Journal: Molecular Medicine Reports

Article Title: Pegylated liposomal-paclitaxel induces ovarian cancer cell apoptosis via TNF-induced ERK/AKT signaling pathway

doi: 10.3892/mmr.2018.8811

Figure Lengend Snippet: ERK/AKT signaling pathway is involved in the activation of the TNF/caspase-3 cascade within ovarian cancer cells. (A) Inhibition of ERKIR decreased caspase-3 expression levels within CAOV-3 cells compared with in the control group. (B) Inhibition of ERKIR inhibited TNF-induced CAOV-3 cell apoptosis compared with in the control group. (C) PL-PTX-induced CAOV-3 cell apoptosis was inhibited via an ERK inhibitor compared with in the control group. **P

Article Snippet: The primary antibodies used were against: Caspase-9 (1:1,200; ab32539), ERK (1:1,000; ab54230) and phosphorylayed (p)ERK 1/2 (phospho-Thr202/Tyr204; 1:1,000; ab214362), caspase-3 (1:1,200; ab2171), protein kinase B (AKT; 1:1,000; ab8805), p-AKT (phosphor-S473; 1:1,000; ab8932) and β-actin (1:500; ab8226; all Abcam) for 12 h at 4°C.

Techniques: Activation Assay, Inhibition, Expressing

Journal: Cell Death & Disease

Article Title: CRLF1 promotes malignant phenotypes of papillary thyroid carcinoma by activating the MAPK/ERK and PI3K/AKT pathways

doi: 10.1038/s41419-018-0352-0

Figure Lengend Snippet: CRLF1 promotes tumorigenesis by activating the MAPK/ERK and PI3K/AKT pathways. a An MAPK phosphorylation antibody array revealed that ERK1/2 and AKT (P-S473) were activated in CRLF1-overexpressing IHH-4 cells. b Western blotting assays showed changes in the levels of ERK1/2 and AKT (P-S473) in CRLF1 knockdown or CRLF1-overexpressing cells. c IHH-4-CRLF1 cells were cultured with 10 μM U0126, 10 μM MK-2206, or 10 μM U0126+10 μM MK-2206 for 24 h. Western blotting analyses were performed to evaluate the effects of these two inhibitors on phosphorylation levels of ERK1/2 and AKT. β-Actin was used as a loading control. d MTT proliferation assay results demonstrating inhibited proliferation in CRLF1-overexpressing IHH-4 cells treated with MK-2206, U0126, or MK-2206+U0126

Article Snippet: The sections were then incubated with diluted rabbit anti-CRLF1 antibody (1:100; HPA041493, Sigma-Aldrich, USA), p-ERK1/2 antibody (1:400; #4370, Cell Signaling Technology, USA), or p-AKT antibody (1:100; #4060, Cell Signaling Technology, USA) at 4 °C overnight.

Techniques: Ab Array, Western Blot, Cell Culture, MTT Assay, Proliferation Assay

Journal: PLoS ONE

Article Title: Substance P Inhibits Hyperosmotic Stress-Induced Apoptosis in Corneal Epithelial Cells through the Mechanism of Akt Activation and Reactive Oxygen Species Scavenging via the Neurokinin-1 Receptor

doi: 10.1371/journal.pone.0149865

Figure Lengend Snippet: Role of redox regulation in the anti-apoptotic effects of SP. Mouse corneal epithelial cells were treated with 100 μM L-BSO and 1 μM SP 2 h before the addition of glucose for 24 h. The phosphorylation of Akt was evaluated by Immunofluorescence staining or Western blot (A). The apoptosis was evaluated by FACS analysis followed by FITC-Annexin V/PI staining (B), and the detection of Bad, Bax, AIF, Ca 2+ and mitochondrial membrane potential (C). The intracellular ROS and glutathione (GSH) were detected by staining with the fluorescence probes (D) and measured by the fluorescence intensity (E). The cellular total antioxidant capacity (TAC) was measured by the ABTS assay (E).

Article Snippet: Immunofluorescence staining Cells were fixed by 4% paraformaldehyde for 15 min, permeabilized with 0.1% Triton X-100 for 5 min and blocked with 5% bovine serum albumin (BSA) for 1 h. The samples were stained with primary antibodies against AIF (Santa Cruz Biotechnology, Santa Cruz, CA), p-Akt (pS473, Epitomics, Burlingame, CA), MnSOD, catalase, NQO1, Hmox1, Bad and Bax (abcam, Cambridge, MA) at a concentration of 1:100 overnight at 4°C, and then incubated with fluorescein-conjugated secondary antibodies (Santa Cruz Biotechnology) at 37°C for 30 min.

Techniques: Immunofluorescence, Staining, Western Blot, FACS, Fluorescence, ABTS Assay

Journal: PLoS ONE

Article Title: Substance P Inhibits Hyperosmotic Stress-Induced Apoptosis in Corneal Epithelial Cells through the Mechanism of Akt Activation and Reactive Oxygen Species Scavenging via the Neurokinin-1 Receptor

doi: 10.1371/journal.pone.0149865

Figure Lengend Snippet: Role of NK-1 receptor in the anti-apoptotic effects of SP. Mouse corneal epithelial cells were treated with 1 μM NK-1 receptor antagonist L-733,060 with 1 μM SP 2 h before the addition of glucose for 24 h. The phosphorylation of Akt was evaluated by Immunofluorescence staining or Western blot (A). The apoptosis was evaluated by FACS analysis followed by FITC-Annexin V/PI staining (B), and the detection of Bad, Bax, AIF, Ca 2+ and mitochondrial membrane potential (C). The intracellular ROS and glutathione (GSH) were detected by staining with the fluorescence probes (D) and measured by the fluorescence intensity (E). The cellular total antioxidant capacity (TAC) was measured by the ABTS assay (E).

Article Snippet: Immunofluorescence staining Cells were fixed by 4% paraformaldehyde for 15 min, permeabilized with 0.1% Triton X-100 for 5 min and blocked with 5% bovine serum albumin (BSA) for 1 h. The samples were stained with primary antibodies against AIF (Santa Cruz Biotechnology, Santa Cruz, CA), p-Akt (pS473, Epitomics, Burlingame, CA), MnSOD, catalase, NQO1, Hmox1, Bad and Bax (abcam, Cambridge, MA) at a concentration of 1:100 overnight at 4°C, and then incubated with fluorescein-conjugated secondary antibodies (Santa Cruz Biotechnology) at 37°C for 30 min.

Techniques: Immunofluorescence, Staining, Western Blot, FACS, Fluorescence, ABTS Assay

Journal: PLoS ONE

Article Title: Substance P Inhibits Hyperosmotic Stress-Induced Apoptosis in Corneal Epithelial Cells through the Mechanism of Akt Activation and Reactive Oxygen Species Scavenging via the Neurokinin-1 Receptor

doi: 10.1371/journal.pone.0149865

Figure Lengend Snippet: Role of Akt reactivation in the anti-apoptotic effects of SP. Mouse corneal epithelial cells were treated with 40 μM Akt inhibitor V and 1 μM SP 2 h before the addition of glucose for 24 h. The phosphorylation of Akt was evaluated by Immunofluorescence staining or Western blot (A). The apoptosis was evaluated by FACS analysis followed by FITC-Annexin V/PI staining (B), and the detection of Bad, Bax, AIF, Ca 2+ and mitochondrial membrane potential (C). The intracellular ROS and glutathione (GSH) were detected by staining with the fluorescence probes (D) and measured by the fluorescence intensity (E). The cellular total antioxidant capacity (TAC) was measured by the ABTS assay (E).

Article Snippet: Immunofluorescence staining Cells were fixed by 4% paraformaldehyde for 15 min, permeabilized with 0.1% Triton X-100 for 5 min and blocked with 5% bovine serum albumin (BSA) for 1 h. The samples were stained with primary antibodies against AIF (Santa Cruz Biotechnology, Santa Cruz, CA), p-Akt (pS473, Epitomics, Burlingame, CA), MnSOD, catalase, NQO1, Hmox1, Bad and Bax (abcam, Cambridge, MA) at a concentration of 1:100 overnight at 4°C, and then incubated with fluorescein-conjugated secondary antibodies (Santa Cruz Biotechnology) at 37°C for 30 min.

Techniques: Immunofluorescence, Staining, Western Blot, FACS, Fluorescence, ABTS Assay