Journal: International Journal of Biological Sciences

Article Title: Inhibition of Heat Shock-Induced H3K9ac Reduction Sensitizes Cancer Cells to Hyperthermia

doi: 10.7150/ijbs.86384

Figure Lengend Snippet: Heat shock-induced H3K9ac downregulation is mainly influenced by HDAC6. A-B AGS and SW480 cells were treated with SAHA (0.01 and 0.02 μM) and 3-TYP (5 and 10 μM) (A) or SAHA (0.1 μM) and LBH589 (0.05 μM) (B), followed by heat shock at 43°C for 1 hour. Changes in H3K9 acetylation were evaluated by WB. C AGS and SW480 cells were treated with a (0.5 μM), b (1 μM), c (0.5 μM), or d (1 μM), followed by heat shock at 43°C for 1 hour. Changes in H3K9 acetylation were analyzed by WB. a: ACY-241, b: TC-H 106, c: ACY-775, d: PCI34051. D AGS and SW480 cells were pre-transfected with control siRNA (siCon) or siRNAs targeting HDAC6 and subjected to heat shock at 43°C for 1 hour. Cells were lysed and evaluated by WB. Two independent siRNAs were used. E AGS and SW480 cells were treated with SAHA (0.01 μM) and YF-2 (10 and 20 μM), followed by heat shock at 43°C for 1 hour. Changes in H3K9ac were evaluated by WB. F-G AGS cells were fixed and subjected to Nuclear cytoplasmic fractionation assay and stained for WB (F) and immunofluorescence microscopy using an anti-HA antibody, in case of AGS cells were transfected with HA-HDAC6 plasmid (G). Scale bars, 10 μm.

Article Snippet: The following compounds were used: Vorinostat (SAHA) (Topscience, Shanghai, China; Cat# T1583), 3-TYP (Sellect, Shanghai, China; Cat# S8628), Citarinostat (ACY-241) (Topscience; Cat# T3661), Pimelic Diphenylamide 106 (TC-H 106) (Topscience; Cat# T3193), ACY-775 (Topscience; Cat# TQ0074), PCI34051 (Topscience; Cat# T6325), Panobinostat (LBH589) (Topscience; Cat# T2383), Erythrosphingosine (Topscience; Cat# T5891), LY317615 (Sellect; Cat# S1055), YF-2 (Sellect; Cat# S0022).

Techniques: Transfection, Control, Fractionation, Staining, Immunofluorescence, Microscopy, Plasmid Preparation

Journal: International Journal of Biological Sciences

Article Title: Inhibition of Heat Shock-Induced H3K9ac Reduction Sensitizes Cancer Cells to Hyperthermia

doi: 10.7150/ijbs.86384

Figure Lengend Snippet: SAHA and LBH589 enhance heat shock-induced apoptosis. A-D Effect of SAHA on heat shock-induced apoptosis in AGS, HGC27, BGC823, and SW480 cells analyzed by CCK8 assay (A), WB (B), and flow cytometry assay (C). Prior to heat shock at 43°C for 1 h, cells were treated with SAHA as the indicated concentration for 24 h (B, C) or 48 h (A). Percentage of apoptotic cells was quantified by flow cytometry assay (D). N=3. E-H Effect of LBH589 on heat shock-induced apoptosis in AGS, HGC27, and BGC823 cells analyzed by CCK8 assay (E), WB (F), and flow cytometry assay (G). Prior to heat shock at 43°C for 1 h, cells were treated with LBH589 as indicated concentration for 24 h. Percentage of apoptotic cells was quantified by flow cytometry assay (H). N=3. Date: mean ± SEM. Statistical analysis: two-way ANOVA. * p -value <0.05, *** p -value <0.001, **** p -value <0.0001. ns, no significance.

Article Snippet: The following compounds were used: Vorinostat (SAHA) (Topscience, Shanghai, China; Cat# T1583), 3-TYP (Sellect, Shanghai, China; Cat# S8628), Citarinostat (ACY-241) (Topscience; Cat# T3661), Pimelic Diphenylamide 106 (TC-H 106) (Topscience; Cat# T3193), ACY-775 (Topscience; Cat# TQ0074), PCI34051 (Topscience; Cat# T6325), Panobinostat (LBH589) (Topscience; Cat# T2383), Erythrosphingosine (Topscience; Cat# T5891), LY317615 (Sellect; Cat# S1055), YF-2 (Sellect; Cat# S0022).

Techniques: CCK-8 Assay, Flow Cytometry, Concentration Assay

Journal: International Journal of Biological Sciences

Article Title: Inhibition of Heat Shock-Induced H3K9ac Reduction Sensitizes Cancer Cells to Hyperthermia

doi: 10.7150/ijbs.86384

Figure Lengend Snippet: Synergistic effects of combining SAHA or LBH589 with hyperthermia on gastric cancer cells growth in vivo . A-D MFC cells were treated with or without heat shock at 43°C for 1 hour and were then detected for WB (A), CCK8 assay (B) and flow cytometry (C) after pre-treatment with SAHA at the indicated concentration for 24 h (A, C) or 48 h (B) and LBH589 for 24 h (A-C). Percentage of apoptotic cells was quantified by flow cytometry assay (D). N=3. E Schedule of animal experiments. 615 mice were subcutaneously (s.c.) injected with MFC cells. SAHA (25 mg/kg body weight) and LBH589 (10 mg/kg body weight) were administered via intraperitoneal (i.p.) injection every day or every other day, respectively, starting on day 4 after cell injection. Mice were exposed to hyperthermia at 40°C for 1 hour every other day starting on day 5 after cell injection. The mice were sacrificed and the tumors were collected on day 22. F Representative images MFC tumor burdens in mice. N=6. G Tumor growth was monitored at the indicated times. N=6. H Tumor weights of MFC tumors on day 22. N=6. I Immunohistochemistry of cleaved caspase-3. Scale bar, 100 μm. Date: mean ± SEM. Statistical analysis: The data in H was analyzed using two-tailed unpaired t test; the others were analyzed using two-way ANOVA. * p -value <0.05, ** p -value <0.01, *** p -value <0.001, **** p -value <0.0001. ns, no significance.

Article Snippet: The following compounds were used: Vorinostat (SAHA) (Topscience, Shanghai, China; Cat# T1583), 3-TYP (Sellect, Shanghai, China; Cat# S8628), Citarinostat (ACY-241) (Topscience; Cat# T3661), Pimelic Diphenylamide 106 (TC-H 106) (Topscience; Cat# T3193), ACY-775 (Topscience; Cat# TQ0074), PCI34051 (Topscience; Cat# T6325), Panobinostat (LBH589) (Topscience; Cat# T2383), Erythrosphingosine (Topscience; Cat# T5891), LY317615 (Sellect; Cat# S1055), YF-2 (Sellect; Cat# S0022).

Techniques: In Vivo, CCK-8 Assay, Flow Cytometry, Concentration Assay, Injection, Immunohistochemistry, Two Tailed Test

Journal: Oncotarget

Article Title: MYB induces the expression of the oncogenic corepressor SKI in acute myeloid leukemia

doi: 10.18632/oncotarget.25051

Figure Lengend Snippet: ( A ) RT-qPCR analysis of three independent experiments performed in duplicates for MYB and SKI transcripts in HL60 transfected with siMYB #1 (siMYB) or siNonsense #5 (siNons). Cells were harvested 24 h after transfection. Values are normalized to GAPDH and plotted relative to siNons (mean ± s.d.). * P < 0.03. ( B ) Western Blot analysis for SKI and MYB in HL60 transfected with indicated amounts of siMyb #1-4 or pool (mixed #1–4) or siNons #1or #5. Cells were harvested 24 h after transfection. β-ACTIN served as loading control. Numbers indicate relative mean values INT of quantified SKI and MYB protein bands normalized to quantified β-ACTIN bands. ( C ) RT-qPCR analysis of four independent experiments performed in duplicates for MYB and SKI transcripts in HL60 and U937 cells treated with 0 (ctrl) or 5 mM VPA for 48 h. Values are normalized to GAPDH and plotted relative to ctrl cells (mean ± s.d.). * P < 0.05, ** P < 0.004. ( D , E ) Western Blot analysis for SKI and MYB in AML cell lines HL60 and U937 treated with 0, 1 or 5 mM VPA (D) or DMSO (solvent control), 20, 30 or 40 nM LBH589 (E) for 48 h. β-ACTIN served as loading control.

Article Snippet: HL60 and U937 cells were treated with the indicated concentrations of valproic acid sodium salt (Sigma-Aldrich, Munich, Germany), LBH589 (Panobinostat) (Biomol, Hamburg, Germany) or dimethyl sulfoxide (DMSO) (Carl Roth, Karlsruhe, Germany) as solvent control.

Techniques: Quantitative RT-PCR, Transfection, Western Blot