Michal Armoni - Academia.edu (original) (raw)
Papers by Michal Armoni
Journal of Biological Chemistry, Aug 15, 2003
The synthetic thiazolidinedione ligands of peroxisome proliferator-activated receptor-␥ (PPAR␥) i... more The synthetic thiazolidinedione ligands of peroxisome proliferator-activated receptor-␥ (PPAR␥) improve insulin sensitivity in type II diabetes and induce GLUT4 mRNA expression in fat and muscle. However, the molecular mechanisms involved are still unclear. We studied the regulatory effects of PPAR␥ and its ligands on GLUT4 gene expression in primary rat adipocytes and CHO-K1 cells cotransfected with PPAR␥ and the GLUT4 promoter reporter. PPAR␥1 and PPAR␥2 repressed the activity of the GLUT4 promoter in a dose-dependent manner. Whereas this repression was augmented by the natural ligand 15⌬-prostaglandin J 2 , it was completely alleviated by rosiglitazone (Rg). Ligand binding-defective mutants PPAR␥1-L468A/E471A and PPAR␥2-L496A/ E499A retained the repression effect, which was unaffected by Rg, whereas the PPAR␥2-S112A mutant exhibited a 50% reduced capacity to repress GLUT4 promoter activity. The ؊66/؉163 bp GLUT4 promoter region was sufficient to mediate PPAR␥ inhibitory effects. The PPAR␥/retinoid X receptor-␣ heterodimer directly bound to this region, whereas binding was abolished in the presence of Rg. Thus, we show that PPAR␥ represses transcriptional activity of the GLUT4 promoter via direct and specific binding of PPAR␥/retinoid X receptor-␣ to the GLUT4 promoter. This effect requires an intact Ser 112 phosphorylation site on PPAR␥ and is completely alleviated by Rg, acting via its ligand-binding domain. These data suggest a novel mechanism by which Rg exerts its antidiabetic effects via detaching PPAR␥ from the GLUT4 gene promoter, thus leading to increased GLUT4 expression and enhanced insulin sensitivity.
Journal of Clinical Investigation, 1986
We examined insulin's effects on glucose transport and on subcellular transporter distribution in... more We examined insulin's effects on glucose transport and on subcellular transporter distribution in isolated human omental adipocytes of various sizes. Insulin stimulated 3-0-methylglucose transport by twofold in small cells, while a smaller and insignificant effect was measured in large cells. In the small cells, basal concentrations of glucose transporters were 2.9 and 17.2 pmol/mg membrane protein in the plasma and the low density microsomal membranes, respectively. Increasing cell size was associated with a 50% decrease in the concentration of transporters in each fraction, with no change in their total number per cell. Insulin stimulated the translocation of transporters from the intracellular pool to the plasma membranes, irrespective of cell size. Thus, insulin resistance at the postreceptor level, observed in human obesity, may be associated with a relative depletion of total transporters per cell together with a reduction in their intrinsic activity at the plasma membrane level.
Diabetologia, 2006
Aims/hypothesis We examined a clinical model of ex vivo transdifferentiation of primary adult hep... more Aims/hypothesis We examined a clinical model of ex vivo transdifferentiation of primary adult hepatocytes to insulinsecreting cells for the treatment of type 1 diabetes. Materials and methods Isolated rat hepatocytes were transduced in primary culture with a human lentivirus containing pancreatic duodenal homeobox 1 (PDX1, now known as insulin promoter factor 1, homeodomain transcription factor [IPF1]). Insulin expression and secretion of the newly engineered cells were assessed in vitro by RT-PCR, in situ hybridisation, immunostaining and radioimmunoassay. PDX1-transduced hepatocytes were further studied in vivo by injecting them under the renal capsule of diabetic SCID mice. Results Isolated rat hepatocytes were efficiently transduced with the lentiviral vector, as assessed by green fluorescent reporter gene expression. The transduced cells exhibited insulin at both mRNA (RT-PCR, in situ hybridisation) and protein levels (immunostaining and radioimmunoassay). Moreover, insulin secretion by the engineered cells was dependent on glucose and sulfonylurea. Other beta cell genes, including those encoding solute carrier family 2 (facilitated glucose transporter), member 2 (Slc2a2), glucokinase (Gck), ATP-binding cassette, sub-family C (CFTR/ MRP), member 8 (Abcc8), the potassium inwardly-rectifying channel, subfamily J, member 11 (Kcnj11) and proprotein convertase subtilisin/kexin type 1 (Pcsk1) were also expressed. The PDX1-transduced hepatocytes expressed several pancreatic transcription factors related to early pancreatic endocrine development (endogenous Pdx1, neurogenic differentiation factor 1 [Neurod1], and NK6 transcription factor related, locus 1 [Nkx6-1]) as well as the late-stage pancreatic transcription factors (paired box gene 4 [Pax4], paired box gene 6 [Pax6], and v-maf musculoaponeurotic fibrosarcoma oncogene homolog A [Mafa]). Transplantation of 3×10 6 transdifferentiated liver cells under the renal capsule of seven streptozotocin-induced diabetic SCID mice resulted in significant reduction of nonfasting blood glucose levels from 30.7 ± 1.3 to 8.7 ± 3.7 mmol/l (mean ± SEM, p=0.01), in 6 to 8 weeks. Removal of the graft resulted in severe hyperglycaemia. Conclusions/interpretation Ex vivo lentiviral-mediated PDX1 expression in isolated adult liver cells represents a potential model for type 1 diabetes mellitus therapy.
American Journal of Physiology-Endocrinology and Metabolism, 2011
Conventional (whole body) CYP2E1 knockout mice displayed protection against high-fat diet-induced... more Conventional (whole body) CYP2E1 knockout mice displayed protection against high-fat diet-induced weight gain, obesity, and hyperlipidemia with increased energy expenditure despite normal food intake and spontaneous locomotor activity. In addition, the CYP2E1 knockout mice displayed a marked improvement in glucose tolerance on both normal chow and high-fat diets. Euglycemic-hyperinsulinemic clamps demonstrated a marked protection against high-fat diet-induced insulin resistance in CYP2E1 knockout mice, with enhanced adipose tissue glucose uptake and insulin suppression of hepatic glucose output. In parallel, adipose tissue was protected against high-fat diet-induced proinflammatory cytokine production. Taken together, these data demonstrate that the CYP2E1 deletion protects mice against high-fat diet-induced insulin resistance with improved glucose homeostasis in vivo.
Hormone and metabolic research = Hormon- und Stoffwechselforschung = Hormones et métabolisme, 2015
AHNAK is a 700 KD phosphoprotein primarily involved in calcium signaling in various cell types an... more AHNAK is a 700 KD phosphoprotein primarily involved in calcium signaling in various cell types and regulating cytoskeletal organization and cell membrane architecture. AHNAK expression has also been associated with obesity. To investigate the role of AHNAK in regulating metabolic homeostasis, we studied whole body AHNAK knockout mice (KO) on either regular chow or high-fat diet (HFD). KO mice had a leaner phenotype and were resistant to high-fat diet-induced obesity (DIO), as reflected by a reduction in adipose tissue mass in conjunction with higher lean mass compared to wild-type controls (WT). However, KO mice exhibited higher fasting glucose levels, impaired glucose tolerance, and diminished serum insulin levels on either diet. Concomitantly, KO mice on HFD displayed defects in insulin signaling, as evident from reduced Akt phosphorylation and decreased cellular glucose transporter (Glut4) levels. Glucose intolerance and insulin resistance were also associated with changes in exp...
The Journal of Clinical Endocrinology & Metabolism, 2002
Current Medicinal Chemistry-Immunology, Endocrine & Metabolic Agents, 2005
The overall objective of this paper is to review the mechanisms by which various metabolic and ce... more The overall objective of this paper is to review the mechanisms by which various metabolic and cellular signals, as well as nuclear transcription factors, regulate the expression and function of the insulin responsive glucose transporter-4 (GLUT4) gene. Reviewing this information will help the reader to understand the molecular processes involved in both glucose homeostasis and the pathogenesis of abnormal metabolic states involving impaired insulin action, such as insulin resistance and diabetes mellitus type 2 (DM2). The same molecular mechanisms are also involved in tumorigenesis. Studies on GLUT4 regulation, its translocation machinery, and intrinsic activity have contributed valuable knowledge that may be useful for developing therapeutic strategies aimed at increasing GLUT4 protein levels, which could potentially improve glucose homeostasis in insulin resistance and diabetes.
Hormone and metabolic research = Hormon- und Stoffwechselforschung = Hormones et métabolisme, 2015
Being a key-factor in glucose homeostasis, PPARγ transcriptional activity (TA) is of high importa... more Being a key-factor in glucose homeostasis, PPARγ transcriptional activity (TA) is of high importance. However, its mediation by ligands and post-translational modifications in insulin target tissues are unclear. We investigated effects of rosiglitazone (Rg) and sumoylation on PPARγ-TA by overexpressing expression vectors and promoter-reporters for PPARγ1 and PPARγ2 in primary rat adipocytes. Wild type (WT) PPARγ1 and PPARγ2 dose-dependently repressed transcription from their promoters to a maximum of 40-50%. PPARγ2 mutants defective in either MAP-kinase phosphorylation (S112A) or the ligand-binding domain (LBD; P495L, L496A/E499A) exhibited decreased repression of PPARγ2 promoter. Rg enhanced repression by S112A, but not by LBD-defective mutants. Sumoylation-defective PPARγ1 mutants K77R and K365R repressed PPARγ2 promoter activity similar to WT, while Rg enhanced repression by K77R but not by K365R. Sumoylation-defective PPARγ2 mutants K107R and K395R exhibited impaired TA and impa...
Physiology & Behavior, 1994
Models studying in vivo insulin animal action usually employ single-use anesthetized animals, mai... more Models studying in vivo insulin animal action usually employ single-use anesthetized animals, mainly for technical reasons. We developed a modification of the euglycemic insulin clamp technique and used it to repeatedly assess in vivo insulin effects in awake streptozotocin-induced diabetic rats, and in weight- and/or age-matched controls. Permanent catheters implanted into the left carotid artery and the right jugular vein were used for miniature blood sampling (20 microliters) and recycling. Insulin was infused at 1, 2, 3, 15, and 30 mU/kg.min. Plasma insulin and C-peptide levels and glucose utilization rate were measured at blood glucose levels of 100 mg/dl. Diabetes was associated with diminished elevation of plasma C-peptide and insulin levels after ad lib feeding, 50% decreased (p < 0.005) insulin sensitivity, 31% decreased (p < 0.001) insulin responsiveness, and unchanged insulin clearance rates. Thus, using repeated clamps of the same rat over a prolonged period of time, we demonstrate that diabetes is associated with unchanged clearance but decreased sensitivity and responsiveness to insulin.
Journal of Biological Chemistry, 2005
Hyperlipidemia (HL) impairs cardiac glucose homeostasis, but the molecular mechanisms involved ar... more Hyperlipidemia (HL) impairs cardiac glucose homeostasis, but the molecular mechanisms involved are yet unclear. We examined HL-regulated GLUT4 and peroxisome proliferator-activated receptor (PPAR) ␥ gene expression in human cardiac muscle. Compared with control patients, GLUT4 protein levels were 30% lower in human cardiac muscle biopsies from patients with HL and/or type 2 diabetes mellitus, whereas GLUT4 mRNA levels were unchanged. PPAR␥ mRNA levels were 30-50% lower in patients with HL and/or diabetes mellitus type 2 than in controls. Reporter studies in H9C2 cardiomyotubes showed that HL in vitro, induced by high levels of arachidonic (AA) stearic, linoleic, and oleic acids (24 h, 200 M) repressed transcription from the GLUT4 promoter; AA also repressed transcription from the PPAR␥1 and PPAR␥2 promoters. Co-expression of PPAR␥2 repressed GLUT4 promoter activity, and the addition of AA further enhanced this effect. 5-Deletion analysis revealed three GLUT4 promoter regions that accounted for AAmediated effects: two repression-mediating sequences at ؊443/؊423 bp and ؊222/؊197 bp, the deletion of either or both of which led to a partial derepression of promoter activity, and a third derepression-mediating sequence at ؊612/؊587 bp that was required for sustaining this derepression effect. Electromobility shift assay further shows that AA enhanced binding to two of the three regions of cardiac nuclear protein(s), the nature of which is still unknown. We propose that HL, exhibited as a high free fatty acid level, modulates GLUT4 gene expression in cardiac muscle via a complex mechanism that includes: (a) binding of AA mediator proteins to three newly identified response elements on the GLUT4 promoter gene and (b) repression of GLUT4 and the PPAR␥ genes by AA.
Journal of Biological Chemistry, 2006
FOXO1 and peroxisome proliferator-activated receptor-␥ (PPAR␥) are crucial transcription factors ... more FOXO1 and peroxisome proliferator-activated receptor-␥ (PPAR␥) are crucial transcription factors that regulate glucose metabolism and insulin responsiveness in insulin target tissues. We have shown that, in primary rat adipocytes, both factors regulate transcription of the insulin-responsive GLUT4 gene and that PPAR␥2 detachment from the GLUT4 promoter upon thiazolidinedione binding up-regulates GLUT4 gene expression, thus increasing insulin sensitivity (
Journal of Biological Chemistry, 1999
We report that the vanadium ligand L-Glu(␥)HXM potentiates the capacity of free vanadium ions to ... more We report that the vanadium ligand L-Glu(␥)HXM potentiates the capacity of free vanadium ions to activate glucose uptake and glucose metabolism in rat adipocytes in vitro (by 4-5-fold) and to lower blood glucose levels in hyperglycemic rats in vivo (by 5-7-fold). A molar ratio of two L-Glu(␥)HXM molecules to one vanadium ion was most effective. Unlike other vanadium ligands that potentiate the insulinomimetic actions of vanadium, L-Glu(␥)HXM partially activated lipogenesis in rat adipocytes in the absence of exogenous vanadium. This effect was not manifested by D-Glu(␥)HXM. At 10-20 M L-Glu(␥)HXM, lipogenesis was activated 9-21%. This effect was approximately 9-fold higher (140 ؎ 15% of maximal insulin response) in adipocytes derived from rats that had been treated with vanadium for several days. Titration of vanadium(IV) with L-Glu(␥)HXM led to a rapid decrease in the absorbance of vanadium(IV) at 765 nm, and 51 V NMR spectroscopy revealed that the chemical shift of vanadium(IV) at ؊490 ppm disappeared with the appearance of a signal characteristic to vanadium(V) (؊530 ppm) upon adding one equivalent of L-Glu(␥)HXM. In summary, L-Glu(␥)HXM is highly active in potentiating vanadium-activated glucose metabolism in vitro and in vivo and facilitating glucose metabolism in rat adipocytes in the absence of exogenous vanadium probably through conversion of trace intracellular vanadium into an active insulinomimetic compound. We propose that the active species is either a 1:1 or 2:1 L-Glu(␥)HXM vanadium complex in which the endogenous vanadium(IV) has been altered to vanadium(V). Finally we demonstrate that L-Glu(␥)HXM-and L-Glu(␥)HXM⅐vanadium-evoked lipogenesis is arrested by wortmannin and that activation of glucose uptake in rat adipocytes is because of enhanced translocation of GLUT4 from low density microsomes to the plasma membrane.
Diabetes, 1987
To determine the role of insulin in reversing the insulin resistance associated with depletion of... more To determine the role of insulin in reversing the insulin resistance associated with depletion of the intracellular pool of glucose transporters, streptozocin-induced diabetic rats were treated with 5 U/day s.c. of insulin for 0, 8, or 14 days. At each time point, adipose cells were isolated, and 3-O-methylglucose transport was measured in the absence and presence of 1000 |xU/ml insulin. With the cytochalasin B-binding assay, concentrations of glucose transporters in the plasma and the low-density microsomal membrane fractions were determined. Eight-day insulin therapy enhanced glucose transport rate (mean ± SE) from 0.2 ± 0.0 to 1.1 ± 0.1 fmol • cell' 1 • min~1 in the basal state and from 0.8 ± 0.1 to 5.5 ± 0.4 fmol • c e l l 1-min 1 in the insulin-stimulated state in untreated and treated diabetic rats, respectively; this is a 3-fold increment of glucose transport rate in both states compared with control rats. After 14-day insulin therapy, glucosetransport activity declined toward normal but still remained ~1.5-and 4-fold higher than control and diabetic rats, respectively. Despite the persistent enhancement of glucose transport rate, concentration of glucose transporters in the intracellular pool was restored only to its prediabetic state. Likewise, the increased concentration of glucose transporters in the plasma membranes after insulin stimulation was similar to that of control rats. Thus, we suggest that 8-14 days of insulin therapy reversed the insulin resistance in diabetic rat adipocytes by at least two mechanisms: restoration of the intracellular pool of glucose transporters and enhancement of glucosetransport activity. The mechanism(s) responsible for this supernormal glucose-transport activity is unknown but may be related to transient appearance
Cancer Research, 2004
Tumorigenesis is associated with enhanced cellular glucose uptake and increased metabolism. Becau... more Tumorigenesis is associated with enhanced cellular glucose uptake and increased metabolism. Because the p53 tumor suppressor is mutated in a large number of cancers, we evaluated whether p53 regulates expression of the GLUT1 and GLUT4 glucose transporter genes. Transient cotransfection of osteosarcoma-derived SaOS-2 cells, rhabdomyosarcoma-derived RD cells, and C2C12 myotubes with GLUT1-P-Luc or GLUT4-P-Luc promoter-reporter constructs and wild-type p53 expression vectors dose dependently decreased both GLUT1 and GLUT4 promoter activity to approximately 50% of their basal levels. PG13-Luc activity, which was used as a positive control for functional p53 expression, was increased up to ∼250-fold by coexpression of wild-type p53. The inhibitory effect of wild-type p53 was greatly reduced or abolished when cells were transfected with p53 with mutations in amino acids 143, 248, or 273. A region spanning −66/+163 bp of the GLUT4 promoter was both necessary and sufficient to mediate the i...
American Journal of Physiology-Endocrinology and Metabolism, 2008
The insulin-responsive glucose transporter 4 (GLUT4) plays a key role in glucose uptake and metab... more The insulin-responsive glucose transporter 4 (GLUT4) plays a key role in glucose uptake and metabolism in insulin target tissues. Being a rate-limiting step in glucose metabolism, the expression and function of the GLUT4 isoform has been extensively studied and found to be tightly regulated at both mRNA and protein levels. Adaptation to states of enhanced metabolic demand is associated with increased glucose metabolism and GLUT4 gene expression, whereas states of insulin resistance such as type 2 diabetes mellitus (DM2), obesity, and aging are associated with impaired regulation of GLUT4 gene expression and function. The present review focuses on the interplay among hormonal, nutritional, and transcription factors in the regulation of GLUT4 transcription in health and sickness.
Trends in Endocrinology & Metabolism, 2007
Journal of Biological Chemistry, Aug 15, 2003
The synthetic thiazolidinedione ligands of peroxisome proliferator-activated receptor-␥ (PPAR␥) i... more The synthetic thiazolidinedione ligands of peroxisome proliferator-activated receptor-␥ (PPAR␥) improve insulin sensitivity in type II diabetes and induce GLUT4 mRNA expression in fat and muscle. However, the molecular mechanisms involved are still unclear. We studied the regulatory effects of PPAR␥ and its ligands on GLUT4 gene expression in primary rat adipocytes and CHO-K1 cells cotransfected with PPAR␥ and the GLUT4 promoter reporter. PPAR␥1 and PPAR␥2 repressed the activity of the GLUT4 promoter in a dose-dependent manner. Whereas this repression was augmented by the natural ligand 15⌬-prostaglandin J 2 , it was completely alleviated by rosiglitazone (Rg). Ligand binding-defective mutants PPAR␥1-L468A/E471A and PPAR␥2-L496A/ E499A retained the repression effect, which was unaffected by Rg, whereas the PPAR␥2-S112A mutant exhibited a 50% reduced capacity to repress GLUT4 promoter activity. The ؊66/؉163 bp GLUT4 promoter region was sufficient to mediate PPAR␥ inhibitory effects. The PPAR␥/retinoid X receptor-␣ heterodimer directly bound to this region, whereas binding was abolished in the presence of Rg. Thus, we show that PPAR␥ represses transcriptional activity of the GLUT4 promoter via direct and specific binding of PPAR␥/retinoid X receptor-␣ to the GLUT4 promoter. This effect requires an intact Ser 112 phosphorylation site on PPAR␥ and is completely alleviated by Rg, acting via its ligand-binding domain. These data suggest a novel mechanism by which Rg exerts its antidiabetic effects via detaching PPAR␥ from the GLUT4 gene promoter, thus leading to increased GLUT4 expression and enhanced insulin sensitivity.
Journal of Clinical Investigation, 1986
We examined insulin's effects on glucose transport and on subcellular transporter distribution in... more We examined insulin's effects on glucose transport and on subcellular transporter distribution in isolated human omental adipocytes of various sizes. Insulin stimulated 3-0-methylglucose transport by twofold in small cells, while a smaller and insignificant effect was measured in large cells. In the small cells, basal concentrations of glucose transporters were 2.9 and 17.2 pmol/mg membrane protein in the plasma and the low density microsomal membranes, respectively. Increasing cell size was associated with a 50% decrease in the concentration of transporters in each fraction, with no change in their total number per cell. Insulin stimulated the translocation of transporters from the intracellular pool to the plasma membranes, irrespective of cell size. Thus, insulin resistance at the postreceptor level, observed in human obesity, may be associated with a relative depletion of total transporters per cell together with a reduction in their intrinsic activity at the plasma membrane level.
Diabetologia, 2006
Aims/hypothesis We examined a clinical model of ex vivo transdifferentiation of primary adult hep... more Aims/hypothesis We examined a clinical model of ex vivo transdifferentiation of primary adult hepatocytes to insulinsecreting cells for the treatment of type 1 diabetes. Materials and methods Isolated rat hepatocytes were transduced in primary culture with a human lentivirus containing pancreatic duodenal homeobox 1 (PDX1, now known as insulin promoter factor 1, homeodomain transcription factor [IPF1]). Insulin expression and secretion of the newly engineered cells were assessed in vitro by RT-PCR, in situ hybridisation, immunostaining and radioimmunoassay. PDX1-transduced hepatocytes were further studied in vivo by injecting them under the renal capsule of diabetic SCID mice. Results Isolated rat hepatocytes were efficiently transduced with the lentiviral vector, as assessed by green fluorescent reporter gene expression. The transduced cells exhibited insulin at both mRNA (RT-PCR, in situ hybridisation) and protein levels (immunostaining and radioimmunoassay). Moreover, insulin secretion by the engineered cells was dependent on glucose and sulfonylurea. Other beta cell genes, including those encoding solute carrier family 2 (facilitated glucose transporter), member 2 (Slc2a2), glucokinase (Gck), ATP-binding cassette, sub-family C (CFTR/ MRP), member 8 (Abcc8), the potassium inwardly-rectifying channel, subfamily J, member 11 (Kcnj11) and proprotein convertase subtilisin/kexin type 1 (Pcsk1) were also expressed. The PDX1-transduced hepatocytes expressed several pancreatic transcription factors related to early pancreatic endocrine development (endogenous Pdx1, neurogenic differentiation factor 1 [Neurod1], and NK6 transcription factor related, locus 1 [Nkx6-1]) as well as the late-stage pancreatic transcription factors (paired box gene 4 [Pax4], paired box gene 6 [Pax6], and v-maf musculoaponeurotic fibrosarcoma oncogene homolog A [Mafa]). Transplantation of 3×10 6 transdifferentiated liver cells under the renal capsule of seven streptozotocin-induced diabetic SCID mice resulted in significant reduction of nonfasting blood glucose levels from 30.7 ± 1.3 to 8.7 ± 3.7 mmol/l (mean ± SEM, p=0.01), in 6 to 8 weeks. Removal of the graft resulted in severe hyperglycaemia. Conclusions/interpretation Ex vivo lentiviral-mediated PDX1 expression in isolated adult liver cells represents a potential model for type 1 diabetes mellitus therapy.
American Journal of Physiology-Endocrinology and Metabolism, 2011
Conventional (whole body) CYP2E1 knockout mice displayed protection against high-fat diet-induced... more Conventional (whole body) CYP2E1 knockout mice displayed protection against high-fat diet-induced weight gain, obesity, and hyperlipidemia with increased energy expenditure despite normal food intake and spontaneous locomotor activity. In addition, the CYP2E1 knockout mice displayed a marked improvement in glucose tolerance on both normal chow and high-fat diets. Euglycemic-hyperinsulinemic clamps demonstrated a marked protection against high-fat diet-induced insulin resistance in CYP2E1 knockout mice, with enhanced adipose tissue glucose uptake and insulin suppression of hepatic glucose output. In parallel, adipose tissue was protected against high-fat diet-induced proinflammatory cytokine production. Taken together, these data demonstrate that the CYP2E1 deletion protects mice against high-fat diet-induced insulin resistance with improved glucose homeostasis in vivo.
Hormone and metabolic research = Hormon- und Stoffwechselforschung = Hormones et métabolisme, 2015
AHNAK is a 700 KD phosphoprotein primarily involved in calcium signaling in various cell types an... more AHNAK is a 700 KD phosphoprotein primarily involved in calcium signaling in various cell types and regulating cytoskeletal organization and cell membrane architecture. AHNAK expression has also been associated with obesity. To investigate the role of AHNAK in regulating metabolic homeostasis, we studied whole body AHNAK knockout mice (KO) on either regular chow or high-fat diet (HFD). KO mice had a leaner phenotype and were resistant to high-fat diet-induced obesity (DIO), as reflected by a reduction in adipose tissue mass in conjunction with higher lean mass compared to wild-type controls (WT). However, KO mice exhibited higher fasting glucose levels, impaired glucose tolerance, and diminished serum insulin levels on either diet. Concomitantly, KO mice on HFD displayed defects in insulin signaling, as evident from reduced Akt phosphorylation and decreased cellular glucose transporter (Glut4) levels. Glucose intolerance and insulin resistance were also associated with changes in exp...
The Journal of Clinical Endocrinology & Metabolism, 2002
Current Medicinal Chemistry-Immunology, Endocrine & Metabolic Agents, 2005
The overall objective of this paper is to review the mechanisms by which various metabolic and ce... more The overall objective of this paper is to review the mechanisms by which various metabolic and cellular signals, as well as nuclear transcription factors, regulate the expression and function of the insulin responsive glucose transporter-4 (GLUT4) gene. Reviewing this information will help the reader to understand the molecular processes involved in both glucose homeostasis and the pathogenesis of abnormal metabolic states involving impaired insulin action, such as insulin resistance and diabetes mellitus type 2 (DM2). The same molecular mechanisms are also involved in tumorigenesis. Studies on GLUT4 regulation, its translocation machinery, and intrinsic activity have contributed valuable knowledge that may be useful for developing therapeutic strategies aimed at increasing GLUT4 protein levels, which could potentially improve glucose homeostasis in insulin resistance and diabetes.
Hormone and metabolic research = Hormon- und Stoffwechselforschung = Hormones et métabolisme, 2015
Being a key-factor in glucose homeostasis, PPARγ transcriptional activity (TA) is of high importa... more Being a key-factor in glucose homeostasis, PPARγ transcriptional activity (TA) is of high importance. However, its mediation by ligands and post-translational modifications in insulin target tissues are unclear. We investigated effects of rosiglitazone (Rg) and sumoylation on PPARγ-TA by overexpressing expression vectors and promoter-reporters for PPARγ1 and PPARγ2 in primary rat adipocytes. Wild type (WT) PPARγ1 and PPARγ2 dose-dependently repressed transcription from their promoters to a maximum of 40-50%. PPARγ2 mutants defective in either MAP-kinase phosphorylation (S112A) or the ligand-binding domain (LBD; P495L, L496A/E499A) exhibited decreased repression of PPARγ2 promoter. Rg enhanced repression by S112A, but not by LBD-defective mutants. Sumoylation-defective PPARγ1 mutants K77R and K365R repressed PPARγ2 promoter activity similar to WT, while Rg enhanced repression by K77R but not by K365R. Sumoylation-defective PPARγ2 mutants K107R and K395R exhibited impaired TA and impa...
Physiology & Behavior, 1994
Models studying in vivo insulin animal action usually employ single-use anesthetized animals, mai... more Models studying in vivo insulin animal action usually employ single-use anesthetized animals, mainly for technical reasons. We developed a modification of the euglycemic insulin clamp technique and used it to repeatedly assess in vivo insulin effects in awake streptozotocin-induced diabetic rats, and in weight- and/or age-matched controls. Permanent catheters implanted into the left carotid artery and the right jugular vein were used for miniature blood sampling (20 microliters) and recycling. Insulin was infused at 1, 2, 3, 15, and 30 mU/kg.min. Plasma insulin and C-peptide levels and glucose utilization rate were measured at blood glucose levels of 100 mg/dl. Diabetes was associated with diminished elevation of plasma C-peptide and insulin levels after ad lib feeding, 50% decreased (p < 0.005) insulin sensitivity, 31% decreased (p < 0.001) insulin responsiveness, and unchanged insulin clearance rates. Thus, using repeated clamps of the same rat over a prolonged period of time, we demonstrate that diabetes is associated with unchanged clearance but decreased sensitivity and responsiveness to insulin.
Journal of Biological Chemistry, 2005
Hyperlipidemia (HL) impairs cardiac glucose homeostasis, but the molecular mechanisms involved ar... more Hyperlipidemia (HL) impairs cardiac glucose homeostasis, but the molecular mechanisms involved are yet unclear. We examined HL-regulated GLUT4 and peroxisome proliferator-activated receptor (PPAR) ␥ gene expression in human cardiac muscle. Compared with control patients, GLUT4 protein levels were 30% lower in human cardiac muscle biopsies from patients with HL and/or type 2 diabetes mellitus, whereas GLUT4 mRNA levels were unchanged. PPAR␥ mRNA levels were 30-50% lower in patients with HL and/or diabetes mellitus type 2 than in controls. Reporter studies in H9C2 cardiomyotubes showed that HL in vitro, induced by high levels of arachidonic (AA) stearic, linoleic, and oleic acids (24 h, 200 M) repressed transcription from the GLUT4 promoter; AA also repressed transcription from the PPAR␥1 and PPAR␥2 promoters. Co-expression of PPAR␥2 repressed GLUT4 promoter activity, and the addition of AA further enhanced this effect. 5-Deletion analysis revealed three GLUT4 promoter regions that accounted for AAmediated effects: two repression-mediating sequences at ؊443/؊423 bp and ؊222/؊197 bp, the deletion of either or both of which led to a partial derepression of promoter activity, and a third derepression-mediating sequence at ؊612/؊587 bp that was required for sustaining this derepression effect. Electromobility shift assay further shows that AA enhanced binding to two of the three regions of cardiac nuclear protein(s), the nature of which is still unknown. We propose that HL, exhibited as a high free fatty acid level, modulates GLUT4 gene expression in cardiac muscle via a complex mechanism that includes: (a) binding of AA mediator proteins to three newly identified response elements on the GLUT4 promoter gene and (b) repression of GLUT4 and the PPAR␥ genes by AA.
Journal of Biological Chemistry, 2006
FOXO1 and peroxisome proliferator-activated receptor-␥ (PPAR␥) are crucial transcription factors ... more FOXO1 and peroxisome proliferator-activated receptor-␥ (PPAR␥) are crucial transcription factors that regulate glucose metabolism and insulin responsiveness in insulin target tissues. We have shown that, in primary rat adipocytes, both factors regulate transcription of the insulin-responsive GLUT4 gene and that PPAR␥2 detachment from the GLUT4 promoter upon thiazolidinedione binding up-regulates GLUT4 gene expression, thus increasing insulin sensitivity (
Journal of Biological Chemistry, 1999
We report that the vanadium ligand L-Glu(␥)HXM potentiates the capacity of free vanadium ions to ... more We report that the vanadium ligand L-Glu(␥)HXM potentiates the capacity of free vanadium ions to activate glucose uptake and glucose metabolism in rat adipocytes in vitro (by 4-5-fold) and to lower blood glucose levels in hyperglycemic rats in vivo (by 5-7-fold). A molar ratio of two L-Glu(␥)HXM molecules to one vanadium ion was most effective. Unlike other vanadium ligands that potentiate the insulinomimetic actions of vanadium, L-Glu(␥)HXM partially activated lipogenesis in rat adipocytes in the absence of exogenous vanadium. This effect was not manifested by D-Glu(␥)HXM. At 10-20 M L-Glu(␥)HXM, lipogenesis was activated 9-21%. This effect was approximately 9-fold higher (140 ؎ 15% of maximal insulin response) in adipocytes derived from rats that had been treated with vanadium for several days. Titration of vanadium(IV) with L-Glu(␥)HXM led to a rapid decrease in the absorbance of vanadium(IV) at 765 nm, and 51 V NMR spectroscopy revealed that the chemical shift of vanadium(IV) at ؊490 ppm disappeared with the appearance of a signal characteristic to vanadium(V) (؊530 ppm) upon adding one equivalent of L-Glu(␥)HXM. In summary, L-Glu(␥)HXM is highly active in potentiating vanadium-activated glucose metabolism in vitro and in vivo and facilitating glucose metabolism in rat adipocytes in the absence of exogenous vanadium probably through conversion of trace intracellular vanadium into an active insulinomimetic compound. We propose that the active species is either a 1:1 or 2:1 L-Glu(␥)HXM vanadium complex in which the endogenous vanadium(IV) has been altered to vanadium(V). Finally we demonstrate that L-Glu(␥)HXM-and L-Glu(␥)HXM⅐vanadium-evoked lipogenesis is arrested by wortmannin and that activation of glucose uptake in rat adipocytes is because of enhanced translocation of GLUT4 from low density microsomes to the plasma membrane.
Diabetes, 1987
To determine the role of insulin in reversing the insulin resistance associated with depletion of... more To determine the role of insulin in reversing the insulin resistance associated with depletion of the intracellular pool of glucose transporters, streptozocin-induced diabetic rats were treated with 5 U/day s.c. of insulin for 0, 8, or 14 days. At each time point, adipose cells were isolated, and 3-O-methylglucose transport was measured in the absence and presence of 1000 |xU/ml insulin. With the cytochalasin B-binding assay, concentrations of glucose transporters in the plasma and the low-density microsomal membrane fractions were determined. Eight-day insulin therapy enhanced glucose transport rate (mean ± SE) from 0.2 ± 0.0 to 1.1 ± 0.1 fmol • cell' 1 • min~1 in the basal state and from 0.8 ± 0.1 to 5.5 ± 0.4 fmol • c e l l 1-min 1 in the insulin-stimulated state in untreated and treated diabetic rats, respectively; this is a 3-fold increment of glucose transport rate in both states compared with control rats. After 14-day insulin therapy, glucosetransport activity declined toward normal but still remained ~1.5-and 4-fold higher than control and diabetic rats, respectively. Despite the persistent enhancement of glucose transport rate, concentration of glucose transporters in the intracellular pool was restored only to its prediabetic state. Likewise, the increased concentration of glucose transporters in the plasma membranes after insulin stimulation was similar to that of control rats. Thus, we suggest that 8-14 days of insulin therapy reversed the insulin resistance in diabetic rat adipocytes by at least two mechanisms: restoration of the intracellular pool of glucose transporters and enhancement of glucosetransport activity. The mechanism(s) responsible for this supernormal glucose-transport activity is unknown but may be related to transient appearance
Cancer Research, 2004
Tumorigenesis is associated with enhanced cellular glucose uptake and increased metabolism. Becau... more Tumorigenesis is associated with enhanced cellular glucose uptake and increased metabolism. Because the p53 tumor suppressor is mutated in a large number of cancers, we evaluated whether p53 regulates expression of the GLUT1 and GLUT4 glucose transporter genes. Transient cotransfection of osteosarcoma-derived SaOS-2 cells, rhabdomyosarcoma-derived RD cells, and C2C12 myotubes with GLUT1-P-Luc or GLUT4-P-Luc promoter-reporter constructs and wild-type p53 expression vectors dose dependently decreased both GLUT1 and GLUT4 promoter activity to approximately 50% of their basal levels. PG13-Luc activity, which was used as a positive control for functional p53 expression, was increased up to ∼250-fold by coexpression of wild-type p53. The inhibitory effect of wild-type p53 was greatly reduced or abolished when cells were transfected with p53 with mutations in amino acids 143, 248, or 273. A region spanning −66/+163 bp of the GLUT4 promoter was both necessary and sufficient to mediate the i...
American Journal of Physiology-Endocrinology and Metabolism, 2008
The insulin-responsive glucose transporter 4 (GLUT4) plays a key role in glucose uptake and metab... more The insulin-responsive glucose transporter 4 (GLUT4) plays a key role in glucose uptake and metabolism in insulin target tissues. Being a rate-limiting step in glucose metabolism, the expression and function of the GLUT4 isoform has been extensively studied and found to be tightly regulated at both mRNA and protein levels. Adaptation to states of enhanced metabolic demand is associated with increased glucose metabolism and GLUT4 gene expression, whereas states of insulin resistance such as type 2 diabetes mellitus (DM2), obesity, and aging are associated with impaired regulation of GLUT4 gene expression and function. The present review focuses on the interplay among hormonal, nutritional, and transcription factors in the regulation of GLUT4 transcription in health and sickness.
Trends in Endocrinology & Metabolism, 2007