Novel mechanisms for hesperetin action in endothelial cells inform effects of oral hesperidin treatment to improve endothelial dysfunction and reduce circulating markers of inflammation in our exploratory clinical trial. Hesperetin has vasculoprotective actions that may explain beneficial cardiovascular effects of citrus consumption.

SynopsisEndothelial dysfunction and insulin resistance are frequently co-morbid states. Vasodilator actions of insulin are mediated by phosphatidylinositol 3-kinase (PI3K)-dependent signaling pathways that stimulate production of nitric oxide from vascular endothelium. This helps to couple metabolic and hemodynamic homeostasis under healthy conditions. In pathological states, shared causal factors including glucotoxicity, lipotoxicity, and inflammation selectively impair PI3K-dependent insulin signaling pathways that contribute to reciprocal relationships between insulin resistance and endothelial dysfunction. We discuss implications of pathway-selective insulin resistance in vascular endothelium, interactions between endothelial dysfunction and insulin resistance, and therapeutic interventions that may simultaneously improve both metabolic and cardiovascular physiology in insulin-resistant conditions. KeywordsNitric Oxide; Insulin Resistance; Endothelial Dysfunction; Metabolic Syndrome IntroductionInsulin resistance plays a major patho-physiological role in type 2 diabetes and is tightly associated with major public health problems including obesity, hypertension, coronary artery disease, dyslipidemias, and a cluster of metabolic and cardiovascular abnormalities that define the metabolic syndrome [1,2]. A global epidemic of obesity is driving the increased incidence and prevalence of insulin resistance and its cardiovascular complications [3]. Insulin regulates glucose homeostasis by promoting glucose disposal in skeletal muscle and adipose tissue and inhibiting gluconeogenesis in liver [4]. In addition to these classical insulin target tissues, insulin has important physiological functions in the brain, pancreatic beta cells, heart, and vascular endothelium that help coordinate and couple metabolic and cardiovascular homeostasis under healthy conditions [5]. For example, vasodilator actions of insulin to stimulate production of nitric oxide (NO) from vascular endothelium lead to increased blood *Corresponding author for proof and reprints: Michael J. Quon, MD, PhD, Chief, Diabetes Unit, NCCAM, NIH, 9 Memorial Drive, Building 9, Room 1N-105 MSC 0920, Bethesda, MD 20892-0920, Tel: (301) Fax: (301) 402-1679, Email: quonm@nih.gov. Coauthors addresses: Ranganath Muniyappa, MD, PhD, Diabetes Unit, NCCAM, NIH, 10 Center Drive, 10-CRC, Room 4-1741, Bethesda, MD 20892-1632, Tel: (301) Fax: (301) 480 3159, Email: muniyapr@mail.nih.gov Micaela Iantorno, MD, Diabetes Unit, NCCAM, NIH, 9 Memorial Drive, Building 9, Room 1N-105 MSC 0920, Bethesda, MD 20892-0920, Tel: (301) Fax: (301) 402-1679, Email: iantornm@mail.nih.gov Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be...

These findings indicate that ghrelin reverses endothelial dysfunction in patients with metabolic syndrome by increasing nitric oxide bioactivity, thereby suggesting that decreased circulating levels of the peptide, such as those found in these patients, might play a role in the pathobiology of atherosclerosis.

Recent studies showed that chronic administration of losartan, an angiotensin II type I receptor antagonist, improved skeletal muscle function in dystrophin-deficient mdx mice. In this study, C57BL/10ScSn-Dmdmdx/J female mice were either untreated or treated with losartan (n = 15) in the drinking water at a dose of 600 mg/L over a 6-month period. Cardiac function was assessed via in vivo high frequency echocardiography and skeletal muscle function was assessed using grip strength testing, Digiscan monitoring, Rotarod timing, and in vitro force testing. Fibrosis was assessed using picrosirius red staining and Image J analysis. Gene expression was evaluated using real-time polymerized chain reaction (RT-PCR). Percentage shortening fraction was significantly decreased in untreated (26.9% ± 3.5%) mice compared to losartan-treated (32.2% ± 4.2%; P < .01) mice. Systolic blood pressure was significantly reduced in losartan-treated mice (56 ± 6 vs 69 ± 7 mm Hg; P < .0005). Percentage cardiac fibrosis was significantly reduced in losartan-treated hearts (P < .05) along with diaphragm (P < .01), extensor digitorum longus (P < .05), and gastrocnemius (P < .05) muscles compared to untreated mdx mice. There were no significant differences in skeletal muscle function between treated and untreated groups. Chronic treatment with losartan decreases cardiac and skeletal muscle fibrosis and improves cardiac systolic function in dystrophin-deficient mdx mice.

Despite on-going evolution and iteration of drug-eluting stent (DES) technology, the prevalence of in-stent restenosis (ISR) remains relatively unchanged, encompassing ≈10% of percutaneous coronary interventions. The mechanism of ISR is multifactorial, including biological, mechanical, patient, and operator-related factors. The main mechanical contributors are stent underexpansion or fracture, while biological factors include local inflammation leading to aggressive neointimal proliferation and late neoatherosclerosis. Intracoronary imaging is critical to identify the mechanism of ISR and tailor therapy accordingly. The presentation of DES-ISR is not benign and is challenging for optimal treatment. Among the proposed treatment modalities are scoring and high-pressure balloons, percutaneous coronary intervention with additional DES, atheroablative therapies by laser or mechanical atherectomy, drug-coated balloons, vascular brachytherapy, and surgical revascularization. We propose a new classification for ISR that differentiates among mechanical, biological, and mixed etiologies. Stratifying ISR by mechanism guides individualized treatment of DES-ISR to improve clinical outcomes. An algorithmic approach, guided by intracoronary imaging, for the treatment of DES-ISR, is recommended based on the specific cause of restenosis.

Abstract-Hypertensive patients have increased endothelin-1-dependent vasoconstrictor tone. This abnormality, however, might not be uniformly present in all forms of hypertension, as suggested by experimental studies showing that endothelin-1 activity is enhanced predominantly in low-renin, high-volume models and in insulin-resistant states. Because hypertension in obesity is commonly associated with both expanded plasma volume and insulin resistance, this study sought to determine whether increased body mass index (BMI) in hypertensive patients relates to activation of the endothelin-1 system. Forearm blood flow (FBF) responses (plethysmography) to intra-arterial infusion of an ET A receptor blocker (BQ-123) were analyzed in hypertensive patients and normotensive control subjects according to BMI. The vasodilator response to BQ-123 was significantly higher in hypertensive patients than in control subjects (PϽ0.001). During BQ-123, a significant increase in FBF from baseline was observed in obese (BMI Ն30 kg/m 2 ; PϽ0.001) and overweight (BMI, 27 to 29.9 kg/m 2 ; Pϭ0.04) but not in lean (BMI Ͻ27 kg/m 2 ; Pϭ0.83) hypertensive patients. In contrast, no significant change in FBF was observed during BQ-123 either in obese (Pϭ0.53), overweight (Pϭ0.76), or lean (Pϭ0.93) normotensive subjects. Moreover, a significant correlation between BMI and the vasodilator response to ET A blockade was observed in hypertensive subjects (Rϭ0.53; Pϭ0.005) but not in control subjects (Rϭ0.11; Pϭ0.58). In human hypertension, increased BMI is associated with enhanced ET A -dependent vasoconstrictor activity, suggesting that this abnormality may play a role in the pathophysiology of obesity-related hypertension and that targeting the endothelin-1 system may be useful in the treatment of these patients. Key Words: endothelin Ⅲ hypertension, obesity Ⅲ vasculature Ⅲ atherosclerosis P revious work in our laboratory with infusion of endothelin (ET) receptor antagonists in the human forearm circulation demonstrated that ET-1-dependent vasoconstrictor tone is increased in patients with essential hypertension and may play a role in the pathophysiology of hypertensionrelated vascular damage. 1 Also, blockade of ET-1 receptors in patients with hypertension enhances endothelium-dependent vasodilation, 2 thereby suggesting that activation of the ET-1 system may contribute to the atherosclerotic process in hypertensive vessels.Because essential hypertension is a heterogeneous condition, an activation of the ET-1 system might not be a generalized finding in all patients. This view is supported by the results of previous studies in animal models, indicating that ET-1 activity is predominantly enhanced in low-renin, high-volume forms of experimental hypertension. 3 Thus, in DOCA-salt hypertensive rats, ET-1 gene expression is enhanced and abolishment of ET-1 overexpression results in lower blood pressure with regression of vascular growth. 4,5 Similar results have been observed in Dahl salt-sensitive rats, another model of low-renin, high-volume hyperte...

Ghrelin is an orexigenic peptide hormone secreted by the stomach. In patients with metabolic syndrome and low ghrelin levels, intra-arterial ghrelin administration acutely improves their endothelial dysfunction. Therefore, we hypothesized that ghrelin activates endothelial nitric oxide synthase (eNOS) in vascular endothelium, resulting in increased production of nitric oxide (NO) using signaling pathways shared in common with the insulin receptor. Similar to insulin, ghrelin acutely stimulated increased production of NO in bovine aortic endothelial cells (BAEC) in primary culture (assessed using NO-specific fluorescent dye 4,5-diaminofluorescein) in a time- and dose-dependent manner. Production of NO in response to ghrelin (100 nM, 10 min) in human aortic endothelial cells was blocked by pretreatment of cells with NG-nitro-L-arginine methyl ester (nitric oxide synthase inhibitor), wortmannin [phosphatidylinositol (PI) 3-kinase inhibitor], or (D-Lys3)-GHRP-6 (selective antagonist of ghrelin receptor GHSR-1a), as well as by knockdown of GHSR-1a using small-interfering (si) RNA (but not by mitogen/extracellular signal-regulated kinase inhibitor PD-98059). Moreover, ghrelin stimulated increased phosphorylation of Akt (Ser473) and eNOS (Akt phosphorylation site Ser1179) that was inhibitable by knockdown of GHSR-1a using siRNA or by pretreatment of cells with wortmannin but not with PD-98059. Ghrelin also stimulated phosphorylation of mitogen-activated protein (MAP) kinase in BAEC. However, unlike insulin, ghrelin did not stimulate MAP kinase-dependent secretion of the vasoconstrictor endothelin-1 from BAEC. We conclude that ghrelin has novel vascular actions to acutely stimulate production of NO in endothelium using a signaling pathway that involves GHSR-1a, PI 3-kinase, Akt, and eNOS. Our findings may be relevant to developing novel therapeutic strategies to treat diabetes and related diseases characterized by reciprocal relationships between endothelial dysfunction and insulin resistance.

Objective Offspring of patients with type 2 diabetes (OPDs) exhibits endothelial dysfunction (ED) associated with a chronic inflammatory state. N-3 polyunsaturated fatty acids (n-3 PUFA) may have antioxidant and anti-inflammatory properties that are beneficial for cardiovascular and metabolic health. Therefore, in the present study, we tested the hypothesis that dietary supplementation with fish oil rich in n-3 PUFA may improve ED in otherwise healthy OPDs. Methods and design A double-blind, placebo-controlled trial was conducted with 50 OPDs. Participants were randomized to treatment with either placebo or n-3 PUFA (2 g/day) for 12 weeks. Before and after treatment we evaluated endothelial function (using flow-mediated dilation (FMD) of the brachial artery), circulating inflammatory markers (adiponectin, TNF-α, and high sensitivity-CRP), and insulin resistance (QUICKI). Results No significant changes were observed in study outcomes in subjects treated with placebo. By contrast, when compared with baseline values, subjects treated with n-3 PUFA had significant improvement in FMD (9.1 ± 5.8% vs. 11.7 ± 4.4%, p = 0.02) that was accompanied by decreased plasma triglycerides (117 ± 73 mg/dl vs. 86 ± 44 mg/dl, p = 0.001) and TNF-α levels (8.9 ± 2.3 pg/ml vs. 6.8 ± 2.7 pg/ml, p = 0.001), and a trend towards increased plasma adiponectin levels (7.8 ± 4.5 μg/ml vs. 9.5 ± 5.1 μg/ml, p = 0.09). When data were analyzed by multiple regression analysis, decreased TNF-α after treatment with n-3 PUFA predicted increased FMD. Conclusion Dietary supplementation with n-3 PUFA significantly improved endothelial function and reduced pro-inflammatory markers in OPDs. Thus, fish oil consumption may have beneficial cardiovascular and metabolic health effects in otherwise healthy subjects predisposed to diabetes and its vascular complications.