Genetic downregulation of AMPK-alpha isoforms uncovers the mechanism by which metformin decreases FA uptake and oxidation in skeletal muscle cells.
Metformin is known to improve insulin sensitivity in part via a rise in AMP-activated protein kinase (AMPK) activity and alterations in muscle metabolism. However, a full understanding of how metformin alters AMPK-α(1) vs. AMPK-α(2) activation remains unknown. To study this question, L6 skeletal muscle cells were treated with or without RNAi oligonucleotide sequences to downregulate AMPK-α(1) or AMPK-α(2) protein expression and incubated with or without 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) or metformin and/or insulin. In contrast to AICAR, which preferentially activated AMPK-α(2), metformin preferentially activated AMPK-α(1) in a dose- and time-dependent manner. Metformin increased (P < 0.05) glucose uptake and plasma membrane (PM) Glut4 in a dose- and time-dependent manner. Metformin significantly reduced palmitate uptake (P < 0.05) and oxidation (P < 0.05), and this was accompanied by a similar decrease (P < 0.05) in PM CD36 content but with no change in acetyl-CoA carboxylase (ACC) phosphorylation (P > 0.05). AICAR and metformin similarly increased (P < 0.05) nuclear silent mating-type information regulator 2 homolog 1 (SIRT1) activity. Downregulation of AMPK-α(1) completely prevented the metformin-induced reduction in palmitate uptake and oxidation but only partially reduced the metformin-induced increase in glucose uptake. Downregulation of AMPK-α(2) had no effect on metformin-induced glucose uptake, palmitate uptake, and oxidation. The increase in SIRT1 activity induced by metformin was not affected by downregulation of either AMPK-α(1) or AMPK-α(2). Our data indicate that, in muscle cells, the inhibitory effects of metformin on fatty acid metabolism occur via preferential phosphorylation of AMPK-α(1), and the data indicate that cross talk between AMPK and SIRT1 does not favor either AMPK isozyme.
Bogachus
Lindsey D
LD
Department of Biological Sciences, College of Letters, Arts and Sciences, University of Southern California, Los Angeles, California 90089-0652, USA.
Turcotte
Lorraine P
LP
D000091162:AMP-Activated Protein Kinase Kinases; D000103:Acetyl-CoA Carboxylase / Q000032:analysis; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D018955:CD36 Antigens / Q000032:analysis; D002460:Cell Line; D015536:Down-Regulation; D005227:Fatty Acids / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D051275:Glucose Transporter Type 4 / Q000378:metabolism; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D007328:Insulin / Q000494:pharmacology; D008687:Metformin / Q000494:pharmacology*; D018482:Muscle, Skeletal / Q000187:drug effects* / Q000201:enzymology; D010168:Palmitates / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D051381:Rats; D012265:Ribonucleotides / Q000494:pharmacology; D056564:Sirtuin 1 / Q000032:analysis
D018955:CD36 Antigens; D005227:Fatty Acids; D051275:Glucose Transporter Type 4; D007004:Hypoglycemic Agents; D007328:Insulin; D010168:Palmitates; D012265:Ribonucleotides; C495694:Slc2a4 protein, rat; D000620:Aminoimidazole Carboxamide; D008687:Metformin; D011494:Protein Kinases; D000091162:AMP-Activated Protein Kinase Kinases; C488368:Sirt1 protein, rat; D056564:Sirtuin 1; D000103:Acetyl-CoA Carboxylase; C031143:AICA ribonucleotide; D005947:Glucose