The cardiovascular changes underlying a low cardiac output with exercise in patients with type 2 diabetes mellitus.

The significant morbidity and premature mortality of type 2 diabetes mellitus (T2DM) is largely associated with its cardiovascular consequences. Focus has long been on the arterial atheromatosis of DM giving rise to early stroke and myocardial infarctions, whereas less attention has been given to its non-ischemic cardiovascular consequences. Irrespective of ischemic changes, T2DM is associated with heart failure (HF) most commonly with preserved ejection fraction (HFpEF). Largely due to increasing population ages, hypertension, obesity and T2DM, HFpEF is becoming the most prevalent form of heart failure. Unfortunately, randomized controlled trials of HFpEF have largely been futile, and it now seems logical to address the important different phenotypes of HFpEF to understand their underlying pathophysiology. In the early phases, HFpEF is associated with a significantly impaired ability to increase cardiac output with exercise. The lowered cardiac output with exercise results from both cardiac and peripheral causes. T2DM is associated with left ventricular (LV) diastolic dysfunction based on LV hypertrophy with myocardial disperse fibrosis and significantly impaired ability for myocardial blood flow increments with exercise. T2DM is also associated with impaired ability for skeletal muscle vasodilation during exercise, and as is the case in the myocardium, such changes may be related to vascular rarefaction. The present review discusses the underlying phenotypical changes of the heart and peripheral vascular system and their importance for an adequate increase in cardiac output. Since many of the described cardiovascular changes with T2DM must be considered difficult to change if fully developed, it is suggested that patients with T2DM are early evaluated with respect to their cardiovascular compromise.

C-type natriuretic peptide induces inotropic and lusitropic effects in human 3D-engineered cardiac tissue: Implications for the regulation of cardiac function in humans.

The role of C-type natriuretic peptide (CNP) in the regulation of cardiac function in humans remains to be established as previous investigations have been confined to animal model systems. Here, we used well-characterized engineered cardiac tissues (ECTs) generated from human stem cell-derived cardiomyocytes and fibroblasts to study the acute effects of CNP on contractility. Application of CNP elicited a positive inotropic response as evidenced by increases in maximum twitch amplitude, maximum contraction slope and maximum calcium amplitude. This inotropic response was accompanied by a positive lusitropic response as demonstrated by reductions in time from peak contraction to 90% of relaxation and time from peak calcium transient to 90% of decay that paralleled increases in maximum contraction decay slope and maximum calcium decay slope. To establish translatability, CNP-induced changes in contractility were also assessed in rat ex vivo (isolated heart) and in vivo models. Here, the effects on force kinetics observed in ECTs mirrored those observed in both the ex vivo and in vivo model systems, whereas the increase in maximal force generation with CNP application was only detected in ECTs. In conclusion, CNP induces a positive inotropic and lusitropic response in ECTs, thus supporting an important role for CNP in the regulation of human cardiac function. The high degree of translatability between ECTs, ex vivo and in vivo models further supports a regulatory role for CNP and expands the current understanding of the translational value of human ECTs. NEW FINDINGS: What is the central question of this study? What are the acute responses to C-type natriuretic peptide (CNP) in human-engineered cardiac tissues (ECTs) on cardiac function and how well do they translate to matched concentrations in animal ex vivo and in vivo models? What is the main finding and its importance? Acute stimulation of ECTs with CNP induced positive lusitropic and inotropic effects on cardiac contractility, which closely reflected the changes observed in rat ex vivo and in vivo cardiac models. These findings support an important role for CNP in the regulation of human cardiac function and highlight the translational value of ECTs.

Genetic evidence implicating natriuretic peptide receptor-3 in cardiovascular disease risk: a Mendelian randomization study.

BACKGROUND: C-type natriuretic peptide (CNP) is a known target for promoting growth and has been implicated as a therapeutic opportunity for the prevention and treatment of cardiovascular disease (CVD). This study aimed to explore the effect of CNP on CVD risk using the Mendelian randomization (MR) framework. METHODS: Instrumental variables mimicking the effects of pharmacological intervention on CNP were identified as uncorrelated genetic variants located in the genes coding for its primary receptors, natriuretic peptide receptors-2 and 3 (NPR2 and NPR3), that associated with height. We performed MR and colocalization analyses to investigate the effects of NPR2 signalling and NPR3 function on CVD outcomes and risk factors. MR estimates were compared to those obtained when considering height variants from throughout the genome. RESULTS: Genetically-proxied reduced NPR3 function was associated with a lower risk of CVD, with odds ratio (OR) 0.74 per standard deviation (SD) higher NPR3-predicted height, and 95% confidence interval (95% CI) 0.64-0.86. This effect was greater in magnitude than observed when considering height variants from throughout the genome. For CVD subtypes, similar MR associations for NPR3-predicted height were observed when considering the outcomes of coronary artery disease (0.75, 95% CI 0.60-0.92), stroke (0.69, 95% CI 0.50-0.95) and heart failure (0.77, 95% CI 0.58-1.02). Consideration of CVD risk factors identified systolic blood pressure (SBP) as a potential mediator of the NPR3-related CVD risk lowering. For stroke, we found that the MR estimate for NPR3 was greater in magnitude than could be explained by a genetically predicted SBP effect alone. Colocalization results largely supported the MR findings, with no evidence of results being driven by effects due to variants in linkage disequilibrium. There was no MR evidence supporting effects of NPR2 on CVD risk, although this null finding could be attributable to fewer genetic variants being identified to instrument this target. CONCLUSIONS: This genetic analysis supports the cardioprotective effects of pharmacologically inhibiting NPR3 receptor function, which is only partly mediated by an effect on blood pressure. There was unlikely sufficient statistical power to investigate the cardioprotective effects of NPR2 signalling.

Liraglutide Lowers Endothelial Vascular Cell Adhesion Molecule-1 in Murine Atherosclerosis Independent of Glucose Levels.

The authors determined the effect of the GLP-1 receptor agonist liraglutide on endothelial surface expression of vascular cell adhesion molecule (VCAM)-1 in murine apolipoprotein E knockout atherosclerosis. Contrast-enhanced ultrasound molecular imaging using microbubbles targeted to VCAM-1 and control microbubbles showed a 3-fold increase in endothelial surface VCAM-1 signal in vehicle-treated animals, whereas in the liraglutide-treated animals the signal ratio remained around 1 throughout the study. Liraglutide had no influence on low-density lipoprotein cholesterol or glycated hemoglobin, but reduced TNF-α, IL-1ß, MCP-1, and OPN. Aortic plaque lesion area and luminal VCAM-1 expression on immunohistology were reduced under liraglutide treatment.

A State of Natriuretic Peptide Deficiency.

Measurement of natriuretic peptides (NPs) has proven its clinical value as biomarker, especially in the context of heart failure (HF). In contrast, a state of partial NP deficiency appears integral to several conditions in which lower NP concentrations in plasma presage overt cardiometabolic disease. Here, obesity and type 2 diabetes have attracted considerable attention. Other factors-including age, sex, race, genetics, and diurnal regulation-affect the NP "armory" and may leave some individuals more prone to development of cardiovascular disease. The molecular maturation of NPs has also proven complex, with highly variable O-glycosylation within the biosynthetic precursors. The relevance of this regulatory step in post-translational propeptide maturation has recently become recognized in biomarker measurement/interpretation and cardiovascular pathophysiology. An important proportion of people appear to have reduced effective net NP bioactivity in terms of receptor activation and physiological effects. The state of NP deficiency both entails a potential for further biomarker development and could also offer novel pharmacological possibilities. Alleviating the state of NP deficiency before development of overt cardiometabolic disease in selected patients could be a future path for improving precision medicine.

Therapeutic potential of the Proprotein Convertase Subtilisin/Kexin family in vascular disease.

Proprotein convertase subtilisin/kexins (PCSKs) constitute a family of nine related proteases: PCSK1-7, MBTPS1, and PCSK9. Apart from PCSK9, little is known about PCSKs in cardiovascular disease. Here, we aimed to investigate the expression landscape and druggability potential of the entire PCSK family for CVD. We applied an integrative approach, combining genetic, transcriptomic and proteomic data from three vascular biobanks comprising carotid atherosclerosis, thoracic and abdominal aneurysms, with patient clinical parameters and immunohistochemistry of vascular biopsies. Apart from PCSK4, all PCSK family members lie in genetic regions containing variants associated with human cardiovascular traits. Transcriptomic analyses revealed that FURIN, PCSK5, MBTPS1 were downregulated, while PCSK6/7 were upregulated in plaques vs. control arteries. In abdominal aneurysms, FURIN, PCSK5, PCSK7, MBTPS1 were downregulated, while PCSK6 was enriched in diseased media. In thoracic aneurysms, only FURIN was significantly upregulated. Network analyses of the upstream and downstream pathways related to PCSKs were performed on the omics data from vascular biopsies, revealing mechanistic relationships between this protein family and disease. Cell type correlation analyses and immunohistochemistry showed that PCSK transcripts and protein levels parallel each other, except for PCSK9 where transcript was not detected, while protein was abundant in vascular biopsies. Correlations to clinical parameters revealed a positive association between FURIN plaque levels and serum LDL, while PCSK6 was negatively associated with Hb. PCSK5/6/7 were all positively associated with adverse cardiovascular events. Our results show that PCSK6 is abundant in plaques and abdominal aneurysms, while FURIN upregulation is characteristic for thoracic aneurysms. PCSK9 protein, but not the transcript, was present in vascular lesions, suggesting its accumulation from circulation. Integrating our results lead to the development of a novel 'molecular' 5D framework. Here, we conducted the first integrative study of the proprotein convertase family in this context. Our results using this translational pipeline, revealed primarily PCSK6, followed by PCSK5, PCSK7 and FURIN, as proprotein convertases with the highest novel therapeutic potential.

Matching of O2 Utilization and O2 Delivery in Contracting Skeletal Muscle in Health, Aging, and Heart Failure.

Skeletal muscle is one of the most dynamic metabolic organs as evidenced by increases in metabolic rate of >150-fold from rest to maximal contractile activity. Because of limited intracellular stores of ATP, activation of metabolic pathways is required to maintain the necessary rates of ATP re-synthesis during sustained contractions. During the very early phase, phosphocreatine hydrolysis and anaerobic glycolysis prevails but as activity extends beyond ∼1 min, oxidative phosphorylation becomes the major ATP-generating pathway. Oxidative metabolism of macronutrients is highly dependent on the cardiovascular system to deliver O2 to the contracting muscle fibres, which is ensured through a tight coupling between skeletal muscle O2 utilization and O2 delivery. However, to what extent O2 delivery is ideal in terms of enabling optimal metabolic and contractile function is context-dependent and determined by a complex interaction of several regulatory systems. The first part of the review focuses on local and systemic mechanisms involved in the regulation of O2 delivery and how integration of these influences the matching of skeletal muscle O2 demand and O2 delivery. In the second part, alterations in cardiovascular function and structure associated with aging and heart failure, and how these impact metabolic and contractile function, will be addressed. Where applicable, the potential of exercise training to offset/reverse age- and disease-related cardiovascular declines will be highlighted in the context of skeletal muscle metabolic function. The review focuses on human data but also covers animal observations.

Fibrotic Signaling in Cardiac Fibroblasts and Vascular Smooth Muscle Cells: The Dual Roles of Fibrosis in HFpEF and CAD.

Patients with heart failure with preserved ejection fraction (HFpEF) and atherosclerosis-driven coronary artery disease (CAD) will have ongoing fibrotic remodeling both in the myocardium and in atherosclerotic plaques. However, the functional consequences of fibrosis differ for each location. Thus, cardiac fibrosis leads to myocardial stiffening, thereby compromising cardiac function, while fibrotic remodeling stabilizes the atherosclerotic plaque, thereby reducing the risk of plaque rupture. Although there are currently no drugs targeting cardiac fibrosis, it is a field under intense investigation, and future drugs must take these considerations into account. To explore similarities and differences of fibrotic remodeling at these two locations of the heart, we review the signaling pathways that are activated in the main extracellular matrix (ECM)-producing cells, namely human cardiac fibroblasts (CFs) and vascular smooth muscle cells (VSMCs). Although these signaling pathways are highly overlapping and context-dependent, effects on ECM remodeling mainly act through two core signaling cascades: TGF-ß and Angiotensin II. We complete this by summarizing the knowledge gained from clinical trials targeting these two central fibrotic pathways.

Semaglutide treatment attenuates vessel remodelling in ApoE-/- mice following vascular injury and blood flow perturbation.

Background and aims: Randomized clinical studies have shown a reduction in cardiovascular outcomes with glucagon-like peptide 1 receptor agonist (GLP-1RA) treatment with the hypothesized mechanisms being an underlying effect on atherosclerosis. Here, we aimed to assess the pharmacological effects of semaglutide in an atheroprone murine model that recapitulates central mechanisms related to vascular smooth muscle cell (VSMC) phenotypic switching and endothelial dysfunction known to operate within the atherosclerotic plaque. Methods: In study A, we employed an electrical current to the carotid artery in ApoE-/- mice to induce severe VSMC injury and death, after which the arteries were allowed to heal for 4 weeks. In study B, a constrictive cuff was added for 6 h at the site of the healed segment to induce a disturbance in blood flow. Results: Compared to vehicle, semaglutide treatment reduced the intimal and medial area by ∼66% (p = 0.007) and ∼11% (p = 0.0002), respectively. Following cuff placement, expression of the pro-inflammatory marker osteopontin and macrophage marker Mac-2 was reduced (p < 0.05) in the semaglutide-treated group compared to vehicle. GLP-1R were not expressed in murine carotid artery and human coronary vessels with and without atherosclerotic plaques, and semaglutide treatment did not affect proliferation of cultured primary human VSMCs. Conclusions: Semaglutide treatment reduced vessel remodelling following electrical injury and blood flow perturbation in an atheroprone mouse model. This effect appears to be driven by anti-inflammatory and -proliferative mechanisms independent of GLP-1 receptor-mediated signalling in the resident vascular cells. This mechanism of action may be important for cardiovascular protection.

Nitrate-rich beetroot juice ingestion reduces skeletal muscle O2 uptake and blood flow during exercise in sedentary men.

Dietary nitrate supplementation has been shown to reduce pulmonary O2 uptake during submaximal exercise and enhance exercise performance. However, the effects of nitrate supplementation on local metabolic and haemodynamic regulation in contracting human skeletal muscle remain unclear. To address this, eight healthy young male sedentary subjects were assigned in a randomized, double-blind, crossover design to receive nitrate-rich beetroot juice (NO3, 9 mmol) and placebo (PLA) 2.5 h prior to the completion of a double-step knee-extensor exercise protocol that included a transition from unloaded to moderate-intensity exercise (MOD) followed immediately by a transition to intense exercise (HIGH). Compared with PLA, NO3 increased plasma levels of nitrate and nitrite. During MOD, leg V̇O2 and leg blood flow (LBF) were reduced to a similar extent (∼9%-15%) in NO3. During HIGH, leg V̇O2 was reduced by ∼6%-10% and LBF by ∼5%-9% (did not reach significance) in NO3. Leg V̇O2 kinetics was markedly faster in the transition from passive to MOD compared with the transition from MOD to HIGH both in NO3 and PLA with no difference between PLA and NO3. In NO3, a reduction in nitrate and nitrite concentration was detected between arterial and venous samples. No difference in the time to exhaustion was observed between conditions. In conclusion, elevation of plasma nitrate and nitrate reduces leg skeletal muscle V̇O2 and blood flow during exercise. However, nitrate supplementation does not enhance muscle V̇O2 kinetics during exercise, nor does it improve time to exhaustion when exercising with a small muscle mass. KEY POINTS: Dietary nitrate supplementation has been shown to reduce systemic O2 uptake during exercise and improve exercise performance. The effects of nitrate supplementation on local metabolism and blood flow regulation in contracting human skeletal muscle remain unclear. By using leg exercise engaging a small muscle mass, we show that O2 uptake and blood flow are similarly reduced in contracting skeletal muscle of humans during exercise. Despite slower V̇O2 kinetics in the transition from moderate to intense exercise, no effects of nitrate supplementation were observed for V̇O2 kinetics and time to exhaustion. Nitrate and nitrite concentrations are reduced across the exercising leg, suggesting that these ions are extracted from the arterial blood by contracting skeletal muscle.

Menopausal transition does not influence skeletal muscle capillary growth in response to cycle training in women.

The influence of the menopausal transition, with a consequent loss of estrogen, on capillary growth in response to exercise training remains unknown. In the present study, we evaluated the effect of a period of intense endurance training on skeletal muscle angiogenesis in late premenopausal and recent postmenopausal women with an age difference of <4 yr. Skeletal muscle biopsies were obtained from the thigh muscle before and after 12 wk of intense aerobic cycle training and analyzed for capillarization, fiber-type distribution, and content of vascular endothelial growth factor (VEGF). At baseline, there was no difference in capillary per fiber ratio (C:F; 1.41 ± 0.22 vs. 1.40 ± 0.30), capillary density (CD; 305 ± 61 vs. 336 ± 52 mm2), muscle fiber area (MFA; 4,889 ± 1,868 vs. 4,195 ± 749), or distribution of muscle fiber type I (47.3% ± 10.1% vs. 49.3% ± 15.1%), between the pre- and postmenopausal women, respectively. There was a main effect of training on the C:F ratio (+9.2% and +12.1%, for the pre- and postmenopausal women, respectively) and the CD (+6.9% and +8.9%, for the pre- and postmenopausal women, respectively). MFA and fiber-type distribution were unaltered by training. Skeletal muscle VEGF protein content was similar between groups at baseline, and there was a main effect of training (+21.1% and +27.2%, for the pre- and postmenopausal women, respectively). In conclusion, the loss of estrogen per se at menopause does not influence the capillary growth response to intense aerobic exercise training.NEW & NOTEWORTHY We evaluated the effect of 12 wk of intense aerobic exercise training on skeletal muscle angiogenesis in late pre- and recent postmenopausal women, with <4 yr of age difference. There was a main effect of training on capillary per fiber ratio, capillary density, and muscle VEGF protein content, with no difference between groups. It is concluded that the loss of estrogen per se at menopause does not influence the capillary growth response to intense aerobic training.

The Impact of Lower Limb Immobilization and Rehabilitation on Angiogenic Proteins and Capillarization in Skeletal Muscle.

PURPOSE: Skeletal muscle vascularization is important for tissue regeneration after injury and immobilization. We examined whether complete immobilization influences capillarization and oxygen delivery to the muscle and assessed the efficacy of rehabilitation by aerobic exercise training. METHODS: Young healthy males had one leg immobilized for 14 d and subsequently completed 4 wk of intense aerobic exercise training. Biopsies were obtained from musculus vastus lateralis, and arteriovenous blood sampling for assessment of oxygen extraction and leg blood flow during exercise was done before and after immobilization and training. Muscle capillarization, muscle and platelet content of vascular endothelial growth factor (VEGF), and muscle thrombospondin-1 were determined. RESULTS: Immobilization did not have a significant impact on capillary per fiber ratio or capillary density. The content of VEGF protein in muscle samples was reduced by 36% (P = 0.024), and VEGF to thrombospondin-1 ratio was 94% lower (P = 0.046). The subsequent 4-wk training period increased the muscle VEGF content and normalized the muscle VEGF to thrombospondin-1 ratio but did not influence capillarization. Platelet VEGF content followed the trend of muscle VEGF. At the functional level, oxygen extraction, blood flow, and oxygen delivery at rest and during submaximal exercise were not affected by immobilization or training. CONCLUSIONS: The results demonstrate that just 2 wk of leg immobilization leads to a strongly reduced angiogenic potential as evidenced by reduced muscle and platelet VEGF content and a reduced muscle VEGF to thrombospondin-1 ratio. Moreover, a subsequent period of intensive aerobic exercise training fails to increase capillarization in the previously immobilized leg, possibly because of the angiostatic condition caused by immobilization.

Effects of High-Intensity Exercise Training on Adipose Tissue Mass, Glucose Uptake and Protein Content in Pre- and Post-menopausal Women.

The menopausal transition is accompanied by changes in adipose tissue storage, leading to an android body composition associated with increased risk of type 2 diabetes and cardiovascular disease in post-menopausal women. Estrogens probably affect local adipose tissue depots differently. We investigated how menopausal status and exercise training influence adipose tissue mass, adipose tissue insulin sensitivity and adipose tissue proteins associated with lipogenesis/lipolysis and mitochondrial function. Healthy, normal-weight pre- (n = 21) and post-menopausal (n = 20) women participated in high-intensity exercise training three times per week for 12 weeks. Adipose tissue distribution was determined by dual-energy x-ray absorptiometry and magnetic resonance imaging. Adipose tissue glucose uptake was assessed by positron emission tomography/computed tomography (PET/CT) by the glucose analog [18F]fluorodeoxyglucose ([18F]FDG) during continuous insulin infusion (40 mU·m-2·min-1). Protein content associated with insulin signaling, lipogenesis/lipolysis, and mitochondrial function were determined by western blotting in abdominal and femoral white adipose tissue biopsies. The mean age difference between the pre- and the post-menopausal women was 4.5 years. Exercise training reduced subcutaneous (~4%) and visceral (~6%) adipose tissue masses similarly in pre- and post-menopausal women. Insulin-stimulated glucose uptake, assessed by [18F]FDG-uptake during PET/CT, was similar in pre- and post-menopausal women in abdominal, gluteal, and femoral adipose tissue depots, despite skeletal muscle insulin resistance in post- compared to pre-menopausal women in the same cohort. Insulin-stimulated glucose uptake in adipose tissue depots was not changed after 3 months of high-intensity exercise training, but insulin sensitivity was higher in visceral compared to subcutaneous adipose tissue depots (~139%). Post-menopausal women exhibited increased hexokinase and adipose triglyceride lipase content in subcutaneous abdominal adipose tissue. Physical activity in the early post-menopausal years reduces abdominal obesity, but insulin sensitivity of adipose tissue seems unaffected by both menopausal status and physical activity.

Deep learning reveals 3D atherosclerotic plaque distribution and composition.

Complications of atherosclerosis are the leading cause of morbidity and mortality worldwide. Various genetically modified mouse models are used to investigate disease trajectory with classical histology, currently the preferred methodology to elucidate plaque composition. Here, we show the strength of light-sheet fluorescence microscopy combined with deep learning image analysis for characterising and quantifying plaque burden and composition in whole aorta specimens. 3D imaging is a non-destructive method that requires minimal ex vivo handling and can be up-scaled to large sample sizes. Combined with deep learning, atherosclerotic plaque in mice can be identified without any ex vivo staining due to the autofluorescent nature of the tissue. The aorta and its branches can subsequently be segmented to determine how anatomical position affects plaque composition and progression. Here, we find the highest plaque accumulation in the aortic arch and brachiocephalic artery. Simultaneously, aortas can be stained for markers of interest (for example the pan immune cell marker CD45) and quantified. In ApoE-/- mice we observe that levels of CD45 reach a plateau after which increases in plaque volume no longer correlate to immune cell infiltration. All underlying code is made publicly available to ease adaption of the method.

Hypertension is associated with blunted NO-mediated leg vasodilator responsiveness that is reversed by high-intensity training in postmenopausal women.

The menopausal transition is associated with increased prevalence of hypertension, and in time, postmenopausal women (PMW) will exhibit a cardiovascular disease risk score similar to male counterparts. Hypertension is associated with vascular dysfunction, but whether hypertensive (HYP) PMW have blunted nitric oxide (NO)-mediated leg vasodilator responsiveness and whether this is reversible by high-intensity training (HIT) is unknown. To address these questions, we examined the leg vascular conductance (LVC) in response to femoral infusion of acetylcholine (ACh) and sodium nitroprusside (SNP) and skeletal muscle markers of oxidative stress and NO bioavailability before and after HIT in PMW [12.9 ± 6.0 (means ± SD) years since last menstrual cycle]. We hypothesized that ACh- and SNP-induced LVC responsiveness was reduced in hypertensive compared with normotensive (NORM) PMW and that 10 wk of HIT would reverse the blunted LVC response and decrease blood pressure (BP). Nine hypertensive (HYP (clinical systolic/diastolic BP, 149 ± 11/91 ± 83 mmHg) and eight normotensive (NORM (122 ± 13/75 ± 8 mmHg) PMW completed 10 wk of biweekly small-sided floorball training (4-5 × 3-5 min interspersed by 1-3-min rest periods). Before training, the SNP-induced change in LVC was lower (P < 0.05) in HYP compared with in NORM. With training, the ACh- and SNP-induced change in LVC at maximal infusion rates, i.e., 100 and 6 µg·min-1·kg leg mass-1, respectively, improved (P < 0.05) in HYP only. Furthermore, training decreased (P < 0.05) clinical systolic/diastolic BP (-15 ± 11/-9 ± 7 mmHg) in HYP and systolic BP (-10 ± 9 mmHg) in NORM. Thus, the SNP-mediated LVC responsiveness was blunted in HYP PMW and reversed by a period of HIT that was associated with a marked decrease in clinical BP.

Proprotein convertase subtilisin/kexin type 9 inhibition as the next statin?

PURPOSE OF REVIEW: Despite the wide use of statins and other LDL-cholesterol (LDL-C)-lowering therapies, atherosclerotic cardiovascular disease remains an important cause of mortality and morbidity. Here, we discuss efficacy, side effects and convenience of current and future therapies inhibiting proprotein convertase subtilisin/kexin type 9 (PCSK9). RECENT FINDINGS: Clinical trials with mAbs administered every 2-4 weeks and small interfering RNAs given two to four times per year have consistently demonstrated substantial LDL-C-lowering (40-60%) and improved outcome when added to existing lipid-lowering therapies. Pleiotropic effects of PCSK9 inhibition are somewhat different from those observed with statin treatment as evidenced by reduced levels of triglycerides and lipoprotein(a) with no apparent effect on inflammatory markers in patients treated with PCSK9 inhibitors. Treatment with mAb and small interfering RNA are associated with a high-cost, however, small molecules and vaccines may improve cost and convenience if development of these are successful. SUMMARY: PCSK9 inhibitors are currently considered to be an add-on therapy and whether these drugs will be used as stand-alone and/or as a first choice is dependent on clinical readouts from ongoing and future trials, real-world evidence, convenience and treatment costs.

A neutralizing antibody against DKK1 does not reduce plaque formation in classical murine models of atherosclerosis: Is the therapeutic potential lost in translation?

BACKGROUND AND AIMS: Clinical interventions targeting nonlipid risk factors are needed given the high residual risk of atherothrombotic events despite effective control of dyslipidemia. Dickkopf-1 (DKK1) plays a lipid-independent role in vascular pathophysiology but its involvement in atherosclerosis development and its therapeutic attractiveness remain to be established. METHODS: Patient data, in vitro studies and pharmacological intervention in murine models of atherosclerosis were utilized. RESULTS: In patients' material (n = 127 late stage plaque specimens and n = 10 control vessels), DKK1 mRNA was found to be higher in atherosclerotic plaques versus control arteries. DKK1 protein was detected in the luminal intimal area and in the necrotic core of plaques. DKK1 was released from isolated primary human platelets (~12 - 21-fold) and endothelial cells (~1.4-2.5-fold) upon stimulation with different pathophysiological stimuli. In ApoE-/- and Ldlr-/- mice, plasma DKK1 concentrations were similar to those observed in humans, whereas DKK1 expression in different atheroprone arterial segments was very low/absent. Chronic treatment with a neutralizing DKK1 antibody effectively reduced plasma concentrations, however, plaque lesion area was not reduced in ApoE-/- and Ldlr-/- mice fed a western diet for 14 and 16 weeks. Anti-DKK1 treatment increased bone volume and bone mineral content. CONCLUSIONS: Functional inhibition of DKK1 with an antibody does not alter atherosclerosis progression in classical murine models. This may reflect the absence of DKK1 expression in plaques and more advanced animal disease models could be needed to evaluate the role and therapeutic attractiveness of DKK1 in late stage complications such as plaque destabilization, calcification, rupture and thrombosis.

Microvascular Function Is Impaired after Short-Term Immobilization in Healthy Men.

PURPOSE: We examined whether 2 wk of one-leg immobilization would impair leg microvascular function and to what extent a subsequent period of intense aerobic cycle training could restore function. METHODS: Study participants were healthy young men (n = 12; 20-24 yr of age). Leg microvascular function was determined before the intervention, after the immobilization period, and after a 4-wk exercise training period. Microvascular function was assessed as the vasodilator response to intra-arterial infusion of acetylcholine and sodium nitroprusside and as the vasoconstrictor response to endogenous noradrenaline release induced by tyramine infusion. Vasodilator enzymes as well as prooxidant and antioxidant enzymes were assessed by protein analysis in skeletal muscle samples: endothelial nitric oxide synthase, NADPH oxidase (NOX p67 and NOX gp91), and superoxide dismutase 2 (SOD2). RESULTS: The acetylcholine-induced change in vascular conductance was reduced after the 2 wk of immobilization (P = 0.003), tended to increase (P = 0.061), and was back to baseline levels after the subsequent 4 wk of exercise training. Plasma prostacyclin levels in response to acetylcholine infusion were lower after immobilization than before (P = 0.041). The changes in vascular conductance with sodium nitroprusside and tyramine were similar during all conditions. Skeletal muscle protein levels of endothelial nitric oxide synthase in the experimental leg were unchanged with immobilization and subsequent training but increased 47% in the control leg with training (P = 0.002). NOX p67, NOX gp91, and SOD2 in the experimental leg remained unaltered with immobilization, and SOD2 was higher than preimmobilization after 4 wk of training (P < 0.001). CONCLUSIONS: The study shows that 2 wk of immobilization impairs leg microvascular endothelial function and prostacyclin formation but that 4 wk of intense aerobic exercise training restores the function. The underlying mechanism may reside in the prostacyclin system.

Essential hypertension is associated with blunted smooth muscle cell vasodilator responsiveness and is reversed by 10-20-30 training in men.

Essential hypertension is associated with impairments in vascular function and sympathetic nerve hyperactivity; however, the extent to which the lower limbs are affected remains unclear. We examined the leg vascular responsiveness to infusion of acetylcholine (ACh), sodium nitroprusside (SNP), and phenylephrine (PEP) in 10 hypertensive men [HYP: age 59.5 ± 9.7 (means ± SD) yr; clinical and nighttime blood pressure: 142 ± 10/86 ± 10 and 141 ± 11/83 ± 6 mmHg, respectively; and body mass index (BMI): 29.2 ± 4.0 kg/m2] and 8 age-matched normotensive counterparts (NORM: age 57.9 ± 10.8 yr; clinical and nighttime blood pressure: 128 ± 9/78 ± 7 and 116 ± 3/69 ± 3 mmHg, respectively; and BMI: 26.3 ± 3.1 kg/m2). The vascular responsiveness was evaluated before and after 6 wk of 10-20-30 training, consisting of 3 × 5 × 10-s sprint followed by 30 and 20 s of low- to moderate-intensity cycling, respectively, interspersed by 3 min of rest. Before training, the vascular responsiveness to infusion of SNP was lower (P < 0.05) in HYP compared with NORM, with no difference in the responsiveness to infusion of ACh and PEP. The vascular responsiveness to infusion of SNP and ACh improved (P < 0.05) with training in HYP, with no change in NORM. With training, intra-arterial systolic blood pressure decreased (P < 0.05) by 9 mmHg in both HYP and NORM whereas diastolic blood pressure decreased (5 mmHg; P < 0.05) in HYP only. We provide here the first line of evidence in humans that smooth muscle cell vasodilator responsiveness is blunted in the lower limbs of hypertensive men. This impairment can be reversed by 10-20-30 training, which is an effective intervention to improve the responsiveness of smooth muscle cells in men with essential hypertension.