c src inhibitor pp1  (Santa Cruz Biotechnology)

Journal: Experimental and Therapeutic Medicine

Article Title: Regulation of transforming growth factor β-mediated epithelial-mesenchymal transition of lens epithelial cells by c-Src kinase under high glucose conditions

doi: 10.3892/etm.2018.6348

Figure Lengend Snippet: Effects of 10 µmol/l c-Src inhibitor PP1 and 10 µmol/l ALK5 inhibitor SB431542 on the protein expression levels of p-Src418, TGF-β1, TGF-β2, ALK5, E-cadherin and α-SMA under HG conditions at 0 to 6 h. Protein expression levels of (A) p-Src418, (B) TGF-β1, (C) TGF-β2, (D) ALK5, (E) E-cadherin and (F) α-SMA were determined using western blot analysis. Data are presented as the mean ± standard deviation. *P<0.05 as indicated. HG, high glucose; TGF-β, transforming growth factor-β; α-SMA, α-smooth muscle actin; ALK5, activin receptor-like kinase 5.

Article Snippet: Stock solutions of PP1, a c-Src inhibitor, and SB431542, an ALK5 inhibitor, (Enzo Life Sciences, Inc., Farmingdale, NY, USA) were prepared in dimethyl sulfoxide and diluted with cell culture media to obtain a final concentration of 10 µmol/l.

Techniques: Expressing, Western Blot, Standard Deviation

Journal: Experimental and Therapeutic Medicine

Article Title: Regulation of transforming growth factor β-mediated epithelial-mesenchymal transition of lens epithelial cells by c-Src kinase under high glucose conditions

doi: 10.3892/etm.2018.6348

Figure Lengend Snippet: Effects of c-Src inhibitor PP1 and ALK5 inhibitor SB431542 on the secretion of TGF-β1 and TGF-β2 in human lens epithelial B3 cells under HG (35.5 mM) conditions at 0 to 6 h. Secretion of (A) TGF-β1 and (B) TGF-β2 was determined using ELISA. Data are presented as the mean ± standard deviation. *P<0.05 as indicated. HG, high glucose; TGF-β, transforming growth factor-β.

Article Snippet: Stock solutions of PP1, a c-Src inhibitor, and SB431542, an ALK5 inhibitor, (Enzo Life Sciences, Inc., Farmingdale, NY, USA) were prepared in dimethyl sulfoxide and diluted with cell culture media to obtain a final concentration of 10 µmol/l.

Techniques: Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: Oncotarget

Article Title: TRAIL induces pro-apoptotic crosstalk between the TRAIL-receptor signaling pathway and TrkAIII in SH-SY5Y cells, unveiling a potential therapeutic “Achilles heel” for the TrkAIII oncoprotein in neuroblastoma

doi: 10.18632/oncotarget.13098

Figure Lengend Snippet: ( A ) Representative phase contrast micrographs demonstrating inhibition of TRAIL-induced TrkAIII SH-SY5Y apoptosis following 24 hour treatment (200 ng/ml) by 10 μM z-VAD-fmk (VAD), 10 μM z-IETD-fmk (IETD) but not 10 μM z-LEHD-fmk (LEHD) and by 1 μM GW441756 (GW), 1 μM PP1 and 1 μM NSC-87877 (NSC) (bar = 100 μm). ( B ) Histograms displaying the mean (+SD) percentage (%) TrkAIII SH-SY5Y survival (white) or death (black) following treatment with TRAIL alone (200 ng/ml), TRAIL plus 10 μM z-VAD-fmk (VAD), TRAIL plus 10 μM z-LEHD-fmk (LEHD), TRAIL plus 10 μM z-IETD-fmk (IETD), TRAIL plus 1 μM GW441756 (GW), TRAIL plus 1 μM PP1 and TRAIL plus 1 μM NSC-87877 (NSC), in three independent AO/EBr experiments, each performed in duplicate (* = significant difference compared to TrkAIII SH-SY5Y cells treated for 24 hours with TRAIL in the absence of inhibitors).

Article Snippet: GW441756 TrkA inhibitor [ ], z-VAD-fmk pan-caspase inhibitor, z-IETD-fmk caspase 8 inhibitor and z-LEHD-fmk caspase 9 inhibitor; human recombinant TRAIL and PP1 c-Src inhibitor were purchased from Sigma-Aldrich (St. Louis Mo).

Techniques: Inhibition

Journal: Oncotarget

Article Title: TRAIL induces pro-apoptotic crosstalk between the TRAIL-receptor signaling pathway and TrkAIII in SH-SY5Y cells, unveiling a potential therapeutic “Achilles heel” for the TrkAIII oncoprotein in neuroblastoma

doi: 10.18632/oncotarget.13098

Figure Lengend Snippet: ( A ) Immunoprecipitation/Western blots demonstrating SHP-1 pull down of c-Src but not TrkAIII from untreated TrkAIII SH-SY5Y whole cell extracts (500 μg) and SHP-1 pull down of both c-Src and TrkAIII in TRAIL-treated (200 ng/ml for 12 hours) TrkAIII SH-SY5Y whole cell extracts (500 μg), plus Western blots showing input levels of SHP-1, c-Src and TrkAIII in TrkAIII SH-SY5Y whole cell extracts (20 μg). ( B ) Western blots demonstrating TRAIL-induced (200 ng/ml for 0, 6 and 12 hours) c-Src Y527 de-phosphorylation and stimulation of c-Src Y416 phosphorylation; 1 μM PP1 inhibition of TRAIL-induced stimulation of c-Src Y416 phosphorylation but not TRAIL-induced c-Src Y527 de-phosphorylation, and 1 μM NSC-87877 (NSC) inhibition of both TRAIL-induced stimulation of c-Src Y416 phosphorylation and c-Src Y527 de-phosphorylation, compared to total c-Src levels in whole cell TrkAIII SH-SY5Y extracts (20 μg). ( C ) Histograms demonstrating the mean (+SD) change in densitometric ratio of total to Y527-phosphorylated c-Src and total to Y416-phosphorylated c-Src associated with TrkAIII SH-SY5Y cells treated with TRAIL (200 ng/ml) alone, TRAIL plus 1 μM PP1 or TRAIL plus 1 μM NSC-87877 (NSC) for 0, 6 and 12 hours, assessed by ImageJ densitometry of 3 independent Western blots of whole TrkAIII SH-SY5Y extracts (20 μg) (* = significant difference compared to untreated (TRAIL 0 hrs controls).

Article Snippet: GW441756 TrkA inhibitor [ ], z-VAD-fmk pan-caspase inhibitor, z-IETD-fmk caspase 8 inhibitor and z-LEHD-fmk caspase 9 inhibitor; human recombinant TRAIL and PP1 c-Src inhibitor were purchased from Sigma-Aldrich (St. Louis Mo).

Techniques: Immunoprecipitation, Western Blot, De-Phosphorylation Assay, Inhibition

Journal: Oncotarget

Article Title: TRAIL induces pro-apoptotic crosstalk between the TRAIL-receptor signaling pathway and TrkAIII in SH-SY5Y cells, unveiling a potential therapeutic “Achilles heel” for the TrkAIII oncoprotein in neuroblastoma

doi: 10.18632/oncotarget.13098

Figure Lengend Snippet: ( A ) Western blots demonstrating no change total and Y674/675 phosphorylated TrkAIII levels in TrkAIII SH-SY5Y cells incubated for 0, 3 and 6 hours in absence of TRAIL, compared to time-dependent TRAIL-induced (200 ng/ml) TrkAIII Y674/675 de-phosphorylation associated with constant TrkAIII levels, in TrkAIII SH-SY5Y cells incubated for 3 and 6 hours plus the inhibition of TRAIL-induced TrkAIII Y674/675 de-phosphorylation by 1 μM PP1 (TRAIL + PPI) and by 1 μM NSC-87877 (TRAIL + NSC) at both 3 and 6 hours. ( B ) Histograms demonstrating the mean (±SD) change the ratio of total to Y674/675-phosphorylated TrkAIII in whole cell extracts (20 μg) from TrkAIII SH-SY5Y cells incubated for 0–6 hours in the absence of TRAIL (control) in the presence of TRAIL (200 ng/ml) alone (TRAIL), with TRAIL plus 1 μM PP1 (TRAIL + PP1) or with TRAIL plus 1 μM NSC-87877 (TRAIL + NSC), assessed by ImageJ densitometry of 3 independent Western blots (* = significant difference compared to untreated 6 hour controls).

Article Snippet: GW441756 TrkA inhibitor [ ], z-VAD-fmk pan-caspase inhibitor, z-IETD-fmk caspase 8 inhibitor and z-LEHD-fmk caspase 9 inhibitor; human recombinant TRAIL and PP1 c-Src inhibitor were purchased from Sigma-Aldrich (St. Louis Mo).

Techniques: Western Blot, Incubation, De-Phosphorylation Assay, Inhibition

Journal: Breast Cancer Research : BCR

Article Title: Trefoil factor 3 promotes metastatic seeding and predicts poor survival outcome of patients with mammary carcinoma

doi: 10.1186/s13058-014-0429-3

Figure Lengend Snippet: Forced expression of TFF3 in MCF7 cells enhanced phosphorylation of c-SRC that subsequently increased STAT3 activity to promote invasion. (A) Western blot analysis was used to assess the levels of p-c-SRC and c-SRC in MCF7 cells with either forced or depleted expression of TFF3 as described in Methods. (B) Western blot analysis was used to assess the levels of pSTAT3, STAT3, and CDH1 in MCF7 cells with forced expression of TFF3 on exposure to PP1 (5 μM), PP2 (2 μM) or PP3 (50 μM) as described in Methods. (C) CDH1 promoter activity in MCF7 cells with either forced or depleted expression of TFF3, on exposure to PP1 (5 μM), PP2 (2 μM) or PP3 (50 μM). The luciferase assay was performed as described in Methods. (D) α2-M promoter activity in MCF7 cells with either forced or depleted expression of TFF3, on exposure to PP1 (5 μM), PP2 (2 μM), or PP3 (50 μM). The luciferase assay was performed as described in Methods. (E) . Invasive capacity of MCF7 cells with either forced or depleted expression of TFF3, on exposure to PP1 (5 μM), PP2 (2 μM) or PP3 (50 μM); and/or in combination with transiently transfected STAT3 CA . Cell invasion was evaluated using a Transwell assay. Statistical significance was assessed by using an unpaired two-tailed Student's t test ( P <0.05 was considered as significant) using GraphPad Prism 5. Columns are the mean of triplicate experiments; bars, ± SD. ** P < 0.001, * P < 0.05.

Article Snippet: STAT3 activity inhibitor, JSI-124 and Stattic, was purchased from Sigma-Aldrich (Singapore). c-SRC family kinase inhibitor PP1, and the specific c-SRC kinase inhibitor PP2 was purchased from Sigma-Aldrich (Singapore); the structurally related non-inhibitory PP3 (50 μM) was purchased from Calbiochem (Singapore).

Techniques: Expressing, Phospho-proteomics, Activity Assay, Western Blot, Luciferase, Transfection, Transwell Assay, Two Tailed Test