Insulin-stimulated glucose transport in muscle. Evidence for a protein-kinase-C-dependent component which is unaltered in insulin-resistant mice

The aim of our work was to investigate a possible role of protein kinase C (PKC) in insulin-stimulated glucose uptake in mouse skeletal muscle, and to search for a defect in PKC activation in insulin resistance found in obesity. In isolated soleus muscle of lean mice, insulin (100 nM) and 12-O-tetradecanoylphorbol 13-acetate (TPA) (1 microM) acutely stimulated glucose uptake 3- and 2-fold respectively. The effects of insulin and TPA were not additive. When PKC activity was down-regulated by long-term (24 h) TPA pretreatment, before measurement of glucose transport, the TPA effect was abolished, but in addition insulin-stimulated glucose transport returned to basal values. Furthermore, polymyxin B, which inhibits PKC in muscle extracts, prevented insulin-stimulated glucose uptake in muscle. In muscle of obese insulin-resistant mice, glucose uptake evoked by insulin was decreased, whereas the TPA effect, expressed as a fold increase, was unaltered. Thus both agents stimulated glucose transport to the same extent. Furthermore, no difference was observed when PKC activation by TPA was measured in muscle from lean and obese mice. These results suggest that: (1) PKC is involved in the insulin effect on glucose transport in muscle; (2) PKC activation explains only part of the insulin stimulation of glucose transport; (3) the defect in insulin response in obese mice does not appear to be due to an alteration in the PKC-dependent component of glucose transport. We propose that insulin stimulation of glucose uptake occurs by a sequential two-step mechanism, with first translocation of transporters to the plasma membrane, which is PKC dependent, and second, activation of the glucose transporters. In obesity only the activation step was decreased, whereas the translocation step was unaltered.

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DAG accumulation from saturated fatty acids desensitizes insulin stimulation of glucose uptake in muscle cells

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.2001.280.2.e229 ◽

2001 ◽

Vol 280 (2) ◽

pp. E229-E237 ◽

Cited By ~ 162

Author(s):

Eulàlia Montell ◽

Marco Turini ◽

Mario Marotta ◽

Matthew Roberts ◽

Véronique Noé ◽

...

Keyword(s):

Insulin Resistance ◽

Fatty Acids ◽

Protein Kinase C ◽

Protein Kinase ◽

Glucose Uptake ◽

Insulin Response ◽

Insulin Stimulation ◽

Kinase C ◽

Basal Glucose Uptake ◽

Stimulation Of

The increased availability of saturated lipids has been correlated with development of insulin resistance, although the basis for this impairment is not defined. This work examined the interaction of saturated and unsaturated fatty acids (FA) with insulin stimulation of glucose uptake and its relation to the FA incorporation into different lipid pools in cultured human muscle. It is shown that basal or insulin-stimulated 2-deoxyglucose uptake was unaltered in cells preincubated with oleate, whereas basal glucose uptake was increased and insulin response was impaired in palmitate- and stearate-loaded cells. Analysis of the incorporation of FA into different lipid pools showed that palmitate, stearate, and oleate were similarly incorporated into phospholipids (PL) and did not modify the FA profile. In contrast, differences were observed in the total incorporation of FA into triacylglycerides (TAG): unsaturated FA were readily diverted toward TAG, whereas saturated FA could accumulate as diacylglycerol (DAG). Treatment with palmitate increased the activity of membrane-associated protein kinase C, whereas oleate had no effect. Mixture of palmitate with oleate diverted the saturated FA toward TAG and abolished its effect on glucose uptake. In conclusion, our data indicate that saturated FA-promoted changes in basal glucose uptake and insulin response were not correlated to a modification of the FA profile in PL or TAG accumulation. In contrast, these changes were related to saturated FA being accumulated as DAG and activating protein kinase C. Therefore, our results suggest that accumulation of DAG may be a molecular link between an increased availability of saturated FA and the induction of insulin resistance.

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Protein kinase C is not required for insulin stimulation of hexose uptake in muscle cells in culture

Biochemical Journal ◽

10.1042/bj2420131 ◽

1987 ◽

Vol 242 (1) ◽

pp. 131-136 ◽

Cited By ~ 61

Author(s):

A Klip ◽

T Ramlal

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Glucose Uptake ◽

Phorbol Esters ◽

Suitable Model ◽

Insulin Stimulation ◽

Intact Cells ◽

L6 Cells ◽

Kinase C ◽

Stimulation Of

The L6 skeletal muscle cell line has been identified as a suitable model to study the action of insulin on glucose uptake in muscle [Klip, Li & Logan (1984) Am. J. Physiol. 247, E291-E296]. The signals that transfer information from occupied insulin receptors to glucose transporters remain unknown. Here we report that activation of protein kinase C by exogenous phorbol esters results in stimulation of glucose uptake. Protein C kinase activity was induced to migrate from the cytosolic fraction to the microsomal fraction after 40 min of exposure of intact cells to 4 beta-phorbol 12,13-dibutyrate. In contrast, incubation with insulin did not alter the subcellular distribution of the kinase. Prolonged preincubation of L6 cells with phorbol esters resulted in depletion of kinase C activity, whereas neither the basal rate of glucose uptake nor its stimulation by insulin were affected. This suggests that protein kinase C is expressed in L6 cells, and that insulin stimulation of hexose transport does not involve protein kinase C.

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Protein kinase C modulates insulin action in human skeletal muscle

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.2000.278.3.e553 ◽

2000 ◽

Vol 278 (3) ◽

pp. E553-E562 ◽

Cited By ~ 67

Author(s):

Ronald N. Cortright ◽

John L. Azevedo ◽

Qian Zhou ◽

Madhur Sinha ◽

Walter J. Pories ◽

...

Keyword(s):

Insulin Resistance ◽

Skeletal Muscle ◽

Protein Kinase C ◽

Protein Kinase ◽

Glucose Uptake ◽

Glucose Transport ◽

Insulin Action ◽

Human Skeletal Muscle ◽

Kinase C ◽

Pkc Activation

There is good evidence from cell lines and rodents that elevated protein kinase C (PKC) overexpression/activity causes insulin resistance. Therefore, the present study determined the effects of PKC activation/inhibition on insulin-mediated glucose transport in incubated human skeletal muscle and primary adipocytes to discern a potential role for PKC in insulin action. Rectus abdominus muscle strips or adipocytes from obese, insulin-resistant, and insulin-sensitive patients were incubated in vitro under basal and insulin (100 nM)-stimulated conditions in the presence of GF 109203X (GF), a PKC inhibitor, or 12-deoxyphorbol 13-phenylacetate 20-acetate (dPPA), a PKC activator. PKC inhibition had no effect on basal glucose transport. GF increased ( P < 0.05) insulin-stimulated 2-deoxyglucose (2-DOG) transport approximately twofold above basal. GF plus insulin also increased ( P < 0.05) insulin receptor tyrosine phosphorylation 48% and phosphatidylinositol 3-kinase (PI 3-kinase) activity ∼50% ( P< 0.05) vs. insulin treatment alone. Similar results for GF on glucose uptake were observed in human primary adipocytes. Further support for the hypothesis that elevated PKC activity is related to insulin resistance comes from the finding that PKC activation by dPPA was associated with a 40% decrease ( P < 0.05) in insulin-stimulated 2-DOG transport. Incubation of insulin-sensitive muscles with GF also resulted in enhanced insulin action (∼3-fold above basal). These data demonstrate that certain PKC inhibitors augment insulin-mediated glucose uptake and suggest that PKC may modulate insulin action in human skeletal muscle.

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Tyrosine phosphorylation of specific protein kinase C isoenzymes participates in insulin stimulation of glucose transport in primary cultures of rat skeletal muscle

Diabetes ◽

10.2337/diabetes.48.10.1922 ◽

1999 ◽

Vol 48 (10) ◽

pp. 1922-1929 ◽

Cited By ~ 77

Author(s):

L. Braiman ◽

L. Sheffi-Friedman ◽

A. Bak ◽

T. Tennenbaum ◽

S. R. Sampson

Keyword(s):

Skeletal Muscle ◽

Protein Kinase C ◽

Protein Kinase ◽

Primary Cultures ◽

Specific Protein ◽

Insulin Stimulation ◽

Rat Skeletal Muscle ◽

Kinase C ◽

Stimulation Of ◽

Specific Protein Kinase

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Effect of protein kinase C activation and depletion on insulin stimulation of glycogen synthesis in cultured hepatoma cells

Cellular and Molecular Life Sciences ◽

10.1007/bf01960235 ◽

1988 ◽

Vol 44 (1) ◽

pp. 34-37 ◽

Cited By ~ 11

Author(s):

M. Caron ◽

G. Cherqui ◽

D. Wicek ◽

J. Capeau ◽

J. Bertrand ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Glycogen Synthesis ◽

Hepatoma Cells ◽

Insulin Stimulation ◽

Kinase C ◽

Protein Kinase C Activation ◽

Stimulation Of

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Selected Contribution: PKC activation inhibits Ca2+ signaling in tracheal epithelial cells kept in simulated microgravity

Journal of Applied Physiology ◽

10.1152/jappl.2000.89.2.855 ◽

2000 ◽

Vol 89 (2) ◽

pp. 855-864 ◽

Cited By ~ 4

Author(s):

Jennifer A. Felix ◽

Ellen R. Dirksen ◽

Michael L. Woodruff

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Simulated Microgravity ◽

Tracheal Epithelial Cells ◽

Kinase C ◽

Tracheal Epithelial ◽

Epithelial Sheets ◽

Pkc Activation ◽

Stimulation Of ◽

In Cells

Microgravity has been shown to alter protein kinase C (PKC) activity; therefore, we investigated whether microgravity influences mechanically stimulated Ca2+signaling and ATP-induced Ca2+ oscillations, both of which are modulated by PKC. Rabbit tracheal epithelial outgrowth cultures or suspended epithelial sheets were rotated in bioreactors to simulate microgravity. Mechanical stimulation of a single cell increased the cytosolic Ca2+ concentration in 35–55 cells of both outgrowth cultures and epithelial sheets kept at unit gravity (G) or in simulated microgravity (sμG). In outgrowth cultures, 12- O-tetradecanoylphorbol-13-acetate (TPA; 80 nM), a PKC activator, restricted Ca2+ “waves” to about 10 cells in unit G and to significantly fewer cells in sμG. TPA only slightly reduced the spread of Ca2+ waves in epithelial sheets kept in sμG but did not inhibit Ca2+ waves of sheets kept in unit G. In both cell preparations from both conditions, TPA inhibited ATP-induced Ca2+ oscillations; however, the effect was more pronounced in cells kept in sμG. These results suggest that PKC activation is more robust in cells subjected to sμG.

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Role of protein kinase C in the regulation of glucose transport in the rat adipose cell. Translocation of glucose transporters without stimulation of glucose transport activity.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)52429-7 ◽

1991 ◽

Vol 266 (1) ◽

pp. 261-267 ◽

Cited By ~ 3

Author(s):

J Saltis ◽

A D Habberfield ◽

J J Egan ◽

C Londos ◽

I A Simpson ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Glucose Transport ◽

Glucose Transporters ◽

Transport Activity ◽

Adipose Cell ◽

Kinase C ◽

Stimulation Of

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Regulation of glucose transporters by insulin and extracellular glucose in C2C12 myotubes

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00194.2006 ◽

2006 ◽

Vol 291 (4) ◽

pp. E817-E828 ◽

Cited By ~ 69

Author(s):

Taku Nedachi ◽

Makoto Kanzaki

Keyword(s):

Skeletal Muscle ◽

Glucose Uptake ◽

Glucose Transport ◽

Glucose Transporter ◽

C2c12 Myotubes ◽

Glut4 Translocation ◽

Insulin Stimulation ◽

Extracellular Glucose ◽

Insulin Dependent ◽

Stimulation Of

It is well established that insulin stimulation of glucose uptake in skeletal muscle cells is mediated through translocation of GLUT4 from intracellular storage sites to the cell surface. However, the established skeletal muscle cell lines, with the exception of L6 myocytes, reportedly show minimal insulin-dependent glucose uptake and GLUT4 translocation. Using C2C12 myocytes expressing exofacial-Myc-GLUT4-enhanced cyan fluorescent protein, we herein show that differentiated C2C12 myotubes are equipped with basic GLUT4 translocation machinery that can be activated by insulin stimulation (∼3-fold increase as assessed by anti-Myc antibody uptake and immunostaining assay). However, this insulin stimulation of GLUT4 translocation was difficult to demonstrate with a conventional 2-deoxyglucose uptake assay because of markedly elevated basal glucose uptake via other glucose transporter(s). Intriguingly, the basal glucose transport activity in C2C12 myotubes appeared to be acutely suppressed within 5 min by preincubation with a pathophysiologically high level of extracellular glucose (25 mM). In contrast, this activity was augmented by acute glucose deprivation via an unidentified mechanism that is independent of GLUT4 translocation but is dependent on phosphatidylinositol 3-kinase activity. Taken together, these findings indicate that regulation of the facilitative glucose transport system in differentiated C2C12 myotubes can be achieved through surprisingly acute glucose-dependent modulation of the activity of glucose transporter(s), which apparently contributes to obscuring the insulin augmentation of glucose uptake elicited by GLUT4 translocation. We herein also describe several methods of monitoring insulin-dependent glucose uptake in C2C12 myotubes and propose this cell line to be a useful model for analyzing GLUT4 translocation in skeletal muscle.

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