Journal: Cancers

Article Title: Euchromatic Histone Lysine Methyltransferase 2 Inhibition Enhances Carfilzomib Sensitivity and Overcomes Drug Resistance in Multiple Myeloma Cell Lines.

doi: 10.3390/cancers15082199

Figure Lengend Snippet: Figure 4. UNC0642/CFZ combination is not toxic for normal cells, and it is effective in the presence of bone marrow milieu. (a) PMBCs were extracted from four healthy donors. Combinatorial treatments with increasing doses of UNC0642 together with 2.5 nM CFZ did not affect the viability of PBMCs compared to myeloma KMS28-BM control cells. Annexin V-PI staining was performed 72 h post- treatment. (b) GFP+ bone marrow stromal cells (HS-5) were seeded at 50,000/mL and subjected to single or double treatment. KMS28-BM cells were plated separately and used as a control. Cell death was monitored with FACS (PI-staining or TMRM-staining) at the indicated time points. (c) The KMS28-BM cell line was cultured on a monolayer of GFP+ HS-5 stromal cells and treated with the indicated doses of UNC0642 and CFZ. Cell viability was estimated by FACS analysis by staining with CD138-APC antibody, and the ratio with GFP+ HS-5 was calculated at the indicated time points. Statistical significance was calculated with the two-way ANOVA test (n = 7). (d) 3D scaffolds were populated with HS-5 cells (GFP+) and RPMI-I8226 cells (tRFP+) and treated with CFZ, UNC0642, or with the combination. RPMI-8226 cells were specifically detected using FACS analysis 6 days after treatment (n = 2 independent replicates) (unpaired t-test CFZ vs. CFZ + UNC0682 p-value = 0.0019). * p <0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Apoptosis was measured by flow cytometry after staining with tetrametylrodamine methyl ester (TMRM; Molecular Probes, Eugene, OR, USA) or Annexin V-FITC Kit (Catalog No. 130-092-052, Miltenyi Biotec, Bergisch Gladbach, Germany), according to the manufacturer’s instructions.

Techniques: Control, Staining, Cell Culture

Journal: Thorax

Article Title: Neutrophil TLR4 expression is reduced in the airways of infants with severe bronchiolitis

doi: 10.1136/thx.2008.107821

Figure Lengend Snippet: Antibodies used for flow cytometry

Article Snippet: TLR2 , FITC mouse monoclonal IgG 2a , Imgenex.

Techniques: Control

Journal: Frontiers in pharmacology

Article Title: Activating Cannabinoid Receptor 2 Protects Against Diabetic Cardiomyopathy Through Autophagy Induction.

doi: 10.3389/fphar.2018.01292

Figure Lengend Snippet: FIGURE 2 | Administration of HU308 enhances cardiac autophagy in heart issues from DCM mice. (A) Representative images of the autophagy-related proteins including beclin-1, LC3-II/I, and p62. (B) Quantitative analysis of the autophagy-related proteins (n = 5). ∗∗P < 0.01 vs. Nor group; $$P < 0.01 vs. DCM group. Nor, normal; DCM, diabetic cardiomyopathy.

Article Snippet: The primary antibodies applied were listed as follows: rabbit anti-Beclin-1 monoclonal antibody (1:1000; Cell Signaling Technology, Danvers, MA, United States), rabbit anti-LC3 polyclonal antibody (1:1000; Novus Biologicals, Littleton, CO, United States), rabbit anti-p62 antibody (1:1000; Cell Signaling Technology, Danvers, MA, United States), rabbit anti-adenosine 5′- monophosphate (AMP)-activated protein kinase (AMPK) antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States), rabbit anti-phosphorylated AMPK antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States) anti-mammalian target of rapamycin rabbit (mTOR) antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States), rabbit anti-phosphorylated mTOR antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States), rabbit anti-p70 ribosomal protein S6 kinase (p70S6K) antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States), rabbit antiphosphorylated p70S6K antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States) and mouse anti-β-actin antibody (1:5000, Beyotime Biotechnology, Shanghai, China).

Techniques:

Journal: Frontiers in pharmacology

Article Title: Activating Cannabinoid Receptor 2 Protects Against Diabetic Cardiomyopathy Through Autophagy Induction.

doi: 10.3389/fphar.2018.01292

Figure Lengend Snippet: FIGURE 4 | Administration of HU308 enhances cardiac autophagy in cardiomyocytes under the challenge of HG. Primary murine ventricular cardiomyocytes were obtained from neonatal C57BL/6J mice and challenged with high glucose concentration (HG; 33 mM glucose) for 24 h as hyperglycemia group or medium with normal glucose concentration (Nor; 5.5 mM glucose) as the control group. In certain groups, HU308 (10 µM) was treated to cells at 10 min before suffering HG. PBS in the same volume was administrated as vehicle. (A) Representative images of the autophagy-related proteins including beclin-1, LC3-II/I, and p62. (B) Quantitative analysis of the autophagy-related proteins (n = 5). ∗∗P < 0.01 vs. Nor group; $$P < 0.01 vs. HG group. (C) Representative images of transmission electron microscopy with arrows marking autophagosomes. (D) Quantitative analysis of the number of autophagosomes in murine ventricular cardiomyocytes (n = 5). ∗∗P < 0.01 vs. Nor group; $$P < 0.01 vs. HG group. Nor, normal; DCM, diabetic cardiomyopathy.

Article Snippet: The primary antibodies applied were listed as follows: rabbit anti-Beclin-1 monoclonal antibody (1:1000; Cell Signaling Technology, Danvers, MA, United States), rabbit anti-LC3 polyclonal antibody (1:1000; Novus Biologicals, Littleton, CO, United States), rabbit anti-p62 antibody (1:1000; Cell Signaling Technology, Danvers, MA, United States), rabbit anti-adenosine 5′- monophosphate (AMP)-activated protein kinase (AMPK) antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States), rabbit anti-phosphorylated AMPK antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States) anti-mammalian target of rapamycin rabbit (mTOR) antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States), rabbit anti-phosphorylated mTOR antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States), rabbit anti-p70 ribosomal protein S6 kinase (p70S6K) antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States), rabbit antiphosphorylated p70S6K antibody (1:1000, Cell Signaling Technology, Danvers, MA, United States) and mouse anti-β-actin antibody (1:5000, Beyotime Biotechnology, Shanghai, China).

Techniques: Concentration Assay, Control, Transmission Assay, Electron Microscopy

Journal: Chromosoma

Article Title: CTCF is essential for proper mitotic spindle structure and anaphase segregation.

doi: 10.1007/s00412-023-00810-w

Figure Lengend Snippet: Fig. 4 Mitotic spindle structure is perturbed in CTCF knock- downs. A Example images of mitosis in wild-type and CTCF knockdowns c13 and c21 with labeled microtubules (green, α-tubulin) and DNA (magenta, Hoechst). Mitotic spindles were classified as abnormal mitotic spindle if they were tri-/ tetrapolar spindles (yellow) or other abnormal (blue) if DNA was substantially behind the spindle pole or no division was observed. For all images, see Supplemental Figure 1. B Graph of the percentage of abnormal mitotic spindles (WT n = 0/48; CTCF knockdown c13 n = 12/42; CTCF knockdown c21 n = 12/57; CENP-E inhibitor GSK923295 72 h n = 11/17). C Box and whisker plot of average and individual measurement of DNA position relative to the spindle pole. Purple dots represent spindles where DNA is > 1 μm behind the spindle pole (WT 0%, c13 30%, c21 12%, GSK923295 29%). Error bar represents standard error. Statistical tests are Student’s t-tests, significance denoted by *p < 0.05, **p < 0.01, and ***p < 0.001, while no signifi- cance is denoted by ns. Scale bar is 10 μm

Article Snippet: Coverslips were placed on 50-μL drops of the primary antibody mixture consisting of rat anti-alpha tubulin at 1:100 dilution (Novus biological); mouse anti-lamin A/C at 1:100 dilution (Cell Signaling Technology); and PBS with 5% BSA.

Techniques: Labeling, Knockdown, Whisker Assay

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: Combined functional genome survey of therapeutic targets for hepatocellular carcinoma.

doi: 10.1158/1078-0432.CCR-09-2214

Figure Lengend Snippet: Fig. 5. Protein expression in HCC. Hematoxylin and eosin (HE) staining (original magnification, × 100) and immunoperoxidase staining (original magnifications, × 100 and × 400) of AKR1B10, HCAP-G, RRM2, and TPX2 proteins in HCC and adjacent nontumorous liver tissue. The specificity of antibodies was determined by immunoblotting of the KIM-1 cell lysate (left). N, nontumorous liver.

Article Snippet: Anti-TPX2 antibody was purchased from Novus Biologicals.

Techniques: Expressing, Staining, Immunoperoxidase Staining, Western Blot