Journal: Gastroenterology

Article Title: Neurotensin signaling activates microRNAs-21 and -155 and Akt, promotes tumor growth in mice, and is increased in human colon tumors.

doi: 10.1053/j.gastro.2011.07.038

Figure Lengend Snippet: Figure 4. NT regulates miR-21 and miR-155 signaling pathways in colon cancer cells. (A) Luciferase activity of the PTEN and SOCS1 3=UTRs after NT treatment (100 nmol/L) for 24 hours in HCT116 and DLD1 cells (untreated, as-miR-NC–treated, as-miR-21–treated, or as-miR-155–treated). (B) PTEN and SOCS1 mRNA levels, assessed by real-time PCR, in NT-treated (100 nmol/L, 6 h) HCT116 and DLD1 cells (untreated, as-miR-NC– treated, as-miR-21–treated, or as-miR-155–treated). (C) PTEN, SOCS1, and -actin protein levels, assessed by Western blot, in NT-treated (100 nmol/L, 6 h) HCT116 and DLD1 cells. (D) AKT phosphorylation (S473) levels, assessed by enzyme-linked immunosorbent assay, in HCT116 and DLD1 cells transfected with as-miR-NC (100 nmol/L), as-miR-21 (100 nmol/L), and/or as-miR-155 (100 nmol/L) for 24 hours and treated with NT (100 nmol/L) for 24 hours.

Article Snippet: AKT phosphorylation (S473) (DuoSet IC nzyme Linked Immunosorbent Assay, DYC887; R&D Systems, inneapolis, MN), phospho-GSK3 (S9) (DYC-1590; R&D Sysems), and phospho-p65(S536) (7834; Cell Signaling) were asessed by enzyme linked immunosorbent assay in cells transected with as-miR-21 (100 nmol/L) and/or as-miR-155 (100 mol/L) for 24 hours and treated with NT (100 nmol/L) for 24 ours.

Techniques: Protein-Protein interactions, Luciferase, Activity Assay, Real-time Polymerase Chain Reaction, Western Blot, Phospho-proteomics, Enzyme-linked Immunosorbent Assay, Transfection

Journal: Gastroenterology

Article Title: Neurotensin signaling activates microRNAs-21 and -155 and Akt, promotes tumor growth in mice, and is increased in human colon tumors.

doi: 10.1053/j.gastro.2011.07.038

Figure Lengend Snippet: Figure 5. NT activates AKT through suppression of PPP2CA by direct interaction with miR-155. (A) miR-155 binding sites in 3=UTRs of PPP2CA predicted by Lever algorithm analysis. (B) Luciferase activity of the PPP2CA 3=UTRs in HCT116 and DLD1 cells transfected with as-miR-155 (100 nmol/L) or as-miR-155 (100 nmol/L) for 24 hours. (C) Luciferase activity of the PPP2CA 3=UTRs in HCT116 and DLD1 cells treated with NT (100 nmol/L). (D) PPP2CA and -actin protein levels, assessed by Western blot, in NT-treated (100 nmol/L, 6 h) HCT116 and DLD1 cells. (E) AKT phosphorylation (S473) levels, assessed by enzyme-linked immunosorbent assay, in HCT116 and DLD1 cells transfected with miR-155 (100 nmol/L) or si-PPP2CA (100 nmol/L) for 24 hours. (F) NF-B/p65 activity, assessed by enzyme-linked immunosorbent assay, in HCT cells transfected with NT (100 nmol/L) and as-miR-NC (100 nmol/L), as-miR-21 (100 nmol/L), as-miR-155 (100 nmol/L), or with 10 nmol/L of an AKT pharmacologic inhibitor (MK-2206).

Article Snippet: AKT phosphorylation (S473) (DuoSet IC nzyme Linked Immunosorbent Assay, DYC887; R&D Systems, inneapolis, MN), phospho-GSK3 (S9) (DYC-1590; R&D Sysems), and phospho-p65(S536) (7834; Cell Signaling) were asessed by enzyme linked immunosorbent assay in cells transected with as-miR-21 (100 nmol/L) and/or as-miR-155 (100 mol/L) for 24 hours and treated with NT (100 nmol/L) for 24 ours.

Techniques: Binding Assay, Luciferase, Activity Assay, Transfection, Western Blot, Phospho-proteomics, Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: Interferon-γ Suppresses Intestinal Epithelial Aquaporin-1 Expression via Janus Kinase and STAT3 Activation

doi: 10.1371/journal.pone.0118713

Figure Lengend Snippet:

Article Snippet: Phosphorylated AKT (Cat# AF887, R & D Systems, Minneapolis, MN) , 5% BSA in TTBS , 1:400 , goat anti-rabbit.

Techniques: Blocking Assay, Concentration Assay