rabbit anti human integrin β3  (Cell Signaling Technology Inc)

Journal: Cancer Cell International

Article Title: Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

doi: 10.1186/s12935-016-0365-7

Figure Lengend Snippet: Identification of integrin β3 as a kallistatin-binding protein. a The pull-down assay with the recombinant human kallistatin. The membrane proteins of NCI-H446 cell were pulled down by recombinant human kallistatin and subjected to immunoblotting with antibody anti-integrinβ3 ( upper panel ) and anti-kallistatin, respectively ( lower panel ). The membrane proteins incubated with Ni–NTA alone were used as control. b Immunoprecipitation of membrane proteins of NCI-H446 cell with anti-kallistatin antibody followed by immunoblotting against integrin β3 and kallistatin, respectively. The extracts of NCI-H446 cells treated without kallistatin were immunoprecipitatedas control. Co-immunoprecipitation sample without treated with Protein A-Sepharose were used to immunoblotting as input. c Immunoprecipitation of membrane proteins of NCI-H446 cell with anti-Integrin β3 antibody followed by immunoblotting against integrin β3 and kallistatin. The extracts of NCI-H446 cells treated without kallistatin were immunoprecipitated as control. Co-immunoprecipitation samples without treated with Protein A-Sepharose were used to immunoblotting as input

Article Snippet: Rabbit anti-human integrin β3, rabbit anti-human phospho-Integrin β3, and mouse anti-human integrin β3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Binding Assay, Pull Down Assay, Recombinant, Membrane, Western Blot, Incubation, Control, Immunoprecipitation

Journal: Cancer Cell International

Article Title: Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

doi: 10.1186/s12935-016-0365-7

Figure Lengend Snippet: The effect of kallistatin on viability of NCI-H446 and CHO cells. a The effects of kallistatin on the viability of NCI-H446 cells in a dose dependent manner. N = 6 × 3 for each group. * P < 0.05. b Detect integrin expression in NCI-H446 and CHO cells by western blot. c Effect of kallistatin on cell viability in CHO cells which is lack of integrin β3 genes as a negative control. N = 6 × 3 for each group

Article Snippet: Rabbit anti-human integrin β3, rabbit anti-human phospho-Integrin β3, and mouse anti-human integrin β3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Expressing, Western Blot, Negative Control

Journal: Cancer Cell International

Article Title: Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

doi: 10.1186/s12935-016-0365-7

Figure Lengend Snippet: Integrin β3 is involved in the effects of kallistatin on viability of NCI-H446 cells. a Integrin β3 expression in NCI-H446 cells was downregulated by siRNA in a time-dependent manner that was analyzed by immunoblotting (N = 3). Housekeeping proteins GAPDH are useful as loading controls for immunoblotting and protein normalization. The data are expressed as mean ± SD from three independent experiments. *P < 0.05. b Inhibition effect of kallistatin on the viability of NCI-H446 cells was attenuated by treatment of siRNA. N = 6×3 for each group. N.C siRNA was used as negative control, respectively. *P < 0.05, # P < 0.05, # comparison of the differences between column 2 and 3 and between column 4 and 5. b Inhibition effect of kallistatin on the viability of NCI-H446 cells was attenuated by antibody against integrin β3. N = 6 × 3 for each group. Mouse IgG were used as negative control. *P < 0.05, # P < 0.05, # comparison of the differences between column 2 and 3 and between column 4 and 5

Article Snippet: Rabbit anti-human integrin β3, rabbit anti-human phospho-Integrin β3, and mouse anti-human integrin β3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Expressing, Western Blot, Inhibition, Negative Control, Comparison

Journal: Cancer Cell International

Article Title: Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

doi: 10.1186/s12935-016-0365-7

Figure Lengend Snippet: Integrin β3 is involved in the inhibition effect of kallistatin on proliferation of NCI-H446 cells. a Proliferation of NCI-H446 cells was inhibited in presence of kallistatin. Proliferating NCI-H446 cells were labeled with EdU ( red ). Cell nuclei were stained with Hoechst 33,342 ( blue ). b The percentage of EdU-positive NCI-H446 cells was quantified. Kallistatin inhibited the proliferation of NCI-H446 cells in a dose-dependent manner. Data were presented as mean ± SD (N = 6). * P < 0.05. c Down-regulation of integrin β3 by siRNA attenuated the proliferation inhibition of NCI-H446 cells at 48 h post kallistatin treatment as measured by EdU assay. d The percentage of EdU-positive NCI-H446 cells was quantified. Data were presented as mean ± SD (N = 6). N.C siRNA was used as negative control. *P < 0.05, # P < 0.05, # comparison of the differences between column 1 and 2 and between column 3 and 4. e Blockage of integrin β3 with antibody (anti-integrin β3) attenuated the viability inhibition of NCI-H446 cells at 48 h post kallistatin treatment as measured by EdU assay. f The percentage of EdU-positive NCI-H446 cells was quantified. Data were presented as mean ± SD (N = 6). Mouse IgG was used as negative control. *P < 0.05, # P < 0.05, # comparison of the differences between column 1 and 2 and between column 3 and 4

Article Snippet: Rabbit anti-human integrin β3, rabbit anti-human phospho-Integrin β3, and mouse anti-human integrin β3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Inhibition, Labeling, Staining, EdU Assay, Negative Control, Comparison

Journal: Cancer Cell International

Article Title: Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

doi: 10.1186/s12935-016-0365-7

Figure Lengend Snippet: Reduction effects of kallistatin on NCI-H446 cells migration can be diminished by inhibition of integrin β3. A Optical microscopy imaging of the cells in experimental transwells. The cells have migrated from upper chamber to underside of the membrane between the two chambers after treatment with kallistatin. a saline; b 12.5 μg/ml kallistatin; c 50 μg/ml kallistatin; d 200 μg/ml kallistatin. B Statistic analysis for the migrated cell numbers. C Either down-regulation of integrin β3 by siRNA or blockage of integrin β3 with antibody (anti-integrin β3) attenuated the reduction effects of kallistatin on NCI-H446 cell migration. a Transfected with negative control siRNA and treated with saline; b transfected with negative control siRNA and treated with 200 μg/ml kallistatin; c transfected with integrin β3 siRNA and treated with saline; d transfected with integrin β3 siRNA and treated with 200 μg/ml kallistatin; e treated with mouse IgG and saline; f treated with mouse IgG and 200 μg/ml kallistatin; g treated with integrin β3 antibody and saline; h treated with integrin β3 antibody and 200 μg/ml kallistatin. D Statistic analysis for the migrated cell numbers. Data were presented as mean ± SD (N = 6). *P < 0.05. # P < 0.05, # comparison of the differences between column 1 and 2 and between column 3 and 4, and the differences between column 5 and 6 and between 7 and 8

Article Snippet: Rabbit anti-human integrin β3, rabbit anti-human phospho-Integrin β3, and mouse anti-human integrin β3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Migration, Inhibition, Microscopy, Imaging, Membrane, Saline, Transfection, Negative Control, Comparison

Journal: Cancer Cell International

Article Title: Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

doi: 10.1186/s12935-016-0365-7

Figure Lengend Snippet: Kallistatin inhibits phosphorylation along the integrin signaling pathway. a 200 μg total cellular protein of each sample was used for immunoblotting analysis. Phosphorylation of integrin β3, FAK and Src was inhibited in a time-dependent manner by treatment of kallistatin, while each of their total protein levels showed undetectable change. (N = 3). The protein levels were normalized with β-actin. b – d The relative levels of phosphorylated integrin β3, phosphorylated FAK and phosphorylated Src proteins were calculated and plotted. The data are expressed as mean ± SD from three independent experiments (N = 3). *P < 0.05

Article Snippet: Rabbit anti-human integrin β3, rabbit anti-human phospho-Integrin β3, and mouse anti-human integrin β3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Phospho-proteomics, Western Blot

Journal: Cancer Cell International

Article Title: Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

doi: 10.1186/s12935-016-0365-7

Figure Lengend Snippet: Kallistatin inhibits integrin β3-mediated PI3-K/AKT pathwaysignaling. a and e Using 200 μg total cellular proteins from each of the samples for immunoblotting analysis. The protein levels were normalized with β-actin. a Phosphorylation of AKT Erk1/2 and PI3 K were inhibited in a time-dependent manner without detectable change of their total AKT and Erk1/2 protein levels after treated with kallistatin (N = 3). ( b – d ) The relative levels of PI3 K, phosphorylated AKT and phosphorylated Erk1/2 proteins were calculated and plotted. The data are expressed as mean ± SD from three independent experiments (N = 3). *P < 0.05. f The relative levels of Grb2 proteins were calculated and plotted. The data are expressed as mean ± SD from three independent experiments (N = 3). *P < 0.05

Article Snippet: Rabbit anti-human integrin β3, rabbit anti-human phospho-Integrin β3, and mouse anti-human integrin β3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Western Blot, Phospho-proteomics

Journal: Cancer Cell International

Article Title: Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

doi: 10.1186/s12935-016-0365-7

Figure Lengend Snippet: The proposed effect sites of kallistatin through integrin signaling pathway. Kallistatin inhibited the both of PI3 K/AKT and Erk/MAPK resulted in a downregulation of Bcl-2 expression for the anti-apoptotic homologs, and an upregulation for the pro-apoptotic homologs, and inhinited the phosphorylation integrin β3 and suppresses the expression of Grb2 for the cell migration

Article Snippet: Rabbit anti-human integrin β3, rabbit anti-human phospho-Integrin β3, and mouse anti-human integrin β3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Expressing, Phospho-proteomics, Migration