Dexamethasone impairs insulin signalling and glucose transport by depletion of insulin receptor substrate-1, phosphatidylinositol 3-kinase and protein kinase B in primary cultured rat adipocytes

OBJECTIVE: Glucocorticoid excess leads to insulin resistance. This study explores the effects of glucocorticoids on the glucose transport system and insulin signalling in rat adipocytes. The interaction between glucocorticoids and high levels of insulin and glucose is also addressed. DESIGN AND METHODS: Isolated rat adipocytes were cultured for 24 h at different glucose concentrations (5 and 15 mmol/l) with or without the glucocorticoid analogue dexamethasone (0.3 micromol/l) and insulin (10(4) microU/ml). After the culture period, the cells were washed and then basal and insulin-stimulated glucose uptake, insulin binding and lipolysis as well as cellular content of insulin signalling proteins (insulin receptor substrate-1 (IRS-1), IRS-2, phosphatidylinositol 3-kinase (PI3-K) and protein kinase B (PKB)) and glucose transporter isoform GLUT4 were measured. RESULTS: Dexamethasone in the medium markedly decreased both basal and insulin-stimulated glucose uptake at both 5 and 15 mmol/l glucose (by approximately 40-50%, P<0.001 and P<0.05 respectively). Combined long-term treatment with insulin and dexamethasone exerted additive effects in decreasing basal, and to a lesser extent insulin-stimulated, glucose uptake capacity (P<0.05) compared with dexamethasone alone, but this was seen only at high glucose (15 mmol/l). Insulin binding was decreased (by approximately 40%, P<0.05) in dexamethasone-treated cells independently of surrounding glucose concentration. Following dexamethasone treatment a approximately 75% decrease (P<0.001) in IRS-1 expression and an increase in IRS-2 (by approximately 150%, P<0.001) was shown. Dexamethasone also induced a subtle decrease in PI3-K (by approximately 20%, P<0.01) and a substantial decrease in PKB content (by approximately 45%, P<0.001). Insulin-stimulated PKB phosphorylation was decreased (by approximately 40%, P<0.01) in dexamethasone-treated cells. Dexamethasone did not alter the amount of total cellular membrane-associated GLUT4 protein. The effects of dexamethasone per se on glucose transport and insulin signalling proteins were mainly unaffected by the surrounding glucose and insulin levels. Dexamethasone increased the basal lipolytic rate (approximately 4-fold, P<0.05), but did not alter the antilipolytic effect of insulin. CONCLUSIONS: These results suggest that glucocorticoids, independently of the surrounding glucose and insulin concentration, impair glucose transport capacity in fat cells. This is not due to alterations in GLUT4 abundance. Instead dexamethasone-induced insulin resistance may be mediated via reduced cellular content of IRS-1 and PKB accompanied by a parallel reduction in insulin-stimulated activation of PKB.     

Insulin resistance induced by high glucose and high insulin precedes insulin receptor substrate 1 protein depletion in human adipocytes

The aim of this study was to investigate whether high glucose and/or high insulin produces cellular insulin resistance in human adipocytes and, if so, to evaluate the time course and content of key proteins in the insulin signaling pathway. Subcutaneous fat biopsies were taken from 27 nondiabetic subjects. Insulin action in vitro was studied by measurement of glucose uptake after incubation at a physiologic glucose level (6 mmol/L) for 24 hours or with the last 2, 6, or 24 hours at a high glucose level (20 mmol/L) with or without high insulin (10(4)microU/mL). High glucose alone for 24 hours produced a small but significant impairment (by approximately 20%, P < .05) of insulin's effect to stimulate glucose transport, whereas nonstimulated glucose uptake was left intact. In contrast, the combination of high glucose and high insulin for 6 hours or more reduced basal glucose uptake by approximately 40% (P < .05). In addition, insulin-stimulated glucose uptake capacity was reduced by approximately 40% already after 2 hours (P < .05) and reached a maximal decline (by approximately 50%, P < .05) after a 6-hour culture in high glucose and high insulin. Treatment with high glucose and high insulin in combination for at least 6 hours reduced cellular insulin receptor substrate (IRS)-1, but not IRS-2, protein content by approximately 45% or more (P < .05). Moreover, after 24 hours, the ability of insulin to activate protein kinase B (ie, the phosphorylated protein kinase B [pPKB]-protein kinase B ratio) was decreased by approximately 50% (P < .05). No significant effects were seen on insulin signaling proteins or glucose transporter 4 after a long-term high-glucose culture. Culture with high insulin alone (and low glucose, 6 mmol/L) decreased basal and insulin-stimulated glucose uptake in conformity with the high-glucose/high-insulin setting. However, IRS-1 protein content remained unchanged. We conclude that, in adipocytes from healthy humans, high insulin alone for 2 hours or more decrease glucose uptake capacity. Likewise, high glucose and high insulin in combination for 2 hours or more decrease glucose uptake to the same extent as when cells were cultured with high insulin alone but, in addition, with a diminishment in IRS-1 protein lagging behind. Thus, IRS-1 depletion appears to be a secondary phenomenon in this model of insulin resistance. High glucose alone induces only a minor insulin resistance in human fat cells.     

Glucose and insulin regulate insulin sensitivity in primary cultured adipocytes without affecting insulin receptor kinase activity

We have previously shown in primary cultured adipocytes that chronic insulin exposure decreases insulin's subsequent ability to maximally restimulate the glucose transport system, and that extracellular glucose potentiates this ligand-induced defect in maximal insulin responsiveness. To examine whether glucose could also modulate insulin sensitivity (i.e. acute insulin effects at submaximal concentrations), adipocytes were cultured for 5 and 24 h in the absence and presence of various glucose and insulin concentrations. Then, after washing cells to remove any insulin and allow for full deactivation of transport, we assessed the dose response of insulin's acute ability to stimulate 2-deoxyglucose transport, bind to cell surface receptors, and activate insulin receptor tyrosine kinase activity. After 5 h, glucose and insulin alone had no chronic regulatory effects; however, in combination, these agents were able to decrease insulin sensitivity. In cells preincubated with 50 ng/ml insulin, the insulin ED50 for acute stimulation of glucose transport was increased by 65% and 116% as medium glucose was raised to 5 and 20 mM, respectively, relative to that at 0 mM glucose. After 24 h, chronic exposure to either glucose (20 mM) or insulin (50 ng/ml) alone increased the ED50 value by 52%, and, together they acted synergistically to increase the ED50 by 183%. While glucose and insulin independently and synergistically impaired insulin sensitivity, both agents were necessary for coregulation of maximal insulin responsiveness (confirming our previous observation). Insulin receptor down-regulation (18%) was observed after 24 h (but not 5 h) in insulin-treated cells; however, the major portion of the decrease in insulin sensitivity was due to uncoupling of occupied insulin receptors from stimulation of the glucose transport system. To further determine the mechanism for postbinding desensitization, we tested for concordant regulation of insulin receptor kinase activity. Insulin's ability to stimulate the receptor tyrosine kinase was assessed by multiple methods, including 1) receptor autophosphorylation and phosphorylation of Glu4-Tyr1 by solubilized insulin receptors activated in vitro, 2) histone-2B phosphorylation by receptors that were stimulated in intact cells and then solubilized under conditions that preserve the in cellulo phosphorylation state, and 3) receptor autophosphorylation and phosphorylation of pp180 in intact cells. Long term treatment (24 h) with glucose (10 mM) and insulin (50 ng/ml) markedly decreased insulin sensitivity (and receptor coupling), but did not affect insulin receptor kinase activity in any of these studies.(ABSTRACT TRUNCATED AT 400 WORDS)     

Polymorphisms of the insulin receptor substrate-2 in patients with type 2 diabetes

We investigated the significance of Gly1057Asp and Leu647Val insulin receptor substrate (IRS)-2 polymorphisms in two Italian cohorts comprising 186 glucose-tolerant subjects and 240 subjects with type 2 diabetes from the Lazio region (i.e. representative of central Italy), and 123 glucose-tolerant subjects from the Sicily region (i.e. representative of south Italy). The allelic frequency of Gly1057Asp variant did not differ between diabetics (32.9%) and nondiabetic subjects, whatever their ethnicity was (35.8% and 33.7% from Lazio and Sicily, respectively). As compared with Gly/Gly subjects within each group, Asp/Asp individuals showed no differences in quantitative traits, including fasting insulin and C-peptide, and several indices of insulin sensitivity and secretion. Only one of the diabetic patients was heterozygous for the Leu647Val variant, and none of the control subjects carried this variant. This patient had three children who were also heterozygous for this variant. They were glucose tolerant, and their insulin sensitivity and insulin secretion indices were within the range of age-matched controls. We also analyzed IRS-2 function in fibroblasts from carriers of Gly1057Asp or Leu647Val variant. No defects in IRS-2 expression, insulin-stimulated phosphorylation, or binding to the p85 subunit of phosphatidylinositol 3-kinase were observed. These results strongly argue against a major role of IRS-2 polymorphisms in the pathogenesis of type 2 diabetes.     

The depletion of cellular mitochondrial DNA causes insulin resistance through the alteration of insulin receptor substrate-1 in rat myocytes

Since the bioenergetic capacity of skeletal muscle mitochondria is decreased in type 2 diabetes and obesity, the reduction of mitochondrial DNA (mtDNA) content may be involved in the development of insulin resistance in skeletal muscle. To elucidate the association of cellular mtDNA content and insulin resistance, we produced L6 GLUT4myc myocytes depleted of mtDNA by long-term treatment with ethidium bromide (EtBr). L6 GLUT4myc cells cultured with 0.2 microg/ml EtBr (termed depleted cells) revealed a marked decrease in cellular mtDNA, concomitant with a lack of mRNAs encoded by mtDNA. Interestingly, the mtDNA-depleted cells showed a drastic decrease in basal and insulin-stimulated glucose uptake, indicating that L6 GLUT4myc cells develop impaired glucose utilization and insulin resistance. The repletion of mtDNA normalized basal and insulin-stimulated glucose uptake. The plasma membrane (PM) GLUT4 in the basal state was decreased, and the insulin-stimulated GLUT4 translocation to the PM was drastically reduced by mtDNA depletion. Interestingly, the expression of IRS-1 associated with insulin signaling was decreased by 90% in the depleted cells, and the insulin-stimulated phosphorylation of IRS-1 and Akt2/PKB were drastically reduced in the depleted cells. Those changes returned to control levels after mtDNA repletion. Taken together, our data suggest that PM GLUT4 content and insulin signal pathway intermediates are modulated by the alteration of cellular mtDNA content, and the reduction in the expression of IRS-1 and insulin-stimulated phosphorylation of IRS-1 and Akt2/PKB are associated with insulin resistance in the mtDNA-depleted L6 GLUT4myc myocytes.     

Insulin stimulation of glucose metabolism in rat adipocytes: possible implication of protein kinase C

The hypothesis that insulin stimulation of glucose and lipid metabolism in rat adipocytes may involve the activation of protein kinase C was evaluated. 4 beta-Phorbol 12 beta-myristate, 13 alpha-acetate (PMA, 0.1-1000 ng/ml), a potent tumor promoter acting as a substitute for diacylglycerol to activate protein kinase C, stimulated 2-deoxyglucose transport in a time- and dose-dependent manner, without affecting passive glucose diffusion. PMA (0.1-1000 ng/ml) also elicited a dose-dependent activation of lipogenesis from [3-3H] glucose. Maximal PMA effects (100 ng/ml) on both processes were 60% of insulin maximal effects. In contrast, PMA (1-1000 ng/ml) failed to mimic the ability of insulin to stimulate lipogenesis from [3H]acetate. 4 beta-Phorbol 12,13 dibutyrate, mezerein, 1-oleyl-2-acetyl-glycerol, 1,2 diolein, known as protein kinase C activators, also markedly stimulated glucose metabolism whereas 4 alpha-phorbol 12,13 didecanoate and 4 beta-phorbol 13-monoacetate, shown not to activate protein kinase C, were ineffective. Mellitin, a cytotoxin-inhibiting protein kinase C, markedly decreased both PMA and insulin stimulation of glucose metabolism. PMA decreased insulin submaximal effects on 2-deoxyglucose transport without inhibiting insulin binding. Maximal PMA and insulin effects on 2-deoxyglucose transport and lipogenesis from [3-3H]glucose were not additive. Both PMA and insulin activated each metabolic process in a calcium-dependent manner. PMA, like insulin, no longer stimulated 2-deoxyglucose transport in fat cells treated with 2,4-dinitrophenol. These data show that PMA exhibited specific insulin-like properties on glucose metabolism in fat cells, without any effect on lipid synthesis from acetate. They indicate that PMA and insulin bioeffects may share a common step implicating a calcium- and energy-dependent process, distal to the initial insulin binding event. Our results suggest that protein kinase C may play a role in insulin regulation of glucose metabolism.     

Sulfonylureas activate glucose transport and protein kinase C in rat adipocytes

Glyburide and tolbutamide, at concentrations of 20 to 40 mumol/L and 1 to 2 mmol/L, respectively, stimulated glucose transport in rat adipocytes. Concomitantly, protein kinase C was activated, as evidenced by translocation of immunoreactive enzyme from cytosol to membranes. Glucose transport effects of the sulfonylureas were blocked by three inhibitors of protein kinase C (H-7, staurosporine, and sangivamycin), and by phorbol ester-induced down-regulation of protein kinase C. These findings suggest that sulfonylureas may stimulate glucose transport in rat adipocytes through activation of protein kinase C.     

Antiinsulin receptor autoantibodies induce insulin receptors to constitutively associate with insulin receptor substrate-1 and -2 and cause severe cell resistance to both insulin and insulin-like growth factor I.

We report here that antiinsulin receptor (anti-IR) autoantibodies (AIRs) from a newly diagnosed patient with type B syndrome of insulin resistance induced cellular resistance not only to insulin but also to insulin-like growth factor I (IGF-I) for the stimulation of phosphatidylinositol 3-kinase and mitogen-activated protein kinase activities and of glycogen and DNA syntheses. The molecular mechanisms of this dual resistance were investigated. Patient AIRs bound the IR at the insulin-binding site and caused insulin resistance at the IR level by inducing a 50% decrease in cell surface IRs and a severe defect in the tyrosine kinase activity of the residual IRs, manifested by a loss of insulin-stimulated IR autophosphorylation and IR substrate-1 (IRS-1)/IRS-2 phosphorylation. In contrast, cell resistance to IGF-I occurred at a step distal to IGF-I receptors (IGF-IRs), as AIRs altered neither IGF-I binding nor IGF-I-induced IGF-IR autophosphorylation, but inhibited the ability of IGF-IRs to mediate tyrosine phosphorylation of IRS-1 and IRS-2 in response to IGF-I. Coimmunoprecipitation assays showed that in AIR-treated cells, IRs, but not IGF-IRs, were constitutively associated with IRS-1 and IRS-2, strongly suggesting that AIR-desensitized IRs impeded IGF-I action by sequestering IRS-1 and IRS-2. Accordingly, AIRs had no effect on the stimulation of mitogen-activated protein kinase activity or DNA synthesis by vanadyl sulfate, FCS, epidermal growth factor, or platelet-derived growth factor, all of which activate signaling pathways independent of IRS-1/IRS-2. Thus, AIRs induced cell resistance to both insulin and IGF-I through a novel mechanism involving a constitutive and stable association of IRS-1 and IRS-2 with the IR.     

Myokines in insulin resistance and type 2 diabetes

Skeletal muscle represents the largest organ of the body in non-obese individuals and is now considered to be an active endocrine organ releasing a host of so-called myokines. These myokines are part of a complex network that mediates communication between muscle, the liver, adipose tissue, the brain and other organs. Recent data suggest that myokines regulated by muscle contraction may play a key role in mediating the health-promoting effects of regular physical activity. Although hundreds of myokines have recently been described in proteomic studies, we currently have a rather limited knowledge of the specific role these myokines play in the prevention of insulin resistance, inflammation and associated metabolic dysfunction. Several myokines are known to have both local and endocrine functions, but in many cases the contribution of physical activity to the systemic level of these molecules remains as yet unexplored. Very recently, novel myokines such as irisin, which is thought to induce a white to brown shift in adipocytes, have gained considerable interest as potential therapeutic targets. In this review, we summarise the most recent findings on the role of myokines in the regulation of substrate metabolism and insulin sensitivity. We further explore the role of myokines in the regulation of inflammation and provide a critical assessment of irisin and other myokines regarding their potential as therapeutic targets.     

吸烟减少大鼠骨骼肌胰岛素受体底物1 mRNA和蛋白表达

应用RT-PCR及免疫组化技术分别检测实验大鼠后肢骨骼肌胰岛素受体底物1 mRNA及蛋白的表达。结果提示正常吸烟组、高脂饲养吸烟组及糖尿病吸烟组大鼠骨骼肌胰岛素受体底物1基因mRNA的表达及蛋白含量显著低于各自对照组（P〈0．05或P〈0．01），这可能是吸烟导致胰岛素抵抗的分子机制之一。     

Catecholamines and tumour promoting phorbolesters inhibit insulin receptor kinase and induce insulin resistance in isolated human adipocytes

Summary The effect of the catecholamine isoprenaline (10^–5mol/l) and of the tumour promoting phorbolester tetradecanoyl--phorbol acetate (10^–9mol/l) on insulin stimulated 3-O-methyl-glucose transport was studied in freshly isolated human adipocytes. Both substances reduced the maximal responsiveness of the glucose transport system to insulin by approximately 50%. To test if this is caused by inhibition of the insulin receptor kinase the receptor from phorbolester and isoprenaline treated cells was solubilized, partially purified and its kinase activity studied in vitro. Insulin stimulated ³²P-incorporation into the -subunit of the insulin receptor of phorbolester or isoprenaline treated cells was reduced to 20–60% of the values found with receptor from control cells at insulin concentrations between 10^–10mol/l and 10^–7mol/l. This inhibition of kinase activity of receptor from phorbolester and isoprenaline treated cells was observed at nonsaturating adenosine triphosphate levels (5 mol/l), and it could be overcome with higher concentrations of -³²P-adenosine triphosphate in the phosphorylation assay. A Lineweaver Burk analysis of the insulin stimulated receptor phosphorylation revealed that the Michaelis constant for adenosine triphosphate of the receptor kinase from phorbolester and isoprenaline treated cells was increased to >100 mol/l compared with <50 mol/l for receptor from control cells. We conclude from the data that catecholamine and phorbolester treatment of human adipocytes modulates the kinase activity of the insulin receptor by increasing its Michaelis constant for adenosine-triphosphate, and propose that this modulation of receptor kinase is a mechanism that can contribute to the pathogenesis of insulin resistance in human fat cells.     

Dexamethasone regulates the glucose transport system in primary cultured adipocytes: different mechanisms of insulin resistance after acute and chronic exposure

We have studied the ability of dexamethasone to regulate the glucose transport system in primary cultured adipocytes and delineated the mechanisms of insulin resistance after both acute and chronic treatment. Acutely, 20 nM dexamethasone led to a 65% decrease in basal and a 31% decrement in maximally insulin-stimulated glucose transport (ED50 = 3-4 nM; t1/2 = 50 min). These effects were maximal by 90-120 min, and a plateau was maintained over an additional 1-1.5 h. Chronic dexamethasone exposure (24 h) led to a more profound decrease in basal (77%; ED50 = 0.4 nM) and maximally stimulated (55%; ED50 = 1.0 nM) rates of glucose transport and shifted the transport: insulin dose-response curve to the right by increasing the half-maximally effective insulin concentration from 0.2 to 0.4 ng/ml. Dexamethasone did not affect cell surface insulin binding over 24 h. Both the short and long term effects of dexamethasone were partially blocked by the combined presence of insulin during preincubation and were not modulated by glucose. We also assessed effects on the number and cellular distribution of glucose transporter proteins using the cytochalasin-B binding assay. After 2 h, dexamethasone (30 nM) decreased the number of glucose transporters in plasma membranes by 30% in basal cells and by 41% in maximally insulin-stimulated cells, while increasing the number of low density microsomal transporters by 22-23% (P = NS). Transporter number in a total cellular membrane fraction was unaltered by short term dexamethasone. Chronic dexamethasone exposure (24 h) decreased plasma membrane and low density microsomal transporters by 30-50% in both basal and insulin-stimulated cells and depleted transporters by 43% in a total cellular membrane fraction. In conclusion, 1) dexamethasone induces progressive insulin resistance by sequentially regulating multiple aspects of the insulin-responsive glucose transport system. At early times (2 h) dexamethasone impairs insulin's ability to translocate intracellular glucose transporters to the cell surface and with more chronic exposure (24 h), depletes the total number of cellular transporters. 2) Glucose modulates desensitization of the glucose transport system by insulin, but not by dexamethasone, and thus, there are both glucose-dependent and -independent mechanisms of insulin resistance. 3) Insulin can heterologously inhibit dexamethasone's effects on glucose transport at both early and late phases of desensitization. These studies highlight the complex hormonal regulation at the glucose transport system.     

Impaired glucose transport and protein kinase B activation by insulin, but not okadaic acid, in adipocytes from subjects with Type II diabetes mellitus

Aims/hypothesis. To study the effects of insulin and okadaic acid, a serine/threonine phosphatase inhibitor which does not increase PI3-kinase activity, on the rate of glucose transport and protein kinase B activation in adipocytes from healthy subjects and subjects with Type II (non-insulin-dependent) diabetes mellitus. Methods. Adipocytes were incubated with or without insulin or okadaic acid or both and glucose transport, protein kinase B activity, phosphorylation and protein expression measured. Results. Insulin and okadaic acid alone increased glucose uptake to a similar degree in adipocytes from healthy subjects and, when combined, exerted a partial additive effect. The effect of insulin was reduced by about 60 % in adipocytes from Type II diabetic patients, whereas the effect of okadaic acid was essentially unchanged and no further increase was seen when okadaic acid and insulin were combined. Okadaic acid increased protein kinase B activity to a greater extent (two to threefold) than insulin but only slightly increased the serine phosphorylation of protein kinase B. Adipocytes from Type II diabetic subjects exhibited both an impaired sensitivity as well as a reduced total serine phosphorylation and activation of protein kinase B in response to insulin but protein kinase B activity in response to okadaic acid was intact. Conclusion/interpretation. These results show that the ability of insulin to increase glucose transport and activate protein kinase B is reduced in fat cells from Type II diabetic subjects. Protein kinase B can, however, be activated by agents like okadaic acid which bypass the upstream defects in the insulin signalling pathway in Type II diabetic cells and, thus, increase glucose uptake. [Diabetologia (1999) 42: 819–825] Received: 18 January 1999 and in revised form: 3 March 1999     

2型糖尿病患者ICIR改变及其影响因素的研究

目的 研究２型糖尿病（ＤＭ）ＩＣＩＲ改变及其影响因素。方法 Ｒｏｄｂｅｌｌ法获取游离脂肪细胞，Ｆａｎｔｕｓ ＩＧ’ｓ法行脂肪组织体外孵育实验，放射受体分析法分别测肥胖２型ＤＭ、非肥胖２型ＤＭ、单纯肥胖及正常对照组的ＩＣＩＲ。结果 １各种研究组ＩＣＩＲ数量较对照组明显减低，两２型ＤＭ组ＩＣＩＲ减少幅度高于单纯肥胖组，而两２型ＤＭ间ＩＣＩＲ减少无显著性差异；２两２例ＤＭ组减少的ＩＣＩＲ主要是低亲和力高     

Proatherosclerotic mechanisms involving protein kinase C in diabetes and insulin resistance

In diabetes and insulin resistance, activation of protein kinase C (PKC) in vascular cells may be a key link between elevated plasma and tissue concentrations of glucose and nonesterified fatty acids and abnormal vascular cell signaling. Initial studies of PKC activation in diabetes focused on microvascular complications, but increasing evidence supports that PKC plays a role in several mechanisms promoting atherosclerosis. This review explains how PKC is thought to be activated in diabetes and insulin resistance through de novo synthesis of diacylglycerol. Furthermore, the review summarizes studies that implicate PKC in promoting proatherogenic mechanisms or inhibiting antiatherogenic mechanisms, including studies of endothelial dysfunction; gene induction and activation of vascular NAD(P)H oxidase; endothelial nitric oxide synthase expression and function; endothelin-1 expression; growth, migration, and apoptosis of vascular smooth muscle cells; induction of adhesion molecules; and oxidized low-density lipoprotein uptake by monocyte-derived macrophages.     

Endothelial dysfunction in insulin resistance and type 2 diabetes

Macrovascular disease is the number one killer in type 2 diabetes patients. The cluster of risk factors in the insulin resistance syndrome (IRS) partly explains this notion. Insulin action in muscle, liver or adipose tissue has been thoroughly described in the literature, whilst this has been less described for the endothelium. Insulin stimulates nitric oxide (NO) production in the endothelium and reduced bioavailability of NO is usually defined as endothelial dysfunction. This impairment might be related to defective insulin signalling in the endothelial cell. Therefore, insulin resistance mechanisms in the endothelial cell will be emphasized in this review. Imbalance between the vasodilating agent NO and the vasoconstrictor endothelin-1 (ET-1) contributes to endothelial dysfunction. Different methods and circulating markers to assess endothelial function will be outlined. Circulating markers of an activated endothelium appear long before type 2 diabetes develops suggesting a unique role of the endothelium in the pathophysiology of the disease. Hampered blood flow in nutritive capillaries due to endothelial dysfunction is coupled with decreased glucose uptake and hyperglycemia. The forearm model combined with muscle microdialysis enables us to measure interstitial glucose and an index for capillary recruitment, the permeability surface area (PS). Available data from this method suggest that capillary recruitment in response of insulin is impaired in insulin resistant human subjects.     

血浆脂联素与2型糖尿病胰岛素抵抗关系的研究

目的　测定 2型糖尿病患者血浆脂联素的水平 ,并分析它与体重指数、血糖、胰岛素、血脂和胰岛素抵抗的关系 ,从而探讨脂联素在糖尿病发病中的作用。方法　健康对照组 2 8例 ,2型糖尿病组 60例 ,根据体重指数又将糖尿病组分为非肥胖糖尿病组 3 0例 (BMI <2 5kg/m2 )和肥胖糖尿病组 3 0例 (BMI >2 5kg/m2 )。用ELISA方法检测空腹血浆脂联素浓度 ,同时测定各组的空腹血糖、胰岛素、血脂的水平 ;根据HOMA模型提出的公式 ,计算分析胰岛素抵抗指数 ,并分析各指标间的相关性。结果　 (1)糖尿病各组血浆脂联素的水平明显低于正常对照组 ,且肥胖糖尿病组脂联素的水平低于非肥胖组 ,差异均有显著性 (P 0 .0 1) ;(2 )血浆脂联素浓度与体重指数、空腹胰岛素、胰岛素抵抗指数 (IR)、甘油三酯呈显著负相关。结论　脂联素参与了胰岛素抵抗的发生过程 ,与糖尿病的发生发展密切相关 ;脂联素可作为评价胰岛素抵抗程度的一种新的敏感指标     

2型糖尿病病人的内脏脂肪性肥胖和胰岛素抵抗

目的 探讨2型糖尿病病人内脏型肥胖和胰岛素抵抗的关系。方法 对50例2型糖尿病病人进行内脏脂肪测定,采用CT影像学检查方法;胰岛素抵抗采用高胰岛素—正血糖钳夹试验,得出葡萄糖利用率。根据有无内脏型肥胖将病人分为内脏肥胖组和非内脏肥胖组。结果 (1)内脏肥胖组的内脏脂肪平均值为127.28cm~2,非内脏肥胖组为70.41cm~2;内脏肥胖组比非肥胖型组有更高的甘油三酯水平和收缩压;内脏肥胖组的葡萄糖利用率平均值为3.54mg/(kg·min),非内脏肥胖组的平均值为9.66mg/(kg·min),二组差异有显著意义。(2)内脏脂肪和皮下脂肪与2型糖尿病病人胰岛素介导的葡萄糖利用率显著相关,内脏脂肪是胰岛素抵抗的影响的最强指数,即使与年龄、性别及体重指数校正以后也不影响结果。结论 内脏脂肪肥胖和心血管危险因子密切相关,是2型糖尿病胰岛素抵抗的独立影响因子。     

The microvasculature in insulin resistance and type 2 diabetes

Insulin resistance of muscle has been attributed to impairment of elements of insulin signaling, glucose transport, and/or metabolism within the muscle cells. This article explores the notion that a component of insulin resistance in vivo may result from impaired hemodynamic effects of this hormone to facilitate access to the muscle cells for itself and other nutrients, including glucose. In chronic situations this may manifest as a decreased capillary density of muscle, but in the acute, there may be impaired mechanisms for increasing total limb blood flow or for achieving optimal microvascular perfusion. Newly developed techniques show that insulin acts to recruit muscle capillary flow to enhance microvascular perfusion in animals and humans. This microvascular effect of insulin correlates closely with muscle glucose uptake, is independent of increases in bulk blood flow, and is impaired in obese insulin-resistant patients. Similarly, there are impaired vasodilatory responses in the skin of diabetic subjects.     

Site of insulin resistance in type 1 diabetes: insulin-mediated glucose disposal in vivo in relation to insulin binding and action in adipocytes in vitro

To evaluate the mechanism of insulin resistance in type 1 diabetes mellitus, we measured insulin sensitivity in vivo and insulin action in adipocytes in vitro. The study groups consisted of 18 insulin-treated type 1 diabetic patients and 14 matched normal subjects. In each subject, insulin-mediated glucose disposal in vivo was measured by the euglycemic clamp technique. An open surgical biopsy was performed in 9 diabetic and 7 healthy subjects to obtain abdominal sc adipose tissue for the measurement of [125I]insulin binding, D-[14C]-glucose transport, oxidation, and lipogenesis. During the euglycemic clamp studies, similar steady state plasma glucose (4.8 mmol/liter) and insulin (80 mU/liter = 700 pM) levels were maintained in both groups. The rate of glucose metabolism (M) was 43% lower in the diabetic patients (4.75 +/- 0.34 mg/kg X min) than in the normal subjects (8.27 +/- 0.43 mg/kg X min; P less than 0.001). [125I]Insulin binding to adipocytes was reduced in the diabetic patients (26% reduction in tracer binding; P less than 0.05) due to a reduction in receptor number. Insulin binding was not related to the M value at any insulin concentration. Basal and insulin-stimulated rates of glucose transport were not significantly different in diabetic and normal subjects. The basal glucose oxidation rate was reduced by 50% (P less than 0.02), and maximal glucose oxidation was reduced by 49% (P less than 0.03) in the diabetic patients (237 +/- 30 vs. 359 +/- 49 pmol/30,000 cells X 90 min, basal vs. maximal glucose oxidation, respectively) compared to those in normal subjects (513 +/- 101 vs. 700 +/- 133 pmol/30,000 cells X 90 min). The percentage responses of glucose oxidation and glucose transport to insulin were similar in both groups. Glucose oxidation rates at basal (r = 0.68; P less than 0.01), half-maximally (ED50; r = 0.70; P less than 0.01), and maximally (r = 0.64; P less than 0.05) effective insulin concentrations were positively related to the M value. Basal and insulin-stimulated rates of lipogenesis were comparable between the diabetic and normal subjects. In conclusion, insulin-mediated glucose disposal in vivo is reduced in conventionally treated type 1 diabetic patients. In vitro, adipocytes from diabetes bound slightly less insulin at tracer insulin concentrations, but the magnitude of this reduction was not related to impairment of glucose metabolism in vivo. Of the pathways of glucose metabolism studied, the rate of glucose oxidation was most affected. A significant relationship was found between the M value and the rate of in vitro glucose oxidation.(ABSTRACT TRUNCATED AT 400 WORDS)