Journal: Advanced Science

Article Title: Cryo‐Inactivated Cancer Cells Derived Magnetic Micromotors for Tumor Immunotherapy

doi: 10.1002/advs.202504986

Figure Lengend Snippet: In vitro anti‐cancer efficacy of CICC@FeMnP and DC maturation. a) Live/dead staining images of 4T1 cells after different treatments. Scale bars, 100 µm. (b, c) FCM analysis b) and corresponding apoptosis rates c) of 4T1 cells after various treatments. d) Analysis of 4T1 cell viability following various treatments using the CCK8 assay. e,f) CRT (e) and HMGB1 (f) expression of 4T1 cells after various treatments by immunofluorescence staining. Scale bars, 50 µm. g) WB analysis of p‐STING, p‐IRF3, and p‐TBK1 expression in 4T1 cells incubated with various formulations. h) Schematic diagram of ICD and DC maturation processes. i,j) FCM analysis of DC maturation (i) and corresponding maturation ratios (j) after various treatments. k,l) Quantitative assessment of cytokine secretion (TNF‐α and IL‐6) by DCs. Data are presented as the mean ± SD. n. s.: no significance, *** p < 0.001.

Article Snippet: DCFH‐DA reactive oxygen species assay kit, CCK8 assay kit, mitochondrial membrane potential assay kit with JC‐1, annexin V‐FITC/PI apoptosis detection kit were purchased from Beyotime Biotechnology Co., Ltd. Prussian blue stain kit was obtained from Solarbio Science & Technology Co., Ltd. C11‐BODIPY 581/591 was bought from Invitrogen.

Techniques: In Vitro, Staining, CCK-8 Assay, Expressing, Immunofluorescence, Incubation

Journal: Molecular Biotechnology

Article Title: miR-324-3p Suppresses Hepatic Stellate Cell Activation and Hepatic Fibrosis Via Regulating SMAD4 Signaling Pathway

doi: 10.1007/s12033-024-01078-w

Figure Lengend Snippet: MiR-324-3p suppresses the activation of transforming growth factor (TGF)-β1- induced LX-2 cells in vitro. A Hepatic stellate cells (HSCs) were transfected with miR-324-3p mimic, and expression of the miR-324-3p was determined by real-time-quantitative polymerase chain reaction (RT-qPCR). B RT-qPCR to analyze the miR-324-3p expression in HSCs transfected with the miR-324-3p inhibitor. C Cell counting kit-8 (CCK-8) to analyze the proliferation of transforming growth factor (TGF)-β1-induced LX-2 cells with indicated treatment. D Flow cytometry to detect the cycle and apoptosis of TGF-β1-induced LX-2 cells. E , G Western blot (WB) assay and RT-qPCR to evaluate α-smooth muscle actin (α-SMA) and Vimentin expression in transfected HSC cells. F , H The α-SMA, and Vimentin levels in transfected cells were analyzed by WB assay as well as RT-qPCR. * P < 0.05 and ** P < 0.01 vs . Control group; # P < 0.05 and ## P < 0.01 vs . NC + TGF-β1 group

Article Snippet: Then, the apoptosis was detected using the Annexin V-FITC/PI Apoptosis Analysis Kit (Bestbio, China) and analyzed using a CytoFLEX flow cytometer (Beckman Coulter, USA).

Techniques: Activation Assay, In Vitro, Transfection, Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Cell Counting, CCK-8 Assay, Flow Cytometry, Western Blot, Control

Journal: Cell Death & Disease

Article Title: 4EBP1 senses extracellular glucose deprivation and initiates cell death signaling in lung cancer

doi: 10.1038/s41419-022-05466-5

Figure Lengend Snippet: A , B A549 and H1299 cells transfected with either control (CTL) siRNA or 4EBP1 siRNAs ( A ), A549 and H1299 cells were transfected with CTL siRNAs or 4EBP1 siRNA or cotransfected with CTL siRNAs and mutant plasmid eIF4E-S209E ( B ). Cells were treated with normal or glucose starvation. Cell growth rate was assessed by MTT assay. The data represent the averages of three independent experiments (mean ± SD). * P < 0.05, ** P < 0.01, *** P < 0.001. C A549 cells transfected with either control siRNA or 4EBP1 siRNAs were treated with normal or glucose starvation for 16 h. The changes of cell morphological was investigated (scale bar = 100 µm, magnification: ×100). D A549 cells transfected with either control siRNA or 4EBP1 siRNAs were treated with normal or glucose starvation for 16 h. Cell apoptosis analysis was done by flow cytometry. E A549 cells transfected with either control siRNA or 4EBP1 siRNAs were treated with normal (0 h) or glucose starvation. The expression levels of the apoptosis related proteins were checked by western blot. C-Caspase3: Cleaved-Caspase3. F , G parental A549 (CON) and A549 stable cell lines with 4EBP1 knockdown (A549-sh4EBP1) were treated with normal or glucose starvation for 12 h. Lactic acid levels were monitored ( F , upper panel). ECAR ( F , lower panel) and OCR ( G ) measurements were done.

Article Snippet: Flow cytometric analysis of apoptotic cells was performed using the Annexin V-FITC / PI Apoptosis Detection Kit (Elabscience).

Techniques: Transfection, Control, Mutagenesis, Plasmid Preparation, MTT Assay, Flow Cytometry, Expressing, Western Blot, Stable Transfection, Knockdown

Journal: Cell Death & Disease

Article Title: 4EBP1 senses extracellular glucose deprivation and initiates cell death signaling in lung cancer

doi: 10.1038/s41419-022-05466-5

Figure Lengend Snippet: A Western blot analysis of 4EBP1 expression in parental A549 and A549-sh4EBP1 cells. B Parental A549 and A549-sh4EBP1 cells were treated with normal or glucose starvation for 48 h. Cell growth rate was assessed by MTT assay. The data represent the averages of three independent experiments (mean ± SD). * P < 0.05. C Parental A549 and A549-sh4EBP1 cells were treated with normal or glucose starvation for 16 h. The changes of cell morphological was investigated (scale bar = 100 µm, magnification: ×100). D Parental A549 and A549-sh4EBP1 cells were treated with normal or glucose starvation for 20 h. Cell apoptosis analysis was done by flow cytometry. E , F In vivo xenograft assay was performed using parental A549 and A549-sh4EBP1 cells, tumors were photographed ( E ), and their weights and volumes were measured ( F ). The P value was calculated by paired t -test. * P < 0.05. G Immunohistochemical staining in the tumor-central region of tumor induced by parental A549 and A549-sh4EBP1 cells for 4EBP1, p-STAT3, BCL2, MCL1, Survivin and Cleaved PARP1 (C-PARP1). Scale bars: 50 μm, magnification: ×200.

Article Snippet: Flow cytometric analysis of apoptotic cells was performed using the Annexin V-FITC / PI Apoptosis Detection Kit (Elabscience).

Techniques: Western Blot, Expressing, MTT Assay, Flow Cytometry, In Vivo, Xenograft Assay, Immunohistochemical staining, Staining

Journal: European Journal of Medical Research

Article Title: Tanshinone IIA induces ferroptosis in colorectal cancer cells through the suppression of SLC7A11 expression via the PI3K/AKT/mTOR pathway

doi: 10.1186/s40001-025-02842-7

Figure Lengend Snippet: Effects of Tan IIA on OUMS23 cell proliferation and apoptosis. OUMS23 cells were exposed to various concentration of Tan IIA. A OUMS23 cells were labeled with EdU and their proliferation was examined. B Flow cytometry analysis of apoptotic cells. ** P < 0.01 vs. the 0 μM Tan IIA treatment group

Article Snippet: After digesting with trypsin, OUMS23 cells were collected by centrifugation at 4 °C for 5 min. Then, the cells were washed twice with PBS (AS1044, ASPEN) and stained using the Annexin V/propidium iodide (PI) Apoptosis Detection Kit (Beyotime).

Techniques: Concentration Assay, Labeling, Flow Cytometry

Journal: Journal of Hematology & Oncology

Article Title: Reductive TCA cycle catalyzed by wild-type IDH2 promotes acute myeloid leukemia and is a metabolic vulnerability for potential targeted therapy

doi: 10.1186/s13045-022-01245-z

Figure Lengend Snippet: Over-expression of wild-type IDH2 in AML and its effect on leukemia cell proliferation. a Comparison of IDH2 mRNA levels in AML ( n = 80) and normal cells (monocytes and neutrophils, n = 6), using the leukemia dataset (Stegmaier Leukemia datasets) available in the Oncomine database. b IDH2 mRNA levels in primary AML samples ( n = 204) in comparison with normal cells (hematopoietic stem cells, n = 6; metamyelocytes, n = 3; band cells, n = 3; polymorphonuclear cells, n = 3; monocytes, n = 4) available in the BloodSpot datasets. c Western blotting of IDH2 in normal human peripheral blood mononuclear cells (PBMC #1–#3), AML cell lines and PBMCs from AML patients with wt-IDH2 (AML#1–#8), β-actin was used as a loading control. d Relative mRNA level of IDH2 in U937 and ML-1 cells transfected with control shRNA (shRNA-Ctrl) or IDH2 shRNA (shIDH2#1, shIDH2#2) was measured by RT-qPCR. e Western blotting analysis of IDH2 protein in U937 and ML-1 cells transfected with shRNA-Ctrl or IDH2 shRNA. f , g Measurement of cell proliferation in U937 and ML-1 cells expressing shRNA-Ctrl or IDH2 shRNA. h Relative IDH2 mRNA level in HL-60 cells transfected with control vector (Vector) or IDH2 over-expression vector (IDH2 OE ). Expression of mRNA was measured by RT-qPCR. i Western blotting analysis of IDH2 protein levels in HL-60 cells transfected with control vector or IDH2 over-expression vector. j Comparison of cell proliferation in HL-60 cells transfected with control vector or IDH2 over-expression vector. k , l Colony formation assay in U937 and ML-1 cells expressing shRNA-Ctrl or IDH2 shRNA. Colonies were counted under inverted microscope. m Comparison of percentage of Annexin V/ PI negative cells in U937 and ML-1 transfected with shRNA-Ctrl or IDH2 shRNA for 48 h. The original flow cytometry analysis data are presented as Additional file : Fig. S1h. n = 3, mean ± SD ; * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: For analysis of apoptosis, C-MYC knockdown cell death, and the number of apoptotic cells after AGI-6780 and dimethyl-αKG treatment, was analyzed by FITC-Annexin V/PI assay kit (BD, Franklin Lakes, NJ, USA).

Techniques: Over Expression, Western Blot, Transfection, shRNA, Quantitative RT-PCR, Expressing, Plasmid Preparation, Colony Assay, Inverted Microscopy, Flow Cytometry

Journal: Journal of Hematology & Oncology

Article Title: Reductive TCA cycle catalyzed by wild-type IDH2 promotes acute myeloid leukemia and is a metabolic vulnerability for potential targeted therapy

doi: 10.1186/s13045-022-01245-z

Figure Lengend Snippet: Cytotoxic effect of α-KG in AML cells with wild-type IDH2. a Induction of apoptosis by cell-permeable DM-αKG in AML cell lines (U937 and ML-1) and primary AML cells isolated from patients with wt-IDH2 (AML#1 and AML#2). Cells were treated with the indicated concentrations of DM-αKG for 48 h, and apoptosis was measured by flow cytometry analysis of annexin-V positivity. The number inside each panel shows the percentage of dead cells. b , c Quantitation of the concentration-dependent apoptosis induced by DM-αKG in U937 and ML-1 cells ( n = 3, mean ± SD ). d Apoptosis of human primary AML cells harboring wild-type ( n = 7) or mutant IDH2 (IDH2-R140Q, n = 2) treated with various concentrations (2, 4, 6, 8 and 10 mM) of DM-αKG for 48 h. Apoptosis was measured by flow cytometry analysis of annexin-V positivity

Article Snippet: For analysis of apoptosis, C-MYC knockdown cell death, and the number of apoptotic cells after AGI-6780 and dimethyl-αKG treatment, was analyzed by FITC-Annexin V/PI assay kit (BD, Franklin Lakes, NJ, USA).

Techniques: Isolation, Flow Cytometry, Quantitation Assay, Concentration Assay, Mutagenesis

Journal: Journal of Hematology & Oncology

Article Title: Reductive TCA cycle catalyzed by wild-type IDH2 promotes acute myeloid leukemia and is a metabolic vulnerability for potential targeted therapy

doi: 10.1186/s13045-022-01245-z

Figure Lengend Snippet: Inhibition of IDH2 by AGI-6780 suppressed AML survival in vitro and in vivo. a , b Effect of AGI-6780 on cell proliferation in U937 and ML-1 cells. Cells were treated with AGI-6780 (10 µM) or solvent (DMSO), and cell numbers were counted at the indicated time points. Data are mean ± SD of three experiments. c Levels of protein expression of IDH2 and C-MYC in U937 and ML-1 cells treated with AGI-6780 (10 µM) or solvent (DMSO) for 24 h. d The indicated cell line were treated with AGI-6780 for 72 h, and cell viability was measured using MTS assay. e Primary leukemia cells from AML patients with wt-IDH2 ( n = 4) and normal human bone marrow cells (HBMC) were treated ex vivo with the indicated concentrations of AGI-6780 for 72 h, and cell viability was measured using MTS assay. f Comparison of percentage of Annexin V/PI-negative cells in primary AML cells and primary normal bone marrow cells treated with 10–20 µM AGI-6780 for 48 h. The original flow cytometry analysis data are presented as Additional file : Fig. S8c, d. g Protein levels of IDH2 and C-MYC in primary AML cells treated with AGI-6780 (20 µM) or with solvent (DMSO) for 24 h. h Growth curves of AML xenografts in athymic nude mice inoculated with ML-1 cells harboring wt-IDH2 ( n = 6 per group, mean ± SEM ). Mice were treated with daily i.p. injection of AGI-6780 or solvent as indicated. i Photographs of gross tumors isolated from mice at the end of the experiment. j Western blot analysis of C-MYC expression in tumor tissues of ML-1 xenografts isolated from mice treated with AGI-6780 or solvent as indicated. k Tumor weights of each group at the end of the experiment ( mean ± SEM ). l Relative C-MYC protein levels in tumor tissues of mice treated with solvent ( n = 6) or AGI-6780 ( n = 6). m Mouse body weights ( mean ± SD ). Two-tailed unpaired student’s t test for a , b , f , h , k and l , * p < 0.05, *** p < 0.001

Article Snippet: For analysis of apoptosis, C-MYC knockdown cell death, and the number of apoptotic cells after AGI-6780 and dimethyl-αKG treatment, was analyzed by FITC-Annexin V/PI assay kit (BD, Franklin Lakes, NJ, USA).

Techniques: Inhibition, In Vitro, In Vivo, Expressing, MTS Assay, Ex Vivo, Flow Cytometry, Injection, Isolation, Western Blot, Two Tailed Test