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Microvascular dysfunction predicts adverse cardiovascular events despite absence of large vessel disease. A shift in the mediator of flow-mediated dilatation (FMD) from nitric oxide (NO) to mitochondrial-derived hydrogen peroxide (H

Objective: Coronary artery disease (CAD) is associated with a compensatory switch in mechanism of flow-mediated dilation (FMD) from nitric oxide (NO) to H 2 O 2 . The underlying mechanism responsible for the pathological shift is not well understood, and recent reports directly implicate telomerase and indirectly support a role for autophagy. We hypothesize that autophagy is critical for shear stress–induced release of NO and is a crucial component of for the pathway by which telomerase regulates FMD. Approach and Results: Human left ventricular, atrial, and adipose resistance arterioles were collected for videomicroscopy and immunoblotting. FMD and autophagic flux were measured in arterioles treated with autophagy modulators alone, and in tandem with telomerase-activity modulators. LC3B II/I was higher in left ventricular tissue from patients with CAD compared with non-CAD (2.8±0.2 versus 1.0±0.2-fold change; P 2 O 2 , while activation of autophagy restored NO-mediated vasodilation in CAD arterioles. In the presence of an autophagy activator, telomerase inhibitor prevented the expected switch (Control: 82±4%; NG-Nitro-l-arginine methyl ester: 36±5%; polyethylene glycol catalase: 80±3). Telomerase activation was unable to restore NO-mediated FMD in the presence of autophagy inhibition in CAD arterioles (control: 72±7%; NG-Nitro-l-arginine methyl ester: 79±7%; polyethylene glycol catalase: 38±9%). Conclusions: We provide novel evidence that autophagy is responsible for the pathological switch in dilator mechanism in CAD arterioles, demonstrating that autophagy acts downstream of telomerase as a common denominator in determining the mechanism of FMD.

Background and purpose NO produces arteriolar flow-induced dilation (FID) in healthy subjects but is replaced by mitochondria-derived hydrogen peroxide (mtH2 O2 ) in patients with coronary artery disease (CAD). Lysophosphatidic acid (LPA) is elevated in patients with risk factors for CAD, but its functional effect in arterioles is unknown. We tested whether elevated LPA changes the mediator of FID from NO to mtH2 O2 in human visceral and subcutaneous adipose arterioles. Experimental approach Arterioles were cannulated on glass micropipettes and pressurized to 60 mmHg. We recorded lumen diameter after graded increases in flow in the presence of either NOS inhibition (L-NAME) or H2 O2 scavenging (Peg-Cat) ± LPA (10 μM, 30 min), ±LPA1 /LPA3 receptor antagonist (Ki16425) or LPA2 receptor antagonist (H2L5186303). We analysed LPA receptor RNA and protein levels in human arterioles and human cultured endothelial cells. Key results FID was inhibited by L-NAME but not Peg-Cat in untreated vessels. In vessels treated with LPA, FID was of similar magnitude but inhibited by Peg-Cat while L-NAME had no effect. Rotenone attenuated FID in vessels treated with LPA indicating mitochondria as a source of ROS. RNA transcripts from LPA1 and LPA2 but not LPA3 receptors were detected in arterioles. LPA1 but not LPA3 receptor protein was detected by Western blot. Pretreatment of vessels with an LPA1 /LPA3 , but not LPA2 , receptor antagonist prior to LPA preserved NO-mediated dilation. Conclusions and implications These findings suggest an LPA1 receptor-dependent pathway by which LPA increases arteriolar release of mtH2 O2 as a mediator of FMD.

Blood flow through healthy human vessels releases NO to produce vasodilation, whereas in patients with coronary artery disease (CAD), the mediator of dilation transitions to mitochondria-derived hydrogen peroxide ( mt H 2 O 2 ). Excessive mt H 2 O 2 production contributes to a proatherosclerotic vascular milieu. Loss of PGC-1α (peroxisome proliferator–activated receptor γ coactivator 1α) is implicated in the pathogenesis of CAD. We hypothesized that PGC-1α suppresses mt H 2 O 2 production to reestablish NO-mediated dilation in isolated vessels from patients with CAD. Isolated human adipose arterioles were cannulated, and changes in lumen diameter in response to graded increases in flow were recorded in the presence of PEG (polyethylene glycol)–catalase (H 2 O 2 scavenger) or L-NAME ( N G -nitro- l -arginine methyl ester; NOS inhibitor). In contrast to the exclusively NO- or H 2 O 2 -mediated dilation seen in either non-CAD or CAD conditions, respectively, flow-mediated dilation in CAD vessels was sensitive to both L-NAME and PEG-catalase after PGC-1α upregulation using ZLN005 and α-lipoic acid. PGC-1α overexpression in CAD vessels protected against the vascular dysfunction induced by an acute increase in intraluminal pressure. In contrast, downregulation of PGC-1α in non-CAD vessels produces a CAD-like phenotype characterized by mt H 2 O 2 -mediated dilation (no contribution of NO). Loss of PGC-1α may contribute to the shift toward the mt H 2 O 2 -mediated dilation observed in vessels from subjects with CAD. Strategies to boost PGC-1α levels may provide a therapeutic option in patients with CAD by shifting away from mt H 2 O 2 -mediated dilation, increasing NO bioavailability, and reducing levels of mt H 2 O 2 . Furthermore, increased expression of PGC-1α allows for simultaneous contributions of both NO and H 2 O 2 to flow-mediated dilation.

Vascular endothelial dysfunction is an early event in the development of atherosclerosis. Flow-mediated dilation (FMD) can be used to identify such endothelial dysfunction which is characterized by a decrease of nitric oxide (NO). Indeed, in subjects with coronary artery disease (CAD), microvascular FMD switches from a NO-mediated to a H 2 O 2 -mediated mechanism with corresponding elevations in mitochondrial reactive oxygen species (ROS). Recent evidence has shown an increase circulating mitochondrial DNA Damage-Associated Molecular Patterns ( mt DAMPs) in Coronary Heart Disease subjects and has been linked to vascular dysfunction. mt DAMPs signaling can activate the inflammatory response directly through activation of Toll Like Receptors 9 (TLR9). The effect of mt DAMPs on the endothelial function in the human microcirculation and the potential role for TLR9- mtDNA signaling axis to affect microvascular FMD, is unknown. In this study, we hypothesized that increased levels of mt DAMPs result in changes in microvascular redox environment that promote a loss of NO mediated dilation and elevation in mitochondrial derived ROS The incubation of cultured endothelial cells (ECs) with mt DAMPs decreased the ratio peNOS:teNOS in a time-dependent manner supporting the idea that mt DAMPs reduce NO bioavailability. This effect was eliminated when the ECs were pre-incubated with TLR9-targeted si-RNA. Prolonged (15-20H) ex vivo exposure to low dose of mt DAMPs (1ug/mL) changed the mechanism of FMD from NO to compensatory increase in H 2 O 2 to preserve overall dilator capacity in non-CAD human adipose micro-arterioles (% Max Diameter: Control 84.7±8.2; + L-NAME: 89.4±5.5; + Peg-Cat 35.4±10.8*; N=5, *pmt DAMPs (2.5ug/mL), the magnitude of the FMD was however significantly altered (% Max Diameter: high dose: 42.8 ±9.1* vs. low dose of mt DAMPs 84.7±8.2; N=4; p Our data suggest a potential pathogenic role of mt DAMPs in microvascular function through TLR9 to reduce NO bioavailability and induce endothelial dysfunction.

KHCAs from patients with and without CAD were assessed for mRNA and protein expression of KAssays of mRNA transcripts, membrane protein expression, and vascular cell-specific localization revealed abundant expression of KK

> “ But I think it [the Model] is more likely to change when, and because, far-reaching changes in the mental temper of our descendants demand that it should. The new Model will not be set up without evidence, but the evidence will turn up when the need for it becomes sufficiently great .” (222–223) The fate of biomedical research lies in the hands of future generations of scientists. In recent decades, the diversity of scientific career opportunities has exploded multidimensionally. However, the educational system for maintaining a pipeline of talented biomedical trainees remains unidimensional and has become outdated. This Viewpoint identifies inadequacies in training and offers potential solutions and implementation strategies to stimulate interest in science at a younger age and to better align individualized training pathways with career opportunities (precision training). Both interventions support of the ultimate goal of attracting the best possible future leaders in biomedical science . In his final published work, the eminent scholar C.S. Lewis describes the medieval model that dominated European intellectual life for centuries and explores how and why this model was replaced—because of discovery of dying stars, development of new scientific theories, and more. Aside from being a testament to an exceptional mind, C.S. Lewis’ The Discarded Image also contains wisdom directly relevant to the scientific milieu of the 21st century. Over the past 100 years, science and society have experienced unprecedented, dramatic change as a result of globalization, the Internet, advances in scientific capability, and an increasingly diverse and expanding scientific workforce. Such a large shift in the fabric of society has necessitated a corresponding shift in the environment in which we as scientists operate. However, scientific education lingers behind, using principles and processes that have not changed for many decades. The educational system responsible for training a new generation of …

The microcirculation is responsible for orchestrating adjustments in vascular tone to match local tissue perfusion with oxygen demand. Beyond this metabolic dilation, the microvasculature plays a critical role in modulating vascular tone by endothelial release of an unusually diverse family of compounds including nitric oxide, other reactive oxygen species, and arachidonic acid metabolites. Animal models have provided excellent insight into mechanisms of vasoregulation in health and disease. However, there are unique aspects of the human microcirculation that serve as the focus of this review. The concept is put forth that vasculoparenchymal communication is multimodal, with vascular release of nitric oxide eliciting dilation and preserving normal parenchymal function by inhibiting inflammation and proliferation. Likewise, in disease or stress, endothelial release of reactive oxygen species mediates both dilation and parenchymal inflammation leading to cellular dysfunction, thrombosis, and fibrosis. Some pathways responsible for this stress-induced shift in mediator of vasodilation are proposed. This paradigm may help explain why microvascular dysfunction is such a powerful predictor of cardiovascular events and help identify new approaches to treatment and prevention.

Most organs are able to modulate both blood flow and oxygen extraction to meet oxygen demand during increases in metabolism. However, the heart extracts oxygen near maximally (60%–80%) at rest and, therefore, relies almost exclusively on changes in perfusion to meet this demand.1 This requires moment-to-moment changes in arteriolar (resistance artery) tone to match changes in oxygen demand with flow. It is generally thought that a metabolic dilator substance released from the myocardium serves as the signal for vasodilation. However, despite several decades of investigation, the chemical mediator of cardiac metabolic dilation remains elusive. Recently, Chilian laboratory has proposed hydrogen peroxide (H2O2) as the link between cardiac metabolism and coronary dilation.2 Although H2O2 had been studied extensively as an endothelium-derived vasodilator, its role in metabolic dilation had not been well-defined. H2O2 has several characteristics that support a role in metabolic dilation. It is a product of cardiac metabolism, is produced in large quantities proportional to mitochondrial respiration, is cell membrane permeable, and has a sufficiently long half-life to serve as an intercellular signaling molecule. H2O2 elicits a dose-dependent dilation in coronary arterioles.3 Article, see p 612 On the basis of these characteristics, H2O2 has been studied as a mediator of metabolic dilation. Chilian laboratory used a bioassay where effluent from freshly isolated cardiomyocytes was dripped onto rat coronary arterioles. The resulting dilation was shown to be catalase-sensitive and related to the metabolic rate of the cultured myocytes.2 A variation of that preparation was used by Shimokawa laboratory, where pressurized coronary arterioles from a rabbit were placed on a canine beating heart.4 Pacing the dog heart induced a …

Mitochondrial dysfunction results in high levels of oxidative stress and mitochondrial damage, leading to disruption of endothelial homeostasis. Recent discoveries have clarified several pathways, whereby mitochondrial dysregulation contributes to endothelial dysfunction and vascular disease burden. One such pathway centers around peroxisome proliferator receptor-γ coactivator 1α (PGC-1α), a transcriptional coactivator linked to mitochondrial biogenesis and antioxidant defense, among other functions. Although primarily investigated for its therapeutic potential in obesity and skeletal muscle differentiation, the ability of PGC-1α to alter a multitude of cellular functions has sparked interest in its role in the vasculature. Within this context, recent studies demonstrate that PGC-1α plays a key role in endothelial cell and smooth muscle cell regulation through effects on oxidative stress, apoptosis, inflammation, and cell proliferation. The ability of PGC-1α to affect these parameters is relevant to vascular disease progression, particularly in relation to atherosclerosis. Upregulation of PGC-1α can prevent the development of, and even encourage regression of, atherosclerotic lesions. Therefore, PGC-1α is poised to serve as a promising target in vascular disease. This review details recent findings related to PGC-1α in vascular regulation, regulation of PGC-1α itself, the role of PGC-1α in atherosclerosis, and therapies that target this key protein.

OBJECTIVES KV channels are important regulators of vascular tone, but the identity of specific KV channels involved and their regulation in disease remain less well understood. We determined the expression of KV 1 channel subunits and their role in cAMP-mediated dilation in coronary resistance arteries from subjects with and without CAD. METHODS HCAs from patients with and without CAD were assessed for mRNA and protein expression of KV 1 channel subunits with molecular techniques and for vasodilator response with isolated arterial myography. RESULTS Assays of mRNA transcripts, membrane protein expression, and vascular cell-specific localization revealed abundant expression of KV 1.5 in vascular smooth muscle cells of non-CAD HCAs. Isoproterenol and forskolin, two distinct cAMP-mediated vasodilators, induced potent dilation of non-CAD arterioles, which was inhibited by both the general KV blocker 4-AP and the selective KV 1.5 blocker DPO-1. The cAMP-mediated dilation was reduced in CAD and was accompanied by a loss of or reduced contribution of 4-AP-sensitive KV channels. CONCLUSIONS KV 1.5, as a major 4-AP-sensitive KV 1 channel expressed in coronary VSMCs, mediates cAMP-mediated dilation in non-CAD arterioles. The cAMP-mediated dilation is reduced in CAD coronary arterioles, which is associated with impaired 4-AP-sensitive KV channel function.