Journal: Cellular signalling

Article Title: Cytotoxic T lymphocyte antigen-2 alpha induces apoptosis of murine T-lymphoma cells and cardiac fibroblasts and is regulated by cAMP/PKA

doi: 10.1016/j.cellsig.2011.05.014

Figure Lengend Snippet: A) and B). WT cells were incubated with PKA-selective (N6-cAMP, N6 100 µM) or Epac-selective (8Me-cAMP, 8Me 100 µM) analogues for 24h A) expression of CTLA-2α was assessed by real-time PCR for 48h B) extent of apoptosis was assayed. The data shown are the representative of those obtained in 3 separate experiments. C) Real-time PCR analysis of CTLA-2α mRNA expression in WT cells treated with the PDE3 inhibitor, milrinone (10 µM) or the PDE4 inhibitor, rolipram (10 µM) for 24h. ** p<0.01 compared to control (vehicle-treated cells).

Article Snippet: Reagents were purchased from Sigma–Aldrich or Invitrogen except Annexin V-fluorescein isothiocyanate (BD Pharmingen); 8-pCPT-2Me-cAMP (8Me-cAMP) and N6-Phenyl-cAMP (N6-cAMP) from Axxora; CTLA-2α primers and GAPDH primers (Integrated DNA Technologies); BCA Protein Assay kit (Pierce); anti-rabbit GAPDH (Abcam); anti-rabbit IgG-horseradish peroxidase (Cell Signaling Technology); Z-FY(tBu)-DMK (Axxora); Cathepsin L inhibitor I (C-I inhibitor, Z-FF-FMK, Calbiochem); Cytobuster protein extraction reagent (Novagene); QPCR Mastermix Plus SYBR Green Kit (Eurogentec); Versagene RNA Cell Kit (Gentra); Control siRNA and CTLA-2α siRNA (Ambion).

Techniques: Incubation, Expressing, Real-time Polymerase Chain Reaction