Journal: Cell Death & Disease

Article Title: Honokiol induces apoptosis and autophagy via the ROS/ERK1/2 signaling pathway in human osteosarcoma cells in vitro and in vivo

doi: 10.1038/s41419-017-0166-5

Figure Lengend Snippet: a The anti-proliferative effect of HNK on osteosarcoma cell lines was determined by MTT. Cells were treated with various concentrations of HNK for 24, 48, and 72 h. Control group contained 0.1% DMSO. Data represented the mean of five replicates. b Colony-formation assay of HOS and U2OS cells with control or HNK. c Comparison of the effect of HNK on two normal human primary skin fibroblast samples with that on osteosarcoma cells for 24 h. d HNK-induced G0/G1 phase arrest. Cells were treated with control or HNK for 24 h and analyzed by flow cytometry. e HOS and U2OS cells were treated with HNK for 24 h. The expressions of cell cycle-regulated proteins were measured by western blot. f Intracellular proteasome activity in HOS and U2OS cells after treatment with HNK. Cells were treated with 5, 10, 20 or 30 μM HNK for 24 h. * p < 0.05, HNK vs. control. g Western blot for GRP78 in HOS and U2OS cells after treatment with HNK. Cells were treated with 5, 10, 20 or 30 μM HNK for 24 h. h The intracellular Ca 2+ concentration in HOS and U2OS after treatment with HNK. Cells were treated with 5, 10, 20 or 30 μM HNK for 24 h. The ratio of 350 to 380 nm was measured by an ELISA reader. * p < 0.05, HNK vs. control. Data are presented as the mean ± s.d. of three independent experiments. Semi-quantification of western blot bands is presented in Figure

Article Snippet: HOS (ATCC: CRL-1543) and U2OS (HTB-96TM, ATCC) human osteosarcoma cells were obtained from the American Type Culture Collection (ATCC).

Techniques: Control, Colony Assay, Comparison, Flow Cytometry, Western Blot, Activity Assay, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: Cell Death & Disease

Article Title: Honokiol induces apoptosis and autophagy via the ROS/ERK1/2 signaling pathway in human osteosarcoma cells in vitro and in vivo

doi: 10.1038/s41419-017-0166-5

Figure Lengend Snippet: a HOS and U2OS cells treated with HNK were stained with annexin V-FITC/PI and analyzed by flow cytometry. The chart illustrates apoptosis proportion from three separate experiments. b The mitochondrial membrane potential was measured with JC-1 fluorescent probe and assessed by flow cytometry. The chart illustrates changes of JC-1 red/green rate from three independent experiments. c Cells were treated with various concentrations of HNK for 24 h. The expressions of cleaved PARP, caspase-3, caspase-9, BCL-2, BCL-xl, and survivin were determined by western blot. Semi-quantification of western blot bands is presented in Figure

Article Snippet: HOS (ATCC: CRL-1543) and U2OS (HTB-96TM, ATCC) human osteosarcoma cells were obtained from the American Type Culture Collection (ATCC).

Techniques: Staining, Flow Cytometry, Membrane, Western Blot

Journal: Cell Death & Disease

Article Title: Honokiol induces apoptosis and autophagy via the ROS/ERK1/2 signaling pathway in human osteosarcoma cells in vitro and in vivo

doi: 10.1038/s41419-017-0166-5

Figure Lengend Snippet: a Cells were treated with HNK for 24 h and then loaded with DCFH-DA for 30 min. The level of ROS was determined by flow cytometry. Representative images are presented. Quantitative analysis of ROS generation is shown in histograms. * P < 0.05 vs. control. b Cells were preincubated with NAC (5 mM) for 2 h, and then treated with HNK (30 μM) for 24 h. Level of ROS was determined by flow cytometry. c HOS cell viability was measured by MTT. d HOS cells were preincubated with NAC (5 mM) for 2 h, and then treated with HNK (30 μM) for 24 h. Levels of PARP, cleaved caspase-3, caspase-9, BCL-2, BCL-xl, and survivin were determined by western blot. e The expressions of HOS cell cycle-regulated proteins were measured by western blot. f Levels of phospho-ERK, ERK , and LC3B-II were determined by western blot. g HOS and U2OS cells were preincubated with PD98059 (40 μM) for 2 h, and then treated with HNK (30 μM) for 24 h. Level of ROS was determined by flow cytometry. Semi-quantification of western blot bands is presented in Figure S

Article Snippet: HOS (ATCC: CRL-1543) and U2OS (HTB-96TM, ATCC) human osteosarcoma cells were obtained from the American Type Culture Collection (ATCC).

Techniques: Flow Cytometry, Control, Western Blot

Journal: Cell Death & Disease

Article Title: Honokiol induces apoptosis and autophagy via the ROS/ERK1/2 signaling pathway in human osteosarcoma cells in vitro and in vivo

doi: 10.1038/s41419-017-0166-5

Figure Lengend Snippet: a Western blot of Atg7 in HOS cells treated with 30 μM honokiol for the indicated times. b Real-time PCR analysis of the expression of Atg7 in HOS cells. c Western blotting showed the repression of LC3B levels in HOS cells after siRNA-Atg7 treatment. d HOS and U2OS cells were preincubated with 3-MA (2.5 mM) or z-VAD-fmk (20 μM) for 2 h, and then treated with HNK (30 μM) for 24 h. Cell viability was measured by MTT. * P < 0.05. e The level of MDC in HOS cells are presented. f HOS cells were preincubated with z-VAD-fmk (20 μM) for 2 h, and then treated with HNK (30 μM) for 24 h. Levels of LC3B, cleaved PARP and caspase-3 were assessed by western blot . *P < 0.05 vs. control. g HOS cells were preincubated with 3-MA (2.5 mM) for 2 h, and then treated with HNK (30 μM) for 24 h. Cells were stained with annexin V-FITC/PI and analyzed by flow cytometry. h HOS cells were preincubated with 3-MA (2.5 mM) for 2 h, and then treated with HNK (30 μM) for 24 h. Levels of LC3B, cleaved PARP and caspase-3 were assessed by western blot. * P < 0.05 vs. control. Semi-quantification of western blot bands is presented in Figure S

Article Snippet: HOS (ATCC: CRL-1543) and U2OS (HTB-96TM, ATCC) human osteosarcoma cells were obtained from the American Type Culture Collection (ATCC).

Techniques: Western Blot, Real-time Polymerase Chain Reaction, Expressing, Control, Staining, Flow Cytometry