Journal: iScience

Article Title: RUNX1 colludes with NOTCH1 to reprogram chromatin in T cell acute lymphoblastic leukemia

doi: 10.1016/j.isci.2023.106795

Figure Lengend Snippet: RUNX1-KD is associated with a genome-wide reduction in H3K27ac (A) Experimental strategy of NOTCH1 inhibition (NOTCH1-INB) and RUNX1 knockdown (RUNX1-KD). NSC means non-silencing control and GSI means gamma secretase inhibitor. The number of replicates is indicated in brackets. (B) Anti-RUNX1 Western blot of protein levels following knockdown by shRUNX1-58 and shRUNX1-59. Numbers below the RUNX1 panel indicate their relative expression level after normalization to β-actin loading control. (C) Growth competition assays showing the fraction of GFP+ cells following transduction with GFP-tagged lentiviruses expressing shRUNX1s and normalized to a scrambled shRNA control (RUNX1-CTL). (D) Normalized read densities of H3K27ac and H3K4me1 peaks following RUNX1-KD (red ≥ 2-fold loss; black ≥2-fold gain; and gray unchanged). Read densities were normalized by sequencing depth. (E) Mean RUNX1 ChIP-seq read densities at gained, lost, and stable H3K27ac regions following RUNX1-KD. RUNX1 ChIP-seq data was derived from CUTLL1 and p values were calculated using KS-test. (F) Western blot of histone H3, H3K27ac, and H3K4me1 levels in control (RUNX1-WT) and RUNX1-KOs (KO1 and KO2) using two CRISPR guides in KOPTK1. (G) Ratio of H3K27ac and H3K4me1 relative to the H3 control was quantified from the protein bands shown in (F). Three independent experiments were performed to calculate p values using unpaired two-tailed t-test where ∗ indicates p value ≤ 0.05 and ∗∗ for p value ≤ 0.01. Data are mean ± s.e.m. of H3K27ac/H3. (H) Western blot analysis of K3K27ac and RUNX1 in control (RUNX1-WT) and RUNX1-KOs (KO1 and KO2) in Jurkat, HL60, and DU145 cell lines. Where shown, percent reduction was calculated by comparing control (RUNX1-WT) to the knockout samples.

Article Snippet: Prostate adenocarcinoma , ATCC (HTB-81TM) , DU145.

Techniques: Genome Wide, Inhibition, Knockdown, Control, Western Blot, Expressing, Transduction, shRNA, Sequencing, ChIP-sequencing, Derivative Assay, CRISPR, Two Tailed Test, Knock-Out

Journal: iScience

Article Title: RUNX1 colludes with NOTCH1 to reprogram chromatin in T cell acute lymphoblastic leukemia

doi: 10.1016/j.isci.2023.106795

Figure Lengend Snippet:

Article Snippet: Prostate adenocarcinoma , ATCC (HTB-81TM) , DU145.

Techniques: Recombinant, Expressing, Plasmid Preparation, Software

Journal: Pharmacological research

Article Title: A selective alpha1D-adrenoreceptor antagonist inhibits human prostate cancer cell proliferation and motility "in vitro".

doi: 10.1016/j.phrs.2015.11.017

Figure Lengend Snippet: Figure 4: Cell cycle analysis and p27 and p21 levels in PC3 and DU145 cells exposed to A175.

Article Snippet: Human androgen independent prostate cancer PC3 and DU145 cell lines and the androgendependent LNCaP cell line were originally obtained from American Type Culture Collection (ATCC, Rockville, MD, USA); CWR22Rv1 (Rv1) cells were originally obtained from LeibnitzInstitut DSMZ (Braunschweig, Germany).

Techniques: Cell Cycle Assay

Journal: Pharmacological research

Article Title: A selective alpha1D-adrenoreceptor antagonist inhibits human prostate cancer cell proliferation and motility "in vitro".

doi: 10.1016/j.phrs.2015.11.017

Figure Lengend Snippet: Figure 5: Effect of A175 on PC3 and DU145 cell motility.

Article Snippet: Human androgen independent prostate cancer PC3 and DU145 cell lines and the androgendependent LNCaP cell line were originally obtained from American Type Culture Collection (ATCC, Rockville, MD, USA); CWR22Rv1 (Rv1) cells were originally obtained from LeibnitzInstitut DSMZ (Braunschweig, Germany).

Techniques:

Journal: PLoS ONE

Article Title: Withaferin A Induces Cell Death Selectively in Androgen-Independent Prostate Cancer Cells but Not in Normal Fibroblast Cells

doi: 10.1371/journal.pone.0134137

Figure Lengend Snippet: (A, B) Western blot analysis to detect c-Fos (FosB) in TIG-1, LNCaP, DU-145, and PC-3 cells at 4, 8, and 24 h after treatment with 4 μM (A) or 2 μM (B) WA. NT, non-treated. (C) Western blot analysis to detect c-Fos, PARP, FLIP and GAPDH in PC-3 cells at 12 h after 4 μM WA treatment in the presence (+) or absence (-) of three different siRNAs (X–Z) from OriGene. (D) Viability of PC-3 cells after siFos treatment. Data are represented as means ± SEM of three independent experiments; pink arrows indicate a statistically significant reduction in cell number following siFos treatment (**, P < 0.01). (E) Population of apoptotic, necrotic, and live cells distinctly stained with Annexin V–EnzoGold, 7-AAD-Red, and GFP. Data are represented as means ± SEM of three independent experiments; red, blue, and green arrows indicate statistically significant changes (**, P < 0.01). (F) Western blot analysis of c-Fos, PARP, FLIP, and GAPDH in DU-145 at 12 h after 4 μM WA treatment in the presence (+) or absence (-) of siRNAs; siFos or siGL2 (siControl). (G) Viability of DU-145 cells after exogenous overexpression of pcDNA3-FLIP or vector alone in the presence of DMSO (solvent) or 4 μM WA.

Article Snippet: Human cells were acquired from the indicated suppliers: PC-3 and DU-145 (RIKEN Bioresource Center), TIG-1 (Japanese Cancer Research Resources Bank), KD (Japanese Collection of Research Bioresources Cell Bank) and LNCaP (ATCC).

Techniques: Western Blot, Staining, Over Expression, Plasmid Preparation

Journal: PLoS ONE

Article Title: Withaferin A Induces Cell Death Selectively in Androgen-Independent Prostate Cancer Cells but Not in Normal Fibroblast Cells

doi: 10.1371/journal.pone.0134137

Figure Lengend Snippet: (A) Western blot analysis to detect HSPA6, HSP40, HSPA70, EGR-1, PAR-4, and GAPDH (loading control) in TIG-1, LNCaP, DU-145, and PC-3 cells at 0, 4, 8 and 24 h after 4 μM WA treatment. Arrow indicates the band for bona fide EGR-1. (B) Influence of siHSPA6 expression on PC-3 cell growth. (i) Schematic for this experiment. (ii) Western blot analysis to confirm the knockdown of HSPA6 protein by siHSPA6, relative to cells treated with siGL2 (negative control). (iii) Influence of siHSPA6 and siGL2 on PC-3 cell growth, with or without WA treatment. Arrow highlights the reduction in PC-3 cell growth. (C) Influence of siHSF-1 expression on PC-3 cell growth. (i) Schematics for this experiment. (ii) Western blot analysis to confirm the knockdown of HSP protein by siHSF-1, relative to cells treated with siGL2 (negative control), and to determine whether HSF-1 knockdown affected HSP protein levels by detection of HSPA6, HSP40, HSPA70, and GAPDH in PC-3 cells. (iii) Influence of siHSF-1 and siGL2 on PC-3 cell growth, with or without WA treatment. Arrow highlights the reduction in PC-3 cell growth. Data are represented as means ± SEM of three independent experiments; pink, green, and blue arrows indicate significant decreases in cell viabilities (*, P < 0.05; **, P < 0.01).

Article Snippet: Human cells were acquired from the indicated suppliers: PC-3 and DU-145 (RIKEN Bioresource Center), TIG-1 (Japanese Cancer Research Resources Bank), KD (Japanese Collection of Research Bioresources Cell Bank) and LNCaP (ATCC).

Techniques: Western Blot, Expressing, Negative Control

Journal: PLoS ONE

Article Title: Withaferin A Induces Cell Death Selectively in Androgen-Independent Prostate Cancer Cells but Not in Normal Fibroblast Cells

doi: 10.1371/journal.pone.0134137

Figure Lengend Snippet: (A) Western blot analysis to detect IRE-1α, PERK, pS51-eIF2α, CHOP, caspase 3, PARP, BiP, and GAPDH (loading control) in TIG-1, LNCaP, DU-145, and PC-3 cells at 4, 8, and 24 h after 4 μM WA treatment. NT: non-treated. (B) Schematic representation of the analyzed proteins in the apoptotic pathway induced by ER stress. (C) Typical fluorescence images of TIG-1 (i), LNCaP (ii), PC-3 (iii), and DU-145 (iv) that were treated with 4 μM WA for 24 h and subsequently treated with ROS detection reagents using the Image-iT LIVE Green ROS Detection Kit. Merged images are shown of cytoplasmic ROS signals (green) and nuclear DNA signals stained with Hoechst33342 (blue). Green bar, 100 μm. Black bar, 10 μm.

Article Snippet: Human cells were acquired from the indicated suppliers: PC-3 and DU-145 (RIKEN Bioresource Center), TIG-1 (Japanese Cancer Research Resources Bank), KD (Japanese Collection of Research Bioresources Cell Bank) and LNCaP (ATCC).

Techniques: Western Blot, Fluorescence, Staining

Journal: PLoS ONE

Article Title: Withaferin A Induces Cell Death Selectively in Androgen-Independent Prostate Cancer Cells but Not in Normal Fibroblast Cells

doi: 10.1371/journal.pone.0134137

Figure Lengend Snippet: (A) Western blot analysis to detect vimentin and actin in TIG-1, LNCaP, PC-3, and DU-145 at 4, 8, and 24 h after treatment with 2 μM or 4 μM WA. NT: non-treated. (B) Typical images of TIG-1 (i) and PC-3 (ii) stained with anti-vimentin antibody, phalloidin (for F-actin), and Hoechst 33342 (for DNA) at 2, 4, and 6 (h) after WA treatment. NT: non-treated. Pink and yellow arrows indicated colocalized vimentin and F-actin aggregates. Merged images are shown in the bottom row. Bar, 10 μm. (C) Percentage of all observed cells containing dots of aggregated or non-aggregated vimentin are shown for TIG-1 (i) and PC-3 (ii). Cells harboring over 10 vimentin aggregates were scored as “aggregated”, whereas cells with non-aggregated vimentin were scored as “non-aggregated”. Data are represented as means ± SEM of three independent experiments; asterisks indicate statistically significant changes in cell viability (**, P < 0.01; ***, P < 0.001). (D) Jasplakinoloide (Jsp) did not influence PC-3 cell viability. (i) Schematic of the experimental design. (ii) Bar graphs showing cell viability (%) in the presence (+) or absence (-) of 4 μM WA or Jasp 8 h after the addition of WA. NT: non-treated. Bar graphs represent means ± SEM for three independent experiments. Pink, blue, and green arrows indicate statistically significant reductions in cell viability (*, P < 0.05; **, P < 0.01).

Article Snippet: Human cells were acquired from the indicated suppliers: PC-3 and DU-145 (RIKEN Bioresource Center), TIG-1 (Japanese Cancer Research Resources Bank), KD (Japanese Collection of Research Bioresources Cell Bank) and LNCaP (ATCC).

Techniques: Western Blot, Staining

Journal: PLoS ONE

Article Title: Withaferin A Induces Cell Death Selectively in Androgen-Independent Prostate Cancer Cells but Not in Normal Fibroblast Cells

doi: 10.1371/journal.pone.0134137

Figure Lengend Snippet: (A) Frequency of spheres larger than 100 μm (i) and typical images (ii) after incubation of TIG-1, LNCaP, PC-3, DU-145, and SAS cells in sphere-formation medium for 10 days. (B) Viability of SAS cells after WA treatment. Bars represent means ± SEM of three independent experiments. Cell viability was measured at 4, 8, and 24 h after treatment with 2 μM (blue bars) or 4 μM (red bars) WA. NT, non-treated. Data are represented as means ± SEM of three independent experiments; red arrows indicate statistically significant reductions in cell viability (**, P < 0.01). (C) Western blot analysis to detect c-Fos, HSPA6, HSP40, HSP70, PARP, and GAPDH (loading control) in SAS cells at 4, 8, and 24 h after treatment with 2 μM or 4 μM WA. NT: non-treated. (D) Comparison of sensitivity to cisplatin between parental cells and spheres of PC-3 (i and ii) and SAS (iii and iv) after 48 h of incubation following treatment with indicated cisplatin concentrations.

Article Snippet: Human cells were acquired from the indicated suppliers: PC-3 and DU-145 (RIKEN Bioresource Center), TIG-1 (Japanese Cancer Research Resources Bank), KD (Japanese Collection of Research Bioresources Cell Bank) and LNCaP (ATCC).

Techniques: Incubation, Western Blot