Journal: Acta Pharmacologica Sinica

Article Title: Long-term ethanol exposure inhibits glucose transporter 4 expression via an AMPK-dependent pathway in adipocytes

doi: 10.1038/aps.2010.11

Figure Lengend Snippet: Long-term ethanol exposure impaired activation, but not expression of AMPK in rat adipose tissue. After feeding rats with ethanol at the daily dosage of 0, 0.5, or 5 g·kg−1·d−1 for 22 weeks, we determined mRNA levels of AMPK α1 and α2 isoforms by RT-PCR (A), protein levels of total-AMPK (T-AMPK) and phosphorylated-AMPK (pAMPK) by Western blotting (B). mRNA expression was normalized by GAPDH, and protein expression was normalized by β-actin. Values are given as means±SD (n=12 for low-dose ethanol and control group, n=10 for high-dose ethanol group). bP<0.05, cP<0.01 vs controls. eP<0.05 vs the low-dose ethanol group.

Article Snippet: All membranes were incubated overnight at 4 °C with primary antibodies against total AMPKα (rabbit polyclonal antibody, 1:1000 dilution, Cell Signaling, Danvers, MA,USA), pAMPKα (rabbit polyclonal antibody, 1:1000 dilution, directed against both α1 and α2 isoforms of the enzyme phosphorylated at Thr172, Cell Signaling, Danvers, MA,USA), pACC (rabbit polyclonal antibody, Ser-79; 1:1000 dilution, Cell Signaling, Danvers, MA, USA), MEF2 (rabbit polyclonal antibody, 1:100 dilution, Santa Cruz, USA), MEF2A (rabbit monoclonal antibody, 1:1000 dilution, Abcam Ltd, Cambridgeshire, UK ), MEF2D (goat polyclonal antibody, 1:200 dilution, Santa Cruz, USA), or GLUT4 (rabbit polyclonal antibody, 1:2500 dilution, Abcam Ltd, Cambridgeshire, UK).

Techniques: Activation Assay, Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, Control