Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Reduction in the developmental potential of intrathymic T cell progenitors with age.

doi: 10.4049/jimmunol.173.1.245

Figure Lengend Snippet: FIGURE 4. Proliferation and survival of ETP from young and old mice. Thymocytes were freshly isolated from young and old mice and stained for ETP in combination with anti-Ki-67 or annexin V labeling to determine their proliferation or survival status, respectively. A, ETP were labeled with anti-Ki-67 or isotype control Abs following fixation and permeabilization. The upper panel shows representative data for single young and old ani- mals. The lower panel indicates data pooled from the indicated number of different aged mice at each time point. B, The level of apoptosis in ETP was evaluated by annexin V labeling. The upper panel shows representative data for single young and old animals. The lower panel indicates data pooled from the indicated number of different aged mice at each time point. Means and SDs are shown for n 2–6 animals as indicated. , p 0.02; , p 0.002; #, p 0.005; ##, p 0.0005.

Article Snippet: The frequency of apoptotic cells in freshly harvested thymocyte populations was assessed by labeling with annexin V using the Apoptosis Detection kit as described by the manufacturer (R&D Systems, Minneapolis, MN) in combination with Abs to various cell surface determinants.

Techniques: Isolation, Staining, Labeling, Control

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: Gene therapy

Article Title: Tumor cells expressing a fusion protein of MULT1 and Fas are rejected in vivo by apoptosis and NK cell activation.

doi: 10.1038/gt.2008.77

Figure Lengend Snippet: Figure 3 Mouse UL16-binding protein-like transcript 1 E (MULT1E)/FasTI induces apoptosis. A total of 1 106 cells of TC-1 and clones L-5, L-7 and L-10 were treated with 1 mg/ml NKG2D/Fc for 16 h. The cells were then analyzed for apoptosis and necrosis using Annexin V assay (a–c) or caspase-3 assay (d) according to the manufacturers’ protocols. (a) An example of the fluorescence-activated cell sorting (FACS) data. (b, c) Summaries of data from three separate experiments. The statistical analyses were conducted between the controls (open bars) and NKG2D/Fc-treated cells (solid bars) using two-way analysis of variance (ANOVA). The difference between NKG2D/Fc-treated L-5 cells and NKG2D/Fc-treated L-10 cells was also compared using Student’s t-test. *Po0.05; **Po0.01 and ***Po0.001.

Article Snippet: Induction of apoptosis in cells expressing the fusion protein To determine if cells expressing the fusion protein can be induced to undergo apoptosis, one million cells of TC-1 and clones L-5, L-7 and L-10 were treated with 1 mg/ml of NKG2D/Fc for 16 h. Apoptosis of the cells was measured using two systems: a TACS Annexin V-FITC Apoptosis Kit (R&D Systems) and a caspase-3 fluorometric assay (R&D Systems).

Techniques: Binding Assay, Clone Assay, Annexin V Assay, Caspase-3 Assay, FACS

Journal: iScience

Article Title: SeSA-HCPT: A dual-targeting agent that induces DNA damage and inhibits repair for castration-resistant prostate cancer therapy

doi: 10.1016/j.isci.2026.114824

Figure Lengend Snippet: SeSA-HCPT induces S-phase cell-cycle arrest and apoptosis in PCa cells Cells were treated with 200 nM SeSA-HCPT, or combined agents (200 nM SeSAHA +400 nM HCPT). DMSO used as control. (A–D) Apoptosis induction in PCa cell lines by the indicated treatments, as determined by flow cytometry. (E–G) Western blot analysis of cleaved caspase-3 levels in PC3, DU145, and LNCaP cells treated with SeSAHA, HCPT, combination therapy, or SeSA-HCPT. (H) Cell cycle distribution analysis of PC3, DU145, and HaCaT cells treated with either SeSA-HCPT alone or the combination therapy. Analysis was performed using flow cytometry. (I–K) Cell-cycle arrest of PC3, DU145, and HaCaT cells by the tested compounds. (L and M) Western blot analysis of cell cycle-related protein expression in PC3 and DU145 cells treated with SeSAHA, HCPT, combination therapy, or SeSA-HCPT. Data are presented as mean ± SD. Statistical significance is denoted as follows: ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: FITC-Annexin V/PI apoptosis kit , Solarbio , Cat#CA1020.

Techniques: Control, Flow Cytometry, Western Blot, Expressing

Journal: Frontiers in Oncology

Article Title: Mutant NPM1-Regulated FTO-Mediated m 6 A Demethylation Promotes Leukemic Cell Survival via PDGFRB/ERK Signaling Axis

doi: 10.3389/fonc.2022.817584

Figure Lengend Snippet: FTO Promotes Leukemic Cell Proliferation by Facilitating Cell Cycle and Suppressing Cell Apoptosis. (A) CCK-8 analysis of cell growth in the FTO-silenced OCI-AML3. (B) EdU analysis of cell proliferation in the FTO-silenced OCI-AML3 cells. The bar graph showed the percentage of EdU positive cells. Scale bar: 50 μm. (C) Flow cytometric analysis of cell cycle distribution in FTO-silenced OCI-AML3 cells. The bar graph showed the percentage of G0/G1, S, and G2/M phase cells. (D) Cell apoptosis was measured by flow cytometric analysis of the Annexin V/DAPI stained cells. The bar graph showed the cell apoptosis rate. (E) Western blot analysis of Cyclin A2, t-Caspase 9, and c-Caspase 9 protein level in the FTO-silenced OCI-AML3. β-actin was used as the internal control. The bar graph showed the relative level of protein. Data were presented as the mean ± SD of three independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The cell apoptosis was determined by an Annexin V-APC/DAPI Apoptosis Detection Kit (Elabscience, Wuhan, China, #E-CK-A258).

Techniques: CCK-8 Assay, Staining, Western Blot, Control

Journal: Frontiers in Oncology

Article Title: Mutant NPM1-Regulated FTO-Mediated m 6 A Demethylation Promotes Leukemic Cell Survival via PDGFRB/ERK Signaling Axis

doi: 10.3389/fonc.2022.817584

Figure Lengend Snippet: The Oncogenic Function of FTO is Dependent on m 6 A RNA Demethylase Activity. (A) CCK-8 analysis of cell proliferation in OCI-AML3 cells following transfection of the HA-FTO-WT and HA-FTO-MUT plasmids. (B) m 6 A dot blot analysis of the m 6 A level of global RNAs in OCI-AML3 cells with MA or DMSO treatment for 24 h. Methylene blue staining served as a loading control. (C) Dose-dependent effect of MA on cell viability. CCK-8 analysis of cell proliferation in OCI-AML3 cells exposed to 0, 10, 25, 50, and 100 μM MA for 24 h. (D) Time-dependent effect of MA on cell viability. CCK-8 analysis of cell proliferation in OCI-AML3 cells exposed to 100 μM MA for 0, 24, and 48 h. (E) EdU analysis of cell proliferation in OCI-AML3 cells upon 50 μM and 100 μM MA treatments of 24 h. The bar graph showed the percentage of EdU positive cells. Scale bar: 50 μm. (F) Flow cytometric analysis of the cell cycle distribution in OCI-AML3 cells upon 50 μM and 100 μM MA treatments of 24 h. The bar graph showed the percentage of G0/G1, S, and G2/M phase cells. (G) Cell apoptosis was measured by flow cytometric analysis of the Annexin V/DAPI stained cells. The bar graph showed the cell apoptosis rate. Data were presented as the mean ± SD of three independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The cell apoptosis was determined by an Annexin V-APC/DAPI Apoptosis Detection Kit (Elabscience, Wuhan, China, #E-CK-A258).

Techniques: Activity Assay, CCK-8 Assay, Transfection, Dot Blot, Staining, Control

Journal: Stem Cell Research & Therapy

Article Title: Intravenous injection of BMSCs modulate tsRNA expression and ameliorate lung remodeling in COPD mice

doi: 10.1186/s13287-024-04066-8

Figure Lengend Snippet: Tail vein injection of BMSCs alleviates lung tissue inflammation and apoptosis in COPD model mice. (A) Images of lung parenchyma and peribronchitis H&E staining, along with semi-quantitative analysis of peribronchial inflammation and quantitative analysis of lung parenchymal inflammation. (scale bar = 200 μm; N = 5; * P < 0.05). (B) TUNEL-FITC fluorescence staining images and AI analysis of mouse lung tissue cells. Green fluorescence was FITC-labeled cells with abundant dUTP nick ends, blue fluorescence was DAPI-labeled total cells, and cells labeled with both blue and green fluorescence indicated apoptotic cells. the apoptosis index (AI) wasThe ratio of the number of apoptotic cells to the total number of cells. (scale bar = 100 μm; N = 3; * P < 0.05) Control: naive - Control; COPD: COPD model; VC: COPD model-Tail Vein-Injected BMSCs

Article Snippet: TUNEL staining was conducted to label DNA-damaged apoptotic cells with Annexin V-FITC apoptosis detection kit (Beyotime Cat# C1062M, China).

Techniques: Injection, Staining, TUNEL Assay, Fluorescence, Labeling, Control