Journal: International Journal of Medical Sciences

Article Title: 17β-Estradiol Inhibits Mesenchymal Stem Cells-Induced Human AGS Gastric Cancer Cell Mobility via Suppression of CCL5- Src/Cas/Paxillin Signaling Pathway

doi: 10.7150/ijms.6851

Figure Lengend Snippet: CCL-5 induces AGS cell mobility via Src signaling pathway. (A) AGS cells were pretreated with vehicle, LY294002 (Akt activation inhibitor, 1μM), U0126 (ERK1/2 activation inhibitor, 1μM), SB203580 (p38 MAPK inhibitor, 1μM), SP600125 (JNK1/2 inhibitor, 1μM) or PP2 (Src inhibitor, 1μM) for 1h, and followed by recombinant CCL-5 (10 ng/ml) administration for 24h. (B) AGS cells were treated with these inhibitors, respectively. AGS cells were harvested and measured for cellular mobility capacity. **, p <0.01 versus control (line 1); ##, p <0.01 versus CCL-5 treatment (line 2) (mean ± SD, n = 3).

Article Snippet: The LY294002 (PI3K inhibitor), U0126 (MEK1/2 inhibitor), SB203680 (p38 MAPK inhibitor), SP600125 (JNK inhibitor), and PP2 (Src inhibitor) were purchased from TOCRIS (Ellisville, Missouri, USA).

Techniques: Activation Assay, Recombinant, Control

Journal:

Article Title: Oncogenic Ras Blocks Anoikis by Activation of a Novel Effector Pathway Independent of Phosphatidylinositol 3-Kinase

doi: 10.1128/MCB.21.16.5488-5499.2001

Figure Lengend Snippet: ERK activation partially contributes to anoikis resistance of Ras-transformed RIE-1 cells. (A) RIE-1 cells expressing activated Ras were suspended for 6 h in the presence of vehicle or 30 μM U0126 and assayed for ERK activation by immunoblotting with phospho-specific ERK antibodies. Blots were stripped and reprobed with anti-ERK antibodies to determine total ERK levels. (B and C) The effects of pharmacologic inhibition of ERK activation on anoikis were assessed by treating suspended cells with either U0126 (dark bars) or vehicle (light bars) for the indicated time (hours) and performing DNA laddering (B) and DNA fragmentation ELISA (C) analyses. For panel C, only H-Ras(12V)-expressing cells were assayed. Data shown are representative of at least two independent experiments. Bars represent the standard deviations resulting from duplicate reading of the same sample.

Article Snippet: Chemical inhibitors used in this study are specific to MEK1/2 (U0126) (provided by J. Trzaskos, Dupont) ( 13 ), PI3K ( {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}} LY294002 ) (A.G. Scientific) ( 55 ), caspase-1-like proteases (Z-VAD-FMK) (Calbiochem) ( 14 ), and the EGFR kinase (PD153035) (Tocris Cookson) ( 19 ).

Techniques: Activation Assay, Transformation Assay, Expressing, Western Blot, Inhibition, DNA Laddering, Enzyme-linked Immunosorbent Assay

Journal:

Article Title: Oncogenic Ras Blocks Anoikis by Activation of a Novel Effector Pathway Independent of Phosphatidylinositol 3-Kinase

doi: 10.1128/MCB.21.16.5488-5499.2001

Figure Lengend Snippet: Coordinate activation of Raf and PI3K is not necessary or sufficient to block anoikis. (A) RIE-1 cells stably infected with empty vector pBabe-puro (light bars) or encoding H-Ras(12V) (dark bars) were treated with either vehicle or 10 μM LY294002 and/or 30 μM U0126 for the indicated time in suspension (hours) prior to evaluating anoikis by DNA fragmentation ELISA. (B) RIE-1 cells stably infected with pBabe-p110-CAAX and pZIP-raf-22W were established by selection in growth medium supplemented with puromycin and G418. To assay for the activation state of ERK and Akt, Western blot analyses were performed with cells suspended for 6 h in the absence of serum. Cells stably infected with the pZIP-NeoSV(x)1 and pBabe-puro empty vectors were established as controls for these analyses. (C) The resulting cell populations were evaluated for anoikis by DNA fragmentation ELISA after suspension for the indicated time (hours). Data shown are representative of at least two independent experiments. Bars represent the standard deviations resulting from duplicate reading of the same sample.

Article Snippet: Chemical inhibitors used in this study are specific to MEK1/2 (U0126) (provided by J. Trzaskos, Dupont) ( 13 ), PI3K ( {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}} LY294002 ) (A.G. Scientific) ( 55 ), caspase-1-like proteases (Z-VAD-FMK) (Calbiochem) ( 14 ), and the EGFR kinase (PD153035) (Tocris Cookson) ( 19 ).

Techniques: Activation Assay, Blocking Assay, Stable Transfection, Infection, Plasmid Preparation, Enzyme-linked Immunosorbent Assay, Selection, Western Blot