Journal: Molecular medicine (Cambridge, Mass.)

Article Title: SIRT1-mediated deacetylation of FOXO3 enhances mitophagy and drives hormone resistance in endometrial cancer.

doi: 10.1186/s10020-024-00915-7

Figure Lengend Snippet: Fig. 3 Effects of silencing or overexpressing SIRT1 on EC cell growth, proliferation, migration, and invasion. A Schematic diagram of the cell experiments; B Viability changes of RL95-2 cells after silencing or overexpressing SIRT1 at 12, 24, 36, and 48 h measured using the CCK-8 assay; C Proliferation capacity of RL95-2 cells after silencing or overexpressing SIRT1 detected using the EdU assay, where EdU-positive cells appear pink, and EdU-negative cells appear blue; D Colony formation assay to measure the colony formation ability of RL95-2 cells after silencing or overexpressing SIRT1; E Transwell assay to evaluate the migration and invasion capacity of RL95-2 cells after silencing or overexpressing SIRT1; F Wound healing assay to assess the migration of RL95-2 cells after silencing or overexpressing SIRT1; G Flow cytometry analysis to detect apoptosis of RL95-2 cells after silencing or overexpressing SIRT1. Data are presented as mean ± SD, and each cell experiment was repeated three times. *p < 0.05 compared to the sh-NC group; #p < 0.05 compared to the oe-NC group

Article Snippet: Apoptosis in EC cells was detected using the Annexin V-FITC/PI apoptosis detection kit (C1062L, Beyotime).

Techniques: Migration, CCK-8 Assay, EdU Assay, Colony Assay, Transwell Assay, Wound Healing Assay, Flow Cytometry

Journal: Molecular medicine (Cambridge, Mass.)

Article Title: SIRT1-mediated deacetylation of FOXO3 enhances mitophagy and drives hormone resistance in endometrial cancer.

doi: 10.1186/s10020-024-00915-7

Figure Lengend Snippet: Fig. 5 Impact of SIRT1 overexpression on EC cell survival via FOXO3. A Schematic diagram of the experimental procedure; B CCK-8 assay to measure the viability changes of EC cells in different intervention groups at 12, 24, 36, and 48 h; C EdU assay to assess the proliferation ability of EC cells in different intervention groups, with EdU-positive cells shown in pink and EdU-negative cells shown in blue; D Colony formation assay to evaluate the colony-forming ability of EC cells in different intervention groups; E Transwell assay to investigate the migration and invasion ability of EC cells in different intervention groups; F Wound healing assay to examine the migration of EC cells in different intervention groups; G Flow cytometry analysis to detect the apoptosis of EC cells in different intervention groups; H JC-1 staining experiment to assess the change in MMP of EC cells in different intervention groups. Data are presented as mean ± SD, with each cellular experiment repeated 3 times. *p < 0.05 compared to the oe-NC + sh-NC group; #p < 0.05 compared to the oe-SIRT1 + sh-NC group

Article Snippet: Apoptosis in EC cells was detected using the Annexin V-FITC/PI apoptosis detection kit (C1062L, Beyotime).

Techniques: Over Expression, CCK-8 Assay, EdU Assay, Colony Assay, Transwell Assay, Migration, Wound Healing Assay, Flow Cytometry, Staining

Journal: Molecular medicine (Cambridge, Mass.)

Article Title: SIRT1-mediated deacetylation of FOXO3 enhances mitophagy and drives hormone resistance in endometrial cancer.

doi: 10.1186/s10020-024-00915-7

Figure Lengend Snippet: Fig. 8 Overexpression of SIRT1 enhances the intracellular growth of EC cells. A Diagram illustrating the procedure of animal experiments (n = 6); B Line graph showing the tumor volume increase in subcutaneous tumor model mice from day 8 to 44 (n = 6); C Dissection of subcutaneous transplanted tumors in mice from each group on day 44 (n = 6); D Statistical analysis of tumor weight in subcutaneous tumor model mice on day 44 (n = 6); E TUNEL staining for apoptosis in mouse tumors from each group (n = 6); F MitoSOX immunofluorescence staining for ROS production in mouse tumor tissues from each group (n = 6). Data are presented as mean mean ± SD, with 6 nude mice in each group. *p < 0.05 compared to sh-NC group; #p < 0.05 compared to oe-NC group

Article Snippet: Apoptosis in EC cells was detected using the Annexin V-FITC/PI apoptosis detection kit (C1062L, Beyotime).

Techniques: Over Expression, Dissection, TUNEL Assay, Staining, Immunofluorescence

Journal: Molecular medicine (Cambridge, Mass.)

Article Title: SIRT1-mediated deacetylation of FOXO3 enhances mitophagy and drives hormone resistance in endometrial cancer.

doi: 10.1186/s10020-024-00915-7

Figure Lengend Snippet: Fig. 10 Overexpression of SIRT1 enhances tumor growth by inducing cell autophagy. A Schematic diagram of the animal experiment process (n = 6); B Line graph showing the growth of subcutaneously transplanted tumors in mice from day 8 to day 36 (n = 6); C Anatomical diagram of subcutaneously transplanted tumors in mice on day 36 (n = 6); D Statistics of tumor weight in mice with subcutaneously transplanted tumors on day 36 (n = 6); E TUNEL staining to detect apoptosis of tumor cells in mice in each group (n = 6); F MitoSOX immunofluorescence staining to detect the generation of ROS in tumor tissues of mice in each group (n = 6). Data are presented as mean ± SD, with 6 nude mice in each group. *p < 0.05 compared to the PBS group; #p < 0.05 between the two groups

Article Snippet: Apoptosis in EC cells was detected using the Annexin V-FITC/PI apoptosis detection kit (C1062L, Beyotime).

Techniques: Over Expression, TUNEL Assay, Staining, Immunofluorescence

Journal: Medical science monitor : international medical journal of experimental and clinical research

Article Title: The Function of the Novel Mechanical Activated Ion Channel Piezo1 in the Human Osteosarcoma Cells.

doi: 10.12659/msm.906959

Figure Lengend Snippet: Figure 7. (A) Images of nuclei shrinking and evident nuclear condensation of osteosarcoma (OS) cells (shown by arrowhead) after mechanical stretch forces of 0, 2, 12, 24, and 48 hours. Notes: (a) 0-hour group 200×. (b) 0-hour plus shPiezo1RNA group 200x. (c) 2-hour group 200×. (d) 2-hour plus shPiezo1RNA group 200×. (e) 12-hour group 200×. (f) 12-hour plus shPiezo1RNA group 200×. (g) 24-hour group 200×. (h) 24-hour plus shPiezo1RNA group 200×. (i) 48-hour group 200×. (j) 48-hour plus shPiezo1RNA group 200×. (B) Results of flow cytometry detection of 0S cell apoptosis in 0, 2, 12, 24, and 48 hours and shPiezo1RNA groups. Q1: (AnnexinV-FITC)-/PI+, necrotic cells. Q2: (AnnexinV-FITC)+/PI+, late stage apoptotic cells. Q3: (AnnexinV-FITC)+/PI–, early stage apoptotic cells. Q4: (AnnexinV-FITC)-/PI-, live cells. Notes: (a) 0-hour group. (b) 0-hour group plus shPiezo1RNA. (c) 2-hour group. (d) 2-hour group plus shPiezo1RNA. (e) 12-hour group. (f) 12-hour group plus shPiezo1RNA. (g) 24-hour group. (h) 24-hour group plus shPiezo1RNA. (i) 48-hour group. (j) 48-hour group plus shPiezo1RNA. (C) Early stage apoptosis of human OS cells in 0, 2, 12, 24, 48 hors and shPiezo1RNA groups. After a 2 hour mechanical stretch, the rate of early stage apoptosis significantly increased. After a 12 hour mechanical stretch, the late stage apoptotic rate increased and peaked in the 24-hour group. The late stage apoptotic rate was lower in the 48-hour group compared to the 24-hour group. The results are presented as the means ±SD, n=3; ns means p>0.05 versus the control group; * p<0.05 versus the control group; ** p<0.01 versus the control group; one-way ANOVA.

Article Snippet: Cells were stained with FITC-conjugated Annexin V and propidium iodide (PI) according to the manufacturer’s instructions for the Apoptosis Detection Kit (R&D Systems, MN, USA).

Techniques: Flow Cytometry, Control

Journal: Frontiers in Oncology

Article Title: AXL Promotes Metformin-Induced Apoptosis Through Mediation of Autophagy by Activating ROS-AMPK-ULK1 Signaling in Human Esophageal Adenocarcinoma

doi: 10.3389/fonc.2022.903874

Figure Lengend Snippet: Downregulation of AXL expression attenuates metformin-induced apoptotic cell death in EAC cells. SK-GT-4-shCtrl and SK-GT-4-shAXL cells (A) or FLO-1-shCtrl and FLO-1-shAXL cells (B) were subjected to CCK-8 cell viability assay following treatment with the indicated metformin concentrations for 72 h. (C) SK-GT-4-shCtrl and SK-GT-4-shAXL cells were treated with vehicle or metformin (15 mM) for 96 h. Apoptosis was determined by Annexin-V/FITC staining and FACS analysis. (D) Immunoblot analysis of AXL, caspase-3, and PARP proteins in cell lysates from SK-GT-4-shCtrl and SK-GT-4-shAXL cells treated with vehicle or 15 mM metformin for 96 h. (E) Apoptosis in FLO-1-shCtrl and FLO-1-shAXL cells after treatment with vehicle or 10 mM metformin for 72 h was assessed as in panel (C) . (F) Western blot analysis of AXL, caspase-3, and PARP proteins in FLO-1-shCtrl and FLO-1-shAXL cells treated with vehicle or 10 mM metformin for 72 h. Gel loading was normalized for equal β-actin. Data are representative of three independent experiments and statistical analysis was evaluated by one-way ANOVA followed by the Newman-Keuls post hoc test.

Article Snippet: Briefly, cells were harvested and stained with Annexin-V fluorescein isothiocyanate (FITC) using TACS Annexin V-FITC Apoptosis Detection Kit (4830-250-K, R&D Systems) and propidium iodide (PI) (P4170, Sigma-Aldrich).

Techniques: Expressing, CCK-8 Assay, Viability Assay, Staining, Western Blot

Journal: Frontiers in Oncology

Article Title: AXL Promotes Metformin-Induced Apoptosis Through Mediation of Autophagy by Activating ROS-AMPK-ULK1 Signaling in Human Esophageal Adenocarcinoma

doi: 10.3389/fonc.2022.903874

Figure Lengend Snippet: Metformin-induced apoptosis requires autophagy in EAC cells. (A) FLO-1 cells stably expressing control shRNA (shCtrl) or Beclin 1 shRNA (shBeclin1) were treated with increasing concentrations of metformin for 72 h and then subjected to CCK-8 cell viability assay. (B) Western blot analysis of Beclin 1, caspase-3, and PARP proteins in FLO-1-shCtrl and FLO-1-shBeclin1 cells treated with vehicle or 10 mM metformin for 72 h. (C) Immunoblotting of ATG7, Caspase-3, and PARP proteins in FLO-1-shCtrl and FLO-1-shATG7 cells treated with vehicle or 10 mM metformin for 72 h. (D) Western blot analysis of ATG7, caspase-3, and PARP proteins in SK-GT-4-shCtrl and SK-GT-4-shATG7 cells treated with 15 mM metformin for 96 h. (E) Western blot analysis of Beclin 1, p-AMPK (T172), AMPK, and LC3B proteins in FLO-1-shCtrl and FLO-1-shBeclin1 cells treated with vehicle or 10 mM metformin for 2 h. (F) Western blot analysis of ATG7, p-AMPK (T172), AMPK, and LC3B proteins in FLO-1-shCtrl and FLO-1-shATG7 cells treated with vehicle or 10 mM metformin for 2 h. (G) Immunoblotting of ATG7, p-AMPK (T172), AMPK, and LC3B proteins in SK-GT-4-shCtrl and SK-GT-4-shATG7 cells treated with 15 mM metformin for 2 h. Gel loading was normalized for equal β-actin.

Article Snippet: Briefly, cells were harvested and stained with Annexin-V fluorescein isothiocyanate (FITC) using TACS Annexin V-FITC Apoptosis Detection Kit (4830-250-K, R&D Systems) and propidium iodide (PI) (P4170, Sigma-Aldrich).

Techniques: Stable Transfection, Expressing, Control, shRNA, CCK-8 Assay, Viability Assay, Western Blot

Journal: Frontiers in Oncology

Article Title: AXL Promotes Metformin-Induced Apoptosis Through Mediation of Autophagy by Activating ROS-AMPK-ULK1 Signaling in Human Esophageal Adenocarcinoma

doi: 10.3389/fonc.2022.903874

Figure Lengend Snippet: Expression of AXL sensitizes tumors to metformin in a xenograft mouse model. Four groups (10 mice per group) of Rag2 mice were injected on both hip flanks with FLO-1-shCtrl (two groups, no treatment and treatment) or FLO-1-shAXL cells (two groups, no treatment and treatment). When tumors reached a size of approximately 250 mm 3 , the mice were randomized and not treated or treated by oral gavage with metformin (250 mg/kg) daily for 12 days. The metformin dose was then increased to 500 mg/kg daily until the end of the experiment (day 28). (A) Treatment with metformin significantly reduced tumor size in shCtrl cells ( P = 0.004) relative to nontreated tumors. Knockdown of AXL expression (shAXL cells) completely abrogated the suppressive effect of metformin on tumor growth. (B) Representative images of endpoint xenografts depicting the effect of metformin on tumor size. Images (200x magnification) of H&E staining (C) and IHC analysis for expression for Ki-67, a marker of proliferation (C , D) ; cleaved caspase-3, a marker of apoptosis (C , E) ; and p-AMPK (T172), a marker of autophagy (C , F) in representative endpoint tumors of all metformin-treated and non-treated animal groups. (G) Real-time PCR data depicting relative AXL mRNA expression in representative endpoint tumors of all the animal groups. n.s., not significant.

Article Snippet: Briefly, cells were harvested and stained with Annexin-V fluorescein isothiocyanate (FITC) using TACS Annexin V-FITC Apoptosis Detection Kit (4830-250-K, R&D Systems) and propidium iodide (PI) (P4170, Sigma-Aldrich).

Techniques: Expressing, Injection, Knockdown, Staining, Marker, Real-time Polymerase Chain Reaction

Journal: Cell Death & Disease

Article Title: Urocortin protects chondrocytes from NO-induced apoptosis: a future therapy for osteoarthritis?

doi: 10.1038/cddis.2013.231

Figure Lengend Snippet: Apoptotic and necrotic chondrocyte cell death assessed as the percentage of Annexin V- and TUNEL-positive cells (apoptosis) and cellular LDH release (necrosis) following treatment of C-20/A4 cells with increasing concentrations of SNAP ( a ) and TNF- α ( b ). * P <0.05. ** P <0.01 compared with control (untreated)

Article Snippet: Apoptosis was assessed by fluorescence microscopy by Annexin V binding (TACS Annexin V-FITC kit, R&D systems) and by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL; in situ cell death detection kit, Roche diagnostics, Lewes, UK).

Techniques: TUNEL Assay, Control

Journal: Cell Death & Disease

Article Title: Urocortin protects chondrocytes from NO-induced apoptosis: a future therapy for osteoarthritis?

doi: 10.1038/cddis.2013.231

Figure Lengend Snippet: Cytoprotective effects of endogenous Ucn. Apoptotic and necrotic chondrocyte cell death assessed as the percentage of Annexin V- and TUNEL-positive cells (apoptosis) and cellular LDH release (necrosis) following the treatment of C-20/A4 cells, with the CRH receptor antagonist α -helical CRH, anti-UCN antibody and anti-albumin antibody, ** P <0.01 compared with control

Article Snippet: Apoptosis was assessed by fluorescence microscopy by Annexin V binding (TACS Annexin V-FITC kit, R&D systems) and by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL; in situ cell death detection kit, Roche diagnostics, Lewes, UK).

Techniques: TUNEL Assay, Control

Journal: Cell Death & Disease

Article Title: Urocortin protects chondrocytes from NO-induced apoptosis: a future therapy for osteoarthritis?

doi: 10.1038/cddis.2013.231

Figure Lengend Snippet: Cytoprotective effects of exogenous Ucn. Apoptotic and necrotic chondrocyte cell death assessed as the percentage of Annexin V- and TUNEL-positive cells (apoptosis) and cellular LDH release (necrosis) following treatment of C-20/A4 cells with Ucn (U) administered concurrently with SNAP (S) or TNF- α (T) in the presence or absence of α -helical CRH (A) * P <0.05, ** P <0.01 compared with control (C).

Article Snippet: Apoptosis was assessed by fluorescence microscopy by Annexin V binding (TACS Annexin V-FITC kit, R&D systems) and by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL; in situ cell death detection kit, Roche diagnostics, Lewes, UK).

Techniques: TUNEL Assay, Control