Defective Akt activation is associated with glucose- but not glucosamine-induced insulin resistance.
3T3-L1 adipocytes develop insulin-resistant glucose transport upon preincubation with high glucose or glucosamine, provided insulin (0.6 nM) is present during preincubation. Insulin receptor substrate-1 (IRS-1)-associated phosphatidylinositol (PI) 3-kinase activity is unaffected (30). Total cellular IRS-1, PI 3-kinase, or Akt concentrations were unchanged. Akt activation in subcellular fractions was assessed by immunoblotting with two phospho-Akt-specific antibodies. Upon acute 100 nM insulin stimulation, plasma membrane (PM)-associated phospho-Akt was highest in cells preincubated in low glucose with no insulin, less in high glucose with no insulin, even less in low glucose+insulin, and lowest in high glucose+insulin. Only high glucose+insulin caused insulin-resistant glucose transport. Acute insulin stimulation increased total PM-Akt about twofold after preincubation without insulin in low or high glucose. Preincubation with 0.6 nM insulin decreased Akt PM translocation by approximately 25% in low and approximately 50% in high glucose. Preincubation with glucosamine did not affect Akt phosphorylation or translocation.


CONCLUSIONS
chronic exposure to high glucose or insulin downregulates acute insulin-stimulated Akt activation, acting synergistically distal to PI 3-kinase. Maximal insulin activates more Akt than required for maximal glucose transport stimulation. Insulin resistance may ensue when PM-associated phospho-Akt decreases below a threshold. High glucose and glucosamine cause insulin resistance by different mechanisms in 3T3-L1 adipocytes.
Nelson
Bryce A
BA
Division of Endocrinology, Diabetes and Medical Genetics, Department of Medicine, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Robinson
Katherine A
KA
Buse
Maria G
MG
D016475:3T3 Cells; D017667:Adipocytes / Q000187:drug effects* / Q000378:metabolism; D000818:Animals; D001692:Biological Transport; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D005944:Glucosamine / Q000494:pharmacology*; D005947:Glucose / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology*; D051275:Glucose Transporter Type 4; D015151:Immunoblotting; D007328:Insulin / Q000008:administration & dosage / Q000494:pharmacology; D055504:Insulin Receptor Substrate Proteins; D007333:Insulin Resistance; D051379:Mice; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D009124:Muscle Proteins; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D010769:Phosphothreonine / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt
D005944:Glucosamine; D005947:Glucose; D051275:Glucose Transporter Type 4; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins; D010768:Phosphoserine; D010769:Phosphothreonine; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; C526295:Irs1 protein, mouse; C495693:Slc2a4 protein, mouse