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(affects Type 1 AND Type 2 Diabetics)

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Pathophysiology

Interleukin-1–Receptor Antagonist in Type 2 Diabetes Mellitus (NEJM 356:1517-1526, 2007)Background The expression of interleukin-1–receptor antagonist is reduced in pancreatic islets of patients with type 2 diabetes mellitus, and high glucose concentrations induce the production of interleukin-1 in human pancreatic beta cells, leading to impaired insulin secretion, decreased cell proliferation, and apoptosis. Methods In this double-blind, parallel-group trial involving 70 patients with type 2 diabetes, we randomly assigned 34 patients to receive 100 mg of anakinra (a recombinant human interleukin-1–receptor antagonist) subcutaneously once daily for 13 weeks and 36 patients to receive placebo. At baseline and at 13 weeks, all patients underwent an oral glucose-tolerance test, followed by an intravenous bolus of 0.3 g of glucose per kilogram of body weight, 0.5 mg of glucagon, and 5 g of arginine. In addition, 35 patients underwent a hyperinsulinemic–euglycemic clamp study. The primary end point was a change in the level of glycated hemoglobin, and secondary end points were changes in beta-cell function, insulin sensitivity, and inflammatory markers. Results At 13 weeks, in the anakinra group, the glycated hemoglobin level was 0.46 percentage point lower than in the placebo group (P=0.03); C-peptide secretion was enhanced (P=0.05), and there were reductions in the ratio of proinsulin to insulin (P=0.005) and in levels of interleukin-6 (P<0.001) and C-reactive protein (P=0.002). Insulin resistance, insulin-regulated gene expression in skeletal muscle, serum adipokine levels, and the body-mass index were similar in the two study groups. Symptomatic hypoglycemia was not observed, and there were no apparent drug-related serious adverse events. Conclusions The blockade of interleukin-1 with anakinra improved glycemia and beta-cell secretory function and reduced markers of systemic inflammation. (ClinicalTrials.gov number, NCT00303394 [ClinicalTrials.gov] )

• The Inflammation Link: Association between psoriasis, diabetes mellitus, and atherosclerosis - A case-control study(JAAD 56: 629-634 ,April 2007) Background Previous reports demonstrated an association between psoriasis and other diseases including heart failure and diabetes mellitus.Objectives Our aim was to describe the association between psoriasis, diabetes mellitus, and atherosclerosis in Israel. Methods A cross-sectional study was performed utilizing the database of Maccabi Healthcare Services (MHS), a large health provider organization in Israel. Case patients were defined as subjects who were diagnosed with psoriasis. Patients with diabetes and atherosclerosis were identified by using the MHS diabetes and cardiovascular registries, respectively. The control group included MHS enrollees without psoriasis. The proportion of diabetes and atherosclerosis among case and control groups was compared. Chi-square tests were used to compare categorical parameters. Logistic regression models were used for multivariate analyses. Results The study included 46,095 patients with psoriasis (case patients) and 1,579,037 subjects without psoriasis (control patients). The age-adjusted proportion of diabetes was significantly higher in psoriasis patients as compared with the control group (odds ratio [OR] 1.27, 95% confidence interval [CI] 1.1-1.48). The age-adjusted proportion of atherosclerosis was significantly higher in psoriasis patients as compared with the control group (OR 1.28, 95% CI 1.04-1.59). In patients with psoriasis, a multivariate logistic regression model demonstrated an association between diabetes and the multiple use of very potent topical steroids (P < .05) or use of systemic medication for psoriasis (methotrexate, cyclosporine or acitretin) (P < .001). A similar model demonstrated an association between atherosclerosis and the use of phototherapy (P < .001). Limitations Our study was based on a computerized database. The diagnosis of psoriasis was based on digitally transmitted data. Therefore overestimation (false-positive cases) and underestimation (false-negative cases) of psoriasis patients may exist, thereby being a source for information bias. A second limitation is selection bias that may occur due to the possibility that reporting of both psoriasis and associated illnesses is higher in individuals who are seeking medical care. A third limitation concerns the causal effect between occurrence of psoriasis and atherosclerosis or diabetes. The dataset of MHS records diagnoses only from 1997 and does not record the date of disease onset. Conclusions Our study supports previous reports for an association between psoriasis and atherosclerosis and psoriasis and diabetes. Further study is needed to support this observation.

Insulin resistance measured by the euglycaemic insulin clamp predicts subsequent CHD in elderly men. Proinsulin provides a better prediction of CHD than insulin. (Diabetologia [2005]48:1432-0428 (Online))  Aims/hypothesis  The association between CHD and insulin sensitivity (Si) measured by the euglycaemic insulin clamp has not been examined previously. Earlier studies found a relationship between CHD and elevated plasma insulin, an analysis that may have been confounded by co-determination of proinsulin, which has evolved as a stronger predictor of CHD. The aim was to determine the longitudinal relationships between Si, intact proinsulin, 32–33 split proinsulin, specific insulin and subsequent CHD. Methods  This was a population-based cohort study of 815 men in Uppsala, Sweden, aged 70 years at baseline with a follow-up of up to 10 years. Baseline insulin sensitivity was determined by euglycaemic insulin clamp. Fasting proinsulin, 32–33 split proinsulin and specific insulin concentrations were analysed using specific two-site immunometric assays. CHD was taken as diagnosed, if stated (in the event of death) on the Cause of Death Registry, or for subjects hospitalised for the first time with CHD, if CHD was recorded in the Hospital-Discharge Registry. The associations were analysed using Cox's proportional hazards, presented as hazard ratios (HRs) with their 95% CIs for a one-SD increase in the predictor. Results  In multivariate analysis, Si (HR:0.80, CI:0.65–0.97) adjusted for serum cholesterol, systolic blood pressure, fasting plasma glucose, BMI and smoking predicted CHD. Intact proinsulin (HR:1.18, CI:1.01–1.38), adjusted as the model above, predicted CHD, whereas 32–33 split proinsulin (HR:1.13, CI:0.95–1.35) or specific insulin (HR:1.07, CI:0.89–1.30) did not. Conclusions/interpretation  Insulin resistance measured by the euglycaemic insulin clamp predicts subsequent CHD in elderly men. Proinsulin provides a better prediction of CHD than insulin.

• Resistin Is an Inflammatory Marker of Atherosclerosis in Humans (Circulation [2005]111:932-939.) Background— Resistin, a plasma protein, induces insulininsulin-resistant rodents and resistance in rodents. Recent reports suggest that circulating levels of resistin are elevated in obese and humans. Whereas rodent resistin is made in adipocytes,adipocyte and macrophage macrophages are a major source of human resistin. Given the convergence of function, resistin mayResults— We examined provide unique insight into links between obesity, inflammation, and atherosclerosis in humans. Methods and whether plasma resistin(CAC), a quantitative index levels were associated with metabolic and inflammatory markers, as well as with coronary artery calcification of atherosclerosis, in 879 asymptomatic subjects in the Study of Inherited Risk of Coronary Atherosclerosis. Resistin levels were positively associated with levels of inflammatoryand lipoprotein-associated phospholipase markers, including soluble tumor necrosis factor-alpha receptor-2 (P<0.001), interleukin-6 (P=0.04), Aratio and 95% confidence interval in ₂ (P=0.002), but not measures of insulin resistance in multivariable analysis. Resistin levels also were associated (odds ordinal regression)1.52], P=0.03) and further control for with increasing CAC after adjustment for age, sex, and established risk factors (OR, 1.23 [CI, 1.03 to metabolic syndrome and plasma C-reactive proteinmetabolic syndrome, resistin levels (CRP) levels (OR, 1.25 [CI, 1.04 to 1.50], P=0.01). In subjects with further predicted CAC,inflammation and are predictive of whereas CRP levels did not. Conclusions— Plasma resistin levels are correlated with markers of coronary atherosclerosisinflammation, and atherosclerosis. in humans, independent of CRP. Resistin may represent a novel link between metabolic signals, Further studies are needed to define the relationship of resistin to clinical cardiovascular disease.

Tumor Necrosis Factor-alpha Receptor 1 Is a Major Predictor of Mortality and New-Onset Heart Failure in Patients With Acute Myocardial Infarction(Circulation [2005]111:863-870)Background— Tumor necrosis factor alpha (TNF-alpha) activation is an independent prognostic indicator of mortality in patients with heart failure (HF). Despite the recognition that several TNF family cytokines are elevated during myocardial infarction, their role in predicting subsequent prognosis in these setting remains poorly understood. Methods and Results— We performed a systematic evaluation of TNF-alpha and its type 1 and 2 soluble receptors, together with interleukin (IL)-6, IL-1 receptor antagonist, and IL-10, in 184 patients (132 men; mean age, 64±12) consecutively admitted for myocardial infarction. We correlated their values to short- and long-term incidence of death and HF (primary outcome). In 10 patients, we also studied the presence of transcardiac gradients for TNF-alpha and its soluble receptors. The control group comprised 45 healthy subjects who were sex and age matched (33 men; mean age, 65±6 years) to the patients. All tested cytokines were increased in patients, and no transcardiac or systemic AV difference was found. After a median follow-up of 406 days (range, 346 to 696 days), 24 patients died and 32 developed HF. Univariate analysis showed that all cytokines were related to outcome, whereas after adjustment for baseline and clinical characteristics, sTNFR-1 remained the only independent predictor of death and HF (hazard ratio, 2.9; 95% CI, 1.9 to 3.8, tertile 1 versus 3), together with left ventricular ejection fraction, Killip class, and creatine kinase-MB at peak. Conclusions— sTNFR-1 is a major short- and long-term predictor of mortality and HF in patients with acute myocardial infarction.

• Impaired insulin-induced vasodilation in small coronary arteries of Zucker obese rats is mediated by reactive oxygen species (Am J Physiol Heart Circ Physiol [2005]288:H854-H860)Insulin resistance (IR) and associated hyperinsulinemia are major risk factors for coronary artery disease. Mechanisms linking hyperinsulinemia to coronary vascular dysfunction in IR are unclear. We evaluated insulin-induced vasodilation in isolated small coronary arteries (SCA; ~225 µm) of Zucker obese (ZO) and control Zucker lean (ZL) rats. Vascular responses to insulin (0.1–100 ng/ml), ACh (10^–9–10^–5 mol/l), and sodium nitroprusside (10^–8–10^–4 mol/l) were assessed in SCA by measurement of intraluminal diameter using videomicroscopy. Insulin-induced dilation was decreased in ZO compared with ZL rats, whereas ACh and sodium nitroprusside elicited similar vasodilations. Pretreatment of arteries with SOD (200 U/ml), a scavenger of reactive oxygen species (ROS), restored the vasorelaxation response to insulin in ZO arteries, whereas ZL arteries were unaffected. Pretreatment of SCA with N-nitro-L-arginine methyl ester (100 µmol/l), an inhibitor of endothelial nitric oxide (NO) synthase (eNOS), elicited a vasoconstrictor response to insulin that was greater in ZO than in ZL rats. This vasoconstrictor response was reversed to vasodilation in ZO and ZL rats by cotreatment of the SCA with SOD or apocynin (10 µmol/l), a specific inhibitor of vascular NADPH oxidase. Lucigenin-enhanced chemiluminescence showed increased basal ROS levels as well as insulin (330 ng/ml)-stimulated production of ROS in ZO arteries that was sensitive to inhibition by apocynin. Western blot analysis revealed increased eNOS expression in ZO rats, whereas Mn SOD and Cu,Zn SOD expression were similar to ZL rats. Thus IR in ZO rats leads to decreased insulin-induced vasodilation, probably as a result of increased production of ROS by vascular NADPH oxidase, leading to decreased NO bioavailability, despite a compensatory increase in eNOS expression.

Circulating Mononuclear Cells In The Obese Found To Be In Proinflammatory State, Contributing To Diabetes And Heart Disease (Circulation [2004] 110:1564-1571) Background— In view of the increase in plasma concentrations of proinflammatory mediators tumor necrosis factor-a (TNF-a), interleukin-6 (IL-6), and C-reactive protein (CRP) in obesity, we investigated whether peripheral blood mononuclear cells (MNC) from obese subjects are in a proinflammatory state. Methods and Results— MNC were prepared from fasting blood samples of obese (n=16; body mass index [BMI]=37.7±5.0 kg/m²) and normal-weight control (n=16; BMI=23.8±1.9 kg/m²) subjects. Nuclear factor kB (NF-kB) binding to DNA in nuclear extracts was elevated (P<0.05) and the inhibitor of NFkB-ß (IkB-ß) was significantly lower (P<0.001) in the obese group. Reverse transcription–polymerase chain reaction revealed elevated levels of migration inhibitor factor (MIF), IL-6, TNF-a, and matrix metalloproteinase-9 (MMP-9) mRNA expression in the obese subjects (P<0.05). Plasma concentrations of MIF, IL-6, TNF-a, MMP-9, and CRP were also significantly higher. Plasma glucose, insulin, and free fatty acids (FFAs) were measured, and homeostasis model assessment of insulin resistance (HOMA-IR) was calculated. Plasma FFA concentration related significantly to BMI, IL-6, and TNF-a mRNA expression and plasma CRP levels but not to HOMA-IR. On the other hand, the inflammatory mediators were significantly related to BMI and HOMA-IR. Conclusions— These data show (1) for the first time that MNC in obesity are in a proinflammatory state with an increase in intranuclear NF-kB binding, a decrease in IkB-ß, and an increase in the transcription of proinflammatory genes regulated by NF-kB; (2) that plasma FFAs are a modulator of inflammation; and (3) that insulin resistance is a function of inflammatory mediators.

• Urinary 20-Hydroxyeicosatetraenoic Acid Is Associated With Endothelial Dysfunction in Humans (Circulation [2004]110:438-443) Background— 20-Hydroxyeicosatetraenoic acid (20-HETE) is a cytochrome P450 (-hydroxylase) metabolite of arachidonic acid with vasoconstrictor activity that may be involved in the pathogenesis of hypertension. In humans, there are few data relating 20-HETE to vascular pathophysiology. This study aimed to determine whether urinary 20-HETE excretion is related to blood pressure or vascular endothelial function in humans. Methods and Results— Sixty-six subjects (37 males, 29 females), including 29 with untreated hypertension, had urinary 20-HETE excretion measured by gas chromatography/mass spectrometry. There was no significant difference for 20-HETE excretion between hypertensive and normotensive subjects. 20-HETE excretion was positively related to body mass index and sodium excretion. There was a significant inverse association between urinary 20-HETE and endothelium-dependent vasodilation measured by flow-mediated dilation of the brachial artery (P=0.006). There was no association with vasodilator responses to nitroglycerin. In multiple regression analysis, 20-HETE remained an independent predictor of endothelium-dependent vasodilation after adjustment for age, body mass index, and blood pressure. When gender was included in the model, the relationship between 20-HETE and flow-mediated dilation was attenuated. Separate analysis by gender revealed that in women, hypertensive subjects had significantly higher 20-HETE excretion than normotensive subjects, but this was not seen in men. In women, 20-HETE was positively related to diastolic and systolic blood pressure. In men, 20-HETE was positively related to body mass index. Conclusions— This is the first demonstration of an association between 20-HETE excretion and in vivo vascular function in humans. Given the negative modulatory role of nitric oxide on -hydroxylase, the present results suggest a potentially important role for 20-HETE in human vascular physiology.

Fish intake may influence risk of Atrial Fibrillation (Circulation [2004]110:368-373) Background— Atrial fibrillation (AF) is the most common arrhythmia in clinical practice and is particularly common in the elderly. Although effects of fish intake, including potential antiarrhythmic effects, may favorably influence risk of AF, relationships between fish intake and AF incidence have not been evaluated. Methods and Results— In a prospective, population-based cohort of 4815 adults >=age 65 years, usual dietary intake was assessed at baseline in 1989 and 1990. Consumption of tuna and other broiled or baked fish correlated with plasma phospholipid long-chain n-3 fatty acids, whereas consumption of fried fish or fish sandwiches (fish burgers) did not. AF incidence was prospectively ascertained on the basis of hospital discharge records and annual electrocardiograms. During 12 years’ follow-up, 980 cases of incident AF were diagnosed. In multivariate analyses, consumption of tuna or other broiled or baked fish was inversely associated with incidence of AF, with 28% lower risk with intake 1 to 4 times per week (HR=0.72, 95% CI=0.58 to 0.91, P=0.005), and 31% lower risk with intake >=5 times per week (HR=0.69, 95% CI=0.52 to 0.91, P=0.008), compared with <1 time per month (P trend=0.004). Results were not materially different after adjustment for preceding myocardial infarction or congestive heart failure. In similar analyses, fried fish/fish sandwich consumption was not associated with lower risk of AF.Conclusions— Among elderly adults, consumption of tuna or other broiled or baked fish, but not fried fish or fish sandwiches, is associated with lower incidence of AF. Fish intake may influence risk of this common cardiac arrhythmia.

• Data suggest that AKT may be involved in the endothelial dysfunction of hypertension (Circulation [2004]109:2587-2593) Background— In hypertension, reduced nitric oxide production and blunted endothelial vasorelaxation are observed. It was recently reported that AKT phosphorylates and activates endothelial nitric oxide synthase and that impaired kinase activity may be involved in endothelial dysfunction. Methods and Results— To identify the physiological role of the kinase in normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR), we used adenoviral vectors to transfer the human AKT1 gene selectively to the common carotid endothelium. In vitro, endothelial vasorelaxations to acetylcholine, isoproterenol, and insulin were blunted in control carotids from SHR compared with WKY rats, and human AKT1 overexpression corrected these responses. Similarly, blood flow assessed in vivo by Doppler ultrasound was reduced in SHR compared with WKY carotids and normalized after AKT1 gene transfer. In primary cultured endothelial cells, we evaluated AKT phosphorylation, activity, and compartmentalization and observed a mislocalization of the kinase in SHR. Conclusions— We conclude that AKT participates in the settings of endothelial dysfunction in SHR rats by impaired membrane localization. Our data suggest that AKT is involved in endothelium dysfunction in hypertension.

Oxidative stress induces insulin resistance by activating the nuclear factor-kB pathway and disrupting normal subcellular distribution of phosphatidylinositol 3-kinase (Diabetologia [2004] 47: 794 - 805) Aims/hypothesis  Oxidative stress is associated with diabetes, hypertension and atherosclerosis. Insulin resistance is implicated in the development of these disorders. We tested the hypothesis that oxidative stress induces insulin resistance in rats, and endeavoured to identify mechanisms linking the two. Methods  Buthionine sulfoximine (BSO), an inhibitor of glutathione synthase, was administered to Sprague-Dawley rats and 3T3-L1 adipocytes. Glucose metabolism and insulin signalling both in vivo and in 3T3-L1 adipocytes were examined. In 3T3-L1 adipocytes, the effects of overexpression of a dominant negative mutant of inhibitory kB (IkB), one role of which is to block oxidative-stress-induced nuclear factor (NF)-kB activation, were investigated. Results  In rats given BSO for 2 weeks, the plasma lipid hydroperoxide level doubled, indicating increased oxidative stress. A hyperinsulinaemic-euglycaemic clamp study and a glucose transport assay using isolated muscle and adipocytes revealed insulin resistance in BSO-treated rats. BSO treatment also impaired insulin-induced glucose uptake and GLUT4 translocation in 3T3-L1 adipocytes. In BSO-treated rat muscle, adipose tissue and 3T3-L1 adipocytes, insulin-induced IRS-1 phosphorylation in the low-density microsome (LDM) fraction was specifically decreased, while that in whole cell lysates was not altered, and subsequent translocation of phosphatidylinositol (PI) 3-kinase from the cytosol and the LDM fraction was disrupted. BSO-induced impairments of insulin action and insulin signalling were reversed by overexpressing the dominant negative mutant of IkB, thereby suppressing NF-kB activation. Conclusions/interpretation  Oxidative stress induces insulin resistance by impairing IRS-1 phosphorylation and PI 3-kinase activation in the LDM fraction, and NF-kB activation is likely to be involved in this process.

• Endothelial dysfunction predicts type 2 diabetes in women independent of other known risk factors, including obesity and subclinical inflammation.(JAMA [2004] 291:1978-1986) Context  Endothelial dysfunction occurs in diagnosed type 2 diabetes mellitus but may also precede development of diabetes. Objective  To determine whether elevated plasma levels of biomarkers reflecting endothelial dysfunction (E-selectin; intercellular adhesion molecule 1 [ICAM-1]; and vascular cell adhesion molecule 1 [VCAM-1]) predict development of type 2 diabetes in initially nondiabetic women. Design and Setting  Prospective, nested case-control study within the Nurses' Health Study, an ongoing US study initiated in 1976. Participants  Of 121 700 women initially enrolled, 32 826 provided blood samples in 1989-1990; of those free of diabetes, cardiovascular disease, or cancer at baseline, 737 developed incident diabetes by 2000. Controls (n = 785) were selected according to matched age, fasting status, and race. Main Outcome Measure  Risk of confirmed clinically diagnosed type 2 diabetes by baseline levels of E-selectin, ICAM-1, and VCAM-1. Results  Baseline median levels of the biomarkers were significantly higher among cases than among controls (E-selectin, 61.2 vs 45.4 ng/mL; ICAM-1, 264.9 vs 247.0 ng/mL; VCAM-1, 545.4 vs 526.0 ng/mL [all P values <=0.004]). Elevated E-selectin and ICAM-1 levels predicted incident diabetes in logistic regression models conditioned on matching criteria and adjusted for body mass index (BMI), family history of diabetes, smoking, diet score, alcohol intake, activity index, and postmenopausal hormone use. The adjusted relative risks for incident diabetes in the top quintile vs the bottom quintiles were 5.43 for E-selectin (95% confidence interval [CI], 3.47-8.50), 3.56 for ICAM-1 (95% CI, 2.28-5.58), and 1.12 for VCAM-1 (95% CI, 0.76-1.66). Adjustment for waist circumference instead of BMI or further adjustment for baseline levels of C-reactive protein, fasting insulin, and hemoglobin A_1c or exclusion of cases diagnosed during the first 4 years of follow-up did not alter these associations. Conclusion  Endothelial dysfunction predicts type 2 diabetes in women independent of other known risk factors, including obesity and subclinical inflammation.

Interleukin-6 is a positive regulator of tumor necrosis factor -induced adipose-related protein in 3T3-L1 adipocytes (FEBS Letters [2004] 560:153-157) Tumor necrosis factor (TNF)alpha induced adipose-related protein (TIARP) is a novel TNF-alpha-stimulated protein in adipocytes. Besides TNFalpha, interleukin (IL)-6 has recently been shown to be another adipocytokine implicated in insulin resistance. Therefore, the impact of IL-6 on TIARP gene expression in 3T3-L1 adipocytes was determined by quantitative real-time reverse transcription-polymerase chain reaction. Interestingly, TIARP mRNA expression was stimulated up to 3.8-fold by IL-6 in a dose-dependent fashion with significant stimulation detectable at effector concentrations as low as 3 ng/ml and maximal effects seen at 100 ng/ml IL-6. Induction of TIARP mRNA by IL-6 was time-dependent with significant upregulation occurring as early as 2 h after effector addition and maximal effects observed at 4 h. In parallel, TIARP protein synthesis was upregulated with maximal effects seen after 8 h of IL-6 treatment. Furthermore, the Janus kinase 2 inhibitor AG490 decreased TIARP mRNA expression. The increase of TIARP mRNA could be reversed by withdrawal of IL-6 for 24 h. Furthermore, TIARP mRNA induction by IL-6 was also seen in brown adipocytes but not in muscle and liver cells. Taken together, these results show that TIARP is acutely regulated in adipose tissue not only by TNFalphabut also by IL-6 which has been shown to be another important cytokine implicated in the pathogenesis of insulin resistance.

• Plasminogen activator inhibitor correlates with coronary wall thickening in patients with angiographically normal coronary arteries (Thrombosis Research [2003] 112: 123-129) Introduction: Angiographically normal coronary arteries have concealed intimal thickening that importantly contribute to coronary arterial disease activity. Increased plasma levels of plasminogen activator inhibitor (PAI) are associated with myocardial infarction and atherosclerosis. However, it remains unclear whether the PAI contributes to vascular wall thickening detected by intravascular ultrasound (IVUS) in normal coronary angiogram. The aim of this study was to evaluate if the PAI activity contributes to the extent of atherosclerotic changes in angiographically normal coronary arteries using IVUS technique. Materials and methods: We studied 33 consecutive patients with normal coronary angiograms. These patients were divided into a high level of plasma PAI activity group (H-PAI; n=12) and a normal range of PAI activity group (N-PAI; n=21), according to the plasma PAI activity levels. Results: The average of "percent intima+media area (%I+M area)" and "maximal intima+media (I+M) thickness" were significantly greater in the H-PAI group as compared with those in the N-PAI group (p<0.05). Minimal lumen diameter and lumen area were comparable between these groups. The plasma PAI activity level was the independent predictor of increase in maximal I+M thickness, in multiple regression analysis with the traditional risk factors as covariates. Conclusions: Thickened intima+media of angiographically normal coronary arteries were associated with high plasma level of PAI activity, independently of other traditional risk factors. PAI may contribute to the pathogenesis of coronary intimal thickening that might increase coronary arterial tone. Author Keywords: Plasminogen activator inhibitor; Coronary wall thickening; Intravascular ultrasound; Normal coronary angiogram; Quantitative coronary angiography; Nitric oxides

Beta-blockers enhance natriuretic peptides' effects during exercise (J Am Coll Cardiol [2004] 43:353-359) In patients with coronary artery disease, natriuretic peptide levels in plasma are increased during exercise, French physicians now show that treatment with beta-blockers enhances the natriuretic peptide release, thus decreasing cardiac load.Cardiac disease is associated with increased resting levels of atrial and brain natriuretic peptide. Under stress, cardiomyocytes release both atrial and brain natriuretic peptide, which exert vasodilator and diuretic effects, which many be expected to reduce cardiac load. However, this process in patients with coronary artery disease has been poorly characterized.Therefore, Dr. Pierre-Yves Marie and colleagues at UPRES EA in Nancy, measured natriuretic peptide levels at rest and peak exercise in 104 patients with chronic coronary artery disease. They describe their findings in the Journal of the American College of Cardiology for February 4th.The best independent predictors of natriuretic peptide concentrations at rest were aging and the extent of scarred left ventricular area. "No previous study has given evidence of the dramatic influence of the extent of scarred myocardium, as determined by myocardial SPECT," the authors write.Beta blocker treatment did not affect natriuretic peptide concentrations at rest. However, during exercise, the average increase in both atrial and brain natriuretic peptide was more than doubled in the 55 patients treated with beta-blockers, even though their heart rate did not increase as much."This enhanced secretion of potent vasodilating and natriuretic agents might constitute an additional and original mechanism of these drugs for further protecting diseased hearts against stress," Dr. Marie's group concludes.

• MCC134 - which inhibits preconditioning - opens cardiac surface k(ATP) channels but blocks mito k(ATP) channels reaffirming the dependency of cardioprotection upon mito rather than surface k(ATP) channels (Circulation [2003]107(8):1183-8)BACKGROUND: MCC-134 (1-[4-(H-imidazol-1-yl)benzoyl] -N-methylcyclobutane-carbothioamide), a newly developed analog of aprikalim, opens surface smooth muscle-type ATP-sensitive potassium (K(ATP)) channels but inhibits pancreatic K(ATP) channels. However, the effects of MCC-134 on cardiac surface K(ATP) channels and mitochondrial K(ATP) (mitoK(ATP)) channels are unknown. A mixed agonist/blocker with differential effects on the two channel types would help to clarify the role of K(ATP) channels in cardioprotection. METHODS AND RESULTS: To index mitoK(ATP) channels, we measured mitochondrial flavoprotein fluorescence in rabbit ventricular myocytes. MCC-134 alone had little effect on basal flavoprotein fluorescence. However, MCC-134 inhibited diazoxide-induced flavoprotein oxidation in a dose-dependent manner (EC(50)=27 micro mol/L). When ATP was included in the pipette solution, MCC-134 slowly activated surface K(ATP) currents with some delay (>10 minutes). These results indicate that MCC-134 is a mitoK(ATP) channel inhibitor and a surface K(ATP) channel opener in native cardiac cells. In cell-pelleting ischemia assays, coapplication of MCC-134 with diazoxide abolished the cardioprotective effect of diazoxide, whereas MCC-134 alone did not alter cell death. These results were reproducible in both rabbit and mouse myocytes. MCC-134 also attenuated the effect of ischemic preconditioning against myocardial infarction in mice, consistent with the results of cell-pelleting ischemia assays. CONCLUSIONS: A single drug, MCC-134, opens surface K(ATP) channels but blocks mitoK(ATP) channels; the fact that this drug inhibits preconditioning reaffirms the primacy of mitoK(ATP) rather than surface K(ATP), channels in the mechanism of cardioprotection.

Stress upregulates arterial matrix metalloproteinase expression and activity via endothelin 1-A receptor activation (Am J Physiol Heart Circ Physiol [2003] 285: H2225 - H2232) Degradation of the extracellular matrix proteins by matrix metalloproteinases (MMP) is an important regulatory step in the vascular remodeling process. Recent studies demonstrated that ET_A receptors regulate cardiac MMP activity and fibrosis in DOCA-salt hypertension. However, little is known about endothelin (ET)-1 regulation of vascular MMP activity in hypertension. Thus early changes in ET-1-mediated regulation of MMP activity were measured in borderline hypertensive rats that develop impaired vasorelaxation and hypertension with chronic exposure to stress. Experiments were performed after 10 days of exposure to the behavioral stressor, air-jet stress, but before the onset of stress-induced hypertension. Study groups were 1) control (n = 8); 2) air-jet stress for 10 days (n = 8); 3) control plus ET_A antagonist ABT-627 (n = 4), and 4) air-jet stress plus ET_A antagonist (n = 4). MMP activity in the thoracic aorta was assessed by gelatin zymography. MMP protein and tissue ET-1 levels were evaluated by immunohistochemistry, and ET receptor density was determined by immunoblotting. Exposure to stress caused a twofold increase in plasma ET-1 levels (P < 0.05), and there was increased ET-1 staining at the tissue level. Total MMP activity and expression of MMP-2 and MMP-9 were increased in the stress group. ET_A receptor antagonism prevented the increase in MMP expression and activation in the stress group. These results provide evidence that the MMP system is activated before the development of hypertension and ET-1 mediates these early events in vascular remodeling.

• Inhibition of NF-kB activation in macrophages increases atherosclerosis in LDL receptor–deficient mice (J. Clin. Invest.[2003]112:1176-1185 doi:10.1172/JCI200318580) Atherosclerosis is now generally accepted as a chronic inflammatory condition. The transcription factor NF-kB is a key regulator of inflammation, immune responses, cell survival, and cell proliferation. To investigate the role of NF-kB activation in macrophages during atherogenesis, we used LDL receptor–deficient mice with a macrophage-restricted deletion of IkB kinase 2 (IKK2), which is essential for NF-kB activation by proinflammatory signals. These mice showed increased atherosclerosis as quantified by lesion area measurements. In addition, the lesions were more advanced and showed more necrosis and increased cell number in early lesions. Southern blotting revealed that deletion of IKK2 was approximately 65% in macrophages, coinciding with a reduction of 50% in NF-kB activation, as compared with controls. In both groups, the expression of differentiation markers, uptake of bacteria, and endocytosis of modified LDL was similar. Upon stimulation with LPS, production of TNF was reduced by approximately 50% in IKK2-deleted macrophages. Interestingly, we also found a major reduction in the anti-inflammatory cytokine IL-10. Our data show that inhibition of the NF-kB pathway in macrophages leads to more severe atherosclerosis in mice, possibly by affecting the pro- and anti-inflammatory balance that controls the development of atherosclerosis.

Tumor Necrosis Factor-alpha Inhibits Insulin’s Stimulating Effect on Glucose Uptake and Endothelium-Dependent Vasodilation in Humans (Circulation [2003]108:1815) Background— Inflammatory mechanisms could be involved in the pathogenesis of both insulin resistance and atherosclerosis. Therefore, we aimed at examining whether the proinflammatory cytokine tumor necrosis factor (TNF)-alpha inhibits insulin-stimulated glucose uptake and insulin-stimulated endothelial function in humans. Methods and Results— Healthy, lean male volunteers were studied. On each study day, 3 acetylcholine (ACh) or sodium nitroprusside (SNP) dose-response studies were performed by infusion into the brachial artery. Before and during the last 2 dose-response studies, insulin and/or TNF-alpha were coinfused. During infusion of insulin alone for 20 minutes, forearm glucose uptake increased by 220±44%. This increase was completely inhibited during coinfusion of TNF-alpha (started 10 min before insulin) with a more pronounced inhibition of glucose extraction than of blood flow. Furthermore, TNF-alpha inhibited the ACh forearm blood flow response (P<0.001), and this inhibition was larger during insulin infusion (P=0.01) but not further increased by N^G-monomethyl-L-arginine acetate (P=0.2). Insulin potentiated the SNP response less than the ACh response and the effect of TNF-alpha was smaller (P<0.001); TNF-alpha had no effect on the SNP response without insulin infusion. Thus, TNF-alpha inhibition of the combined response to insulin and ACh was likely mediated through inhibition of NO production. ^f Conclusion— These results support the concept that TNF-^alpha could play a role in the development of insulin resistance in humans, both in muscle and in vascular tissue.

• Atherosclerosis is well-begun preceding the clinical onset of diabetes (Arteriosclerosis, Thrombosis, and Vascular Biology [2003] 23:1845) Objective— We examined whether B-mode ultrasound–detected carotid artery intima-media thickness (IMT) was elevated before the onset of clinical diabetes. Methods and Results— The study population for these analyses included 1127 nondiabetic participants, 66 prediabetic participants, and 303 diabetic participants with a mean age of 49.8 years who participated in the Mexico City Diabetes Study, a prospective cohort study. Common carotid artery (CCA) and internal carotid artery (ICA) IMTs were measured bilaterally by B-mode ultrasound. Age- and sex-adjusted mean ICA and CCA IMTs were both significantly higher among prediabetic individuals {0.81 mm [95% confidence interval (CI), 0.75–0.88] and 0.72 mm [95% CI, 0.69–0.75], respectively} than in individuals who remained free of diabetes [0.71 mm (95% CI, 0.69–0.72) and 0.69 mm (95% CI, 0.68–0.69), respectively]. However, after adjustment for established cardiovascular risk factors, ICA IMT, but not CCA IMT, remained significantly higher among prediabetic individuals [0.81 mm (95% CI, 0.75–0.88) and 0.71 mm (95% CI, 0.68–0.74)] than in individuals who remained free of diabetes [0.71 mm (95% CI, 0.69–0.72) and 0.69 mm (95% CI, 0.68–0.70)]. Conclusions— The present study provides direct evidence at the vascular level that atherosclerosis levels are elevated before the clinical onset of diabetes.

Stearoyl-CoA desaturase 1 deficiency elevates insulin-signaling components and down-regulates protein-tyrosine phosphatase 1B in muscle (PNAS [2003] 100:11110-11115)

• Macrophage-Specific p53 Expression Plays a Crucial Role in Atherosclerosis Development and Plaque Remodeling (Arteriosclerosis, Thrombosis, and Vascular Biology [2003]23:1608) Objective— We first showed that absence of p53 accelerates atherosclerosis development in apoE-deficient mice. In this study, we investigated how macrophage-specific loss of p53 function might modulate atherosclerosis development in LDL receptor-deficient mice, a model for familial hypercholesterolemia. Methods and Results— We transferred bone marrow cells isolated from p53^+/+ and p53^-/- mice to lethally irradiated LDL receptor^-/- mice and evaluated the aortic atherosclerotic lesion areas in the recipients at different times afterward. At 15 weeks and again at 20 weeks, we found larger aortic lesion size in mice receiving p53^-/- cells compared with those that received p53^+/+ cells. By measuring the rate of bromodeoxyuridine incorporation, we found that the absence of p53 in macrophages stimulates cellular proliferation. In contrast, the rate of apoptosis in the atheromatous lesion was similar in the two groups of mice. Furthermore, we found that the absence of macrophage-specific p53 expression was associated with vulnerable-appearing lesions marked by increased tissue necrosis and reduced collagen deposition. Conclusions— p53 plays a crucial role in atherosclerosis lesion development and remodeling, and macrophage-specific p53 deficiency stimulates cellular proliferation leading to a vulnerable-appearing phenotype of lesions in a mouse model of familial hypercholesterolemia. Key Words: atherosclerosis • apoptosis • proliferation • necrosis • macrophage

IGT and DM2 associate with increased arterial stiffness occurring before the onset of DM2 and explained neither by conventional CV risk factors nor by hyperglycemia nor by hyperinsulinemia (Circulation [2003]107:2089.) Background— Type 2 diabetes (DM-2) and impaired glucose metabolism (IGM) are associated with an increased cardiovascular disease risk. In nondiabetic individuals, increased arterial stiffness is an important cause of cardiovascular disease. Associations between DM-2 and IGM and arterial stiffness have not been systematically investigated. Methods and Results— In a population-based cohort (n=747; 278 with normal glucose metabolism, 168 with IGM, and 301 with DM-2; mean age, 68.5 years), arterial stiffness was ultrasonically estimated by distensibility and compliance of the carotid, femoral, and brachial arteries and by the carotid elastic modulus. After adjustment for age, sex, and mean arterial pressure, DM-2 was associated with increased carotid, femoral, and brachial stiffness, whereas IGM was associated only with increased femoral and brachial stiffness. Carotid but not femoral or brachial stiffness increased from IGM to DM-2. Standardized ßs (95% CI) for IGM and DM-2, compared with normal glucose metabolism, were -0.06 (-0.23 to 0.10) and -0.37 (-0.51 to -0.23) for carotid distensibility; -0.02 (-0.18 to 0.18) and -0.25 (-0.40 to -0.09) for carotid compliance; -0.05 (-0.23 to 0.13) and 0.25 (0.10 to 0.40) for carotid elastic modulus; -0.70 (-0.89 to -0.51) and -0.67 (-0.83 to -0.52) for femoral distensibility; and -0.62 (-0.80 to -0.44) and -0.79 (-0.94 to -0.63) for femoral compliance. The brachial artery followed a pattern similar to that of the femoral artery. Increases in stiffness indices were explained by decreases in distension, increases in pulse pressure, an increase in carotid intima-media thickness, and, for the femoral artery, a decrease in diameter. Hyperglycemia or hyperinsulinemia explained only 30% of the arterial changes associated with glucose tolerance. Adjustment for conventional cardiovascular risk factors did not affect these findings. Conclusions— IGM and DM-2 are associated with increased arterial stiffness. An important part of the increased stiffness occurs before the onset of DM-2 and is explained neither by conventional cardiovascular risk factors nor by hyperglycemia or hyperinsulinemia.

• In-hospital LVF and cardiac mortality imply admission glycemia in ACS [independent of diabetic status] (Heart [2003] 24:309-321) Objectives: To analyse the relation between serum glucose concentration and hospital outcome across the whole spectrum of acute coronary syndromes. Methods: This was a prospective cohort study of 2127 patients presenting with acute coronary syndromes. The patients were stratified into quartile groups (Q1 to Q4) defined by serum glucose concentrations of 5.8, 7.2, and 10.0 mmol/l. The relation between quartile group and major in-hospital complications was analysed. Results: The proportion of patients with acute myocardial infarction increased incrementally across the quartile groups, from 21.4% in Q1 to 47.9% in Q4 (p < 0.0001). The trend for frequency of in-hospital major complications was similar, particularly left ventricular failure (LVF) (Q1 6.4%, Q4 25.2%, p < 0.0001) and cardiac death (Q1 0.7%, Q4 6.1%, p < 0.0001). The relations were linear, each glucose quartile increment being associated with an odds ratio of 1.46 (95% confidence interval (CI) 1.27 to 1.70) for LVF and 1.52 (95% CI 1.17 to 1.97) for cardiac death. Although complication rates were higher for a discharge diagnosis of acute myocardial infarction than for unstable angina, there was no evidence that the effects of serum glucose concentration were different for the two groups, there being no significant interaction with discharge diagnosis in the associations between glucose quartile and LVF (p = 0.69) or cardiac death (p = 0.17). Similarly there was no significant interaction with diabetic status in the associations between glucose quartile and LVF (p = 0.08) or cardiac death (p = 0.09). Conclusion: Admission glycaemia stratified patients with acute coronary syndromes according to their risk of in-hospital LVF and cardiac mortality. There was no detectable glycaemic threshold for these adverse effects. The prognostic correlates of admission glycaemia were unaffected by diabetic status and did not differ significantly between patients with acute myocardial infarction and those with unstable angina.

S447X (serine) substitution on lipoprotein lipase (LPL) gene associates frequently with a high-HDL/low-LDL phenotype and infrequently with a low-HDL/high-LDL phenotype (Genetic Epidemiology [2003] 24:309-321) S447X, a serine substitution by a stop codon on base 99 of exon 9 of the lipoprotein lipase (LPL) gene, has beneficial effects on blood lipids. Other LPL alleles are associated with lipid levels, but whether one of these variants predominates remains elusive. We performed a systematic survey to identify single-nucleotide polymorphisms (SNPs) in all 10 LPL exons and flanking regions by resequencing the gene in 95 subjects. Of 24 variants, 14 were common (>=3%). We assayed the common SNPs in 186 cases with atherogenic lipid profiles (low HDL, high LDL) and 185 nonatherogenic controls (high HDL, low LDL). Only S447X and exons 6 (base +73) and 10 (base -11) were individually associated with case-control status (P<0.05, adjusted for major nongenetic covariates with known lipid effects). There were no significant SNP×gender interactions. In adjusted multi-SNP and haplotypic analyses, S447X was interpretable as the sole predictor, with a 2-3-fold reduction in the odds of being atherogenic vs. nonatherogenic (adjusted OR, 0.39; 95% CI, 0.21-0.73). S447X and base -11 of exon 10 were statistically interchangeable because they are strongly associated (r=0.92, P<0.0001), but we posit that the LPL association with lipid profile is more likely attributable to the functional S447X rather than the nonfunctional exon 10 SNP. It appears that the S447X variant of LPL may be another rare example (like APOE4, factor V-Leiden, and PPAR-gamma Pro12Ala) of a common variant predisposing to a common disorder

• Fasting glucose levels appear to increase in dysmetabolic hypertensive patients who develop MI and to decrease in normotensives who do not (BMJ [2003] 326:681) Objective: To investigate the impact of an increase in blood glucose on the risk of developing myocardial infarction, with particular emphasis on people taking antihypertensive drugs. Design: Prospective population based cohort study. Setting: Uppsala, Sweden. Participants: 1860 men who had participated in 1970-3 at age 50 in a health survey aimed at identifying risk factors for cardiovascular disease and were re-examined at age 60 and then followed for 17.4 years. Main outcome measure: Myocardial infarction after age 60. Results: The incidence of myocardial infarction was significantly higher in men treated for hypertension than in those without such treatment (23% v 13.5%, P<0.0001). Participants who developed myocardial infarction after the age of 60 (n=253) showed a significantly larger increase in blood glucose between age 50 and 60 than did those without myocardial infarction. In multivariate Cox proportional hazard models increase in blood glucose was an independent risk factor for myocardial infarction (P=0.0001) in men receiving antihypertensive treatment at age 60 (n=291, mainly beta- blockers and thiazide diuretics) but not in those without such treatment. The impact of increase in blood glucose declined after inclusion of serum proinsulin concentrations at baseline but was still significant. A significant interaction existed between proinsulin concentration (a marker of insulin resistance) at baseline and antihypertensive treatment on increase in blood glucose. Conclusions: Increase in blood glucose between the ages of 50 and 60 and baseline proinsulin concentration were important risk factors for myocardial infarction in men receiving antihypertensive treatment, indicating that both an insulin resistant state and the metabolic impact of beta- blockers and diuretics increase the risk of myocardial infarction.
  

Antileukotriene drugs may be an effective treatment regimen in late-stage atherogenesis (Proc. Natl. Acad. Sci. USA, [2003] 10.1073/pnas.242716099) Oxidation products of low-density lipoproteins have been suggested to promote inflammation during atherogenesis, and reticulocyte-type 15-lipoxygenase has been implicated to mediate this oxidation. In addition, the 5-lipoxygenase cascade leads to formation of leukotrienes, which exhibit strong proinflammatory activities in cardiovascular tissues. Here, we studied both lipoxygenase pathways in human atherosclerosis. The 5-lipoxygenase pathway was abundantly expressed in arterial walls of patients afflicted with various lesion stages of atherosclerosis of the aorta and of coronary and carotid arteries. 5-lipoxygenase localized to macrophages, dendritic cells, foam cells, mast cells, and neutrophilic granulocytes, and the number of 5-lipoxygenase expressing cells markedly increased in advanced lesions. By contrast, reticulocyte-type 15-lipoxygenase was expressed at levels that were several orders of magnitude lower than 5-lipoxygenase in both normal and diseased arteries, and its expression could not be related to lesion pathology. Our data support a model of atherogenesis in which 5-lipoxygenase cascade-dependent inflammatory circuits consisting of several leukocyte lineages and arterial wall cells evolve within the blood vessel wall during critical stages of lesion development. They raise the possibility that antileukotriene drugs may be an effective treatment regimen in late-stage disease.

• Diabetic Macrovascular Disease -The Glucose Paradox? (Circulation. 2002;106:2760) -editorial by Peter Libby The specific interventions used to lower glycemia may contribute to the inability to show decreases in macrovascular end points. With some antidiabetic treatments, untoward effects may counterbalance potential benefits. Generally, interventions that increase insulin supply (eg, insulin itself and sulfonylureas) have proven less promising for limiting cardiovascular complications than those that improve glucose utilization or reduce insulin resistance. Indeed, in one arm of UKPDS, metformin monotherapy decreased MI by 39% (P0.01) in an overweight subgroup, a benefit not seen in patients requiring metformin plus sulfonylureas or insulin. Thiazolidinediones (the "glitazones") hold considerable promise as insulin sensitizers and merit careful clinical evaluation for cardiovascular benefit.

Elevated levels of acute-phase proteins and plasminogen activator inhibitor-1 predict the development of type 2 diabetes: the insulin resistance atherosclerosis study.(Diabetes 2002 Apr;51(4):1131-7) Festa A, D'Agostino R Jr, Tracy RP, Haffner SM; The Insulin Resistance Atherosclerosis Study.Elevated serum levels of acute-phase proteins, indicating chronic subclinical inflammation, have been associated with cardiovascular disease as well as the insulin resistance syndrome. Chronic inflammation may also be a risk factor for developing type 2 diabetes. We studied the concentrations of C-reactive protein (CRP), fibrinogen, and plasminogen activator inhibitor-1 (PAI-1) in 1,047 nondiabetic subjects in relation to incident diabetes within 5 years in the Insulin Resistance Atherosclerosis Study. Subjects with diabetes at follow-up (n = 144) had higher baseline levels of fibrinogen (mean +/- SD; 287.8 +/- 58.8 vs. 275.1 +/- 56.0 mg/dl; P = 0.013) as well as of CRP (median [interquartile range]; 2.40 [1.29, 5.87] vs. 1.67 mg/l [0.75, 3.41]; P = 0.0001) and PAI-1 (24 [15, 37.5] vs. 16 ng/ml [9, 27]; P = 0.0001) than nonconverters. The odds ratio (OR) of converting to diabetes was significantly increased with increasing baseline concentrations of the inflammatory markers. In contrast to PAI-1, the association of CRP and fibrinogen with incident diabetes was significantly attenuated after adjustment for body fat (BMI or waist circumference) or insulin sensitivity (S(I)), as assessed by a frequently sampled intravenous glucose tolerance test. In a logistic regression model that included age, sex, ethnicity, clinical center, smoking, BMI, S(I), physical activity, and family history of diabetes, PAI-1 still remained significantly related to incident type 2 diabetes (OR [95% CI] for 1 SD increase: 1.61 [1.20-2.16]; P = 0.002). Chronic inflammation emerges as a new risk factor for the development of type 2 diabetes; PAI-1 predicts type 2 diabetes independent of insulin resistance and other known risk factors for diabetes.

• Depolarization of Endothelial Cells Enhances Platelet Aggregation Through Oxidative Inactivation of Endothelial NTPDase (Arteriosclerosis, Thrombosis, and Vascular Biology 2002, 10.1161/01) Objective: The objective of this study was to investigate whether depolarization of cultured endothelial cells (human umbilical vein endothelial cells, HUVECs) affects their ectonucleotidase activity through superoxide (O2-) production. Methods and Results—Depolarization by the cation channel gramicidin (100 nmol/L) or tetrabutylammonium chloride (1 mmol/L) induced O2- release from HUVECs (n=4), which was decreased by superoxide dismutase (SOD, 500 U/mL). The activity of endothelial ectonucleotidases was assessed by the production of inorganic phosphate from ADP, which was decreased by chronic depolarization by 25% (n=6, P<0.05) and the amount of AMP derived from ADP in the presence of the 5'-nucleotidase inhibitor ,ß-methylene-5'-diphosphate (00 µmol/L). AMP was decreased by chronic depolarization from 0.54±0.16 to 0.39±0.11 µmol/min/mg protein (n=6, P<0.05). This was abolished in the continuous presence of SOD (n=6). NTPDase protein was predominantly expressed in HUVECs (n=4). Protein abundance, viability of cells, and apoptosis rates were not altered by depolarization (n=10). Only in the presence of depolarized HUVECs, but not with control cells or with HUVECs depolarized in the presence of SOD, did 5 µmol/L of ADP cause irreversible platelet aggregation. Increases in transmural pressure induced endothelial depolarization in intact hamster small arterioles. Conclusions—Depolarization causes the endothelial production of O2-, which inhibits the activity of endothelial ectonucleotidases. Increases in transmural pressure induce endothelial depolarization. In chronically hypertensive diseases, depolarization might favor platelet aggregation.

Nutritionally Induced Obesity Is Attenuated in Transgenic Mice Overexpressing Plasminogen Activator Inhibitor-1 (Arteriosclerosis, Thrombosis, and Vascular Biology 2002) Objective—The objective of this study was to investigate the role of plasminogen activator inhibitor-1 (PAI-1) in adipose tissue development in vivo. Methods and Results—Transgenic (Tg) mice overexpressing murine PAI-1 under control of the adipocyte promoter aP2 and wild-type (WT) controls were kept on standard food (SFD) or on high-fat diet (HFD) for 15 weeks. The body weight and the weight of the isolated subcuttaneus and gonadal fat deposits of the Tg mice kept on the HFD were significantly lower than those of the WT mice. The number of adipocytes in the adipose tissue was similar for Tg and WT mice on the HFD, but adipocyte hypotrophy and a significantly lower ratio of stroma cells/adipocytes were observed in the Tg mice. A significant negative correlation (P<0.01) was observed between expression of preadipocyte factor-1, which blocks adipocyte differentiation, and adipose tissue weight. Fasting insulin and total cholesterol levels on the HFD were lower in Tg than in WT mice. Conclusions—High circulating PAI-1 levels attenuate nutritionally induced obesity. This may be related to modifications in adipose tissue cellularity affecting weight and plasma metabolic parameters.

• Increased apoA-I catabolism is found in dysmetabolic women and is consistent with the increased risk of atherosclerosis in this population. (Int J Obes Relat Metab Disord [2002] Sep;26(9):1151-8) Pont f , Duvillard l , Florentin e , Gambert p , Verges b AIMS/HYPOTHESIS: Mechanisms responsible for the decreased high-density lipoprotein (HDL) cholesterol level associated with insulin resistance in obese patients are not clearly understood. To determine the influence of insulin resistance at an early stage on HDL metabolism, we performed a stable isotope kinetic study of apolipoprotein (apo) A-I, in five obese insulin resistant women with normal fasting triglycerides and without impaired glucose tolerance, and in five age-matched control women. METHODS: Each subject received a 16 h constant infusion of L-[1-(13)C]leucine at 0.7 mg/kg/h following a primed bolus of 0.7 mg/kg. RESULTS: ApoA-I fractional catabolic rate (FCR) was significantly increased in insulin-resistantTan en compared to controls (0.316+/-0.056 vs 0.210+/-0.040 per day, P<0.01), indicating a significant 50% increase of apoA-I catabolism, leading to an important Tan ction of plasma apoA-I residence time (3.25+/-0.59 vs 4.92+/-1.11, P<0.01). ApoA-I production rate tended to be higher in insulin resistant wTan than in controls (364+/-77 vs 258+/-60 mg/l/day, P=0.13), but the difference was not statistically significant. ApoA-I FCR was correlated with triglycerides during the fed state (r=0.69; P=0.026) and HDL triglycerides-esterified cholesterol ratio (r=0.73; P=0.016), suggesting that alteration of apoA-I metabolism in insulin resistance mTan e partly related to HDL enrichment in triglycerides. CONCLUSIONS: Our kinetic study shows that patients, at an early stage of insulin resistance (without impaired glucose tolerance nor fasting hypertriglyceridaemia), already have a significant alteration of apoA-I metabolism (increased apoA-I catabolism), which is consistent with the increased risk of atherosclerosis in this population.

Serum homocysteine, creatinine, and glucose are predictors of the severity and extent of
coronary artery disease in asymptomatic members of high-risk families (Eur J Clin Invest [2002] 32:472) Pajunen P , Syvänne Mikko , Nieminen M S, Kareinen Anu , Viitanen Laura , Lehto Seppo , Laakso Markku Background There has been no previous study to determine the severity and extent of coronary artery disease (CAD) in subjects with no diagnosis or symptoms of CAD at the time of the angiography. Methods Fifty-three subjects, who were siblings of patients with early onset CAD, underwent coronary angiography. Indices to describe per-patient characteristics of CAD were calculated, based on computer-aided quantitative coronary angiography. Clinical and laboratory characteristics were correlated to the angiographic parameters. ResultsSerum total homocysteine ( = 0·29, P < 0·05) and creatinine ( = 0·47, P = 0·001) levels were related to the global atheroma burden index. The median of the atheroma burden index was two times higher in the top homocysteine quartile compared to the lowest quartile. The overall atheroma burden index correlated significantly with the fasting blood glucose level in all subjects. Diabetes, especially when albuminuria was present, was a powerful risk factor. In a multivariate analysis, only age and sex were independent predictors of atheroma burden. ConclusionsSerum homocysteine and creatinine concentrations, and diabetes with albuminuria were found to be markers of the severity and extent of CAD in subjects of high-risk families without symptoms of CAD.

• Unsaturated fatty acids and their oxidation products stimulate CD36 gene expression in human macrophages (Atherosclerosis [2002] 164:45-56) Fatty acids (FA) have been implicated in the control of expression of several atherosclerosis-related genes. Similarly, the CD36 receptor has recently been shown to play an important role in atherosclerosis and other pathologies. The aim of the present study was to evaluate the direct effect of FA and their oxidation products (aldehydes), on the expression of CD36 in both THP-1 macrophages and human monocyte-derived macrophages (HMDM). The FA tested included the saturated FA (SFA) lauric, myristic, palmitic and stearic acid; the monounsaturated FA oleic acid; and the unsaturated FA (UFA) linoleic, arachidonic acid (AA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Aldehydes used were malondialdehyde (MDA), hexanal, 2,4-decadienal (DDE) and 4-hydroxynonenal (HNE). CD36 expression was measured by RT-PCR, Western blot and immunofluorescence. Incubation of THP-1 macrophages for 24 h with non-cytotoxic concentrations of UFA significantly increased CD36 mRNA expression. By contrast, exposure of THP-1 macrophages to SFA did not affect the levels of CD36 mRNA. Among all UFAs tested, EPA and DHA were the strongest inducers of CD36 mRNA levels, followed by oleic and linoleic acid. Incubation of HMDM with either oleic or linoleic acid significantly increased steady-state CD36 mRNA in a dose-dependent manner. Consistent with the increase of CD36 mRNA expression, incubation of THP-1 macrophages with oleic and linoleic acid for 24 h markedly increased CD36 protein expression. Treatment of THP-1 macrophages with MDA or hexanal for 24 h significantly increased CD36 mRNA expression in a dose dependent manner. In contrast, DDE and HNE significantly decreased this parameter. The data provide evidence for a direct regulatory effect of UFA on CD36 gene expression and support a role for aldehydes in the regulation of CD36 expression by FA.

Fasting and postchallenge hyperglycaemia in the early phase of an acute myocardial infarction could be used as early markers of high-risk individuals (Lancet [2002] 359: 2140-44)Background Glycometabolic state at hospital admission is an important risk marker for long-term mortality in patients with acute myocardial infarction, whether or not they have known diabetes mellitus. Our aim was to ascertain the prevalence of impaired glucose metabolism in patients without diagnosed diabetes but with myocardial infarction, and to assess whether such abnormalities can be identified in the early course of a myocardial infarction. Methods We did a prospective study, in which we enrolled 181 consecutive patients admitted to the coronary care units of two hospitals in Sweden with acute myocardial infarction, no diagnosis of diabetes, and a blood glucose concentration of less than 11·1 mmol/L. We recorded glucose concentrations during the hospital stay, and did standardised oral glucose tolerance tests with 75 g of glucose at discharge and again 3 months later. Findings The mean age of our cohort was 63·5 years (SD 9) and the mean blood glucose concentration at admission was 6·5 mmol/L (1·4). The mean 2-h postload blood glucose concentration was 9·2 mmol/L (2·9) at hospital discharge, and 9·0 mmol/L (3·0) 3 months later. 58 of 164 (35%, 95% CI 28-43) and 58 of 144 (40%, 32-48) individuals had impaired glucose tolerance at discharge and after 3 months, respectively, and 51 of 164 (31%, 24-38) and 36 of 144 (25%, 18-32) had previously undiagnosed diabetes mellitus. Independent predictors of abnormal glucose tolerance at 3 months were concentrations of HbA_1c at admission (p=0·024) and fasting blood glucose concentrations on day 4 (p=0·044). Interpretation Previously undiagnosed diabetes and impaired glucose tolerance are common in patients with an acute myocardial infarction. These abnormalities can be detected early in the postinfarction period. Our results suggest that fasting and postchallenge hyperglycaemia in the early phase of an acute myocardial infarction could be used as early markers of high-risk individuals.

• Interleukin-18 Is a Strong Predictor of Cardiovascular Death in Stable and Unstable Angina (Circulation [2002]106:24) Background— Interleukin (IL)-18 plays a central role in orchestrating the cytokine cascade and accelerates atherosclerosis and plaque vulnerability in animal models. However, epidemiological data evaluating the role of IL-18 levels in atherosclerosis are lacking. M ethods and Results— In a prospective study of 1229 patients with documented coronary artery disease, we measured baselinemarkers of serum concentrations of IL-18 and other inflammation. During the follow-up period (median, 3.9 years), 95 patients died of cardiovascular causes. Median serum concentrations of IL-18 were significantly higher among patients who had a fatal cardiovascular event than among those who did not (68.4 versus 58.7 pg/mL; P<0.0001). The hazard risk ratio of future cardiovascular death increased with increasing quartiles of IL-18 (hazard risk ratio, 1.46; 95% CI 1.21 to 1.76; P for trend <0.0001). After adjustment for most potential confounders, including the strong predictor ejection fraction as well as the inflammatory variables IL-6, high-sensitive C-reactive protein, and fibrinogen, this relation remained almost unchanged, such that patients within the highest quartile of IL-18 had a 3.3-fold increase in hazard risk compared with those in the first quartile (95% CI, 1.3 to 8.4, P=0.01). This relation was observed in patients with stable angina and patients with unstable angina at baseline. Conclusions— Serum IL-18 level was identified as a strong independent predictor of death from cardiovascular causes in patients with coronary artery disease regardless of the clinical status at admission. This result strongly supports recent experimental evidence of IL-18–mediated inflammation leading to acceleration and vulnerability of atherosclerotic plaques.

Ang II provokes TNF biosynthesis in the adult mammalian heart through a PKC-dependent pathway. (Circulation [2002] 105:2198.)Background— Previous studies suggest that angiotensin II (Ang II) upregulates the expression of tumor necrosis factor (TNF) in nonmyocyte cell types; however, the effect of Ang II on TNF expression in the adult mammalian heart is not known. Methods and Results— To determine whether Ang II was sufficient to provoke TNF biosynthesis in the adult heart, we examined the effects of Ang II in isolated buffer-perfused Langendorff feline hearts. Ang II (10^-7 mol/L) treatment resulted in a time- and dose-dependent increase in myocardial TNF mRNA and protein biosynthesis in the heart as well as in cultured adult cardiac myocytes. The effects of Ang II on myocardial TNF mRNA and protein synthesis were mediated through the angiotensin type 1 receptor (AT₁R), insofar as an AT₁R antagonist (AT₁a) blocked the effects of Ang II, whereas an angiotensin type 2 receptor (AT₂R) antagonist (AT₂a) had no effect. Stimulation with Ang II led to the activation of nuclear factor-kappaB and activator protein-1 (AP-1), two transcription factors that are important for TNF gene expression. Nuclear factor-kappa-B-alpha activation was accompanied by phosphorylation of Ikappa B^alpha on serine 32 as well as degradation of IkappaB-alpha, suggesting that the effects of Ang II were mediated through an IkappaBalpha-dependent pathway. The important role of protein kinase C (PKC) was suggested by studies in which a phorbol ester triggered TNF biosynthesis, and a PKC inhibitor abrogated Ang II–induced TNF biosynthesis. Conclusions— These studies suggest that Ang II provokes TNF biosynthesis in the adult mammalian heart through a PKC-dependent pathway.

• Proinsulin Is an Independent Predictor of Coronary Heart Disease (Circulation [2002] 105:2153) Background— Some, but not all, studies have reported a relationship between plasma insulin and coronary heart disease (CHD). Conventional nonspecific insulin assays are also measuring various fractions of proinsulin-like molecules due to cross-reactivity. The long-term relationship between proinsulin-like molecules and CHD is largely unknown. For this reason, the longitudinal relationships between intact proinsulin, split proinsulin, specific insulin, immunoreactive insulin, and CHD, were studied in a population-based cohort of 50-year-old men (n=874), with a follow-up of 27 years. Methods and Results— Fasting proinsulin-like molecule and specific-insulin concentrations were measured in plasma (stored frozen since baseline 1970 to 1973) by specific 2-site immunometric assays. Immunoreactive insulin concentrations were determined at baseline. The associations between proinsulin-like molecules, specific insulin, immunoreactive insulin, and CHD mortality (International Classification of Diseases [9th revision] codes 410 to 414) were analyzed using Cox’s proportional hazards regression and presented as hazard ratios (HRs) with their 95% confidence intervals (CIs) for a 1-SD increase in a predictor variable. In the univariate analysis, intact proinsulin (HR, 1.69; 95% CI, 1.41 to 2.01) was the strongest predictor of death from CHD. In the multivariate analysis, smoking (HR, 1.57; 95% CI, 1.03 to 2.38), intact proinsulin (HR, 1.47; 95% CI, 1.18 to 1.82), systolic blood pressure (HR, 1.38; 95% CI, 1.14 to 1.66), and LDL/HDL cholesterol ratio (HR, 1.31; 95% CI, 1.12 to 1.53) were independent predictors of CHD mortality (adjusted for body mass index, triglycerides, and fasting glucose), whereas specific insulin and immunoreactive insulin were not (HR, 1.12; 95% CI, 0.90 to 1.40). The increased risk was restricted to the upper third of the proinsulin distribution. Conclusion— Increased proinsulin concentrations predict death and morbidity caused by CHD over a period of 27 years, independent of other major cardiovascular risk factors

Atrial Natriuretic Peptide Reduces TNF-Induced Actin Polymerization and Endothelial Permeability (Circulation Research [2002]90:874) The atrial natriuretic peptide (ANP) is a cardiovascular hormone possessing antiinflammatory potential due to its inhibitory action on the production of inflammatory mediators, such as tumor necrosis factor-alpha (TNF-alpha). The aim of this study was to determine whether ANP is able to attenuate inflammatory effects of TNF-alpha on target cells. Human umbilical vein endothelial cells (HUVECs) were treated with TNF-alpha in the presence or absence of ANP. Changes in permeability, cytoskeletal alterations, phosphorylation of p38 MAPK and HSP27, and expression of MKP-1 were determined by macromolecule permeability assay, fluorescence labeling, RT-PCR, and immunoblotting. Antisense studies were done by transfecting cells with MKP-1 antisense oligonucleotides. Activation of HUVECs with TNF-alpha lead to a significant increase of macromolecule permeability and formation of stress fibers. Treatment of cells with ANP (10(-8) to 10(-6) mol/L) significantly reduced the formation of stress fibers and elevated permeability. Both TNF-alpha-induced effects were shown to be mediated via the activation of p38 using SB203580, a specific inhibitor of p38. ANP significantly reduced the TNF-alpha-induced activation of p38 and attenuated the phosphorylation of HSP27, a central target downstream of p38. ANP showed no effect on p38 upstream kinases MKK3/6. However, a significant induction of the MAPK phosphatase MKP-1 mRNA and protein could be observed in ANP-treated cells. Antisense experiments proved a causal role for MKP-1 induction in the ANP-mediated inhibition of p38. These data show the inhibitory action of ANP on TNF-alpha-induced changes in endothelial cytoskeleton and macromolecule permeability involving an MKP-1-induced inactivation of p38 MAPK. These effects point to an antiinflammatory and antiatherogenic potential of this cardiovascular hormone.

• The Kir6.1-containing K+ channel is critical in the regulation of vascular tonus, especially in the coronary arteries, and its disruption may cause Prinzmetal angina. (Nature Med [2002] 8:466-472) The inwardly rectifying K⁺ channel Kir6.1 forms K⁺ channels by coupling with a sulfonylurea receptor in reconstituted systems, but the physiological roles of Kir6.1-containing K⁺ channels have not been determined. We report here that mice lacking the gene encoding Kir6.1 (known as Kcnj8) have a high rate of sudden death associated with spontaneous ST elevation followed by atrioventricular block as seen on an electrocardiogram. The K⁺ channel opener pinacidil did not induce K⁺ currents in vascular smooth-muscle cells of Kir6.1-null mice, and there was no vasodilation response to pinacidil. The administration of methylergometrine, a vasoconstrictive agent, elicited ST elevation followed by cardiac death in Kir6.1-null mice but not in wild-type mice, indicating a phenotype characterized by hypercontractility of coronary arteries and resembling Prinzmetal (or variant) angina in humans. The Kir6.1-containing K⁺ channel is critical in the regulation of vascular tonus, especially in the coronary arteries, and its disruption may cause Prinzmetal angina.

Insulin resistance in moderate chronic heart failure is related to hyperleptinaemia, but not to norepinephrine or TNF-alpha (International Journal of Cardiology [2002] 83:73-81) Objectives: Chronic heart failure (CHF) has emerged as an insulin-resistant state, independently of ischaemic aetiology. The underlying mechanisms of this finding are not known. Catecholamines, tumor necrosis factor alpha (TNF) and leptin, the adipocyte specific hormone, have all been implicated as mediators of impaired insulin sensitivity. The purpose of this study was to examine in patients with CHF and in comparison to healthy controls subjects whether norepinephrine, TNF or leptin relate to insulin sensitivity. Design: 41 patients with CHF (age 60±2 years, NYHA I/II/III/IV 4/12/22/3, peak oxygen consumption 17.6±1.0 ml/kg per min) and 21 healthy controls of similar age and total and regional fat distribution were studied in a cross-sectional study. Insulin sensitivity was assessed by intravenous glucose tolerance testing using the minimal model approach; catecholamines, TNF and soluble TNF receptors 1 and 2 were also measured. Total and regional body fat mass was assessed by dual energy X-ray absorptiometry. Results: Insulin sensitivity was reduced in CHF patients compared to controls by 31% (P<0.01) and fasting insulin was higher in patients than in controls (79.1±9.7 vs. 41.4±6.0 pmol/l, P<0.01). Patients had, compared to healthy controls, elevated serum leptin levels (8.28±0.84 vs. 4.83±0.68 ng/ml), norepinephrine (3.45±0.34 vs. 1.87±0.16 nmol/l, both P<0.01) and soluble TNF-receptors 1 (1280±141 vs. 639±52 pg/ml) and 2 (2605±184 vs. 1758±221 pg/ml, both P<0.01). Leptin levels corrected for total body fat mass were higher in CHF patients than in controls (41.3±3 vs. 24.3±2 pg/ml per 100 g, P<0.001). TNF was not significantly different between the groups. In both groups there was an inverse correlation between insulin sensitivity and serum leptin (r=-0.65, P<0.0001 for pooled subjects); in contrast, no significant relation was found between insulin sensitivity and norepinephrine or TNF. In multivariate regression analysis, leptin emerged as the only significant predictor of insulin sensitivity (standardised COEFFICIENT=-0.59, P<0.001), independent of body fat mass, age and peak V₂. Conclusion: In moderate CHF, elevated leptin levels directly and independently predict insulin resistance. Elevated serum leptin levels could play a role in the impaired regulation of energy metabolism in CHF. In contrast to observations in other conditions, TNF and norepinephrine are not related to insulin resistance in moderate CHF.

• Leptin contributes to arterial thrombosis following vascular injury in vivo (JAMA. [2002] 287:1706-1709) Context  Complications of atherosclerosis are the leading cause of morbidity and mortality in industrialized societies. Obesity has emerged as an independent risk factor for complications of atherosclerotic vascular disease. Leptin, a hormone produced by the adipocyte, increases with obesity and appears to modulate energy balance and food intake. In addition, other actions of leptin have been proposed, including an in vitro effect on platelet aggregation. Thus, the elevated plasma leptin levels in obese individuals may promote vascular thrombosis. Objective  To test the hypothesis that leptin contributes to in vivo thrombosis via the leptin receptor. Design and Materials  Between September 2000 and September 2001, a vascular thrombosis model was used to test male 10- to 12-week-old mice completely deficient in leptin or the leptin receptor and mice with platelet leptin-receptor deficiency. Main Outcome Measure  Time to formation of an occlusive thrombus in the common carotid artery following experimentally induced endothelial injury. Results  Following onset of vascular injury, wild-type mice (n = 8) formed occlusive thrombosis in a mean (SD) of 42.2 (4.6) minutes, whereas leptin-deficient (n = 5) and leptin receptor–deficient mice (n = 7) formed occlusive thrombosis in 75.2 (10.1) and 68.6 (10.3) minutes, respectively (leptin deficient vs wild-type mice, P = .008; leptin-receptor–deficient vs wild-type, P = .03). When recombinant murine leptin was administered to leptin-deficient mice (n = 4), the time to occlusion was reduced to 41.8 (6.6) minutes (P = .035 vs vehicle control). Following bone marrow transplantation from leptin receptor–deficient (donor) mice to wild-type (recipient) mice, the time to occlusion was prolonged from 22.3 (2.8) minutes in wild-type mice receiving wild-type marrow (n = 3) to 56.8 (5.0) minutes in wild-type mice receiving leptin receptor–deficient bone marrow (n = 5) (P = .003). Conclusions  Leptin contributes to arterial thrombosis following vascular injury in vivo and these prothrombotic effects appear to be mediated through the platelet leptin receptor.

Increased Serum Levels of Macrophage Migration Inhibitory Factor-Related Protein Is a Sensitive Marker for Acute Coronary Syndrome in Patients With Coronary Artery Disease (ACC 2002) BACKGROUND: In patients with coronary artery disease (CAD), it is important to discriminate acute coronary syndrome (ACS). There is accumulating data that ACS relates to recent activation of inflammation affecting atherosclerotic plaques. Macrophage migration inhibitory factor-related protein (MRP) is a calcium-binding protein (heterodimer), which is expressed in infiltrate macrophages during inflammatory reactions. The purpose of this study was to investigate whether MRP is useful for the diagnosis of ACS. Methods: We studied 92 patients with angiographically proven CAD. They comprised two groups [Group ACS, Braunwald's subclass II or III of unstable angina, and acute myocardial infarction, n=53; Group SA, stable angina, n=39]. We purified MRP from human leukocytes and then prepared rabbit monoclonal antibodies against the MRP. Serum concentrations of MRP were measured using a newly-developed enzyme-linked immunosorbent assay system. In addition, we measured serum concentrations of C-reactive protein (CRP). RESULTS: There were no significant differences in age, gender and coronary risk factors between the two groups. Serum MRP levels were significantly higher in Group ACS than in Group SA [3.25±3.08 (SD) microgram/ml vs. 0.77±0.31 microgram/ml, p < 0.0001]. Serum CRP levels were also significantly higher in Group ACS than in Group SA (3.11±6.14 mg/dl vs. 0.19±0.20 mg/dl, p=0.0039). Sensitivity and specificity of positive serum MRP levels (> 1.2 microgram/ml) for detection of ACS were 84.9% and 89.7%, respectively. On the other hand, sensitivity and specificity of positive serum CRP levels (> 0.5mg/dl) for detection of ACS were 52.8% and 92.3%, respectively. Sensitivity was significantly (p < 0.05) higher in the measurements of MRP than in the measurements of CRP, while specificity was comparable in the two markers. Conclusion: The measurement of serum MRP levels is useful for the discrimination of ACS in patients with CAD.

• High fibrinogen levels associated with high Lp(a) levels significantly increases the risk of CHD. (Am J Cardiol [2002] 89:662-666)Fibrinogen has been prospectively found to correlate with coronary heart disease (CHD) but a similar association has not been well established for lipoprotein (a) (Lp(a)). Plasma lipids, Lp(a), and fibrinogen levels were measured in 2,125 men (aged 47 to 76 years) who were free of clinical CHD. During a 5-year follow-up period, 116 first CHD events were documented. Men with CHD were older, smoked more, had a higher prevalence of diabetes, and higher levels of systolic blood pressure, cholesterol, low-density lipoprotein cholesterol, Lp(a), and fibrinogen, and lower plasma high-density lipoprotein cholesterol levels. Only fibrinogen levels in the upper tertile of the distribution compared with the lower tertiles were associated with a significant risk of CHD (adjusted risk ratio 2.5; 95% confidence interval [CI] 1.4 to 4.2; P = 0.0010). Such an association was not observed with Lp(a). To assess a possible relation between fibrinogen and Lp(a) to the risk of CHD events, men were assigned to 1 of 4 groups according to fibrinogen median levels and a Lp(a) cut-off level of 300 mg/L: group 1: fibrinogen <4.05 g/L and Lp(a) <300 mg/L; group 2: fibrinogen <4.05 g/L and Lp(a) 300 mg/L; group 3: fibrinogen 4.05 g/L and Lp(a) <300 mg/L; and group 4: fibrinogen 4.05 g/L and Lp(a) 300 mg/L. Using group 1 as a reference, a significant risk ratio was only documented in group 4 (2.5; 95% CI 1.2 to 5.1; P = 0.0132). In this population, high fibrinogen levels associated with high Lp(a) levels significantly increased the risk of CHD

Patients with chronic heart failure exhibit significant metabolic insulin resistance. (Am J Cardiol [2002] 89:696-703) Chronic heart failure (HF) is associated with insulin resistance. Putative mechanisms of insulin resistance are abnormal skeletal muscle blood flow and antagonism of insulin action due to sympathetic nervous system activation. We measured insulin sensitivity, the vasoactive properties of insulin, and the association between insulin resistance and markers of neurohormonal activation in 10 patients with chronic HF and in 9 healthy controls. Noninvasive hemodynamic measurements and an hyperinsulinemic, euglycemic clamp were used. Patients were insulin resistant compared with the controls (p <0.05 for area under insulin dose-response curve). Insulin infusion led to a selective increase in forearm blood flow accompanied by a decrease in mean arterial pressure and superior mesenteric blood flow. Heart rate decreased in patients but not in controls; however, when baseline measurements were controlled for, there was no difference in the overall hemodynamic response to insulin infusion between the study groups. In univariate analysis, age, serum creatinine, fasting insulin, and triglyceride levels correlated inversely with insulin sensitivity (p <0.05 for all). Cardiac output had a significant correlation with insulin sensitivity (p <0.05). On stepwise multiple linear regression analysis, only age and fasting plasma insulin emerged as significant predictors of insulin sensitivity (R² 0.613, P = 0.001). In particular, we found no evidence of a relation between insulin sensitivity and plasma noradrenaline. Patients with chronic HF exhibit significant metabolic insulin resistance. Insulin resistance is not secondary to failure of insulin-mediated vasodilatation or sympathetic nervous system activation and is likely due to abnormalities at the level of the skeletal myocyte

• A significant relationship exists between insulin resistance and plasma concentrations of asymmetric dimethylarginine (ADMA) (JAMA [2002] 287: 1420-1426) Context  Increased levels of asymmetric dimethylarginine (ADMA) are associated with endothelial dysfunction and increased risk of cardiovascular disease. Several cardiovascular risk factors are associated with reduced sensitivity to insulin, but elevated ADMA concentrations have not been fully linked to the metabolic syndrome.Objective  To evaluate the relationship between insulin sensitivity and plasma ADMA concentrations, and to determine whether a pharmacological treatment that increases insulin sensitivity would also modulate ADMA concentrations.Design, Setting, and Subjects  Cross-sectional study, containing a nonrandomized controlled trial component, of 64 healthy volunteers without diabetes (42 women, 22 men; 48 with normal blood pressure and 16 with hypertension), which was conducted at a university medical center between October 2000 and July 2001.Intervention  Rosiglitazone (4 mg/d for 4 weeks and then 4 mg twice daily for 8 weeks), an insulin-sensitizing agent, was given to 7 insulin-resistant subjects with hypertension. These subjects were studied before and after 12-week treatment.Main Outcome Measures  Insulin sensitivity measured by the insulin suppression test, and fasting plasma levels of low-density lipoprotein cholesterol, triglycerides, high-density lipoprotein cholesterol, glucose, insulin, and ADMA concentrations.Results  Plasma ADMA concentrations were positively correlated with impairment of insulin-mediated glucose disposal in nondiabetic, normotensive subjects (r = 0.73; P<.001). Consistent with the metabolic syndrome, ADMA levels were also positively correlated with fasting triglyceride levels (r = 0.52; P<.001) but not with low-density lipoprotein cholesterol levels (r = 0.19; P = .20). Plasma ADMA concentrations increased in insulin-resistant subjects independent of hypertension. Pharmacological treatment improved insulin sensitivity and reduced mean (SD) plasma ADMA concentrations from 1.50 (0.30) to 1.05 (0.33) µmol/L (P = .001).Conclusion  A significant relationship exists between insulin resistance and plasma concentrations of ADMA. Pharmacological intervention with rosiglitazone enhanced insulin sensitivity and reduced ADMA levels. Increases in plasma ADMA concentrations may contribute to the endothelial dysfunction observed in insulin-resistant patients.

The concentrations of proinsulin-like molecules provide a better way to predict the incidence of CHD than those of insulin (Diabetologia [2002] 45:327-336 Aims/hypothesis: Higher concentrations of insulin correlate with several coronary heart disease (CHD) risk factors and have been shown to predict incident CHD in several studies, leading to hypotheses concerning the proatherogenic properties of insulin. However, in cross-sectional studies, relationships of concentrations of the insulin precursor molecules, proinsulin and des 31, 32 proinsulin, relate as strongly, or more strongly, to levels of risk factors and to (prevalent) CHD. Methods: We investigated the relationship between concentrations of insulin, measured with a specific assay, and of proinsulin-like molecules, and risk factors in 1181 non-diabetic men 50-64 years old during Phase II of the Caerphilly Study. We also related concentrations of these molecules to incident CHD during the 10-14 years follow-up. Results: The relationship between concentrations of insulin, of proinsulin and of des 31, 32 proinsulin and BMI (r = 0.36-0.45), triglyceride (r = 0.25-0.31), high density lipoprotein- (HDL-) cholesterol (r = -0.17 to -0.21), systolic (r = 0.05-0.11) and diastolic blood pressure (r = 0.11-0.15) were similarly close, those with risk factors being somewhat and similarly reduced after adjustment for BMI. The correlation between insulin and of proinsulin-like molecules and those plasminogen activator inhibitor-1 (PAI-1) antigen was also similar (r = 0.28-0.29). There was a negative correlation between concentrations of proinsulin-like molecules - but not insulin - and birth weight. Insulin concentrations correlated positively with height (r = 0.12). In logistic regression models, concentrations of proinsulin-like molecules, but not insulin, predicted incident of CHD over a follow-up of 10-14 years (insulin - standardised odds ratio (SOR) 1.30 (95 %-CI) 0.91, 1.85), p = 0.15; des 31, 32 proinsulin - SOR 1.38 (95 %-CI 1.02, 1.85), p = 0.034; sum of proinsulin-like molecules - SOR 1.54 (95 %-CI 1.07, 2.20), p = 0.019 after adjusting for age and BMI. The predictive ability of these molecules was reduced by around one third after adjustment for standard risk factors and concentrations of tryglyceride and HDL-cholesterol, and by about half after further adjustment for PAI-1 concentrations. Conclusion/ interpretation: We conclude that concentrations of proinsulin-like molecules provide a better way to predict the incidence of CHD than those of insulin. However, the lack of biological evidence for a causative relationship suggests an association through a common antecedent, and this antecedent is not likely to be intrauterine growth retardation. [Diabetologia (2002) 45: 327-336]

• Proinsulin is more strongly and consistently associated with CHD than is intact insulin in older nondiabetic men and women (Circulation. [2002]105:1311.) Background— Insulin or insulin resistance is considered a coronary heart disease (CHD) risk factor, but proinsulin may have a stronger association with CHD than insulin. The role of sex differences in this association is unclear. We examined the cross-sectional association of proinsulin and insulin with CHD in older men and women without diabetes. Methods and Results— A cross-sectional study of community-dwelling men (n=554) and women (n=902), 50 to 97 years of age, without diabetes by history or oral glucose tolerance test, was done between 1992 and 1996; plasma levels of intact insulin, proinsulin, and C-peptide were measured by radioimmunoassay. Based on questionnaire, medical history, or ECG abnormalities, 25% of men (n=136) and 24% of women (n=214) had prevalent CHD. All insulin variables were positively correlated with CHD risk factors. Compared with those without CHD, men and women with CHD had significantly higher levels of proinsulin. Women but not men with CHD also had higher levels of C-peptide and fasting and postchallenge insulin. Only proinsulin was significantly and independently associated with prevalent CHD in both men (OR=2.41, 1.42 to 4.11) and women (OR=1.80, 1.22 to 2.64) (adjusted for age, body mass index, systolic blood pressure, and HDL cholesterol). Similar analyses for fasting and postchallenge intact insulin and for C-peptide showed that among these three variables, only postchallenge insulin was significantly associated with CHD, and only in women. Conclusions— In older nondiabetic men and women, proinsulin was more strongly and consistently associated with CHD than was intact insulin.

Chronic vitamin C deficiency does not influence the initiation or progression of atherosclerotic plaques but severely compromises collagen deposition and induces a type of plaque morphology that is potentially vulnerable to rupture (Circulation. [2002];105:1485.) Background— Oxidative stress is thought to play an important role in atherogenesis, suggesting that antioxidants could prevent coronary artery disease. However, the efficacy of vitamin C in reducing atherosclerosis is debatable in humans and has not been tested rigorously in animals. Methods and Results— Gulo^-/-Apoe^-/- mice were used to test a hypothesis that chronic vitamin C deficiency enhances the initiation and development of atherosclerosis. These mice are dependent on dietary vitamin C because of the lack of L-gulonolactone-gamma-oxidase and are prone to develop atherosclerosis because of lacking apolipoprotein E. Beginning at 6 weeks of age, the Gulo^-/-Apoe^-/- mice were fed regular chow or Western-type diets containing high fat and supplemented with either 0.033 g or 3.3 g/L of vitamin C in their drinking water. This regimen produced mice with chronically low vitamin C (average 1.5 µg/mL in plasma) or high vitamin C (average 10 to 30 µg/mL in plasma). Morphometric analysis showed that within each sex, age, and diet group, the sizes of the atherosclerotic plaques were not different between low vitamin C mice and high vitamin C mice. However, advanced plaques in the low vitamin C mice had significantly reduced amounts of Sirius red–staining collagen (36.4±2.2% versus 54.8±2.3%, P<0.0001), larger necrotic cores within the plaques, and reduced fibroproliferation and neovascularization in the aortic adventitia. Conclusions— Chronic vitamin C deficiency does not influence the initiation or progression of atherosclerotic plaques but severely compromises collagen deposition and induces a type of plaque morphology that is potentially vulnerable to rupture.

• TNF-[alpha] concentration is associated with degrees of early atherosclerosis (European Heart Journal [2002] 23(5):376-383) Aims Tumour necrosis factor-[alpha] (TNF-[alpha]) is a proinflammatory cytokine, which is implicated in some metabolic disorders and may play a role in the development of cardiovascular disease. We examined whether plasma TNF-[alpha] is related to established cardiovascular risk indicators, plasma levels of soluble cellular adhesion molecules and carotid artery intima-media thickness determined by ultrasound examination in a population-based cohort of 96 healthy 50-year-old men. Methods and Results TNF-[alpha] and cellular adhesion molecules were measured with enzyme-linked immunosorbent assays. Plasma TNF-[alpha] concentration was associated with systolic and diastolic blood pressure, degrees of alimentary lipaemia, plasma very low density lipoprotein triglyceride, low density lipoprotein (LDL) cholesterol concentrations and peak LDL particle size. Two indices of insulin resistance as well as all soluble cellular adhesion molecules correlated positively with TNF-[alpha]. The plasma TNF-[alpha] concentration was associated with common carotid intima-media thickness in univariate analysis. In contrast, soluble E-selectin and postprandial triglycerides, but not TNF-[alpha], were independent determinants of common carotid intima-media thickness. Conclusion The plasma TNF-[alpha] concentration is associated with degrees of early atherosclerosis and correlates with metabolic and cellular perturbations that are considered important for the vascular process.

In CHD, vWF and tPAag -but not thrombomodulin - are independent markers of atherosclerosis. (Thrombosis Research [2002] 105(1):25-31) von Willebrand factor (vWF), thrombomodulin and tissue plasminogen activator antigen (tPAag) are regarded as markers of endothelial activation and/or damage. Elevated levels have been associated with atherosclerotic disease states. The aim of the present study was to compare the levels of vWF, thrombomodulin and tPAag in patients with coronary heart disease (CHD) and matched healthy individuals to see if they discriminated significantly between the study groups also after adjustment for established CHD risk factors. Patients (n=193) in various stages of CHD and matched controls (n=193) were included. To evaluate possible influence of acute phase reaction, reinvestigation was performed after 6 months. We observed elevated levels of vWF (P<.001) and tPAag (P<.001) but not thrombomodulin (P=.082) in CHD patients when compared to controls, still statistically significant after 6 months and also after adjustment for established risk factors. Our results indicate that vWF and tPAag but not thrombomodulin in the present population are independent markers of atherosclerosis.

• Perilipin Gene is Potentially Involved in the Rupture of Human Atherosclerotic Plaques (Circulation Research. [2001] 89:547.) Although rupture of an atherosclerotic plaque is the major cause of acute vascular occlusion, the exact molecular mechanisms underlying this process are still poorly understood. In this study, we used suppression subtractive hybridization to make an inventory of genes that are differentially expressed in whole-mount human stable and ruptured plaques. Two libraries were generated, one containing 3000 clones upregulated and one containing 2000 clones downregulated in ruptured plaques. Macroarray analysis of 500 randomly chosen clones showed differential expression of 45 clones. Among the 25 clones that showed at least a 2-fold difference in expression was the gene of perilipin, upregulated in ruptured plaques, and the genes coding for fibronectin and immunoglobulin lambda-chain, which were downregulated in ruptured plaques. Reverse transcriptase–polymerase chain reaction analysis on 10 individual ruptured and 10 individual stable plaques showed a striking consistency of expression for the clones SSH6, present in 8 ruptured and 2 stable plaques, and perilipin, expressed in 8 ruptured plaques and completely absent in stable plaques. Localization studies of both perilipin mRNA and protein revealed expression in cells surrounding the cholesterol clefts and in foam cells of ruptured atherosclerotic plaques. No expression was observed in nondiseased artery, and only a few cells in the shoulder region of stable plaques tested positive for perilipin. In conclusion, this study shows that it is possible to identify genes that are differentially expressed in whole-mount stable or ruptured atherosclerotic plaques. This approach may yield several potential regulators of plaque destabilization.
  

Perilipin ablation results in a lean mouse with aberrant adipocyte lipolysis, enhanced leptin production [per adipocyte mass], a resistance to diet-induced obesity, as well as a tendency toward insulin resistance (Proc. Natl. Acad. Sci. USA [2001] 98: 6494-6499) Perilipin coats the lipid droplets of adipocytes and is thought to have a role in regulating triacylglycerol hydrolysis. To study the role of perilipin in vivo, we have created a perilipin knockout mouse. Perilipin null (peri(-/-)) and wild-type (peri(+/+)) mice consume equal amounts of food, but the adipose tissue mass in the null animals is reduced to approximately 30% of that in wild-type animals. Isolated adipocytes of perilipin null mice exhibit elevated basal lipolysis because of the loss of the protective function of perilipin. They also exhibit dramatically attenuated stimulated lipolytic activity, indicating that perilipin is required for maximal lipolytic activity. Plasma leptin concentrations in null animals were greater than expected for the reduced adipose mass. The peri(-/-) animals have a greater lean body mass and increased metabolic rate but they also show an increased tendency to develop glucose intolerance and peripheral insulin resistance. When fed a high-fat diet, the perilipin null animals are resistant to diet-induced obesity but not to glucose intolerance. The data reveal a major role for perilipin in adipose lipid metabolism and suggest perilipin as a potential target for attacking problems associated with obesity.

• Absence of perilipin results in leanness and reverses obesity in Lepr(db/db) mice (Nat Genet [2000] 26::474-9) Obesity is a disorder of energy balance. Hormone-sensitive lipase (HSL) mediates the hydrolysis of triacylglycerol, the major form of stored energy in the body. Perilipin (encoded by the gene Plin), an adipocyte protein, has been postulated to modulate HSL activity. We show here that targeted disruption of Plin results in healthy mice that have constitutively activated fat-cell HSL. Plin -/- mice consume more food than control mice, but have normal body weight. They are much leaner and more muscular than controls, have 62% smaller white adipocytes, show elevated basal lipolysis that is resistant to beta-adrenergic agonist stimulation, and are cold-sensitive except when fed. They are also resistant to diet-induced obesity. Breeding the Plin -/- alleles into Leprdb/db mice reverses the obesity by ncreasing the metabolic rate of the mice. Our results demonstrate a role for perilipin in reining in basal HSL activity and regulating lipolysis and energy balance; thus, agents that inactivate perilipin may prove useful as anti-obesity medications.
  

Triglycerides >200 mg/dl (But Not Triglyceride Reduction) Strongly Associate with Stroke Risk in BIP (Circulation [2001] 104:2892)Background— Despite unclear associations between blood lipids, including fractionated cholesterol and triglycerides, and stroke, recent evidence demonstrates that lipid-modifying agents decrease the risk of stroke in patients with coronary heart disease (CHD). Methods and Results— Patients with documented CHD who were screened for but not included in the Bezafibrate Infarction Prevention study and had no history of stroke or transient ischemic attack (TIA) (n=11 177) were followed up. At baseline, medical histories were obtained and blood lipids assessed at a central study laboratory. During a 6- to 8-year follow-up period, 941 patients were identified as having nonhemorrhagic cerebrovascular disease, of whom 487 had verified ischemic stroke (per clinical findings and brain CT) or TIA. Patients experiencing an ischemic stroke/TIA had higher mean levels of triglycerides, lower levels of HDL cholesterol, and lower percentages of cholesterol contained in the HDL cholesterol moiety (%HDL; P<0.01 for all). In a logistic regression model, the adjusted ORs for developing an ischemic stroke/TIA were 1.27 (95% CI 1.01 to 1.60) associated with triglycerides >200 mg/dL and 0.87 (95% CI 0.78 to 0.97) associated with a 5% decrease in %HDL. The increased risk associated with high triglycerides was found across subgroups of age, sex, patient characteristics, and cholesterol fractions. Conclusions— High triglycerides constitute an independent risk factor for ischemic stroke/TIA across subgroups of age, sex, patient characteristics, and cholesterol fractions, whereas high %HDL was an independent protective factor among patients with CHD. These findings support the role of blood lipids, including triglycerides, as important modifiable stroke risk factors.

• A major Israeli study of 3090 patients - the Bezafibrate Infarction Prevention trial - showed no significant reduction in fatal or non-fatal MI or sudden death from triglyceride lowering or HDL raising (Circulation [2000] 102:21)Background—Coronary heart disease patients with low high-density lipoprotein cholesterol (HDL-C) levels, high triglyceride levels, or both are at an increased risk of cardiovascular events, but the clinical impact of raising HDL-C or decreasing triglycerides remains to be confirmed. Methods and Results—In a double-blind trial, 3090 patients with a previous myocardial infarction or stable angina, total cholesterol of 180 to 250 mg/dL, HDL-C 45 mg/dL, triglycerides <300 mg/dL, and low-density lipoprotein cholesterol <180 mg/dL were randomized to receive either 400 mg of bezafibrate per day or a placebo; they were followed for a mean of 6.2 years. The primary end point was fatal or nonfatal myocardial infarction or sudden death. Bezafibrate increased HDL-C by 18% and reduced triglycerides by 21%. The frequency of the primary end point was 13.6% on bezafibrate versus 15.0% on placebo (P=0.26). After 6.2 years, the reduction in the cumulative probability of the primary end point was 7.3%, (P=0.24). In a post hoc analysis in the subgroup with high baseline triglycerides (>200 mg/dL), the reduction in the cumulative probability of the primary end point by bezafibrate was 39.5% (P=0.02). Total and noncardiac mortality rates were similar, and adverse events and cancer were equally distributed. Conclusions—Bezafibrate was safe and effective in elevating HDL-C levels and lowering triglycerides. An overall trend in a reduction of the incidence of primary end points was observed. The reduction in the primary end point in patients with high baseline triglycerides (>200 mg/dL) requires further confirmation.

Elevated plasma CRP levels significantly predict the risk of future ischemic stroke and TIA in the elderly and independently of other cardiovascular risk factors. (Stroke. [2001] 32:2575) Background—— The role of C-reactive protein (CRP) as a novel plasma marker of atherothrombotic disease is currently under investigation. Previous studies have mostly related CRP to coronary heart disease, were often restricted to a case-control design, and failed to include pertinent risk factors to evaluate the joint and net effect of CRP on the outcome. We related plasma CRP levels to incidence of first ischemic stroke or transient ischemic attack (TIA) in the Framingham Study original cohort. Methods—— There were 591 men and 871 women free of stroke/TIA during their 1980 to 1982 clinic examinations, when their mean age was 69.7 years. CRP levels were measured by using an enzyme immunoassay on previously frozen serum samples. Analyses were based on sex-specific CRP quartiles. Risk ratios (RRs) were derived, and series of trend analyses were performed. Results—— During 12 to 14 years of follow-up, 196 ischemic strokes and TIAs occurred. Independent of age, men in the highest CRP quartile had 2 times the risk of ischemic stroke/TIA (RR=2.0, P=0.027), and women had almost 3 times the risk (RR=2.7, P=0.0003) compared with those in the lowest quartile. Assessment of the trend in risk across quartiles showed unadjusted risk increase for men (RR=1.347, P=0.0025) and women (RR=1.441, P=0.0001). After adjustment for smoking, total/HDL cholesterol, systolic blood pressure, and diabetes, the increase in risk across CRP quartiles remained statistically significant for both men (P=0.0365) and women (P=0.0084). Conclusions—— Independent of other cardiovascular risk factors, elevated plasma CRP levels significantly predict the risk of future ischemic stroke and TIA in the elderly.

• Pravastatin Treatment Removes the Increased Effect of LARGE Low-Density Lipoprotein Size upon Coronary Events (JAMA. [2001] 286:1468-1474) Context  Small low-density lipoprotein (LDL) particle size has been hypothesized to be a risk factor for coronary heart disease (CHD). Animal models link large LDL to atherosclerosis. However, the strong association between small LDL and other risk factors, particularly triglyceride levels, impedes determining whether LDL size independently predicts CHD in humans.Objective  To examine whether LDL size is an independent predictor of recurrent coronary events in patients with known CHD, as opposed to a marker for other lipid abnormalities.Design and Setting  Prospective, nested case-control study in the Cholesterol and Recurrent Events (CARE) trial, a randomized placebo-controlled trial of pravastatin conducted in 1989-1996.Participants  Survivors of myocardial infarction with typical LDL concentrations (416 cases and 421 controls).Main Outcome Measure  Subsequent myocardial infarction or coronary death during the 5-year follow-up, analyzed by quintile of LDL particle size and by treatment group. Results  Overall, the mean LDL size was identical in cases and controls (25.6 nm). In patients in the placebo group, large LDL predicted coronary events in models adjusted only for age (relative risk [RR], 1.79; 95% confidence interval [CI], 1.01-3.17) and for age and lipid and nonlipid risk factors (RR, 4.00; 95% CI, 1.81-8.82), comparing those in the highest (mean, 26.6 nm) and lowest (mean, 24.5 nm) quintiles of LDL size. This increased risk was not present in those taking pravastatin (age-adjusted analysis: RR, 0.98; 95% CI, 0.47-2.04; P = .046 for interaction for a difference in the effect of LDL size on coronary events between the placebo and treatment groups; multivariable analysis: RR, 1.33; 95% CI, 0.52-3.38; P = .11 for interaction).Conclusions  Large LDL size was an independent predictor of coronary events in a typical population with myocardial infarction, but the adverse effect was not present among patients who were treated with pravastatin. Identifying patients on the basis of LDL size may not be useful clinically, since effective treatment for elevated LDL cholesterol concentrations also effectively treats risk associated with large LDL.

Plasma leptin concentration is a novel and independent risk factor for CHD in WOSCOPS (Diabetologia [2001] 44 (Suppl 1): A315. EASD 37th Annual Meeting, #1209)

• Fasting plasma IL-6 concentrations are positively related to adiposity and negatively related to insulin action (Obesity Research [2001] 9:414-417 )

IKK-ß May Underly the Inflammatory Etiology of the Dysmetabolic Syndrome (Science [2001] 293:1673-1677)

• Increased PAI-1 in the arterial wall associates with diabetes. (Arteriosclerosis, Thrombosis, and Vascular Biology. [2001]21:1378-1382.)

Augmentation of the synthesis of plasminogen activator inhibitor type-1 by precursors of insulin. A potential risk factor for vascular disease. (Circulation [1994] 89:321-330) BACKGROUND: Both vascular disease and elevated concentrations in plasma of plasminogen activator inhibitor type-1 (PAI-1) are prominent in patients with non-insulin-dependent diabetes mellitus (NIDDM). We and others have hypothesized that the increased PAI-1 may contribute to acceleration of atherosclerosis in this condition and in other states characterized by insulin resistance as well. Surprisingly, however, elevations of PAI-1 decrease when type II diabetic patients are treated with exogenous insulin, as do circulating concentrations of the precursor of insulin, proinsulin, in plasma. Accordingly, the increased PAI-1 in patients with NIDDM may reflect effects of precursors of insulin rather than or in addition to those of insulin itself. To assess this possibility directly, this study was performed to identify potential direct effects of proinsulin and proinsulin split products on synthesis of PAI-1 in liver cells, thought to be the major source of circulating PAI-1 in vivo. METHODS AND RESULTS: Hep G2 cells (highly differentiated human hepatoma cells) were exposed to human proinsulin, des(31,32)proinsulin and des(64,65)proinsulin (split products of proinsulin), or C-peptide. Accumulation of PAI-1 in conditioned media increased in a time- and concentration-dependent fashion in response to the two des-intermediates [3.3-fold with des(31,32)proinsulin and 4.5-fold with des(64,65)proinsulin]. C-peptide elicited no increase. Stimulation was transduced at least in part by the insulin receptor as shown by inhibition of stimulation by insulin receptor antibodies, mediated at the level of PAI-1 gene expression as shown by the 2.2- to 2.9-fold increases in steady-state concentrations of PAI-1 mRNA, and indicative of newly synthesized protein as shown by results in metabolic labeling experiments. CONCLUSIONS: Our results are consistent with the hypothesis that precursors of insulin (proinsulin and proinsulin split products), known to be present in relatively high concentrations in plasma in patients with NIDDM and conditions characterized by insulin resistance, may directly stimulate PAI-1 synthesis, thereby attenuating fibrinolysis and accelerating atherogenesis.

• Circulating Non-Esterified Fatty Acids (NEFA) concentration is an independent risk factor for sudden death in middle-aged men. Fish oils are protective, however. (Circulation [2001]104:756-761)

Diabetes and ischemic heart disease interact to accelerate the progression of myocardial dysfunction (JACC [2000] 38:421-428)

• Trans fatty acids impair endothelial function (flow mediated dilation) in healthy individuals (Arteriosclerosis, Thrombosis, and Vascular Biology. [200]1:21:1233.)

Dyslipidemia Is Predictive of Macrovascular Events in Type 2 Diabetes While Glycemia Is Not (ADA 61st SS, Abstract #1910-PO, June 2001)

• Pre-DM subjects may have an increased risk of CVD even prior to the onset of DM (ADA 61st SS, Abstract #118-OR, Sunday, 24 June 2001 )

Losing Weight Returns Compromised T-Cell Function To Normal Via Reduced a-TNF (Clin Endocrinol [2001] 54(3):347-354 )

• The degree of glycemic control among diabetic patients at the time of their index intervention does not have an impact on long-term outcomes after successful Percutaneous Coronary Revascularization (PCR). (Am Heart J [2001]141:117-23.)

Opening of sarcolemmal K_ATPchannels underlies ST elevation during ischemia" (Circulation Res. [2000] 87:837)

• Polycystic Ovary Syndrome Is Associated With Endothelial Dysfunction" (Circulation [2001}103:1410)

High dietary glycemic index and high glycemic load are associated with a lower concentration of plasma HDL-C.(Arch Intern Med. [2001]161:572-576

• Results support the hypothesis that adenosine receptor activation primes the opening of mitochondrial KATP channels in a cardioprotective, protein kinase C-dependent manner" (Circulation [2000] 102:800)

Protein Kinase C action decreases the Hill coefficient of ATP binding to cardiac KATP channels, thereby increasing their open probability at physiological ATP concentrations (PNAS [2000] 10:1073)

• A high dietary glycemic load from refined carbohydrates increases the risk of CHD, independent of known coronary disease risk factors (American J Clin Nutrition [2000] 71:1455-1461)

CLA-1/SR-BI Is Expressed in Atherosclerotic Lesion Macrophages and Regulated by Activators of Peroxisome Proliferator-Activated Receptors (Circulation[2000]101:2411)

• Atherogenic CD36 expression by oxidized LDL and IL-4 by a common signaling pathway is dependent on protein kinase C and PPAR-gamma (J.Lipid Res. [2000] 41:688-696)Acute hyperglycemia increases circulating sICAM-1 levels in normal subjects, whereas the correction of hyperglycemia with insulin or l-arginine supplementation restored to normal levels the increased plasma sICAM-1 levels of type 2 diabetic patients (Circulation [2000] 101:2247)

Glycation of CD59 may form the molecular basis for a link between complement and the vascular complications of diabetes (PNAS [2000] 97:5450-5455)

• Serum leptin is elevated in patients with coronary artery disease (International Journal of Obesity and related metabolic disorders [ 2000] 24 :Pages 369 - 375]

Hyperinsulinemia modest association with CV mortality is largely explained by obesity, hypertension, and dyslipidemia" (Arch Intern Med. [2000]160:1160-1168)

• Microalbuminuria and gross proteinuria significantly
  associate with subsequent mortality from all causes (Arch Intern Med. [2000]160:1093-1100)The odds ratio for 5-year mortality for hyperhomocysteinemia inthe Hoorn Study was 1.34 (0.87 to 2.06) in nondiabetic subjects and 2.51 (1.07 to 5.91) in diabetic subjects (P=0.08 for interaction). Circulation [2000]101:1506

• TGF-beta-1 appears to correlate with hypertension in blacks" Proc. Natl. Acad. Sci. [2000] 97: 3479-3484

Prolonged hyperlipidemia - independent of insulin - may contribute to increased fat metabolism and storage as a result of FAT/CD36, PPAR-2, leptin, UCP-2, UCP-3, and TNF-alpha induction. Diabetes [2000] 49:319–324

• New data suggest that atherogenic changes in the prediabetic state are mainly seen in insulin-resistant subjects and that strategies to prevent type 2 diabetes might focus on insulin-sensitizing interventions rather than interventions that increase insulin secretion because of potential effects on cardiovascular risk. (Circulation [2000] 101:975)

JAMA publishes *VA VOODOO* study - utilizing such inane concepts as *utility index* of blindness and *quality adjusted life years* as well as innumerable assumptions predicting *days of blindness* - to suggest that yearly visual screens in type 2 patients are not cost-effective....(JAMA [2000]283:889-896)

• The change from resting values in diastolic but not systolic blood pressure during exercise correlates with serum cholesterol (R>0.47, P<0.0001 for each workload) and insulin resistance (R>0.38, P<0.01 for each workload)....(Circulation[2000] 101:611)

In multivariate Cox models, the age-adjusted hazard ratio for insulin resistance factor during the 22-year Helsinki Policemen Study follow-up was 1.28 (95% CI 1.10 to 1.50) with regard to CHD risk and 1.64 (95% CI 1.29 to 2.08) with regard to stroke risk. Lipid factor predicted the risk of CHD but not that of stroke, and lifestyle factor predicted a reduced CHD risk. Factor analysis including only 6 risk factor variables proposed to be central components of insulin resistance syndrome (body mass index, subscapular skinfold, AUC insulin, AUC glucose, mean blood pressure, and triglycerides) produced only a single insulin resistance factor that predicted the risk of CHD and stroke independently of other risk factors....(Arteriosclerosis,Thrombosis and Vascular Biology[2000]20:538)

• Lectin-like Oxidized-LDL (OxLDL) Receptor (LOX-1) is a major receptor for OxLDL in vascular smooth muscle cells as in endothelial cells (VSMC). Lysophosphatidyl choline (LPC) enhances the expression of LOX-1 which suggests that OxLDL and LPC might progressively change the function of VSMC and accelerate atherogenesis in vivo.....(FEBS [2000] 467:217-220)

Platelet-derived thrombogenesis in stable coronary patients associates weakly with blood glucose (R² = 0.19, p < 0.008), and apo-B (R² = 0.18, p = 0.002) at a threshold glucose value of 88.2 mg/dl (JACC[2000]35:300-307)

• LDL particle size and LDL vitamin E may associate with conduit and resistance vessel endothelial vasodilator function in type 1 diabetes. Further work will be required to prove cause and effect (JACC [2000]35:292-299)

In normal glucose tolerant offspring of type 2 diabetics, PAI-1 activity was correlated with plasma PAI-1 antigen level (r = 0.40, P = 0.02), fibrinogen (r = 0.45, P = 0.01), and HDL cholesterol (r = –0.36, P = 0.04). However, tPA antigen level, fasting and postload plasma glucose and insulin, total cholesterol, triglycerides, WHR, and BMI did NOT correlate with PAI-1 activity

• Perturbed handling of postprandial triglycerides in normolipidemic CAD patients involves the accumulation of apo C-I–rich large VLDL particles and the generation of small, apo C-I– and cholesterol-rich VLDL remnants

Physiologic elevation in the local plasma insulin concentration increases coronary blood flow in the absence of any increase in myocardial oxygen demand or consumption. These observations are consistent with insulin-mediated elaboration of vasoactive and/or paracrine factors within the coronary circulation.

• A common 4G allele of a 4G/5G polymorphism in the promoter region of the plasminogen activator inhibitor-1 (PAI-1) gene is associated with increased transcription of the PAI-1 protein, which may lead to decreased fibrinolysis. It has even been proposed as a candidate risk factor for myocardial infarction or stroke. Therefore the relationship between PAI-1 4G/5G genotype and the risk of cardiovascular mortality was studied in a prospective cohort study among 12 239 women initially aged between 52 and 67 years, with a maximum follow-up time of 18 years (153 732 follow-up years). However, the PAI-1 4G/5G genotype was not associated with risk of myocardial infarction or other cardiovascular mortality. Moreover, PAI-1 4G4G homozygotes had a markedly reduced risk of cerebrovascular mortality compared with PAI-1 5G5G homozygotes: the relative risk was 0.4, with a 95% CI of 0.2 to 0.7, whereas the relative risk of cerebrovascular mortality in PAI-1 4G5G heterozygotes compared with PAI-1 5G5G homozygotes was 0.7, with a 95% CI of 0.4 to 1.1

Fasting Insulinemia Appears to Associate with Measures of BOTH Increased (t-PA antigen) AND Decreased (PAI-1 antigen) Fibrinolytic Potential

• Results suggest that mitoK-atp channels are downstream of PKC in the mechanism of infarct-size limitation by A₁-receptor activation and that the anti-infarct tolerance afforded by opening of mitoK-atp channels is associated with preservation of mitochondrial function during ischemia/reperfusion

Diabetic subjects have significantly (P<0.001) more increased Dicarboxylic Acid (DA) excretion in urine than corresponding healthy subjects. The yields of DA's in each group decreased in the order adipic>suberic>sebacic acid. Being stable and easily detectable compounds, DA's may be considered potential markers of oxidative attack on PUFA in diabetes

• Data suggest that LDL particle size should probably be added to the cluster of risk factors that characterize the insulin resistance syndrome. The association of 'insulin resistance' and LDL particle diameter, however, is not mediated directly through the level of insulinemia but via alterations in lipid metabolism

Coronary Disease in Type 1's Correlates with Depression, NOT Glucose. Lower Extremity Arterial Disease, however, Does Correlate with Blood Glucose in Type 1's

• A Common Mutation of the Insulin Receptor Substrate-1 Gene Is A Risk Factor for Coronary Artery Disease

LDL size is not a predictor of CHD events in elderly white subjects after controlling for diabetes status

• Fiber consumption predicts (fasting and postprandial) insulin levels, weight gain, and other CVD risk factors more strongly than total or saturated fat consumption

Nonfasting Serum Glucose and Insulin Concentrations Associate Significantly with the Risk of Stroke

• Proteinuria Is an Independent Risk Factor for Ischemic Stroke in Non–Insulin-Dependent Diabetes Mellitus

HDL (HDL₂) from poorly-controlled diabetics is abnormal and does not inhibit LDL-oxidation or Apo-A1 fragmentation or HDL₂-associated platelet-activating factor acetylhydrolase activity as well as non-diabetic HDL(HDL₂) in vitro

• Even though secondarily associating with increased cardiovascular disease in the elderly, higher fasting plasma insulin appears to correlate with improved survival in that population

Should we recognize another class of diabetes called "Prandial Diabetes" with different risk factors? (The new fasting ADA criteria seem to be less predictive than the WHO criteria for the burden of cardiovascular disease associated with abnormal glucose in the elderly.)

• In the fasting state [in middle-aged men with apo-E3/apo-E3 phenotype], LDL cholesterol (P<0.05) and basal proinsulin (P<0.05) were significantly related to IMT, whereas HDL cholesterol, plasma triglycerides, and insulin were not. In the postprandial state, plasma triglycerides at 1 to 4 hours (P<0.01 at 2 hours), total triglyceride area under the curve (AUC) (P<0.05), incremental triglyceride AUC (P<0.01), and the large VLDL (Sf 60 to 400 apo B-100) concentration at 3 hours (P<0.05) were significantly related to Intimal Medial Thickness (IMT). Multivariate analyses showed that plasma triglycerides at 2 hours, LDL cholesterol, and basal proinsulin were consistently and independently related to IMT when cumulative tobacco consumption, alcohol intake, waist-to-hip circumference ratio, and systolic blood pressure were included as confounders.
  
  These results provide further evidence for postprandial triglyceridemia as an independent risk factor for early atherosclerosis and also suggest that the postprandial triglyceridemia is a better predictor of IMT than particle concentrations of individual TRLs.

Protein-Rich Diets May Reduce Heart Disease Risk

• NHLBI Investigators Identify HDL ("Good Cholesterol") Gene

Results suggest a dose-response relation of glucose intolerance at baseline with CHD incidence, CHD mortality, and total mortality, independent of other risk factors, in this cohort of middle-aged and older Japanese-American men.Diabetes Care 22:1262–1265, 1999

• High-Fructose Diets, NOT Insulin-Resistance or Western-Diets, Correlates with Atherosclerosis in LDL-Receptor Deficient Mice with high LDL-cholesterol

Western-Diet (high in meats and fats) protective against stroke dementia: Honolulu-Asia Aging Study

• Levels of LDL cholesterol significantly associated with an increased risk of dementia with stroke - especially when correcting for Lp(a)

Hypertension correlates with Renal Disease and NOT Cardiovascular Disease in the Pima Diabetic

• Homocysteine levels too high, but still significantly lower in diabetics than the general population 

Homocysteine significantly predicts atherosclerotic events in the post-menopausal period independent of diabetes

• Long-Term Exercise appears to decrease cytokines promulgating atherogenesis

Adipose-Cells secrete IL-6 which correlates with BMI, TNF-alfa, insulin resistance, markers of endothelial dysfunction, and the inflammatory marker C-Reactive Protein (CRP)

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