Insulin stimulates glucose uptake by recruiting glucose transporter 4 (GLUT4) from an intracellular compartment to the cell surface; this phenomenon is defective in type 2 diabetes. Here we examine the involvement of actin filaments in GLUT4 translocation and their possible defects in insulin resistance, using L6 myotubes expressing myc-tagged GLUT4. Insulin caused membrane ruffling, a dynamic distortion of the myotube dorsal surface. Fluorescence microscopy and immunogold staining of surface GLUT4myc coupled to backscatter electron microscopy revealed a high density of this protein in membrane ruffles. The t-SNAREs syntaxin4 and SNAP-23 were also abundant in these regions. Below the membrane, GLUT4 and the vesicular protein VAMP2, but not VAMP3, colocalized with the actin structures supporting the membrane ruffles. GLUT4myc externalization and membrane ruffles were reduced by jasplakinolide and by swinholide-A, drugs that affect actin filament stability and prevent actin branching, respectively. Insulin resistance generated by prolonged (24 hours) exposure of myotubes to high glucose and insulin diminished the acute insulin-dependent remodeling of cortical actin and GLUT4myc translocation, reminiscent of the effect of swinholide-A. We propose that GLUT4 vesicle incorporation into the plasma membrane involves insulin-dependent cortical actin remodeling and that defective actin remodeling contributes to insulin resistance.

SummaryCytokine-induced inflammation is involved in the pathogenesis of type 2 diabetes mellitus (DM). We investigated plasma concentrations and ex vivo production of cytokines and chemokines, and intracellular signalling molecules, mitogen-activated protein kinases (MAPK) in T helper (Th) cells and monocytes in 94 type 2 diabetic patients with or without nephropathy and 20 healthy controls. Plasma concentrations of inflammatory cytokines tumour necrosis factor (TNF)-a, interleukin (IL)-6, IL-18 and chemokine CCL2 in patients with diabetic nephropathy (DN) were significantly higher than control subjects, while IL-10, CXCL8, CXCL9, CXCL10 and adiponectin concentrations of DN were significantly higher than patients without diabetic nephropathy (NDN) and control subjects (all P < 0·05). Plasma concentrations of TNF-a, IL-6, IL-10, IL-18, CCL2, CXCL8, CXCL9, CXCL10 and adiponectin exhibited significant positive correlation with urine albumin : creatinine ratio in DN patients. The percentage increases of ex vivo production of IL-6, CXCL8, CXCL10, CCL2 and CCL5 upon TNF-a activation were significantly higher in both NDN and DN patients than controls (all P < 0·05). The percentage increases in IL-18-induced phosphorylation of extracellular signal-regulated kinase (ERK) in Th cells of NDN and DN were significantly higher than controls (P < 0·05), while the percentage increase in TNF-a-induced phosphorylation of p38 MAPK in monocytes and IL-18-induced phosphorylation of p38 MAPK in Th cells and monocytes were significantly higher in NDN patients than controls. These results confirmed that the aberrant production of inflammatory cytokines and chemokines and differential activation of MAPK in different leucocytes are the underlying immunopathological mechanisms of type 2 DM patients with DN.

Diabetes and obesity are complex diseases associated with insulin resistance and fatty liver. The latter is characterized by dysregulation of the Akt, AMP-activated protein kinase (AMPK), and IGF-I pathways and expression of microRNAs (miRNAs). In China, multicomponent traditional Chinese medicine (TCM) has been used to treat diabetes for centuries. In this study, we used a three-herb, berberine-containing TCM to treat male Zucker diabetic fatty rats. TCM showed sustained glucose-lowering effects for 1 week after a single-dose treatment. Two-week treatment attenuated insulin resistance and fatty degeneration, with hepatocyte regeneration lasting for 1 month posttreatment. These beneficial effects persisted for 1 year after 1-month treatment. Two-week treatment with TCM was associated with activation of AMPK, Akt, and insulin-like growth factor-binding protein (IGFBP)1 pathways, with downregulation of miR29-b and expression of a gene network implicated in cell cycle, intermediary, and NADPH metabolism with normalization of CYP7a1 and IGFBP1 expression. These concerted changes in mRNA, miRNA, and proteins may explain the sustained effects of TCM in favor of cell survival, increased glucose uptake, and lipid oxidation/catabolism with improved insulin sensitivity and liver regeneration. These novel findings suggest that multicomponent TCM may be a useful tool to unravel genome regulation and expression in complex diseases.

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Protein kinase C (PKC) has been implicated in insulin-induced glucose uptake in skeletal muscle cell, although the underlying mechanism remains unknown. In this study, we investigated the effect of PKC on actin remodeling and glucose transport in differentiated rat L6 muscle cells expressing myc-tagged glucose transporter 4 (GLUT4). On insulin stimulation, PKC translocated from low-density microsomes to plasma membrane accompanied by increase in GLUT4 translocation and glucose uptake. Z-scan confocal microscopy revealed a spatial colocalization of relocated PKC with the small GTPase Rac-1, actin, and GLUT4 after insulin stimulation. The insulin-mediated colocalization, PKC distribution, GLUT4 translocation, and glucose uptake were inhibited by wortmannin and cell-permeable PKC pseudosubstrate peptide. In stable transfected cells, overexpression of PKC caused an insulin-like effect on actin remodeling accompanied by a 2.1-fold increase in GLUT4 translocation and 1.7-fold increase in glucose uptake in the absence of insulin. The effects of PKC overexpression were abolished by cell-permeable PKC pseudosubstrate peptide, but not wortmannin. Transient transfection of constitutively active Rac-1 recruited PKC to new structures resembling actin remodeling, whereas dominant negative Rac-1 prevented the insulin-mediated PKC translocation. Together, these results suggest that PKC mediates insulin effect on glucose transport through actin remodeling in muscle cells. INTRODUCTIONInsulin stimulates glucose uptake into skeletal muscle tissue mainly through GLUT4 translocation from intracellular pools to the plasma membrane (Klip et al., 1993;Bryant et al., 2002;Saltiel and Pessin, 2002). Tyrosine phosphorylation of insulin receptor substrate-1 by insulin activates phosphatidylinositol 3-kinase (PI3-K) and induces activation of downstream signal molecules protein kinase B (PKB/Akt) (Kohn et al., 1996;Tanti et al., 1997;Hill et al., 1999;Wang et al., 1999) and atypical PKCs (aPKCs) and / (Bandyopadhyay et al., 1997a(Bandyopadhyay et al., ,b, 1999Standaert et al., 1997;Kotani et al., 1998). aPKCs have been implicated in insulin action in adipocytes and muscle tissues (Kotani et al., 1998;Kim et al., 1999;Sajan et al., 2004), although this notion is not supported consistently by some studies (Tsuru et al., 2002). Evidence indicates that activation of aPKCs by insulin in skeletal muscles is defective in type 2 diabetic patients, monkeys, and rodents, and this defect seems to contribute significantly to the diminution in insulin-stimulated glucose disposal and muscle-dependent insulin resistance seen in these diabetic states (Bandyopadhyay et al., 1997a(Bandyopadhyay et al., , 1999Standaert et al., 2002;Beeson et al., 2003;Kim et al., 2003).We have previously shown that insulin causes a rapid and dynamic remodeling of actin into a cortical mesh (Khayat et al., 2000;Tong et al., 2001). Within the submembrane mesh, insulin-effective molecules such as glucose transporter (GLUT) isoform 4, vesicle-associated membrane protein (VAMP) 2,...

In Chinese Type 2 diabetic patients, lean subjects had predominant insulin deficiency and obese subjects had features of metabolic syndrome. Clinicians should have low threshold to initiate insulin therapy in lean Type 2 diabetic patients with suboptimal glycaemic control. In obese diabetic patients, aggressive control of multiple cardiovascular risks is of particular importance.

Insulin stimulates glucose uptake by recruiting glucose transporter 4 (GLUT4) from an intracellular compartment to the cell surface; this phenomenon is defective in type 2 diabetes. Here we examine the involvement of actin filaments in GLUT4 translocation and their possible defects in insulin resistance, using L6 myotubes expressing myc-tagged GLUT4. Insulin caused membrane ruffling, a dynamic distortion of the myotube dorsal surface. Fluorescence microscopy and immunogold staining of surface GLUT4myc coupled to backscatter electron microscopy revealed a high density of this protein in membrane ruffles. The t-SNAREs syntaxin4 and SNAP-23 were also abundant in these regions. Below the membrane, GLUT4 and the vesicular protein VAMP2, but not VAMP3, colocalized with the actin structures supporting the membrane ruffles. GLUT4myc externalization and membrane ruffles were reduced by jasplakinolide and by swinholide-A, drugs that affect actin filament stability and prevent actin branching, respectively. Insulin resistance generated by prolonged (24 hours) exposure of myotubes to high glucose and insulin diminished the acute insulin-dependent remodeling of cortical actin and GLUT4myc translocation, reminiscent of the effect of swinholide-A. We propose that GLUT4 vesicle incorporation into the plasma membrane involves insulin-dependent cortical actin remodeling and that defective actin remodeling contributes to insulin resistance.

Background: Obesity is now a global epidemic. In this study, we aimed to assess the rates of obesity using several major diagnostic criteria in Chinese school adolescents in Hong Kong.