Sterol regulatory element binding protein 1c srebp 1c

The chemicals for Western blotting were purchased from Amersham (Bucks, UK). RNAzol^TM B was purchased from TEL-TEST Inc. (Friendwood, TX, USA). Primary antibodies against FoxO1, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), transcription factor IIB (TFIIB), tyrosine reside phosphorylation of insulin receptor substrate (p-IRS (Tyr632)), p-IRS (Ser307), phosphorylation-serine/threonine protein kinase (p-Akt), total Akt, peroxisome proliferator-activated receptor-alpha (PPARα), PPARβ, PPARγ, β-arrestin, phosphorylation-AMP-activated protein kinase (p-AMPK), AMPK, sterol regulatory element-binding protein 1c (SREBP-1c), and PAR2 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Primary antibodies against p-FoxO1 (Thr24) and p-FoxO1 (Ser256) were manufactured by Cell Signaling Technology, Inc. (Danvers, MA, USA). Secondary antibodies, both anti-rabbit and anti-mouse immunoglobulin G (IgG)-horseradish peroxidase-conjugated antibodies, were purchased from Amersham. Horseradish peroxidase-conjugated donkey anti-sheep/goat IgG was obtained from Serotec (Oxford, UK). Polyvinylidene difluoride (PVDF) transfer membranes were obtained from manufacturer Millipore Corporation (Bedford, MA, USA).

Samples were prepared from AML-12 cells, which were treated as described above, and subjected to immunoblotting, as described previously (23 (link)). The proteins were resolved on a 10% gel and transferred to polyvinylidine difluoride membrane (PVDF) (Millipore, Bedford, MA, USA). Then membrane was incubated with primary antibodies of SIRT1 (Cell Signaling Technology, Canvers, MA, USA), AMPK (Cell Signaling Technology), phospho-AMPK (pAMPK) (Cell Signaling Technology), sterol regulatory element-binding protein-1c (SREBP-1c, Santa Cruz Biotechnology, Dallas, TX, USA), peroxisome proliferator-activated receptor alpha (PPARα, Abcam, Cambridge, UK) and β-actin (Cell Signaling Technology) at 4°C for overnight, and secondary antibodies of Mouse IgG (Cell signaling Technology) and Rabbit IgG (Cell signaling Technology) were incubated at room temperature for 1 hr. The bands detected using an enhanced chemiluminescence (ECL) system (Amersham, Buckinghamshire, UK).