wga agarose slurry  (Vector Biolabs)

Journal: Journal of Biological Chemistry

Article Title: Insulin-like Growth Factor-1 Increases Synthesis of Collagen Type I via Induction of the mRNA-binding Protein LARP6 Expression and Binding to the 5′ Stem-loop of COL1a1 and COL1a2 mRNA

doi: 10.1074/jbc.m113.518951

Figure Lengend Snippet: FIGURE 4. IGF-1 regulation of LARP6 expression. A, real-time RT-PCR determined expression of LARP6 mRNA after 6 h treatment with IGF-1 (100 ng/ml). n 4, Mean S.E. B–D, Western blot of lysate from HASMCs treated for 6 h with IGF-1 (100 ng/ml) in presence of indicated chemical inhibitor: (B) 1 g/ml actinomycin D (act d), (C) 1 g/ml cycloheximide (ch), and (D) 50 M LY29004 (PI3K-inhibitor) or 250 nM Akt VIII (Akt1/2-inhibitor). E, Western blot of lysate from HASMCs overexpressing constitutively active Akt1 and Akt2. Cells were infected with ad-CMV-Akt1 (Myr) (CA-Akt1) and ad-CMV-Akt2 (Myr) (CA-Akt2) at an MOI of 20 for 48 h. F, Western blot of lysate from HASMCs treated for 6 h with IGF-1 (100 ng/ml) in presence of 160 nM PF4708671 (PF, p70S6k inhibitor). Western blot results (B–F) are representative of three independent experiments.

Article Snippet: Adenoviral vectors to express constitutively active Akt1 (ad-CMV-Akt1(Myr)) and Akt2 (ad-CMV-Akt2(Myr)) were purchased from Vector BioLabs (Philadelphia, PA) and cells were incubated at a multiplicity of infection (MOI) of 20 for 48 h. Animals—All animal experiments were performed according to protocols approved by the Institutional Animal Care and Use Committee.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Infection