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Am J Physiol Endocrinol Metab 285: E295-E302, 2003; doi:10.1152/ajpendo.00044.2003
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Increased glucose uptake promotes oxidative stress and PKC-{delta} activation in adipocytes of obese, insulin-resistant mice

Ilana Talior,1 Merav Yarkoni,1 Nava Bashan,2 and Hagit Eldar-Finkelman1

1Department of Human Genetics and Molecular Medicine, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978; and 2Department of Clinical Biochemistry, Faculty of Health, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel

Submitted 31 January 2003 ; accepted in final form 20 April 2003


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURE
 REFERENCES
 
Increased oxidative stress is believed to be one of the mechanisms responsible for hyperglycemia-induced tissue damage and diabetic complications. In these studies, we undertook to characterize glucose uptake and oxidative stress in adipocytes of type 2 diabetic animals and to determine whether these promote the activation of PKC-{delta}. The adipocytes used were isolated either from C57Bl/6J mice that were raised on a high-fat diet (HF) and developed obesity and insulin resistance or from control animals. Basal glucose uptake significantly increased (8-fold) in HF adipocytes, and this was accompanied with upregulation of GLUT1 expression levels. Insulin-induced glucose uptake was inhibited in HF adipocytes and GLUT4 content reduced by 20% in these adipocytes. Reactive oxygen species (ROS) increased twofold in HF adipocytes compared with control adipocytes and were largely reduced with decreased glucose concentrations. At zero glucose, ROS levels were reduced to the normal levels seen in control adipocytes. The activity of PKC-{delta} increased twofold in HF adipocytes compared with control adipocytes and was further activated by H2O2. Moreover, PKC-{delta} activity was inhibited in HF adipocytes either by glucose deprivation or by treatment with the antioxidant N-acetyl-L-cysteine. In summary, we propose that increased glucose intake in HF adipocytes increases oxidative stress, which in turn promotes the activation of PKC-{delta}. These consequential events may be responsible, at least in part, for development of HF diet-induced insulin resistance in the fat tissue.

oxidative stress; protein kinase C-{delta}; insulin resistance; glucose metabolism; C57Bl/6J mice

REACTIVE OXYGEN SPECIES (ROS) and the oxidative damage that they cause are thought to play a key role in the pathogenesis of diabetes and its complications (5, 18). It has been proposed that hyperglycemia contributes to the generation in various tissues of free radicals that, in turn, can lead to multiple complications and, at least in part, to the induction of muscular and adipocyte insulin resistance (10, 15, 39). It has been demonstrated that chronic exposure of cells (such as endothelial cells, mesangial cells, or smooth muscle cells) to high glucose increases intercellular ROS (11, 22, 45, 47, 56). On the other hand, oxidative stress could impair insulin action. Treatment of cells such as L6 myotubes or 3T3-L1 adipocytes with hydrogen peroxide (H2O2) inhibited insulin-induced glucose uptake and altered the translocation of GLUT4 to the plasma membrane (6, 32, 52). The mechanism by which oxidative stress causes insulin resistance is not fully clear. Part of the explanation may stem from the fact that ROS affects various components of the insulin-signaling cascade, as well as the gene regulation of the glucose transporters GLUT1 and GLUT4 (5, 9, 32, 52, 63). In addition, ROS has been shown to activate Syk (49, 55), Lck (23), and the stress-activated protein kinases p38 (6) and c-Jun NH2-terminal kinases (36) and protein kinase C (PKC) (30) in various cell types.

The novel PKC-{delta} is one of the multifunctional isoenzymes of protein kinase C, a family of serine/threonine kinases involved in intracellular signals regulating growth and metabolism, differentiation, and apoptosis. PKC-{delta} itself was implicated in multiple cellular processes, some of which were related to diabetes. In cultured muscle cells, PKC-{delta} was shown to be a major signal in insulin-induced glucose transport (8) and to interact with the insulin receptor in response to insulin (51). Furthermore, overexpression of PKC-{delta} in skeletal muscle cultures abrogated the inhibitory effect of tumor necrosis factor-{alpha} (TNF-{alpha}) on the insulin-induced insulin receptor autophosphorylation (51). Overexpression of PKC-{delta} in 3T3-L1 adipocytes enhanced basal glucose uptake (61) but did not promote GLUT4 translocation when expressed in primary rat adipocytes (2). In mesangial cells, hyperglycemia increased the membrane- and particulate-associated PKC-{delta} (28), and in vascular cells of diabetic rats its cellular localization differed from that of normal animals (25). Recently, PKC-{delta} was implicated in oxidative stress and shown to be activated by H2O2. This activation was accompanied by enhanced phosphorylation of serine and tyrosine residues (30, 31).

Oxidative stress is increased in cultured primary adipocytes exposed to high glucose (37) and was shown both to inhibit insulin sensitivity and to alter the expression levels of glucose transporters in 3T3-L1 adipocytes (52). To the best of our knowledge, however, increased ROS production in adipocytes isolated from diabetic animals has not been documented. In the present study, we undertook to characterize glucose metabolism in adipocytes isolated from obese, insulin-resistant C57Bl/6J mice, an inbred mouse strain susceptible to diet-induced obesity and diabetes (60), and to examine whether this is linked to increased oxidative stress and changes in PKC-{delta} activity. The present studies indicate that glucose uptake is significantly increased in high-fat (HF) adipocytes and this, in turn, promotes oxidative stress and activation of PKC-{delta}.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 DISCLOSURE
 REFERENCES
 
Materials. Insulin was a gift from Novo Nordisk (Bagsværd, Denmark). The radioactive materials were purchased from Amersham Pharmacia Biotech (Piscataway, NJ). GLUT1 and GLUT4 antibodies were from Chemicon International (Temecula, CA), PKC-{delta} and PKC-{alpha} antibodies were from Santa Cruz Biotechnology (Santa Cruz, CA), and the phospho-PKC-{delta} (Ser643) and PKB antibody were from Cell Signaling (Beverly, MA). Insulin receptor and phospho-PKC-{alpha} (Ser657) antibodies were from Upstate Biotechnology (Lake Placid, NY). All other reagents were purchased from Sigma (Rehovot, Israel).

Animal and diets. Four-week-old mice were randomly assigned to receive either standard laboratory food or a high-fat diet containing 35% lard (Bioserve, Frenchtown, NJ), in which 55% of the calories came from fat. The animals were housed in individual cages with free access to water in a temperature-controlled facility with a 12:12-h light-dark cycle; the animals were weighed periodically. Animals were used for testing after 16 wk on their designated diets. Animals were starved for 6 h before experiments and then killed. Blood samples were taken immediately from the aorta. Plasma glucose was measured by the Sugar Accutrend Sensor (Roche, Mannheim, Germany), and plasma insulin was assayed with a radioimmunoassay kit (INSIK-5, DiaSorin, Saluggia, Italy). Animal care followed the institutional animal care and use committee guidelines.

Preparation of adipocytes. To obtain an equal number of cells, the protocol required pooled cells harvested from three to four lean mice for each obese mouse. Adipocytes were isolated from the epididymal fat pad by digestion with 0.4 mg/ml collagenase (Worthington Biochemical, Lakewood, NJ), as described previously (33). Digested fat pads were passed through nylon mesh, and the cells were washed three times with Krebs-bicarbonate buffer (pH 7.4) that contained 1% bovine serum albumin (fraction V; Boehringer Mannheim, Mannheim, Germany), 10 mM HEPES (pH 7.3), 5 mM glucose, and 200 nM adenosine. Aliquots of cells were used to determine the cell concentrations, as described (14), and to determine the amount of genomic DNA (Wizard Genomic; Promega, Madison, WI).

Histology and microscopy. Epididymal fat tissues were fixed in 4% buffered formalin and embedded in paraffin. The paraffin-embedded tissue was sectioned (4 µm) and stained with hematoxylin and eosin (Bio Optica, Milan, Italy). Micro-graphs were taken at x100 magnification.

Glucose transport. Cells (1 x 106) in 1- to 1.5-ml aliquots were divided in plastic vials and incubated with shaking at 37°C. For glucose transport measurements, cells were incubated with insulin at the indicated concentrations for 1 h, followed by the addition of 2-deoxy-[3H]glucose (0.5 µCi/vial) for 30 min. The assay was terminated by the centrifugation of cells through dinonylphthalate (Irvine, CA), and 3H was quantitated with a liquid scintillation analyzer (Packard). Nonspecific uptake of 2-deoxy-[3H]glucose was determined by the addition of cytochalasin B (50 µM) 30 min before the addition of the radioactive material.

Total membrane and cell lysate preparations. Proteins were extracted from the adipocytes or from fat tissue with buffer G (25 µg/ml each of 50 mM Tris, pH 7.5, 150 mM NaCl, 1 mM EDTA, 1 mM EGTA, 10 mM {beta}-glycerophosphate, 50 mM NaF, 5% glycerol, 1% Triton X-100, and leupeptin, aprotinin, and pepstatin A). After centrifugation, supernatants were collected and subjected to gel electrophoresis followed by immunoblot analysis with the indicated antibodies according to the manufacturer's procedures. Total membranes were prepared from adipocytes of control and diabetic animals (5 x 106 cells) as previously described (46), with some modifications. In brief, adipocytes were homogenized in buffer C (50 mM HEPES, pH 7.3, 10 mM NaF, 10 mM KCl, and 0.1 mM Na3VO4 and 25 µg/ml each of protease inhibitors leupeptin, aprotinin, and pepstatin A) by 30 strokes and centrifuged at 1,000 g at 4°C to remove nuclear pellet. The resulting supernatants were centrifuged at high speed (100,000 g) at 4°C to precipitate membranes. Membranes were resuspended with buffer C (100 µl), and equal aliquots of membranes (30 µl) were subjected to gel electrophoresis followed by immunoblot analysis.

Measurement of intracellular ROS generation. The determination of ROS was based on the oxidation of 2',7'-dichlorodihydrofluorescin (DCHF) by peroxide, as previously described, with some modifications (59). In brief, adipocytes were incubated either with various concentrations of glucose (0–5 mM), or with N-acetyl-L-cysteine (NAC; 0.1, 0.2, or 0.4 mM) for 1 h and then incubated with DCHF (30 µM) for an additional 40 min. Incubation with H2O2, a ROS donor, served as a positive control, and known concentrations of DCHF incubated with 20 mM NaOH were used as standards. Cells were washed, and the fluorescence of 2',7'-dichlorofluorescein (DCF) was measured in triplicate samples in a multiplate fluorometer (Fl-600; excitation at 488 nm, emission at 530 nm).

PKC-{delta} activity. PKC-{delta} activity was assayed as previously described (30). In brief, epididymal fat tissues were removed from both the control and the HF mice and immediately homogenized with buffer H (25 µg/ml each of 20 mM Tris, pH 7.5, 50 mM {beta}-glycerophosphate, 1 mM EDTA, 1 mM EGTA, 1 mM DTT, 10 mM NaF, 5% glycerol, 1% Triton X-100, and leupeptin, aprotinin, and pepstatin A). A similar procedure was performed in isolated adipocytes, except that cells were not homogenized. The lysates were centrifuged, and equal protein aliquots were used for PKC-{delta} assays. The enzyme was immunoprecipitated for 2 h with anti-PKC-{delta} antibody bound to protein Sepharose. The immunocomplexes were washed extensively, and the kinase reaction was performed at 30°C in 20 mM Tris · HCl, pH 7.3, with 10 mM MgCl2, 100 µM [{gamma}-32P]ATP, 40 µM phosphatidylserine, and 3 µg histone H1 (per reaction). Reactions were terminated after 20 min by the addition of SDS-PAGE load buffer and were analyzed in 15% acrylamide gels by autoradiography. Autoradiography was quantitated by densitometry.

Expression of results. Results are expressed as means ± SE for the number of cell preparations or animals, as indicated. Statistical analysis was performed by Origin Professional 6.0, using one-way ANOVA to compare the control and HF results. A difference was considered to be statistically significant when P < 0.05.


   RESULTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 DISCLOSURE
 REFERENCES
 
Characterization of HF mice and HF adipocytes. Table 1 characterizes C57BL/6J mice, which were fed the HF diet, and their respective controls, which were fed standard laboratory food. After 16 wk on their designated diets, the animals on the HF diet developed hyperglycemia and hyperinsulinemia. The HF mice increased their weight by 30–40% compared with control animals. Notably, there was about a fourfold increase in the number of adipocytes in each epididymal fat pad of HF mice compared with their respective controls. Determination of genomic DNA further confirmed this result and indicated that DNA content increased about fourfold in adipocytes isolated from HF fat pads compared with that of control (0.2 vs. 0.05 µg/fat pad of HF or control, respectively). Determination of the protein content of the fat tissues (Table 1) indicated that protein content also increased approximately fourfold in the HF tissue compared with control. Hence, protein content per cell did not significantly change in HF adipocytes compared with control adipocytes (calculated value 4 ng/cell), suggesting that the increase in the HF adipocytes size [~2.5-fold (cell diameter); Fig. 1] results mainly from accumulation of lipids.


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  		Table 1. Characteristics of control and HF mice

 


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  		Fig. 1. Fat tissue morphology of high-fat diet-fed and control mice. Representative adipose tissue paraffin sections of the control (C) or obese diabetic (HF) mice stained with hematoxylin and eosin. Micro-graphs were taken at initial magnification of x100.

 

Increased glucose uptake in HF adipocytes is accompanied by GLUT1 upregulation. Basal glucose uptake was strikingly higher in adipocytes from the HF mice compared with the controls (8.6-fold; Fig. 2A) and was blocked with cytochalasin B, indicating that glucose uptake is specific (data not shown). Insulin-stimulated glucose uptake is approximately threefold higher in control adipocytes, but insulin had no stimulatory effect in adipocytes from diabetic animals (Fig. 2A). This is consistent with studies performed in adipocytes derived from type 2 diabetic animals, showing a significant increase in basal glucose uptake and inhibition in insulin-stimulated glucose uptake (13, 50). Analysis of GLUT1 expression levels in total membranes prepared from an equal number of adipocytes indicated that GLUT1 is consistently higher in HF adipocytes (Fig. 2B). The elevation in GLUT1 in HF adipocytes seems to be specific, because the amount of another membrane protein, the insulin receptor, was not elevated but rather was reduced in HF adipocytes compared with controls (Fig. 2C). The expression level of GLUT4, which is known to be located in intracellular membranes, was reduced by 20 ± 4%, whereas no change in the cytosolic PKB was observed in either control or HF adipocytes (Fig. 2, D and E). Notably, alterations in GLUT1 and GLUT4 content were also observed in adipocytes of Zucker diabetic rats (46). All together, our results suggest that the increase in basal glucose uptake in HF adipocytes stems from specific upregulation of GLUT1 and is not due to a general phenomenon, namely increased protein expression levels, because other proteins were not elevated.



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  		Fig. 2. Glucose uptake, GLUT1, GLUT4, insulin receptor (IR), and protein kinase B content in adipocytes from HF and control animals. A: 2-deoxy-[3H]glucose uptake in absence and presence of various insulin concentrations. Results are presented as 2-deoxyglucose incorporation into 106 cells in 1 min and are means ± SE of 5 independent experiments. B: GLUT1 protein content was measured in total membrane prepared from 5 x 106 adipocytes by Western blot analysis as described in MATERIALS AND METHODS. Shown are representative blot (top) and densitometry analysis (bottom) of independent experiments in which GLUT1 level in controls was assigned a value of 1. *P < 0.05 C vs. HF. C: IR protein content in total membrane preparation. D: GLUT4 protein content in total cell lysates was determined by Western blot analysis as described in MATERIALS AND METHODS. E: PKB protein content in total cell lysate was determined by Western blot analysis as described in MATERIALS AND METHODS. In C, D, and E, representative blots performed 4 times are shown. Cell lysate and membranes were prepared as described in MATERIALS AND METHODS, and equal amounts of membranes (corresponding to 1.6 x 106 cells) or cell lysates (corresponding to 1.50 x 105 cells) were loaded onto gels.

 

Intracellular ROS is increased in HF adipocytes. We next examined whether oxidative stress is increased in HF adipocytes and whether this may be related to the enhanced glucose uptake of these adipocytes. Intracellular ROS measurements presented in Fig. 3A indicate that ROS levels increased twofold in HF adipocytes compared with the control cells. Incubation of cells with a glucose-free medium resulted in a 50% decrease in ROS levels in the HF adipocytes (Fig. 3A). Figure 3B demonstrates the production of ROS in HF adipocytes as a function of medium glucose concentrations. As can be seen, elevation in ROS production correlates with elevation in medium glucose concentration, suggesting that elevation in glucose uptake in HF adipocytes is a major contributor to ROS production.



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  		Fig. 3. Reactive oxygen species (ROS) levels in HF and control adipocytes. DCF, 2',7'-dichlorofluorescein. A: ROS levels were determined in control cells after 1-h incubation in glucose-containing medium and in HF adipocytes incubated for 1 h in presence or absence of glucose, as described in MATERIALS AND METHODS. *P < 0.05 C vs. HF or HF incubated in absence vs. HF presence of glucose. Results represent means ± SE of 5 independent experiments. B: same as A, except that HF adipocytes were incubated with various concentrations of glucose (1 h) as indicated. Results represent means ± SE of 3 independent experiments.

 

PKC-{delta} activity is increased in HF adipocytes. Previous studies demonstrated that PKC-{delta} is activated in various cell types by oxidative stress (30, 31). Thus we evaluated both PKC-{delta} activity (using histone H1 as substrate) and the abundance of PKC-{delta} and its phosphorylation state in adipocytes derived from both HF and control mice (Fig. 4). As can be evaluated (Fig. 4A), PKC-{delta} activity was higher in HF adipocytes compared with controls. Immunoblot analysis with the anti-phospho-PKC-{delta} antibody, which recognizes the phosphorylated Ser643, a site known to be essential for the activation of PKC-{delta} when phosphorylated (35) and is phosphorylated in response to H2O2 (31), indicated that phosphorylation of Ser643 incresaed in HF adipocytes compared with controls with no change in the expression of PKC-{delta} (Fig. 4, B and C). The phosphorylation of the conventional PKC-{alpha} isoform on Ser657, a site known to be crucial for the enzyme activity when phosphorylated (7), and its expression levels were similar in HF and control adipocytes (Fig. 4D).



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  		Fig. 4. PKC-{delta} activity is increased in HF adipocytes. A: PKC-{delta} activity measured after immunoprecipitation by phosphorylation of histone H1, as described in MATERIALS AND METHODS. Amount of phosphorylated (p-PKC-{delta}; B) and total PKC-{delta} (C) was determined by immunoblot analysis with either anti-phospho-PKC-{delta} (B) or anti-PKC-{delta} antibody (C) for control or HF samples prepared from equal amounts of protein from control or HF tissue (30 µg each). Bottom: means ± SE of densitometry analysis of 5 independent experiments. Value obtained for control was designated 1, and values obtained for HF are presented as fold increase over control. *P < 0.05 C vs. HF. Immunoblot analysis with anti-phospho-PKC-{alpha} (D) or anti-PKC-{alpha} antibody (E) for control or HF samples prepared from equal amounts of proteins (20 µg each). Immunoblots of 2 of 4 independent experiments are shown.

 

PKC-{delta} activity is activated by H2O2 and inhibited by depletion of glucose and NAC. To further investigate the link between ROS and PKC-{delta}, the effect of H2O2 on PKC-{delta} in HF adipocytes was studied. As shown in Fig. 5A, exposure to H2O2 (200 µM or 2 mM) activated PKC-{delta}, suggesting that the increase in ROS production can contribute to the activation of PKC-{delta} in HF adipocytes. Furthermore, when cells were incubated with glucose-free medium (1 h), PKC-{delta} activity was greatly inhibited (Fig. 5B). To further asses the role of ROS in activating PKC-{delta}, we used NAC, a known free-radical scavenger previously shown to deplete intracellular ROS levels in adipocytes (37). Treatment of HF adipocytes with increasing NAC concentrations reduced ROS production (Fig. 5C) and basal PKC-{delta} activity (Fig. 5D). Taken together, these results suggest that increased oxidative stress in HF adipocytes is dependent on glucose efflux and promotes the activation of PKC-{delta}.



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  		Fig. 5. PKC-{delta} is activated by ROS in HF adipocytes. A: HF adipocytes were treated with H2O2 at indicated concentrations, and immunoprecipitated PKC-{delta} activity assays of the phosphorylation of histone H1 are shown. Bottom: densitometry analysis of 3 independent experiments, in which phosphorylated histone H1 level in controls was assigned a value of 1. B: HF adipocytes were incubated with glucose-free medium (1 h), and PKC-{delta} activity was assayed as described in A. Bottom: densitometry analysis of 3 independent experiments, in which phosphorylated histone H1 level in controls was assigned a value of 1. C: ROS levels were determined for HF adipocytes incubated for 1 h with indicated concentrations of antioxidant N-acetyl-L-cysteine (NAC). Results represent means ± SE of 3 independent experiments. D: HF adipocytes were incubated with NAC at indicated concentrations for 1 h. PKC-{delta} activity was assayed as described in A. Bottom: densitometry analysis of 3 independent experiments, in which phosphorylated histone H1 level in controls was assigned a value of 1. *P < 0.05 no treatment vs. treatment with H2O2 glucose or NAC, respectively.

 


   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
DISCLOSURE
 REFERENCES
 
Numerous reports have shown that oxidative stress is increased in animal models of diabetes and in patients with diabetes (5, 18). In this study, we used an ex vivo system of isolated adipocytes from diabetic mice (HF adipocytes) to investigate whether oxidative stress is increased in these adipocytes and whether this may be related to hyperglycemia and/or activation of PKC. We showed that glucose uptake and ROS levels increased significantly in HF adipocytes compared with control adipocytes. Subsequently, ROS levels were reduced by decreased medium glucose concentrations in HF adipocytes, and in the absence of glucose its levels were reduced twofold, reaching the "normal" ROS levels observed in the control adipocytes. In addition, PKC-{delta} activity was higher in HF adipocytes compared with controls and could be further activated by H2O2. Moreover, the enzyme activity was inhibited by either the antioxidant NAC or the depletion of glucose. Taken together, these results enabled us to propose that the increase in glucose efflux into HF adipocytes may increase intracellular ROS production that, in turn, promoted the activation of PKC-{delta}. These intracellular events could represent, at least in part, one of the mechanisms responsible for induction of insulin resistance in the fat tissue. It should be noted, however, that our studies do not eliminate the role of additional in vivo factors such as hyperinsulinemia, hyperlipidemia, and adipocytokines that could contribute to the increased oxidative stress during the HF diet feeding period, as these factors were shown to elevate ROS levels in various cell systems (1, 5, 17, 20, 29, 42, 44, 64).

GLUT1 levels were significantly increased in HF adipocytes (Fig. 2B). Previous studies demonstrated that GLUT1 expression levels might be affected by stress conditions such as hyperglycemia, hyperinsulinemia, or TNF-{alpha} (24, 34, 40). Although we do not know the exact mechanisms leading to increased GLUT1 expression in HF adipocytes, we suggest that the elevation in GLUT1 was responsible for the increased glucose uptake of HF adipocytes, which in turn led to the elevation of ROS. On the other hand, ROS production could further lead to increased GLUT1 transcription rates. The latter supposition is based on previous studies that indicated that induction of oxidative stress in 3T3-L1 adipocytes or L6 muscle cells resulted in elevation of GLUT1 (32, 52). Thus it is suggested that a vicious cycle may be operating between GLUT1 content and increased oxidative stress in HF adipocytes.

The generation of ROS as a byproduct of the mitochondrial electron transport chain has long been attributed to the high rates of glucose metabolism (4, 16, 54, 63). Nevertheless, additional mechanisms may be operating to produce ROS under hyperglycemic conditions, including the formation of advanced glycation end products (5, 42) or activation of oxidases such as NADPH oxidase (26, 41, 62). We do not know at this point the precise source of the production of ROS in HF adipocytes. Further studies should elucidate this problem.

Our studies indicated that insulin receptor levels significantly decreased in HF adipocytes (Fig. 2C). This phenomenon was reported in various tissues and cells in conditions associated with insulin resistance (21, 38, 65). Hyperinsulinemia has been suggested to be a prominent factor responsible for this phenomenon (27, 48), thereby indicating that downregulation of insulin receptors is a secondary effect in insulin resistance. Nevertheless, it is still possible that reduced levels of insulin receptors in HF adipocytes could limit the sensitivity of the cells to insulin. On the other hand, it should be noted that the remaining insulin receptors (~50%) could still transmit intracellular insulin signaling, as the receptors are most likely not defected (38).

The novel PKC isoform PKC-{delta} was shown to be activated by oxidative stress in various cell systems. Initially, it was shown that H2O2 led to activation in COS7 cells of PKC-{delta}, which was constitutively active and independent of the lipid coactivator, diacylglycerol (30, 31). Additionally, oxidative stress was shown to stimulate PKC-{delta} and c-Abl tyrosine kinase association, facilitating the activation of c-Abl by PKC-{delta} (58). In vascular smooth muscle cells, H2O2 activation of PKC-{delta} was required for the activation of platelet-derived growth factor receptor (53). PKC-{delta} was also shown to mediate oxidative stress-induced activation of several signaling components, including the nonreceptor tyrosine kinases JAK2 and PYK2 (19), as well as the serine/threonine protein kinase MAPK (12). Thus PKC-{delta} may play a central role in oxidative stress-induced cellular processes important in the development of insulin resistance in HF adipocytes. Further studies are needed to examine the downstream targets activated by PKC-{delta} in these cells.

PKC was initially shown to play a role in glucose transport in primary rat adipocytes (43, 57). Overexpression of PKC-{delta} in 3T3-L1 adipocytes enhanced both basal and insulin-induced glucose transport (61). Another study reported that PKC-{alpha}, PKC-{beta}, PKC-{delta}, and PKC-{epsilon} were not involved in insulin-induced GLUT4 translocation (3). Thus it may be concluded that elevated PKC-{delta} activity could contribute to increased basal glucose uptake in HF adipocytes; on the other hand, and in agreement with previous findings (3), it has no effect on insulin-induced GLUT4 translocation in these adipocytes.

In summary, we suggest that the consequential events of glucose uptake, ROS production, and activation of PKC-{delta} may represent one of the mechanisms responsible for the development of insulin resistance in HF adipocytes in response to HF diet feeding.


   DISCLOSURE
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURE
REFERENCES
 
This work was supported by the Israeli Diabetes Foundation and by the Annual Award of the Hendrik and Irene Gutwirth Research Prize in Diabetes Mellitus, which was awarded to H. Eldar-Finkelman.


   ACKNOWLEDGMENTS
 
We thank Prof. Zvi Nevo from the Department of Clinical Biochemistry for conducting the histological studies.


   FOOTNOTES
 

Address for reprint requests and other correspondence: H. Eldar-Finkelman, Dept. of Human Genetics and Molecular Medicine, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel (E-mail: heldar{at}post.tau.ac.il).

Submitted 31 January 2003

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 DISCLOSURE
 REFERENCES
 

  1. Araujo FB, Barbosa DS, Hsin CY, Maranhao RC, and Abdalla DS. Evaluation of oxidative stress in patients with hyperlipidemia. Atherosclerosis 117: 61–71, 1995.[ISI][Medline]
  2. Bandyopadhyay G, Standaert ML, Kikkawa U, Ono Y, Moscat J, and Farese RV. Effects of transiently expressed atypical (zeta, lambda), conventional (alpha, beta) and novel (delta, epsilon) protein kinase C isoforms on insulin-stimulated translocation of epitope-tagged GLUT4 glucose transporters in rat adipocytes: specific interchangeable effects of protein kinases C-zeta and C-lambda. Biochem J 337: 461–470, 1999.[ISI][Medline]
  3. Bandyopadhyay G, Standaert ML, Zhao L, Yu B, Avignon A, Galloway L, Karnam P, Moscat J, and Farese RV. Activation of protein kinase C (alpha, beta, and zeta) by insulin in 3T3/L1 cells. Transfection studies suggest a role for PKC-zeta in glucose transport. J Biol Chem 272: 2551–2558, 1997.[Abstract/Free Full Text]
  4. Batandier C, Fontaine E, Keriel C, and Leverve XM. Determination of mitochondrial reactive oxygen species: methodological aspects. J Cell Mol Med 6: 175–187, 2002.[ISI][Medline]
  5. Baynes JW. Role of oxidative stress in development of complications in diabetes. Diabetes 40: 405–412, 1991.[Abstract]
  6. Blair AS, Hajduch E, Litherland GJ, and Hundal HS. Regulation of glucose transport and glycogen synthesis in L6 muscle cells during oxidative stress. Evidence for cross-talk between the insulin and SAPK2/p38 mitogen-activated protein kinase signaling pathways. J Biol Chem 274: 36293–36299, 1999.[Abstract/Free Full Text]
  7. Bornancin F and Parker PJ. Phosphorylation of protein kinase C-alpha on serine 657 controls the accumulation of active enzyme and contributes to its phosphatase-resistant state. J Biol Chem 272: 3544–3549, 1997.[Abstract/Free Full Text]
  8. Braiman L, Alt A, Kuroki T, Ohba M, Bak A, Tennenbaum T, and Sampson SR. Protein kinase C-{delta} mediates insulin-induced glucose transport in primary cultures of rat skeletal muscle. Mol Endocrinol 13: 2002–2012, 1999.[Abstract/Free Full Text]
  9. Burdon RH. Superoxide and hydrogen peroxide in relation to mammalian cell proliferation. Free Radic Biol Med 18: 775–794, 1995.[ISI][Medline]
  10. Ceriello A. Oxidative stress and glycemic regulation. Metab Clin Exper 49: 27–29, 2000.[ISI][Medline]
  11. Ceriello A, dello Russo P, Amstad P, and Cerutti P. High glucose induces antioxidant enzymes in human endothelial cells in culture. Evidence linking hyperglycemia and oxidative stress. Diabetes 45: 471–477, 1996.[Abstract]
  12. Chen N, Ma W, Huang C, and Dong Z. Translocation of protein kinase Cepsilon and protein kinase C-{delta} to membrane is required for ultraviolet B-induced activation of mitogen-activated protein kinases and apoptosis. J Biol Chem 274: 15389–15394, 1999.[Abstract/Free Full Text]
  13. Czech MP, Richardson DK, Becker SG, Walters CG, Gitomer W, and Heinrich J. Insulin response in skeletal muscle and fat cells of the genetically obese Zucker rat. Metabolism 27: 1967–1981, 1978.[ISI][Medline]
  14. Di Girolamo M, Mendlinger S, and Fertig J. A simple method to determine fat cell size and number in four mammalian species. Am J Physiol 221: 850–858, 1971.[Free Full Text]
  15. Edelstein D, Dimmeler S, Ju Q, Sui C, and Brownlee M. Biochemistry and molecular cell biology of diabetic complications. J Clin Invest 108: 1341–1348, 2001.[ISI][Medline]
  16. Edelstein D, Du XL, Kaneda Y, Guzman M, and Brownlee M. Hyperglycemia-induced mitochondrial superoxide overproduction activates the hexosamine pathway and induces plasminogen activator inhibitor-1 expression by increasing Sp1 glycosylation. J Biol Chem 276: 25096–250100, 2001.[Abstract/Free Full Text]
  17. Esposito K, Nappo F, Marfella R, Giugliano G, Giugliano F, Ciotola M, Quagliaro L, Ceriello A, and Giugliano D. Inflammatory cytokine concentrations are acutely increased by hyperglycemia in humans: role of oxidative stress. Circulation 106: 2067–2072, 2002.[Abstract/Free Full Text]
  18. Fiala E, Westendorf J, and West IC. Radicals and oxidative stress in diabetes. Toxicology 146: 83–92, 2000.[ISI][Medline]
  19. Frank GD, Saito S, Motley ED, Sasaki T, Ohba M, Kuroki T, Inagami T, and Eguchi S. Requirement of Ca(2+) and PKC-{delta} for Janus kinase 2 activation by angiotensin II: involvement of PYK2. Mol Endocrinol 16: 367–377, 2002.[Abstract/Free Full Text]
  20. Glosli H, Tronstad KJ, Wergedal H, Muller F, Svardal A, Aukrust P, Berge RK, and Prydz H. Human TNF-alpha in transgenic mice induces differential changes in redox status and glutathione-regulating enzymes. FASEB J 16: 1450–1452, 2002.[Abstract/Free Full Text]
  21. Grunberger G, Taylor SI, Dons RF, and Gorden P. Insulin receptors in normal and disease states. Clin Endocrinol Metab 12: 191–219, 1983.[ISI][Medline]
  22. Ha H and Lee HB. Reactive oxygen species as glucose signaling molecules in mesangial cells cultured and under high glucose. Kidney Int 77, Suppl: S19–S25, 2000.
  23. Hardwick JS and Sefton BM. Activation of the Lck tyrosine protein kinase by hydrogen peroxide requires the phosphorylation of Tyr-394. Proc Natl Acad Sci USA 92: 4527–4531, 1995.[Abstract/Free Full Text]
  24. Heilig CW, Liu Y, England RL, Freytag SO, Gilbert JD, Heilig KO, Zhu M, Concepcion LA, and Brosius FC III. D-glucose stimulates mesangial cell GLUT1 expression and basal and IGF-I-sensitive glucose uptake in rat mesangial cells: implications for diabetic nephropathy. Diabetes 46: 1030–1039, 1997.[Abstract]
  25. Igarashi M, Wakasaki H, Takahara N, Ishii H, Jiang ZY, Yamauchi T, Kuboki K, Meier M, Rhodes CJ, and King GL. Glucose or diabetes activates p38 mitogen-activated protein kinase via different pathways. J Clin Invest 103: 185–195, 1999.[ISI][Medline]
  26. Inoguchi T, Li P, Umeda F, Yu HY, Kakimoto M, Imamura M, Aoki T, Etoh T, Hashimoto T, Naruse M, Sano H, Utsumi H, and Nawata H. High glucose level and free fatty acid stimulate reactive oxygen species production through protein kinase C-dependent activation of NAD(P)H oxidase in cultured vascular cells. Diabetes 49: 1939–1945, 2000.[Abstract]
  27. Kaplan SA. The insulin receptor. J Pediatr 104: 327–336, 1984.[ISI][Medline]
  28. Kapor-Drezgic J, Zhou X, Babazono T, Dlugosz JA, Hohman T, and Whiteside C. Effect of high glucose on mesangial cell protein kinase C-delta and -epsilon is polyol pathway-dependent. J Am Soc Nephrol 10: 1193–1203, 1999.[Abstract/Free Full Text]
  29. Kashiwagi A, Shinozaki K, Nishio Y, Maegawa H, Maeno Y, Kanazawa A, Kojima H, Haneda M, Hidaka H, Yasuda H, and Kikkawa R. Endothelium-specific activation of NAD(P)H oxidase in aortas of exogenously hyperinsulinemic rats. Am J Physiol Endocrinol Metab 277: E976–E983, 1999.[Abstract/Free Full Text]
  30. Konishi H, Tanaka M, Takemura Y, Matsuzaki H, Ono Y, Kikkawa U, and Nishizuka Y. Activation of protein kinase C by tyrosine phosphorylation in response to H2O2. Proc Natl Acad Sci USA 94: 11233–11237, 1997.[Abstract/Free Full Text]
  31. Konishi H, Yamauchi E, Taniguchi H, Yamamoto T, Matsuzaki H, Takemura Y, Ohmae K, Kikkawa U, and Nishizuka Y. Phosphorylation sites of protein kinase C-{delta} in H2O2-treated cells and its activation by tyrosine kinase in vitro. Proc Natl Acad Sci USA 98: 6587–6592, 2001.[Abstract/Free Full Text]
  32. Kozlovsky N, Rudich A, Potashnik R, and Bashan N. Reactive oxygen species activate glucose transport in L6 myotubes. Free Radic Biol Med 23: 859–869, 1997.[ISI][Medline]
  33. Lawrence, JC Jr, Guinovart JJ, and Larner J. Activation of rat adipocyte glycogen synthase by insulins. J Biol Chem 252: 444–450, 1977.[Abstract/Free Full Text]
  34. Laybutt DR, Thompson AL, Cooney GJ, and Kraegen EW. Selective chronic regulation of GLUT1 and GLUT4 content by insulin, glucose, and lipid in rat cardiac muscle in vivo. Am J Physiol Heart Circ Physiol 273: H1309–H1316, 1997.[Abstract/Free Full Text]
  35. Li W, Zhang J, Bottaro DP, and Pierce JH. Identification of serine 643 of protein kinase C-delta as an important autophosphorylation site for its enzymatic activity. J Biol Chem 272: 24550–24555, 1997.[Abstract/Free Full Text]
  36. Lo YY, Wong JM, and Cruz TF. Reactive oxygen species mediate cytokine activation of c-Jun NH2-terminal kinases. J Biol Chem 271: 15703–15707, 1996.[Abstract/Free Full Text]
  37. Lu B, Ennis D, Lai R, Bogdanovic E, Nikolov R, Salamon L, Fantus C, Le-Tien H, and Fantus IG. Enhanced sensitivity of insulin-resistant adipocytes to vanadate is associated with oxidative stress and decreased reduction of vanadate (+5) to vanadyl (+4). J Biol Chem 276: 35589–35598, 2001.[Abstract/Free Full Text]
  38. Ludwig S, Muller-Wieland D, Goldstein BJ, and Kahn CR. The insulin receptor gene and its expression in insulin-resistant mice. Endocrinology 123: 594–600, 1988.[Abstract]
  39. Marfella R, Quagliaro L, Nappo F, Ceriello A, and Giugliano D. Acute hyperglycemia induces an oxidative stress in healthy subjects. J Clin Invest 108: 635–636, 2001.[ISI][Medline]
  40. McGowan KM, Police S, Winslow JB, and Pekala PH. Tumor necrosis factor-alpha regulation of glucose transporter (GLUT1) mRNA turnover. Contribution of the 3'-untranslated region of the GLUT1 message. J Biol Chem 272: 1331–1337, 1997.[Abstract/Free Full Text]
  41. Mohazzab KM, Kaminski PM, and Wolin MS. NADH oxidoreductase is a major source of superoxide anion in bovine coronary artery endothelium. Am J Physiol Heart Circ Physiol 266: H2568–H2572, 1994.[Abstract/Free Full Text]
  42. Mullarkey CJ, Edelstein D, and Brownlee M. Free radical generation by early glycation products: a mechanism for accelerated atherogenesis in diabetes. Biochem Biophys Res Commun 173: 932–939, 1990.[ISI][Medline]
  43. Nishimura H and Simpson IA. Staurosporine inhibits phorbol 12-myristate 13-acetate- and insulin-stimulated translocation of GLUT1 and GLUT4 glucose transporters in rat adipose cells. Biochem J 302: 271–277, 1994.[Medline]
  44. Paolisso G, Gambardella A, Tagliamonte MR, Saccomanno F, Salvatore T, Gualdiero P, D'Onofrio MV, and Howard BV. Does free fatty acid infusion impair insulin action also through an increase in oxidative stress? J Clin Endocrinol Metab 81: 4244–4248, 1996.[Abstract]
  45. Park SH, Choi HJ, Lee JH, Woo CH, Kim JH, and Han HJ. High glucose inhibits renal proximal tubule cell proliferation and involves PKC, oxidative stress, and TGF-beta 1. Kidney Int 59: 1695–1705, 2001.[ISI][Medline]
  46. Pedersen O, Kahn CR, and Kahn BB. Divergent regulation of the Glut 1 and Glut 4 glucose transporters in isolated adipocytes from Zucker rats. J Clin Invest 89: 1964–1973, 1992.[ISI][Medline]
  47. Peiro C, Lafuente N, Matesanz N, Cercas E, Llergo JL, Vallejo S, Rodriguez-Manas L, and Sanchez-Ferrer CF. High glucose induces cell death of cultured human aortic smooth muscle cells through the formation of hydrogen peroxide. Br J Pharmacol 133: 967–974, 2001.[ISI][Medline]
  48. Pollet RJ. Insulin receptors and action in clinical disorders of carbohydrate tolerance. Am J Med 75: 15–22, 1983.[ISI][Medline]
  49. Qin S, Kurosaki T, and Yamamura H. Differential regulation of oxidative and osmotic stress induced Syk activation by both autophosphorylation and SH2 domains. Biochemistry 37: 5481–5486, 1998.[Medline]
  50. Rondinone CM, Carvalho E, Wesslau C, and Smith UP. Impaired glucose transport and protein kinase B activation by insulin, but not okadaic acid, in adipocytes from subjects with type II diabetes mellitus. Diabetologia 42: 819–825, 1999.[ISI][Medline]
  51. Rosenzweig T, Braiman L, Bak A, Alt A, Kuroki T, and Sampson SR. Differential effects of tumor necrosis factor-alpha on protein kinase C isoforms alpha and delta mediate inhibition of insulin receptor signaling. Diabetes 51: 1921–1930, 2002.[Abstract/Free Full Text]
  52. Rudich A, Tirosh A, Potashnik R, Hemi R, Kanety H, and Bashan N. Prolonged oxidative stress impairs insulin-induced GLUT4 translocation in 3T3-L1 adipocytes. Diabetes 47: 1562–1569, 1998.[Abstract]
  53. Saito S, Frank GD, Mifune M, Ohba M, Utsunomiya H, Motley ED, Inagami T, and Eguchi S. Ligand-independent trans-activation of the platelet-derived growth factor receptor by reactive oxygen species requires protein kinase C-{delta} and c-Src. J Biol Chem 277: 44695–44700, 2002.[Abstract/Free Full Text]
  54. Sakai K, Matsumoto K, Nishikawa T, Suefuji M, Nakamaru K, Hirashima Y, Kawashima J, Shirotani T, Ichinose K, Brownlee M, and Araki E. Mitochondrial reactive oxygen species reduce insulin secretion by pancreatic beta-cells. Biochem Biophys Res Commun 300: 216–222, 2003.[ISI][Medline]
  55. Schieven GL, Kirihara JM, Burg DL, Geahlen RL, and Ledbetter JA. p72syk Tyrosine kinase is activated by oxidizing conditions that induce lymphocyte tyrosine phosphorylation and Ca2+ signals. J Biol Chem 268: 16688–16692, 1993.[Abstract/Free Full Text]
  56. Sharpe PC, Yue KK, Catherwood MA, McMaster D, and Trimble ER. The effects of glucose-induced oxidative stress on growth and extracellular matrix gene expression of vascular smooth muscle cells. Diabetologia 41: 1210–1219, 1998.[ISI][Medline]
  57. Standaert ML, Buckley DJ, Ishizuka T, Hoffman JM, Cooper DR, Pollet RJ, and Farese RV. Protein kinase C inhibitors block insulin and PMA-stimulated hexose transport in isolated rat adipocytes and BC3H-1 myocytes. Metabolism 39: 1170–1179, 1990.[ISI][Medline]
  58. Sun X, Majumder P, Shioya H, Wu F, Kumar S, Weichselbaum R, Kharbanda S, and Kufe D. Activation of the cytoplasmic c-Abl tyrosine kinase by reactive oxygen species. J Biol Chem 275: 17237–17240, 2000.[Abstract/Free Full Text]
  59. Sundaresan M, Yu ZX, Ferrans VJ, Irani K, and Finkel T. Requirement for generation of H2O2 for platelet derived growth factor signal transduction. Science 270: 296–299, 1995.[Abstract/Free Full Text]
  60. Surwit RS, Kuhn CM, Cochrane C, McCubbin JA, and Feinglos MN. Diet-induced type II diabetes in C57BL/6J mice. Diabetes 37: 1163–1167, 1988.[Abstract]
  61. Tsuru M, Katagiri H, Asano T, Yamada T, Ohno S, Ogihara T, and Oka Y. Role of PKC isoforms in glucose transport in 3T3-L1 adipocytes: insignificance of atypical PKC. Am J Physiol Endocrinol Metab 283: E338–E345, 2002.[Abstract/Free Full Text]
  62. Warnholtz A, Nickenig G, Schulz E, Macharzina R, Brasen JH, Skatchkov M, Heitzer T, Stasch JP, Griendling KK, Harrison DG, Bohm M, Meinertz T, and Munzel T. Increased NADH-oxidase-mediated superoxide production in the early stages of atherosclerosis: evidence for involvement of the renin-angiotensin system. Circulation 99: 2027–2033, 1999.[Abstract/Free Full Text]
  63. West IC. Radicals and oxidative stress in diabetes. Diabet Med 17: 171–180, 2000.[ISI][Medline]
  64. Xu L and Badr MZ. Enhanced potential for oxidative stress in hyperinsulinemic rats: imbalance between hepatic peroxisomal hydrogen peroxide production and decomposition due to hyperinsulinemia. Horm Metab Res 31: 278–282, 1999.[ISI][Medline]
  65. Zick Y. The insulin receptor structure and function. Crit Rev Biochem Mol Biol 24: 217–269, 1989.[ISI][Medline]



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