p < 0.01; **** p < 0.0001; Kruskal–Wallis with Dunn’s post hoc test for ( B ), and one-way ANOVA with Tukey’s test for ( D , E ). VEGF: vascular endothelial growth factor, PDL: periodontal ligament, AB: alveolar bone.

Article Snippet: CD31 , Endothelial cell marker , Bioss Antibodies (Shanghai, China) , bs-0195R , BF647 , 1:100.

Techniques: Expressing, Immunofluorescence, Staining

Journal: Biology

Article Title: Force-Dependent Presence of Senescent Cells Expressing Vascular Endothelial Growth Factor During Orthodontic Tooth Movement

doi: 10.3390/biology15020187

Figure Lengend Snippet: Presence and distribution of senescent cells during OTM. ( A ) Representative p21 and p16 immunofluorescence staining images with DAPI. ( B ) Quantitative analysis of p21 + area in PDL. ( C ) Quantitative analysis of p16 + area in PDL. ( D , E ) Distribution of senescence marker–positive areas in the 60 g and 180 g groups. Scale bars: 100 µm (low magnification), 10 µm (high magnification). Data are mean ± SD (n = 4). * p < 0.05; *** p < 0.001; **** p < 0.0001; one-way ANOVA with Tukey’s test. PDL: periodontal ligament, AB: alveolar bone.

Article Snippet: p21 , Senescence marker , Bioss Antibodies (Shanghai, China) , bs-10129R , AF555 , 1:100.

Techniques: Immunofluorescence, Staining, Marker

Journal: Biology

Article Title: Force-Dependent Presence of Senescent Cells Expressing Vascular Endothelial Growth Factor During Orthodontic Tooth Movement

doi: 10.3390/biology15020187

Figure Lengend Snippet: Cellular senescence of vascular endothelial cells (ECs) induced by OTM. ( A ) Representative p21 and CD31 immunofluorescence staining images. p21/CD31 staining without DAPI ( left ) and merged images with DAPI ( right ) are shown. ( B ) Quantitative analysis of p21 + CD31 + area in PDL. ( C ) Representative p16 and CD31 immunofluorescence staining images. p16/CD31 staining without DAPI ( left ) and merged images with DAPI ( right ) are shown. ( D ) Quantitative analysis of p16 + CD31 + area in PDL. Scale bars: 100 µm (low magnification), 10 µm (high magnification). Data are mean ± SD (n = 4). ns: not significant, * p < 0.05; ** p < 0.01; Kruskal–Wallis with Dunn’s post hoc test. PDL: periodontal ligament, AB: alveolar bone.

Article Snippet: p21 , Senescence marker , Bioss Antibodies (Shanghai, China) , bs-10129R , AF555 , 1:100.

Techniques: Immunofluorescence, Staining