Journal: Oncology Letters

Article Title: Long non-coding RNA zinc finger antisense 1 functions as an oncogene in acute promyelocytic leukemia cells

doi: 10.3892/ol.2019.11014

Figure Lengend Snippet: ZFAS1 regulates APL cell apoptosis through the modulation of Bcl-2 family proteins. (A) APL cells were transfected with si-NC, si-ZFAS1-1 or si-ZFAS1-2 for 48 h. The levels of Bcl-2, Mcl-1, XIAP and Bax were examined by western blot analyses with the indicated antibodies. Densitometry analysis of the western blots is presented on the right. **P<0.01, ***P<0.001 vs. si-NC. (B) APL cells were transfected with si-NC, si-ZFAS1-1 or si-ZFAS1-2 for 48 h. Cytosolic fractions were subjected to western blotting with the indicated antibodies and densitometry analysis is presented on the right. ***P<0.001 vs. si-NC. Values are presented as the mean ± standard deviation (n=3). ZFAS1, long non-coding RNA zinc finger antisense 1; APL, acute promyelocytic leukemia; si- small interfering RNA; NC, negative control; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; Bcl-2, B-cell lymphoma-2; Mcl-1, induced myeloid leukemia cell differentiation protein Mcl-1; XIAP, E3 ubiquitin-protein ligase XIAP; Bax, apoptosis regulator BAX; Smac/DIABLO, Diablo homolog mitochondrial.

Article Snippet: The membranes were blocked with 5% skimmed milk for 1 h at room temperature, and incubated at 4°C overnight with primary antibodies diluted at 1:1,000 against Mcl-1 (cat. no. ab32087; Abcam, Cambridge, MA, USA), caspase-9 (cat. no. 9502), cleaved caspase-9 (cat. no. 9505), caspase-3 (cat. no. 9662), cleaved caspase-3 (cat. no. 9661), caspase-8 (cat. no. 9746), Bcl-2 (cat. no. 4223), cytochrome c (cat. no. 12963), Smac/DIABLO (cat. no. 15108), GAPDH (cat. no. 5174), PARP (cat. no. 9532), XIAP (cat. no. 2045) and BAX (cat. no. 5023) (all Cell Signaling Technology, Inc., Danvers, MA, USA).

Techniques: Transfection, Western Blot, Standard Deviation, Small Interfering RNA, Negative Control, Cell Differentiation

Journal: Cell Death & Disease

Article Title: Restoring mitochondrial cardiolipin homeostasis reduces cell death and promotes recovery after spinal cord injury

doi: 10.1038/s41419-022-05369-5

Figure Lengend Snippet: A – C Biomarkers of apoptosis following SCI. Time course of cytochrome c release ( A ), Smac/DIABLO release ( B ), and active caspase-3 expression ( C ) following SCI. * P < 0.05, ** P < 0.01 vs sham (One-way ANOVA, Dunnett post-test, n = 4 rats/group). Data represent mean ± s.e.m. Cyt. c cytochrome c, Smac/D. Smac/DIABLO, A. casp.-3 active casepase-3. D – I Lipidomic analysis after SCI. Lipid extracts of spinal cord from sham, 3 h, and 24 h SCI rats were prepared by using the methyl-tert-butyl ether (MTBE) extraction. D , E Expanded negative-ion ESI mass spectra of rat spinal cord lipid extracts obtained using a Q Exactive™ Hybrid Quadrupole-Orbitrap mass spectrometer. The asterisks indicate the identified CL plus-one isotopologues, which were characteristic of the doubly charged CL molecular species and were used to quantify individual CL molecular species. Each spectrum is displayed after being normalized to the intensity of internal standard 1’, 3’-bis [1, 2-dimyristoyl-sn-glycero-3-phospho]- sn -glycerol (tetra14:0 CL, [CL-2H + 1] 2- = 619.91791) ion. The x -axis is mass-to-charge ratio ( m/z ). A significant decrease in CL was detected at 3 and 24 h post-injury ( D – F ) while a corresponding increase in lyso-CL and ratio of lyso-CL/CL was observed at 24 h post-injury ( G , H ). 4-HNE, a marker of lipid peroxidation, was significantly increased at 3 and 24 h after SCI ( I ). Data represent mean ± s.e.m. * P < 0.05, ** P < 0.01 (One-way ANOVA, Tukey’s multiple comparisons test, n = 4 rats/groups). CL cardiolipin, Lyso-CL lyso-cardiolipin, 4-HNE 4-hydroxy Nonenal.

Article Snippet: The primary antibodies included rabbit polyclonal anti-caspase-3 antibody (Cat# 9664, cleaved, 1:1000, 0.04 µg/ml, Cell Signaling Technology, Boston, MA), mouse anti-cytochrome c antibody (Cat# 556433, 1:300, 1.67 µg/ml, 7H8.2C12, BD Pharmingen, San Jose, CA), mouse anti-Smac/DIABLO antibody (Cat# 612246, 1:1000, 0.25 µg/ml, BD Transduction Laboratories, San Jose, CA), rabbit anti-PARP-1 (Cat# 9542, 1:500, 0.0976 µg/ml, Cell Signaling Technology, Boston, MA), rabbit anti-Bcl-2 (Cat# sc-492, 1:100, 2 µg/ml, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-Bax (Cat# sc-526, 1:100, 2 µg/ml, Santa Cruz Biotechnology, Santa Cruz, CA), and mouse anti-β-tubulin antibody (Cat# 5293, 1:1000, 4.7 µg/ml, Sigma, St. Louis, MO).

Techniques: Expressing, Mass Spectrometry, Marker

Journal: Cell Death & Disease

Article Title: Restoring mitochondrial cardiolipin homeostasis reduces cell death and promotes recovery after spinal cord injury

doi: 10.1038/s41419-022-05369-5

Figure Lengend Snippet: A Time course of mitochondrial cPLA 2 expression and activation after SCI. Data represent mean ± s.e.m., * P < 0.05, ** P < 0.01 vs sham (Two-way ANOVA, Tukey’s multiple comparisons test, n = 6 rats/group). B An electron microscopic image shows phosphorylated cPLA 2 (p-cPLA 2 ) immunopositive peroxidase reaction products in a neuronal dendrite in the rat spinal ventral horn. p-cPLA 2 immunoreactivity (IR) was observed to localize in mitochondria (*) including both the outer (yellow arrow) and inner (red arrows) mitochondrial membranes, and on the cell membrane (green arrow) at 3 days after SCI. Bar = 0.25 μm. C cPLA 2 activation-induced CL loss was reversed by AACOCF 3 , a cPLA 2 inhibitor. D SCI induced a decrease in the ratio of red/green in the MMP (Δψm) assay, which was reversed by AACOCF 3. ( E – H ) AACOCF 3 , a cPLA 2 inhibitor, reversed SCI-induced cytochrome c release ( E ), Smac/DIABLO release ( F ), active caspase-3 expression ( G ), and cleaved PAPRP expression ( H ). Data represent mean ± s.e.m. * P < 0.05, ** P < 0.01 (One-way ANOVA, Tukey’s multiple comparisons test, n = 6 rats/group). Cyt. c cytochrome c, Smac/D. Smac/DIABLO, A. casp-3 active caspase-3. I – M Oxidative stress induces cPLA 2 activation after SCI. I Immunohistochemistry shows a marked increase in p-cPLA 2 -expression, a marker of cPLA 2 activation, at the injury epicenter and 3 mm rostral to it after a single injection of H 2 O 2 into the T9 spinal cord. Right column images show that H 2 O 2 -induced p-cPLA 2 expression was substantially reversed by mepacrine (5 mg/kg), a PLA 2 inhibitor. The expression of p-cPLA 2 was found in neurons (arrows), swelling axons (open arrows), and glial cells (arrowheads). Bar, 100 μm. I – L Administration of XJB (10 mg/kg, i.p.) at 30 min post-injury significantly reduced SCI-induced mitochondrial cPLA 2 activation. J Representative images of phosphorylated cPLA 2 (p-cPLA 2 ), cPLA 2 , and VDAC expression. K Compiled results in a bar graph for the ratio of p-cPLA 2 /cPLA 2 expression. L Compiled results in a bar graph for the ratio of p-cPLA 2 /VDAC expression. M Compiled results in a bar graph for the ratio of cPLA 2 /VDAC expression. * P < 0.05, ** P < 0.01 (One-way ANOVA, Tukey’s multiple comparisons test, n = 6). Data represent the mean ± s.e.m.

Article Snippet: The primary antibodies included rabbit polyclonal anti-caspase-3 antibody (Cat# 9664, cleaved, 1:1000, 0.04 µg/ml, Cell Signaling Technology, Boston, MA), mouse anti-cytochrome c antibody (Cat# 556433, 1:300, 1.67 µg/ml, 7H8.2C12, BD Pharmingen, San Jose, CA), mouse anti-Smac/DIABLO antibody (Cat# 612246, 1:1000, 0.25 µg/ml, BD Transduction Laboratories, San Jose, CA), rabbit anti-PARP-1 (Cat# 9542, 1:500, 0.0976 µg/ml, Cell Signaling Technology, Boston, MA), rabbit anti-Bcl-2 (Cat# sc-492, 1:100, 2 µg/ml, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-Bax (Cat# sc-526, 1:100, 2 µg/ml, Santa Cruz Biotechnology, Santa Cruz, CA), and mouse anti-β-tubulin antibody (Cat# 5293, 1:1000, 4.7 µg/ml, Sigma, St. Louis, MO).

Techniques: Expressing, Activation Assay, Immunohistochemistry, Marker, Injection

Journal: Cell Death & Disease

Article Title: Restoring mitochondrial cardiolipin homeostasis reduces cell death and promotes recovery after spinal cord injury

doi: 10.1038/s41419-022-05369-5

Figure Lengend Snippet: A – D lipidomic analysis at 1 day after SCI showed that XJB-5-131 (XJB, 10 mg/kg) attenuated SCI-induced mitochondrial 4-HNE expression ( A ), oxidized CL ( B ), CL loss ( C ), and lyso-CL ( D ) in adult rats. * P < 0.05, ** P < 0.01 (One-way ANOVA, Tukey’s multiple comparisons test, n = 4 rats/group). Data represent mean ± s.e.m. E – H XJB attenuated mitochondrial dysfunction ( E ), cytochrome c release ( F ), Smac/DIABLO release ( G ), and active caspase-3 expression ( H ) at 1 day after SCI in adult rats. Data represent mean ± s.e.m. * P < 0.05, ** P < 0.01 (One-way ANOVA, Tukey’s multiple comparisons test, n = 4–6 rats/group).

Article Snippet: The primary antibodies included rabbit polyclonal anti-caspase-3 antibody (Cat# 9664, cleaved, 1:1000, 0.04 µg/ml, Cell Signaling Technology, Boston, MA), mouse anti-cytochrome c antibody (Cat# 556433, 1:300, 1.67 µg/ml, 7H8.2C12, BD Pharmingen, San Jose, CA), mouse anti-Smac/DIABLO antibody (Cat# 612246, 1:1000, 0.25 µg/ml, BD Transduction Laboratories, San Jose, CA), rabbit anti-PARP-1 (Cat# 9542, 1:500, 0.0976 µg/ml, Cell Signaling Technology, Boston, MA), rabbit anti-Bcl-2 (Cat# sc-492, 1:100, 2 µg/ml, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-Bax (Cat# sc-526, 1:100, 2 µg/ml, Santa Cruz Biotechnology, Santa Cruz, CA), and mouse anti-β-tubulin antibody (Cat# 5293, 1:1000, 4.7 µg/ml, Sigma, St. Louis, MO).

Techniques: Expressing

Journal: Cell metabolism

Article Title: Systematic dissection of the metabolic-apoptotic interface in AML reveals heme biosynthesis to be a regulator of drug sensitivity

doi: 10.1016/j.cmet.2019.01.011

Figure Lengend Snippet:

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit monoclonal anti-β-Actin (clone 13E5) antibody Cell Signaling Technology Cat# 4970, RRID:AB_2223172 Rabbit monoclonal anti-AIF (clone D39D2) antibody Cell Signaling Technology Cat# 5318, RRID:AB_10634755 Rabbit monoclonal anti-Smac/Diablo (D20A5) antibody Cell Signaling Technology Cat# 9745, RRID:AB_11220423 Rabbit monoclonal anti-HtrA2/Omi (clone D20A5) antibody Cell Signaling Technology Cat# 9745, RRID:AB_11220423 Rabbit monoclonal anti-Bax (D2E11) antibody Cell Signaling Technology Cat# 5023, RRID:AB_10557411 Rabbit monoclonal anti-Bak (clone D4E4) antibody Cell Signaling Technology Cat# 12105, RRID:AB_2716685 Mouse monoclonal anti-cleaved-PARP (Asp214) (clone 19F4) antibody Cell Signaling Technology Cat# 9546, RRID:AB_2160593 Mouse monoclonal anti-cleaved-caspase 3 (Asp175) (clone 5A1E) antibody Cell Signaling Technology Cat# 9664, RRID:AB_2070042 Rabbit polyclonal anti-Na,K-ATPase antibody Cell Signaling Technology Cat# 3010, RRID:AB_2060983 Rabbit monoclonal anti-cytochrome c (clone D18C7) antibody Cell Signaling Technology Cat# 11940, RRID:AB_2637071 Rabbit monoclonal anti-X-IAP (clone 3B6) antibody Cell Signaling Technology Cat# 2045, RRID:AB_2214866 Rabbit monoclonal anti-VDAC1 (clone {"type":"entrez-protein","attrs":{"text":"EPR10852","term_id":"523376397","term_text":"EPR10852"}} EPR10852 (B)) antibody Abcam Cat# ab154856, RRID:AB_2687466 Total OXPHOS Human WB Antibody Cocktail: 1.

Techniques: Virus, Expressing, Plasmid Preparation, Recombinant, Cell Fractionation, Membrane, Cell Culture, Cell Viability Assay, Activity Assay, Isolation, shRNA, Metabolic Labelling, Software