Journal: Cancer medicine

Article Title: Direct effect of dasatinib on signal transduction pathways associated with a rapid mobilization of cytotoxic lymphocytes.

doi: 10.1002/cam4.925

Figure Lengend Snippet: Figure 1. Flow cytometric analysis of each lymphocyte subset and expression of phosphorylated proteins. Representative data are shown. Lymphocyte fractions are classified according to surface antibodies including CD3, CD8, and CD56. Natural killer (NK) cells were defined as the CD3-CD56+ immunophenotype and cytotoxic T lymphocytes (CTLs) were CD3+ CD8+ (A). Cells were costained with phospho-specific antibodies, including antibodies targeting pJAK1, pJAK2, pSTAT1, pSTAT3, pERK, pJNK, pp38, and pAKT, and expression levels of the isotype control and phosphorylated proteins in NK cells are presented (B). The values for the isotype control and each phosphorylated protein are shown as the median fluorescence intensity (MFI).

Article Snippet: Fluorescein isothiocyanate (FITC)- labeled polyclonal rabbit IgG antibodies for phosphorylated proteins, including pJAK1 (Y1034, #3238R), pJAK2 (Y1007, #2485R), pSTAT1 (Y701, #1657R), pSTAT3 (Y705, #1658R), pERK (T202/Y204, #1646R), pJNK (T183/Y185, #1640R), pp38 (T180/Y182, #2210R), pAKT (Y315, #5193R), and isotype control (#0295P) were purchased from Bioss (Wobun, MA, USA).

Techniques: Expressing, Control, Fluorescence

Journal: Cancer medicine

Article Title: Direct effect of dasatinib on signal transduction pathways associated with a rapid mobilization of cytotoxic lymphocytes.

doi: 10.1002/cam4.925

Figure Lengend Snippet: Figure 3. Constitutive levels of phosphorylated proteins including pJAK1, pJAK2, pSTAT1, pSTAT3, pERK, pJNK, pp38, and pAKT in natural killer (NK) cells (A) and cytotoxic T lymphocytes (CTLs) (B) grouped according to treatment (dasatinib [n = 18] or other TKI [n = 12]) are shown. The values for phosphorylated proteins in each fraction are shown as the median fluorescence intensity (MFI).

Article Snippet: Fluorescein isothiocyanate (FITC)- labeled polyclonal rabbit IgG antibodies for phosphorylated proteins, including pJAK1 (Y1034, #3238R), pJAK2 (Y1007, #2485R), pSTAT1 (Y701, #1657R), pSTAT3 (Y705, #1658R), pERK (T202/Y204, #1646R), pJNK (T183/Y185, #1640R), pp38 (T180/Y182, #2210R), pAKT (Y315, #5193R), and isotype control (#0295P) were purchased from Bioss (Wobun, MA, USA).

Techniques: Fluorescence

Journal: PLoS ONE

Article Title: Acetylcholine Acts on Androgen Receptor to Promote the Migration and Invasion but Inhibit the Apoptosis of Human Hepatocarcinoma

doi: 10.1371/journal.pone.0061678

Figure Lengend Snippet: (A–D) Western blots showed the effects of Ach and MEC on STAT3 phosphorylation (A, B) and Akt phosphorylation (C, D) and in SNU-449 cells. Phosphorylation of STAT3 (A) and Akt (C) was enhanced by Ach in SNU-449 cells, whereas acetylcholine receptor antagonists MEC blocked the effect of Ach on STAT3 (B) and AKT (D) phosphorylation.

Article Snippet: Primary antibodies used here included rabbit monoclonal anti-AR (D6F11) (3H8; Cell Signaling Technology, Danvers, MA), rabbit monoclonal anti-nicotinic acetylcholine receptor alpha 7 (ab10096, abcam, USA), rabbit monoclonal anti-pSTAT3, anti-pAKT (195-14; BioCheck, Foster City, CA), and monoclonal anti-β-actin (Sigma).

Techniques: Western Blot

Journal: Endocrinology

Article Title: Nutritional programming affects hypothalamic organization and early response to leptin.

doi: 10.1210/en.2009-0893

Figure Lengend Snippet: FIG. 5. pSTAT3-IR-positive cells evaluated 90 min after ip saline or leptin (10 mg/kg) injections at PND 5 and 12 for ARC (A) and DMD (B) and at PND 12 for VMH (C) and DMV (D): CC (white bars), RC (gray bars), and RR (black bars). Values are means SEM; n 3–4 rats per group. Significant variations between means were tested by Mann-Whitney U test. *, P 0.05 for leptin vs. NaCl; , P 0.05 for RC vs. CC; $, P 0.05 for RR vs. CC in leptin conditions. n.s., Not significant; u, undetectable. Photomicrographs are presented in supplemental Fig. 2.

Article Snippet: Twenty-micrometer coronal sections were realized between 0.60 and 4.08 mm bregma coordinates, according to the Paxinos atlas (24), with a HM560 cryotome (MM, Francheville, France), collected in six series and stored at 20 C under dry atmosphere until further use. pSTAT3 immunoreactivity (IR) was performed as described previously (26) using the rabbit anti-pSTAT3 antibody from Cell Signaling (Ozyme, St. Quentin en Yvelines, France) and a biotinylated goat antirabbit secondary antibody from Jackson (AbCys S.A., Paris, France).

Techniques: Saline, MANN-WHITNEY

Journal: Oncogene

Article Title: The broad-range cyclin-dependent kinase inhibitor UCN-01 induces apoptosis in colon carcinoma cells through transcriptional suppression of the Bcl-x(L) protein.

doi: 10.1038/sj.onc.1207842

Figure Lengend Snippet: Figure 6 Variable response of NF-kB to UCN-01 treatment. The transcriptional activity of NF-kB is suppressed by UCN-01 treatment (1 mM, 48 h) in SW48 cell line, but it is activated in WiDr, HT-29 and LS513 cells Figure 7 UCN-01 treatment can inhibit both Ser727 and Tyr705 phosphorylation of STAT3 a. (a) Constitutive STAT3 phosphor- ylation at Ser727 is detectable in all cell lines. It is inhibited by UCN-01 (1 mM, 48 h) only in the apoptosis-performing cell lines. (b) STAT3 phosphorylation at Tyr705 is not detectable in any of the cell lines (shown LS513). It is inducible by IFNa and is inhibited within 1.5 h by UCN-01 treatment

Article Snippet: The following antibodies were employed: rabbit anti-BAX antibody (N20), goat anti-Akt1 antibody (C-20), mouse monoclonal anti-phosphotyrosine STAT3 antibody (clone B-7) (all from Santa Cruz Biotechnology, Heidelberg, Germany); mouse monoclonal anti-PARP antibody (clone C2-10), and mouse monoclonal anti-Bcl-xL (clone 2H12) (all from BD Pharmingen, Heidelberg, Germany); mouse monoclonal anti-b-actin antibody (clone AC-15) (Sigma-Aldrich Chemie, Taufkirchen, Germany).

Techniques: Activity Assay, Phospho-proteomics

Journal: Oncogene

Article Title: The broad-range cyclin-dependent kinase inhibitor UCN-01 induces apoptosis in colon carcinoma cells through transcriptional suppression of the Bcl-x(L) protein.

doi: 10.1038/sj.onc.1207842

Figure Lengend Snippet: Figure 8 Activation of STAT3 does not lead to Bcl-xL transcrip- tion. (a) Exogenous STAT3 is phosphorylated at Tyr705 after stimulation of SW48 cells with IL-6. (b) IL-6 stimulation activates the transcriptional activity of STAT3 as demonstrated by the increased activity of the luciferase reporter. (c) Bcl-xL transcription is not affected by the transcriptionally active STAT3. Total RNA was isolated from the same cells as used in (b), and the semiquantitative RT–PCR was carried out with the cycle number in the linear range of the amplification. b-actin was amplified for the control of the amounts of mRNA used in each lane

Article Snippet: The following antibodies were employed: rabbit anti-BAX antibody (N20), goat anti-Akt1 antibody (C-20), mouse monoclonal anti-phosphotyrosine STAT3 antibody (clone B-7) (all from Santa Cruz Biotechnology, Heidelberg, Germany); mouse monoclonal anti-PARP antibody (clone C2-10), and mouse monoclonal anti-Bcl-xL (clone 2H12) (all from BD Pharmingen, Heidelberg, Germany); mouse monoclonal anti-b-actin antibody (clone AC-15) (Sigma-Aldrich Chemie, Taufkirchen, Germany).

Techniques: Activation Assay, Activity Assay, Luciferase, Isolation, Reverse Transcription Polymerase Chain Reaction, Control

Journal: Molecular cancer therapeutics

Article Title: Targeted inhibition of Src kinase signaling attenuates pancreatic tumorigenesis

doi: 10.1158/1535-7163.MCT-09-1212

Figure Lengend Snippet: Cells from BxPC3 (A), and PANC1 (B) were treated with DMSO or dasatinib (0-1000 nmol/L) for 12 h, lysed, and analyzed by Western blotting with indicated antibodies. Blots are representative of at least two separate experiments with similar results. There is a lack of inhibition of pSTAT3 and pMAPK signaling with dasatinib in PANC1 cells. (C) BxPC3 cells were treated with dasatinib (5 nmol/L) for 12 h. There was a significant decrease in expression of c-Myc and cyclin-D1 by immunofluorescence.

Article Snippet: Mouse monoclonal antibodies against STAT3 and Src; rabbit monoclonal antibodies against MAPK, pSrc (Tyr416), pAKT (Ser473) and pSTAT3 (Ser727); rabbit polyclonal antibodies against pFAK (Tyr925), pPaxillin (Tyr118), pAKT (Ser473), pMAPK (Thr202/Tyr204), AKT, paxillin and FAK were purchased from Cell Signaling Technology (Beverly, MA).

Techniques: Western Blot, Inhibition, Expressing, Immunofluorescence