Journal: International Journal of Molecular Sciences

Article Title: Enhanced Anti-Cancer Effects of Conditioned Medium from Hypoxic Human Adult Dermal Fibroblasts on Cervical Cancer Cells

doi: 10.3390/ijms23095134

Figure Lengend Snippet: Nodes and edges in the PPIs of intracellular proteins up- and down-regulated (> 2-fold) in HeLa cells treated with H-CM compared with N-CM.

Article Snippet: The membranes were blocked with 4% skim milk for 2 h at room temperature and then incubated with rabbit anti-human CD19 (1:3000 dilution; cat. no. CSB-RA780821A0HU; CUSABIO, Wuhan, China), CHEK1 (1:3000 dilution; cat. no. CSB-RA176809A0HU; CUSABIO), ESR1 (1:3000 dilution; cat. no. CSB-PA11399A0Rb; CUSABIO), LCK (1:3000 dilution; cat. no. CSB-PA009798; CUSABIO), MCL1 (1:3000 dilution; cat. no. CSB-PA03829A0Rb; CUSABIO), PCNA (1:3000 dilution; cat. no. CSB-PA-208009; CUSABIO), POLA1 (1:3000 dilution; cat. no. CSB-PA002170; CUSABIO), TBP (1:3000 dilution; cat. no. CSB-RA821481A0HU; CUSABIO), or TNF antibody (1:3000 dilution; cat. no. CSB-PA07427A0Rb; CUSABIO) and mouse anti-human ꞵ-actin antibody (1:10,000 dilution; cat. no. A1978; Sigma) overnight at −4 °C.

Techniques:

Journal: BMC Cancer

Article Title: Homoharringtonine interacts synergistically with bortezomib in NHL cells through MCL-1 and NOXA-dependent mechanisms

doi: 10.1186/s12885-018-5018-x

Figure Lengend Snippet: The homoharringtonine/bortezomib regimen induces caspase activation and alters the NOXA/MCL1 ratio. a , b OCI-LY18, Carnaval, HBL1 and NCEB cells were treated with HHT (10 to 20 nM) alone or with bortezomib (2 to 3 nmol/L) for 24 h after which cells were lysed and proteins extracted. Expression of the indicated proteins was determined by Western blotting using the indicated antibodies. Each lane was loaded with 25 μg of protein; blots were stripped and re-probed with tubulin/actin to ensure equivalent loading and transfer. Results are representative of three replicate experiments. c SU-DHL-4 cells were exposed the HHT (25 nmol/L) and bortezomib (4 nmol/L) individually or together for 24 h after which cells were lysed and subjected to immunoprecipitation using BAK or NOXA Abs. The immunoprecipitates were separated by SDS-PAGE and immunoblotted with MCL-1 (left and right panel)

Article Snippet: Primary antibodies used in these studies were: cleaved PARP, cleaved caspase-3, BCL- XL , BIM (Cell Signaling Technology, Danvers, MA), MCL-1 (BD Biosciences, San Jose, CA), α-tubulin (EMD Millipore, Billerica, MA), BAX (N20), BAK (G23), actin (Sigma-Aldrich, St. Louis, MO), NOXA (Enzo Life Sciences, Farmingdale, NY).

Techniques: Activation Assay, Expressing, Western Blot, Immunoprecipitation, SDS Page

Journal: BMC Cancer

Article Title: Homoharringtonine interacts synergistically with bortezomib in NHL cells through MCL-1 and NOXA-dependent mechanisms

doi: 10.1186/s12885-018-5018-x

Figure Lengend Snippet: Genetic inhibition of MCL-1 or overexpression of NOXA renders cells significantly more sensitive to bortezomib . a SU-DHL-4 cells were transfected with shRNA constructs designed against MCL-1. Two clones of shRNA MCL-1 were selected. These 2 clones of SU-DHL-4/shMCL1 and shControl cells were exposed to bortezomib (4 nM) for 48 h after which cells were analyzed by flow cytometry and western blot. b Similarly, SU-DHL-16 cells were transfected with shRNA constructs designed against MCL-1. SU-DHL-16/shMCL1 and shControl cells were exposed to bortezomib (1.5 nM) for 48 h after which cells were analyzed flow cytometry and Western blot. c SU-DHL-16 cells were transfected with NOXA/Flag constructs. Three clones with overexpression of NOXA were selected. These clones were exposed to HHT (10 nM) for 48 h then analyzed by flow cytometry. d SU-DHL-16 cells were transfected with shNOXA constructs. Two clones with knockdown of NOXA were selected. These clones were exposed to HHT (10 nM) for 48 h then analyzed by flow cytometry

Article Snippet: Primary antibodies used in these studies were: cleaved PARP, cleaved caspase-3, BCL- XL , BIM (Cell Signaling Technology, Danvers, MA), MCL-1 (BD Biosciences, San Jose, CA), α-tubulin (EMD Millipore, Billerica, MA), BAX (N20), BAK (G23), actin (Sigma-Aldrich, St. Louis, MO), NOXA (Enzo Life Sciences, Farmingdale, NY).

Techniques: Inhibition, Over Expression, Transfection, shRNA, Construct, Clone Assay, Flow Cytometry, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Response of Myeloid Leukemia Cells to Luteolin is Modulated by Differentially Expressed Pituitary Tumor-Transforming Gene 1 (PTTG1) Oncoprotein

doi: 10.3390/ijms19041173

Figure Lengend Snippet: Effects of luteolin on the apoptosis of PTTG1-knockdown THP-1 cells. The THP1 shLacZ and shPTTG1 cells were treated with vehicle or luteolin (25–100 μM) for 24 h. ( a ) Cell population in the sub-G1 phase was determined by flow cytometric analysis. The experiments were triplicated, and the data are presented as the means ± SD from three independent experiments. ** p < 0.01 represents a significant difference compared to the vehicle group (veh). # p < 0.05 and ## p < 0.01 represent a significant difference compared to the THP1 shLacZ control cells. ( b ) The protein expression levels of pro-caspase 3, cleaved caspase 3, PARP1, cleaved PARP1 and β-actin proteins were analyzed by Western blot analysis. The experiments were triplicated, and a representative blot is shown. ( c ) The Mcl-1, Bcl-2, p21 and β-actin proteins were analyzed by Western blot analysis. The experiments were triplicated, and a representative blot is shown.

Article Snippet: The proteins were separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), transferred onto a polyvinylidene difluoride (PVDF) membrane (PerkinElmer, Boston, MA, USA) and incubated with the following specific antibodies: anti-caspase 3, anti-PARP, anti-Mcl-1, and anti-Bcl-2 (GeneTex, Irvine, CA, USA); anti-cleaved caspase 3 and anti-p21Waf1/Cip1 (Cell Signaling Technology, Danvers, MA, USA); and anti-PTTG1 (Thermo Fisher Scientific, Rockford, IL, USA) and anti-β-actin (Sigma-Aldrich, St. Louis, MO, USA).

Techniques: Expressing, Western Blot