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Ramipril retards development of aortic valve stenosis in a rabbit model: Mechanistic considerations
British Journal of Pharmacology
Abstract
Aortic valve stenosis (AVS) is associated with significant cardiovascular morbidity and mortality. To date, no therapeutic modality has been shown to be effective in retarding AVS progression. We evaluated the effect of angiotensin-converting enzyme inhibition with ramipril on disease progression in a recently developed rabbit model of AVS.
The effects of 8 weeks of treatment with either vitamin D₂ at 25,000 IU for 4 days a week alone or in combination with ramipril (0.5 mg·kg⁻¹) on aortic valve structure and function were examined in New Zealand white rabbits. Echocardiographic aortic valve backscatter (AV(BS)) and aortic valve:outflow tract flow velocity ratio were utilized to quantify changes in valve structure and function.
Treatment with ramipril significantly reduced AV(BS) and improved aortic valve :outflow tract flow velocity ratio. The intravalvular content of the pro-oxidant thioredoxin-interacting protein was decreased significantly with ramipril treatment. Endothelial function, as measured by asymmetric dimethylarginine concentrations and vascular responses to ACh, was improved significantly with ramipril treatment.
Ramipril retards the development of AVS, reduces valvular thioredoxin-interacting protein accumulation and limits endothelial dysfunction in this animal model. These findings provide important insights into the mechanisms of AVS development and an impetus for future human studies of AVS retardation using an angiotensin-converting enzyme inhibitor.
... These negative results can be attributed to differences between these two entities including a greater extent of calcification in late-stage AV stenosis [7]. Indeed, recent studies have failed to show any effectiveness of cholesterol-lowering therapy on the progression of AV stenosis or even any association between hypercholesterolaemia and AV disease in an aging population [12][13][14][15][16]. Similarly, experimental studies with systemic or local application of therapeutic agents either failed or showed limited results as they lack the in vivo proof of attenuation of calcification [12,[17][18][19]. ...
... These negative results can be attributed to differences between these two entities including a greater extent of calcification in late-stage AV stenosis [7]. Indeed, recent studies have failed to show any effectiveness of cholesterol-lowering therapy on the progression of AV stenosis or even any association between hypercholesterolaemia and AV disease in an aging population [12][13][14][15][16]. Similarly, experimental studies with systemic or local application of therapeutic agents either failed or showed limited results as they lack the in vivo proof of attenuation of calcification [12,[17][18][19]. In a recent study, ACE inhibitors have proven to have a beneficial effect on the left ventricular mass of asymptomatic patients with moderate to severe AV A Β C stenosis, together with an attenuation of the progression of AV stenosis. ...
... The current experimental model is the most widely used as it has important similarities but also differences with the human AV [10,12,34,35]. The main advantages are the triple layer of the valve and the similar lipoprotein metabolism. ...
The aim of this study was to evaluate in an experimental model of aortic valve (AV) stenosis the effectiveness of zoledronate on the inhibition of calcification. Sixteen New Zealand rabbits were placed on vitamin D-enriched diet for 3 weeks. All animals underwent PET/CT at baseline and before euthanasia to assess calcification. Thereafter, the AVs of eight animals were treated with local delivery of 500 μg/l zoledronate. A placebo mixture was administered in the remaining eight animals. Standardized uptake values were corrected for blood pool activity, providing mean tissue to background ratios (TBRmean). In the zoledronate group, there was no progression of AV calcification (TBRmean 1.20 ± 0.12 vs 1.17 ± 0.78,p = 0.29), while AV calcification progressed in the placebo group (1.22 ± 0.15 vs 1.53 ± 0.23,p = 0.006). Ascending aorta (AA) calcification progressed in both zoledronate and placebo groups. Histology confirmed the results of the PET/CT. Inhibition of AV calcification by local delivery of zoledronate is feasible and effective.
... Blockade of the renin-angiotensin-aldosterone system has been suggested recently as a potential therapy for aortic valve sclerosis and arterial calcification [9][10][11][12][13]. Angiotensin receptor blockade can inhibit arterial calcification by disrupting vascular osteogenesis. ...
... Human heart valves can be successfully replaced with bioprosthetic valves; however, eventual degeneration of the valve leaflet due to calcification is a concern. Several studies have reported a relationship between Ang II and degenerative aortic stenosis with calcification [9][10][11][13][14][15]. To our knowledge, there are no reports about the relationship between Ang II and bovine pericardial xenograft calcification. ...
... Many studies have investigated calcification of the aortic valve and arteries to elucidate the relationship between Ang II and valve calcification [9][10][11][12][13], and to evaluate a preventive role for ACE inhibitors or ARBs on calcification [13,19]. Ang II has a number of potential lesion-promoting effects, such as those mediated by the Ang II type 1 receptor. ...
OBJECTIVES
There is evidence that angiotensin II type I receptor blocker (ARB) could reduce structural valve deterioration. However, the anticalcification effect on the bioprosthetic heart valve (BHV) has not been investigated. Thus, we investigated the effects of losartan (an ARB) on calcification of implanted bovine pericardial tissue in a rabbit intravascular implant model.
METHODS
A total of 16 male New Zealand White rabbits (20 weeks old, 2.98–3.34 kg) were used in this study. Commercially available BHV leaflet of bovine pericardium was trimmed to the shape of a 3-mm triangle and implanted to both external jugular veins of the rabbit. The ARB group (n = 8) was given 25 mg/kg of powdered losartan daily until 6 weeks after surgery by direct administration in the buccal pouch of the animals. The control group (n = 8) was given 5 ml of normal saline by the same method. After 6 weeks, quantitative calcium determination, histological evaluation and western blot analysis of interleukin-6 (IL-6), osteopontin and bone morphogenetic protein 2 (BMP-2) were performed to investigate the mechanisms of the anticalcification effect of losartan.
RESULTS
No deaths or complications such as infection or haematoma were recorded during the experiment. All animals were euthanized on the planned date. The calcium measurement level in the ARB group (2.28 ± 0.65 mg/g) was significantly lower than that in the control group (3.68 ± 1.00 mg/g) (P = 0.0092). Immunohistochemistry analyses revealed that BMP-2-positive reactions were significantly attenuated in the ARB group. Western blot analysis showed that losartan suppressed the expression of IL-6, osteopontin and BMP-2.
CONCLUSIONS
Our results indicate that losartan significantly attenuates postimplant degenerative calcification of a bovine pericardial bioprosthesis in a rabbit intravascular implant model. Further studies are required to assess the effects of ARBs on BHV tissue in orthotopic implantations using a large animal model.
... Сегодня РААС остается главной терапевтической мишенью в поиске медикаментозного лечения КАС [35]. Результаты экспериментальных работ на животных показывают, что ингибирование РААС способствует уменьшению неблагоприятных гистопатологических событий, включающих развитие эндотелиальной дисфункции и фиброза, накопление липидов и формирование лейкоцитарных инфильтратов [49,100,101]. Кроме того, использование иАПФ способствует замедлению гемодинамической прогрессии КАС у подопытных кроликов [100]. ...
... Результаты экспериментальных работ на животных показывают, что ингибирование РААС способствует уменьшению неблагоприятных гистопатологических событий, включающих развитие эндотелиальной дисфункции и фиброза, накопление липидов и формирование лейкоцитарных инфильтратов [49,100,101]. Кроме того, использование иАПФ способствует замедлению гемодинамической прогрессии КАС у подопытных кроликов [100]. ...
Calcific aortic valve stenosis (CAVS) is a serious socio-economic problem in developed countries because this disease is the most common indication for aortic valve replacement. Currently, there are no methods for non-invasive treatment of CAVS. Nevertheless, it is assumed that effective drug therapy for CAVS can be developed on the basis of modulators of the renin-angiotensin-aldosterone system (RAAS), which is involved in the pathogenesis of this disease. The purpose of this paper is to compile and analyze current information on the role of RAAS in the CAVS pathophysiology. Recent data on the effectiveness of RAAS inhibition are reviewed.
... In addition to reducing systemic blood pressure, RAAS inhibition reduces LV hypertrophy and improves survival and other important patient-oriented clinical outcomes in patients with heart failure and reduced LV ejection fraction (LVEF). 75-82 RAAS inhibition is recommended in AS because it slows AS progression, 29,83,84 attenuates myocardial remodeling in response to increased afterload, 85,86 and exerts protective effects on the vasculature ( Figure 2). 87,88 Human aortic valves express prorenin, renin, ACE, and angiotensin II type 2 receptors. ...
... Angiotensin II and chymase (another enzyme involved in the synthesis of angiotensin II) 84 expression is upregulated in stenotic aortic valves, 89 and blockade of the RAAS prevents valve calcification and hemodynamic progression of AS both in experimental models of AS and in humans. 31,83 RAAS inhibition attenuates cardiomyocyte hypertrophy and myocardial fibrosis to a greater extent than other vasodilators for a given reduction in systolic blood pressure. 19 This suggests that RAAS inhibition has pleiotropic effects on the myocardium that go beyond afterload reduction. ...
Untreated, severe, symptomatic aortic stenosis is associated with a dismal prognosis. The only treatment shown to improve survival is aortic valve replacement; however, before symptoms occur, aortic stenosis is preceded by a silent, latent phase characterized by a slow progression at the molecular, cellular, and tissue levels. In theory, specific medical therapy should halt aortic stenosis progression, reduce its hemodynamic repercussions on left ventricular function and remodeling, and improve clinical outcomes. In the present report, we performed a systematic review of studies focusing on the medical treatment of patients with aortic stenosis. Lipid-lowering therapy, antihypertensive drugs, and anticalcific therapy have been the main drug classes studied in this setting and are reviewed in depth. A critical appraisal of the preclinical and clinical evidence is provided, and future research avenues are presented.
... For example, in vitamin D-treated rabbits, aortic valve calcification was associated with increased [75]. Concomitant treatment with ramipril diminished valve narrowing, and was associated with a reduction in intra-valvular accumulation of TXNIP (Fig. 6), as well as with salvage of NO signaling [76]. Furthermore, in humans at increased risk of coronary events, 2 weeks' therapy with ramipril decreased intra-platelet TXNIP expression while potentiating NO response [38]. ...
... Specifically: Fig. 6 Section of micrographs: rabbit aortic valve A treated with vitamin D 2 alone, and B treated with vitamin D 2 plus ramipril. Note suppression of TXNIP expression (brown pigment) with ramipril treatment (Reproduced with permission from reference [76]) a) PARP-1 inhibitors (e.g. rucaparib, olaparib) are currently under investigation predominantly as adjuncts to antineoplastic therapy (for review see reference [89]). ...
The thioredoxin system, which consists of thioredoxin (Trx), nicotinamide adenine dinucleotide phosphate (NADPH) and thioredoxin reductase (TrxR), has emerged as a major anti-oxidant involved in the maintenance of cellular physiology and survival. Dysregulation in this system has been associated with metabolic, cardiovascular, and malignant disorders. Thioredoxin-interacting protein (TXNIP), also known as vitamin D-upregulated protein or thioredoxin-binding-protein-2, functions as a physiological inhibitor of Trx, and pathological suppression of Trx by TXNIP has been demonstrated in diabetes and cardiovascular diseases. Furthermore, TXNIP effects are partially Trx-independent; these include direct activation of inflammation and inhibition of glucose uptake. Many of the effects of TXNIP are initiated by its dissociation from intra-nuclear binding with Trx or other SH-containing proteins: these effects include its migration to cytoplasm, modulating stress responses in mitochondria and endoplasmic reticulum, and also potentially activating apoptotic pathways. TXNIP also interacts with the nitric oxide (NO) signaling system, with apparent suppression of NO effect. TXNIP production is modulated by redox stress, glucose levels, hypoxia and several inflammatory activators. In recent studies, it has been shown that therapeutic agents including insulin, metformin, angiotensin converting enzyme inhibitors and calcium channel blockers reduce TXNIP expression, although it is uncertain to what extent TXNIP suppression contributes to their clinical efficacy. This review addresses the role of TXNIP in health and in cardiovascular and metabolic disorders. Finally, the potential advantages (and disadvantages) of pharmacological suppression of TXNIP in cardiovascular disease and diabetes are summarized.
... We have recently demonstrated in a rabbit model of mild AS with histological features similar to that of human disease, increased intravalvular concentration of TXNIP compared to control animals [53] . Furthermore , co-treatment with ramipril retarded the development of AS in this model, as measured by reduction both in transvalvular velocity and valve echogenicity on echocardiography [59]. This retardation further correlated with reduction in valvular calcium and macrophage infiltration and a reduction in TXNIP accumulation within the valve matrix. ...
... Unfortunately, no hemodynamic valvular measures of AS progression were performed. We have demonstrated hemodynamic retardation of AS, concomitantly with reduction in calcification, macrophage infiltration, redox stress and improvement in endothelial function, with ACEI ramipril treatment in a rabbit model of AS [59]. ...
Aortic valve stenosis (AS) is the commonest form of valvular heart disease in the Western world. Its prevalence increases exponentially with age and it is present in 2-7% of all patients over 65 years of age. In view of the considerable cardiovascular morbidity and mortality associated not only with AS, but even its earlier stage, aortic sclerosis, many investigations have been directed towards better understanding of its pathogenesis, with the ultimate objective of developing strategies to retard its progression. Although risk factors and downstream mediators appear similar for AS and atherosclerosis (older age, male sex, hypertension, smoking, hypercholesterolemia, and diabetes, as many as 50% of patients with AS do not have clinically significant atherosclerosis. On the basis both of recent experimental evidence and clinical trials, it appears that atherogenesis is not pivotal to the pathogenesis of AS. On the other hand, there is increasing evidence of active involvement of aortic valve fibroblasts with resultant increased production of reactive oxygen species, active pro-inflammatory and pro-fibrotic processes culminating in calcification. We also discuss the evidence of involvement of the nitric oxide system in the pathogenesis of AS. The renin-angiotensin system has also emerged as a major player in the pathogenesis of AS. Histologically, there is increased ACE expression and elevated angiotensin II levels in stenotic valves, while we have just demonstrated amelioration of AS with the use of ACE inhibitors in an animal model. We further discuss intervention studies aimed at retarding AS progression, including recent failures of statins to retard progression of AS in large randomized clinical studies. Finally, we discuss the special case of bicuspid aortic valve, including its genetics and unique associated features.
... Candesartan, although well tolerated by patients with severe symptomatic CAS, was not shown to improve LV geometry or function, or symptoms, compared with placebo (The Potential of Candesartan to Retard the Progression of Aortic Stenosis-ROCK-AS trial) [50], while two studies by Côté and co-workers showed that ARBs contribute to reduction of fibrocalcific remodeling of the aortic valves and inflammation [51,52]. In an experimental rabbit model, ramipril was reported to retard the development of CAS [53], but in a study of 211 patients with CAS, ACEI were not shown to slow the disease progression [54]. A Canadian report on 338 subjects with CAS showed that hypertension was associated with faster CAS progression and higher incidence of clinical events, which were abolished by ARBs, but not ACEI [55]. ...
Calcific aortic stenosis (CAS) is the most common cause of acquired valvular heart disease in adults with no available pharmacological treatment to inhibit the disease progression to date. This review provides an up-to-date overview of current knowledge of molecular mechanisms underlying CAS pathobiology and the related treatment pathways. Particular attention is paid to current randomized trials investigating medical treatment of CAS, including strategies based on lipid-lowering and antihypertensive therapies, phosphate and calcium metabolism, and novel therapeutic targets such as valvular oxidative stress, coagulation proteins, matrix metalloproteinases, and accumulation of advanced glycation end products.
... The efficacy of ACEIs in the treatment of aortic stenosis has been investigated in animals and has produced some promising effects [16]. However, the findings of large retrospective studies have been contradictory, showing both protective and neutral effects in the progression of calcified valvular degeneration [17,18]. ...
It is essential to study the factors associated with the evolution of aortic stenosis progression (ASP) to develop therapies that could reduce it. We studied 283 patients 6 months after acute coronary syndrome (ACS). ASP was defined as an increase in the maximum aortic velocity of at least 0.5 m/s between the echocardiogram performed during ACS hospitalization and the last one recorded in the electronic medical registry. The median follow-up was 72.4 months. Twenty patients (7%) had ASP. A multivariate binary logistic regression analysis was performed showing that PCSK9 plasma levels (OR, 0.668 CI (0.457–0.977); p = 0.038), HS-CRP (OR, 1.034 CI (1.005–1.063); p = 0.022), the presence of dyslipidemia (OR, 4.622 CI (1.285–16.618); p = 0.019), the history of PAD (OR, 9.453 CI (1.703–52.452); p = 0.010), and GFR (OR, 0.962 CI (0.939–0.986); p = 0.002) were independent predicting factors of ASP. In patients with ischemic heart disease, low plasma levels of PCSK9 and elevated levels of HS-CRP are independent predictors of ASP.
... Animal studies suggested that administration of ACE inhibitors enalapril and ramipril or angiotensin II receptor blocker olmesartan improves endothelial function and prevents endothelial injury, LDL deposition, macrophage infiltration, and fibrosis [266,269,270]. Randomised clinical trials on use of enalapril, trandolapril (NCT00252317), and ramipril in patients with CAVD demonstrated moderate efficacy including decrease in left ventricular end-systolic volume and left ventricular mass as well as improved effort tolerance [271][272][273]. ...
Calcific aortic valve disease (CAVD), previously thought to represent a passive degeneration of the valvular extracellular matrix (VECM), is now regarded as an intricate multistage disorder with sequential yet intertangled and interacting underlying processes. Endothelial dysfunction and injury, initiated by disturbed blood flow and metabolic disorders, lead to the deposition of low-density lipoprotein cholesterol in the VECM further provoking macrophage infiltration, oxidative stress, and release of pro-inflammatory cytokines. Such changes in the valvular homeostasis induce differentiation of normally quiescent valvular interstitial cells (VICs)into synthetically active myofibroblasts producing excessive quantities of the VECM and proteins responsible for its remodeling. As a result of constantly ongoing degradation and re-deposition, VECM becomes disorganised and rigid, additionally potentiating myofibroblastic differentiation of VICs and worsening adaptation of the valve to the blood flow. Moreover, disrupted and excessively vascularised VECM is susceptible to the dystrophic calcification caused by calcium and phosphate precipitating on damaged collagen fibers and concurrently accompanied by osteogenic differentiation of VICs. Being combined, passive calcification and biomineralisation synergistically induce ossification of the aortic valve ultimately resulting in its mechanical incompetence requiring surgical replacement. Unfortunately, multiple attempts have failed to find an efficient conservative treatment of CAVD; however, therapeutic regimens and clinical settings have also been far from the optimal. In this review, we focused on interactions and transitions between aforementioned mechanisms demarcating ascending stages of CAVD, suggesting a predisposing condition (bicuspid aortic valve)and drug combination (lipid-lowering drugs combined with angiotensin II antagonists and cytokine inhibitors)for the further testing in both preclinical and clinical trials.
... в экспериментальной модели показано, что прием иАПФ уменьшает кальцификацию и макрофагальную инфильтрацию, улучшает функцию эндотелия и замедляет формирование аортального стеноза [36]. Похожие результаты были получены в ретроспективном клиническом исследовании, включавшем 194 пациентов с аортальным стенозом легкой степени (v max = 2,36 ± 0,79 м/с). ...
The calcined aortic stenosis is one of the most frequently occurring cardio-vascular diseases with unfavorable prognosis of treatment. Te actual conceptions of risk factors of development, pathogenesis and course of disease starting from inflammation and endothelial dysfunction and completing with bone metaplasia give an opportunity of determining prospective directions of medicinal therapy favoring deceleration of progressing of calcined aortic stenosis. The review summarizes main data of publications concerning attempts of pathogenetically medicinal impact on progressing of calcined aortic stenosis.
... A number of studies have documented an increased intravalvular content of several pro-oxidants, either in experimental models or in clinical models of AS [23,[43][44][45][46]. In particular, Billaud et al. [29] demonstrated that in patients with BAV, the proximal aorta is characterized by heightened superoxide anion production and oxidative damage [29]. ...
Introduction:
Bicuspid aortic valve (BAV) is the most common congenital valvular heart defect resulting from abnormal aortic cusp formation during heart development, where two of the three normal and equal sized cusps fuse into a single large cusp resulting in a two cusps aortic valve. Over the past years, much interest has been given in understanding the pathogenesis of BAV and its complications. In this review, we focused on the role of inflammation, involved in the degeneration of BAV and the development of its complications.
Role of inflammation:
From a pathophysiological point of view, BAV may rapidly progress into aortic stenosis (AS) and is related to aortopathy. Several histopathologic studies have demonstrated that the development and progression of alterations in bicuspid aortic valve are related to an active process that includes: oxidative stress, shear stress, endothelial dysfunction, disorganized tissue architecture, inflammatory cells and cytokines. These factors are closely related one to each other, constituting the basis of the structural and functional alterations of the BAV.
Conclusion:
Chronic inflammation plays a key role in the degeneration of BAV. Severe aortic stenosis in bicuspid aortic valves is associated with a more aggressive inflammatory process, increased inflammatory cells infiltration and neovascularization when compared to tricuspid AS. These findings might help to explain the more frequent onset and rapid progression of AS and the heavy aortic valve calcification seen in patients with BAV.
... A number of currently available therapeutic agents for human diseases have shown to have the potential to control Txnip expression to some degree. These agents include calcium channel blockers, 95,96 angiotensin-converting enzyme inhibitors, 6,97 angiotensin-II receptor blockers, 98 glucagon-like peptide-1 agonists, 99 and insulin. 20 To date, however, there is no pharmacological therapy specifically designed to inhibit Txnip expression. ...
Myocardial ischemia/reperfusion injury represents a major threat to human health and contributes to adverse cardiovascular outcomes worldwide. Despite the identification of numerous molecular mechanisms, understanding of the complex pathophysiology of this clinical syndrome remains incomplete. Thioredoxin-interacting protein (Txnip) has been of great interest in the past decade since it has been reported to be a critical regulator in human diseases with several important cellular functions. Thioredoxin-interacting protein binds to and inhibits thioredoxin, a redox protein that neutralizes reactive oxygen species (ROS), and through its interaction with thioredoxin, Txnip sensitizes cardiomyocytes to ROS-induced apoptosis. Interestingly, evidence from recent studies also suggests that some of the effects of Txnip may be unrelated to changes in thioredoxin activity. These pleiotropic effects of Txnip are mediated by interactions with other signaling molecules, such as nod-like receptor pyrin domain-containing 3 inflammasome and glucose transporter 1. Indeed, Txnip has been implicated in the regulation of inflammatory response and glucose homeostasis during myocardial ischemia/reperfusion injury. This review attempts to make the case that in addition to interacting with thioredoxin, Txnip contributes to some of the pathological consequences of myocardial ischemia and infarction through endogenous signals in multiple molecular mechanisms.
... The lack of efficacy of the lipid lowering strategy suggests that once mineral deposition in the leaflet is initiated, the calcium accrual might occur independently from further cholesterol accumulation in the valve. Efficacy of ACE-inhibitors on valve calcific degeneration has been investigated in animal studies, which showed some promising effects [78]. However, findings from large retrospective studies have been contradictory showing both protective and neutral effects ACE-inhibitors on the progression of calcific valve degeneration [79,80]. ...
Several clinical studies reported an increased prevalence and accelerated progression of aortic valve calcification among patients with end-stage renal disease when compared with subjects with normal kidney function. Recently, mechanisms of calcific valve degeneration have been further elucidated and many of the pathways involved could be amplified in patients with decreased renal function. In particular, calcium-phosphate balance, MGP metabolism, OPG/RANK/RANKL triad, fetuin-A mineral complexes and FGF-23/Klotho axis have been shown to be impaired among patients with advanced chronic kidney disease and could play a role during vascular/valve calcification. The scope of the present review is to summarize the clinical data and the pathophysiological mechanisms potentially involved in the link between renal function decline and the progression of aortic valve disease.
... We have previously shown that with IB calcific deposits of the aortic valve can be quantitatively and reproducibly assessed in rats [7,8]. Others have used IB for the quantitative detection of aortic valve calcifications in vitamin D treated rabbits, which could be delayed with ramipril [24,25]. Preliminary clinical studies have demonstrated that higher backscatter scores are seen in sclerotic compared to non-sclerotic aortic valves, with a further increase in stenotic valves [26,27]. ...
Background
Calcification is an independent predictor of mortality in calcific aortic valve disease (CAVD). The aim of this study was to evaluate the use of non-invasive, non-ionizing echocardiographic calibrated integrated backscatter (cIB) for monitoring progression and subsequent regression of aortic valvular calcifications in a rat model of reversible renal failure with CAVD, compared to histology.
Methods
28 male Wistar rats were prospectively followed during 21 weeks. Group 1 (N=14) was fed with a 0.5% adenine diet for 9 weeks to induce renal failure and CAVD. Group 2 (N=14) received a standard diet. At week 9, six animals of each group were killed. The remaining animals of group 1 (N=8) and group 2 (N=8) were kept on a standard diet for an additional 12 weeks. cIB of the aortic valve was calculated at baseline, 9 and 21 weeks, followed by measurement of the calcified area (Ca Area) on histology.
Results
At week 9, cIB values and Ca Area of the aortic valve were significantly increased in the adenine-fed rats compared to baseline and controls. After 12 weeks of adenine diet cessation, cIB values and Ca Area of group 1 decreased compared to week 9, while there was no longer a significant difference compared to age-matched controls of group 2.
Conclusions
cIB is a non-invasive tool allowing quantitative monitoring of CAVD progression and regression in a rat model of reversible renal failure, as validated by comparison with histology. This technique might become useful for assessing CAVD during targeted therapy.
... It was reported that activity or expression of angiotensin II (Ang II)producing systems in human calcific aortic valves [10e12], suggesting the role of renin-angiotensin system (RAS) in the aggravation of destruction of valvular morphology. Until now, no prospective trials exists examining the effects of angiotensin converting enzyme inhibitor (ACEI) or Ang II type 1 (AT1) receptor blocker on AS progression; however, effects of RAS inhibitors have been tested in several previous animal studies [13]. We also showed previously that olmesartan suppressed atherosclerotic degeneration of the aortic valve in hypercholesterolemic rabbits q This work was partly presented at the American Heart Association (AHA) 2010 Scientific Sessions, Chicago, Illinois, November 13 to 17, 2010, and published in abstract form (Circulation. 2010; 122: A19135). ...
Background:
Valvular aortic stenosis (AS) is not an infrequent condition in the aged population. Activation of renin-angiotensin system (RAS) is presumed to be involved in the development of AS; however, direct evidence seems to be limited. We herein examined the effect of the administration of angiotensin II (Ang II) on the development of aortic valve thickening in apolipoprotein-E (ApoE)-deficient mice.
Methods and results:
Male ApoE-deficient mice were divided into three groups: control (saline, n = 8), mice that were administered low-dose Ang II (500 ng/kg/min, n = 11), and those with high-dose Ang II (1000 ng/kg/min, n = 11) administration for 4 weeks. Administration of high-dose, but not low-dose, Ang II significantly induced aortic valve thickening. It was found that in the aortic valve leaflets of high-dose Ang II group, integrity of endothelial cells was impaired and the number of myofibroblasts was increased. These phenomena induced by high-dose Ang II were suppressed by Ang II type 1 receptor blocker olmesartan (n = 15), but not by the dilatator, hydralazine (n = 13). Olmesartan also suppressed dilatation of aortic diameter, although it did not significantly affect the plaque area, in the abdominal aorta in ApoE-deficient mice.
Conclusion:
Administration of Ang II to genetically hyperlipidemic mice induced aortic valve thickening by a pressor-independent mechanism. Role of RAS activation in the development of AS in dyslipidemic patients should further be investigated.
... ACE inhibitors have shown mixed results in retrospective clinical studies with one showing a strong association between the use of ACE inhibitors and decreased valve calcification [168] and the other finding no effects [169]. In NZWR, treatment with the ACE inhibitor ramipril concomitantly with induction of CAVD by VitD2 at 25000 IU (four days per week) for eight weeks retarded the development of CAVD [99]. The ARB olmesartan decreased macrophage and myofibroblast accumulation, decreased OPN, ACE, and Runx2 mRNA expression, and maintained endothelial integrity when administered to NZWR during the last four weeks of an eight-week 1% cholesterol diet treatment [95]. ...
Calcific aortic valve disease (CAVD), once thought to be a degenerative disease, is now recognized to be an active pathobiological process, with chronic inflammation emerging as a predominant, and possibly driving, factor. However, many details of the pathobiological mechanisms of CAVD remain to be described, and new approaches to treat CAVD need to be identified. Animal models are emerging as vital tools to this end, facilitated by the advent of new models and improved understanding of the utility of existing models. In this paper, we summarize and critically appraise current small and large animal models of CAVD, discuss the utility of animal models for priority CAVD research areas, and provide recommendations for future animal model studies of CAVD.
Calcific Aortic Valve Disease (CAVD) is prevalent among the elderly as the most common valvular heart disease. Currently, no pharmaceutical interventions can effectively reverse or prevent CAVD, making valve replacement the primary therapeutic recourse. Extensive research spanning decades has contributed to the establishment of animal and in vitro cell models, which facilitates a deeper understanding of the pathophysiological progression and underlying mechanisms of CAVD. In this review, we provide a comprehensive summary and analysis of the strengths and limitations associated with commonly employed models for the study of valve calcification. We specifically emphasize the advancements in three-dimensional culture technologies, which replicate the structural complexity of the valve. Furthermore, we delve into prospective recommendations for advancing in vivo and in vitro model studies of CAVD.
Degenerative aortic stenosis is a growing clinical problem owing to the high incidence in an aging population and its significant morbidity and mortality. Currently, aortic valve replacement remains the only treatment. Despite promising observational data, pharmacological management to slow or halt progression of aortic stenosis has remained elusive. Nevertheless, with a greater understanding of the mechanisms which underpin aortic stenosis, research has begun to explore novel treatment strategies. This review will explore the historical agents used to manage aortic stenosis and the emerging agents that are currently under investigation.
Introduction
Recommended therapy for calcific degenerative aortic stenosis (AS) is still aortic valve replacement (AVR), either transcatheter or surgical, since no conclusive efficacy has been determined in slowing the degenerative process by medical therapy.
Areas covered
This paper offers a brief overview of molecular mechanisms leading to calcification of aortic valve. It is then focused on potential markers of disease progression, as observed in many observational studies. Finally it provides a comprehensive review of drugs already tested in in vitro and human studies in order to slow aortic valve stenosis process.
Expert opinion
Despite research providing numerous molecular pathways underlying the calcification process, further efforts must be made to understand risk factors linked to disease progression. Some existing treatments that have already provided survival benefits in many features of cardiovascular diseases are currently being tested with promising results. In the near future new drugs acting on specific pathways by techniques such as monoclonal antibodies and RNA interference, are expected to provide better medical solutions for this ever growing number of patients.
Монографія присвячена одній з найактуальніших і складних проблем
клінічної медицини – діагностиці та лікуванню серцево-судинних захворювань. Розглянуті питання епідеміології гіпертонічної хвороби з урахуванням поліморфізму окремих генів в різних регіонах України, висвітлені
вікові зміни ендотеліальної функції при цьому захворюванні, сучасні стратегії лікування артеріальної гіпертензії. Представлені сучасні підходи до
діагностики вторинної артеріальної гіпертензії при ендокринних захворюваннях, а також до менеджменту вагітних із прееклампсією. Детально розглянуті сучасні підходи до ведення хворих з серцево-судинною патологією
та коморбідними станами та питання фармакотерапії серцево-судинних
захворювань.
Для кардіологів, лікарів загальної практики – сімейної медицини, нефрологів та ендокринологів.
Aortic valve stenosis has become the most common valvular heart disease on account of aging population and increasing life expectancy. Echocardiography is the primary diagnosis tool for this, but it still has many flaws. Therefore, advanced cardiovascular multimodal imaging techniques are continuously being developed in order to overcome these limitations. Cardiac magnetic resonance imaging (CMR) allows a comprehensive morphological and functional evaluation of the aortic valve and provides important data for the diagnosis and risk stratification in patients with aortic stenosis. CMR can functionally assess the aortic flow using two-dimensional and time-resolved three-dimensional velocity-encoded phase-contrast techniques. Furthermore, by late gadolinium enhancement and T1-mapping, CMR can reveal the presence of both irreversible replacement and diffuse interstitial myocardial fibrosis. Moreover, its role in guiding aortic valve replacement procedures is beginning to take shape. Recent studies have rendered the importance of active and passive biomechanics in risk stratification and prognosis prediction in patients with aortic stenosis, but more work is required is just in its infancy, but data are promising. In addition, cardiac computed tomography is particularly useful for the diagnosis of aortic valve stenosis, and in preprocedural evaluation of the aorta, while positron emission tomography can be also used to assess valvular inflammation and active calcification. The purpose of this review is to provide a comprehensive overview of current available data regarding advanced cardiovascular multimodal imaging in aortic stenosis.
Монографія присвячена одній з найактуальніших і складних проблем клінічної медицини – діагностиці та лікуванню серцево-судинних захво-рювань. Розглянуті питання епідеміології гіпертонічної хвороби з ураху-ванням поліморфізму окремих генів в різних регіонах України, висвітлені вікові зміни ендотеліальної функції при цьому захворюванні, сучасні стра-тегії лікування артеріальної гіпертензії. Представлені сучасні підходи до діагностики вторинної артеріальної гіпертензії при ендокринних захворю-ваннях, а також до менеджменту вагітних із прееклампсією. Детально роз-глянуті сучасні підходи до ведення хворих з серцево-судинною патологією та коморбідними станами та питання фармакотерапії серцево-судинних захворювань.
Для кардіологів, лікарів загальної практики – сімейної медицини, не-фрологів та ендокринологів
Aortic stenosis is one of the most common cardiovascular diseases in the world. Extensive work on the underlying pathophysiology responsible for calcific aortic valve disease and its progression to aortic stenosis, have described a complex process involving inflammation, lipid deposition, mineralization, and genetic factors such as elevated lipoprotein(a). With the advancement of gene silencing technology and development of novel therapeutic agents, we may now be closer than ever to having medical therapies that prevent, or at least slow the progression of aortic stenosis. In this review, we highlight the pathophysiology and risk factors of calcific aortic valve disease, along with current, potential, and emerging novel medical therapies. We also provide potential explanations for the failure of statin trials and suggest new avenues for research, and new randomized trials, in this area.
Le rétrécissement aortique calcifie (RAC) est la valvulopathie la plus fréquente dans les pays occidentaux et touche environ 2% des sujets de plus de 65 ans. Initialement considérée comme une dégénérescence passive de la valve aortique liée à l’âge, le RAC est actuellement considéré comme une pathologie complexe et hautement régulée et qui aboutit à un épaississement et à une calcification des feuillets valvulaires aortiques. A ce jour les mécanismes initiateurs et favorisant la progression du RAC ne sont pas complètement élucidés, et d’autre part, aucun traitement ne s’est avéré efficace pour ralentir ou stopper son évolution. Le remplacement valvulaire aortique (chirurgical ou percutané) reste le seul traitement en cas de rétrécissement aortique serré. D’autre part, il n’existe aucun modèle animal fiable et reproductible du RAC permettant de mieux comprendre la physiopathologie de cette valvulopathie et de tester de nouvelles cibles thérapeutiques. Dans le laboratoire, nous avons fait l’hypothèse que la dysfonction de l’endothélium valvulaire jouait un rôle important dans l’initiation et la progression du RAC, et nous souhaitons utiliser un modèle animal permettant d’évaluer l’effet de nouvelles thérapeutiques. Ce travail porte donc sur l’évaluation du rôle du système endothélinergique dans la sténose aortique calcifiée et le développement d’un nouveau modèle animal murin de RAC. Dans notre première étude, nous nous sommes intéressés au modèle de souris EgfrWa2/Wa2. Ce modèle est induit par la substitution du résidu amine glycine par une valine. Les souris EgfrWa2/Wa2 homozygotes pour la mutation voient leur activité tyrosine kinase diminuée de 90%. Ce modèle induit un épaississement fibreux des feuillets valvulaires avec peu de calcifications. Cependant, l’augmentation du flux transaortique n’est pas liée a un RAC mais a une insuffisance aortique (IA). Nous avons évalué dans ce modèle l’effet d’un régime enrichi en graisse et d’une supplémentation en vitamine D3. Malgré une augmentation des taux sériquesdu cholestérol, de la vitamine D3 et de la calcémie, nous n’avons pas observé d’augmentation de la calcification. Le modèle EgfrWa2/Wa2 reste essentiellement un modèle d’IA avec le développement d’un RAC qui reste rare ou absent. Dans notre seconde étude, nous avons évalué le rôle du système endothélinergique dans des cultures primaires de cellules valvulaires interstitielles (hVICs) et endothéliales (hVECs) humaines, prélevées lors d’un remplacement valvulaire aortique chirurgical chez des patients ayant un RAC serré. Les valves de patients traités par une intervention de Bentall ont été utilisées comme groupe contrôle. Nous avons tout d’abord observé, par RT-PCR quantitative, que les hVECs issues de valves sténosées présentaient une augmentation non significative de l’ARNm codant pour l’endothéline et une augmentation significative de l’ARNm codant pour le récepteur ET-B, ainsi qu’une diminution significative de l’enzyme de conversion de l’endothéline dans les hVECs par rapport aux hVECs issues de valves contrôles. Le récepteur ET-B était également surexprimé dans les hVICs par rapport aux valves contrôles. En revanche, nous n’avons pas retrouvé de variation d’expression du récepteur ET-A dans les hVICs. Nous avons ensuite évalué l’effet de l’endotheline-1 (ET-1) dans les hVICs. Nous avons retrouvé que les hVICs calcifient lorsqu’elles sont en présence d’ET-1. Ces données suggèrent une implication du système endothélinergique dans la calcification valvulaire aortique. Des études complémentaires sont nécessaires pour mieux comprendre son implication, notamment en évaluant l’effet d’antagoniste des récepteurs de l’endothéline dans des cultures de hVICs puis dans un modèle animal fiable mimant cette pathologie.
Previously, antihypertensive treatment in severe aortic stenosis was considered a relative contraindication. However, recent studies have shown that antihypertensive treatment may be safe and even beneficial in terms of reducing the progression of left ventricular pressure overload and even retarding the progression of valvular aortic stenosis. To date, no randomized clinical trials have been performed and no definite treatment guideline exist for the proper antihypertensive regimens. Antihypertensive treatment with β-blockers has generally been avoided in patients with severe aortic stenosis (AS) due to the concerns for inducing left ventricular dysfunction and hemodynamic compromise in the presence of severe outflow tract obstruction. Although it remains unclear whether antihypertensive treatment with a β-blocker is associated with increased risk of cardiovascular events in patients with AS, recent studies have shown that the use of β-blockers may be safe and may even be beneficial. Renin-angiotensin system (RAS) are upregulated in AS and have been shown to be involved in valve calcification and progression in both experimental models and in human trials. As such, theoretically, RAS inhibition would have benefit in retarding the progression of valvular stenosis as well as have benefit in left ventricle remodeling. Recent clinical studies are indeed showing that use of RAS inhibition may be beneficial in patients with AS. Future clinical trials to establish the ideal target blood pressure and antihypertensive regimens in severe AS is essential.
Calcific aortic valve disease (CAVD) represents a spectrum of disease spanning from milder degrees of calcification of valve leaflets, i.e., aortic sclerosis, to severe calcification i.e., aortic stenosis (AS) with hemodynamic instability. The prevalence of CAVD is increasing rapidly due to the aging of the population, being up to 2.8% among patients over 75 years of age. Even without significant aortic valve stenosis, aortic sclerosis is associated with a 50% increased risk of myocardial infarction and death from cardiovascular causes. To date, there is no pharmacological treatment available to reverse or hinder the progression of CAVD. So far, the cholesterol-lowering therapies (statins) and renin–angiotensin system (RAS) blocking drugs have been the major pharmacological agents investigated for treatment of CAVD. Especially angiotensin receptor blockers (ARB)s and angiotensin convertase enzyme inhibitors (ACEI)s, have been under active investigation in clinical trials, but have proven to be unsuccessful in slowing the progression of CAVD. Several studies have suggested that other vasoactive hormones, including endothelin and apelin systems are also associated with development of AS. In the present review, we discuss the role of vasoactive factors in the pathogenesis of CAVD as novel pharmacological targets for the treatment of aortic valve calcification.
• Key messages
• Vasoactive factors are involved in the progression of calcific aortic valve disease.
• Endothelin and renin–angiotensin systems seem to be most prominent targets for therapeutic interventions in the view of valvular pathogenesis.
• Circulating vasoactive factors may provide targets for diagnostic tools of calcified aortic valve disease.
The mechanisms for pathogenesis of cardiac valve calcification were explored by studying the regulation of the Wnt signaling pathway during the transformation from cardiac valvular myofibroblasts to osteoblast-like phenotype.
Studies were carried on primary cultured porcine aortic valvular myofibroblasts. The cells were randomly divided into four groups and treated with angiotensin II (Ang II) according to the following: Ang II (10(-6) mol/l), Valsartan (Val) (10(-5) mol/l), Ang II plus Val (Ang II 10(-6) mol/l + Val 10(-5) mol/l) or mock treated as the control. Protein expression of Bone morphogenetic protein 2 (BMP2), Alkaline phosphatase (ALP), and Wnt pathway components, Wnt3a and β-catenin, was investigated to assess the activation of the Wnt signaling pathway and determine whether cells undergo the transformation to osteoblast-like phenotype.
Ang II treatment of myofibroblasts led to significant up-regulation of α-SMA expression and activation of the cells. Neither the BMP2 or ALP proteins, nor the mRNA was detectable in the control group or the Val-treated group; however, there was a significant increase in Ang II-treated group (P < 0.01). The Wnt/β-catenin signaling ligand, Wnt3a, was not expressed in the control or Val-treated groups, whereas in Ang II-treated cells, both Wnt3a and β-catenin gene expression were enhanced (P < 0.01).The effect of Ang II can be inhibited by the addition of Val (P < 0.05).
Ang II might act on the Ang II receptor on valvular interstitial cells (VICs) and lead to activation of the Wnt/β-catenin pathway and hence cause the activation, differentiation and proliferation of myofibroblasts, and finally, osteoblast-like phenotype transformation, leading to calcification of heart valves.
Background
Aortic valve sclerosis (AVS) is a chronic, progressive disease involving lipid infiltration, inflammation, and tissue calcification. Despite its high prevalence, there are currently no clinically-approved pharmaceuticals for the management of AVS. The objective of the current study was to elucidate the effects of an angiotensin II type 1 receptor blocker, alone or in combination with statin therapy, on the progression of AVS.
Methods
Male New Zealand White rabbits were fed an atherogenic diet for a period of 12 months to induce AVS. Once disease was established, rabbits were block randomly assigned to receive either no treatment, olmesartan medoxomil, atorvastatin calcium, or a combination of both drugs for a period of 6 months. Disease progression was monitored in vivo using clinically-relevant magnetic resonance imaging and aortic valve cusps were examined ex vivo using histological and immunohistochemical methods.
Results
Cusp thickness significantly increased (0.58 ± 0.03 versus 0.39 ± 0.03 mm for Cholesterol and Control, respectively; P <0.0001) and all classic hallmarks of disease progression — including lipid infiltration, inflammation, and tissue calcification — were observed after 12 months. Unfortunately, neither olmesartan medoxomil nor atorvastatin calcium were able to reverse or delay disease progression during the 6 month treatment period. However, several histological changes were observed in the valvular microenvironment.
Conclusions
The current study suggests that angiotensin receptor blockers, alone or in combination with statin therapy, may not be suitable for management of clinical AVS.
Impaired tissue responsiveness to nitric oxide (NO) occurs in many cardiovascular diseases as well as with advanced age and is a correlate of poor outcomes. This phenomenon results from oxidative stress, with NO "scavenging" and dysfunction of soluble guanylate cyclase (sGC). Thioredoxin-interacting protein (TXNIP) is a major intracellular regulator of inflammatory activation and redox stress, but its interactions with NO/sGC are poorly understood. We have now evaluated the relationship between platelet TXNIP expression and function of the NO/sGC axis in subjects of varying age and during therapy with ramipril.
Young (n=42) and aging (n=49) subjects underwent evaluation of platelet TXNIP content. Aging subjects additionally had measurements of platelet NO responsiveness and routine biochemistry. Platelet TXNIP content was greater (376±33units) in the aging compared to younger subjects (289±13units; p<0.05). In the aging subjects there was a significant negative correlation (r=-0.50, p<0.001) between platelet TXNIP content and NO responsiveness. In a separate cohort of 15 subjects two week treatment with ramipril, which reversed platelet NO resistance and potentiated sGC activity, also decreased platelet TXNIP content by 40% (p=0.011).
Platelet TXNIP content increases with aging, varies inversely with responsiveness to NO, and diminishes rapidly following treatment with ramipril. These data suggest that TXNIP-induced oxidative stress may be a critical modulator of tissue resistance to NO, a fundamental basis for cardiovascular disease. Analogously suppression of TXNIP expression can potentially be utilized as an index of restoration of cardiovascular homeostasis.
Several similarities exist between atherosclerosis in the vasculature and chronic degenerative changes in valvular structures. It has been suggested that aortic valve sclerosis (AVS) and mitral annulus calcification (MAC) are manifestations of a generalized atherosclerosis, have similar pathogenesis, share common risk factors and are observed with higher prevalence in patients with different forms of atherosclerotic vascular disease. Moreover, recent studies have shown a close relation of MAC and AVS with adverse cardiovascular and cerebrovascular outcomes. However, many patients with AVS or MAC do not have coexisting peripheral vascular atherosclerosis and vice versa. Lipid-reducing therapy has been uniformly unsuccessful in slowing valvular calcification. Thus, whether valve calcifications are the result of a more generalized atherosclerosis, or reflect a primary degenerative process, progressing with advancing age, still remains. From a clinical point of view, it is of great importance to identify common links between valve calcification and vascular atherosclerosis with a view to assess whether the detection of AVS, MAC or both are indicative of subclinical atherosclerosis and predicts cardiovascular or cerebrovascular events. Thus, in this article, the authors review current evidence regarding the association between AVS and MAC with vascular atherosclerosis.
Systemic hypertension and aortic valve stenosis (AVS) are both age-related diseases. The pathophysiology of AVS shares some similarities with essential hypertension, which might be the link between the two diseases. Although AS is usually related with low blood pressure levels, approximately one third of patients with severe AS suffer from arterial hypertension, a percentage that can increase up to 50 % according some studies. This review will summarize various aspects regarding the prevalence the pathophysiology and the natural history of those two diseases that seems to be linked, as well as the effect of blood pressure and antihypertensive treatment on various echocardiographic parameters in patients with AVS.
Aortic stenosis is the most prevalent valve pathology and calcific aortic valve disease (CAVD) is its most frequent etiology in developed countries. There is extensive evidence that CAVD represents an active disease process similar to that of atherosclerosis with similar classical cardiovascular risk factors and pathological mechanisms. Given that in the vast majority of situations the only treatment available is valve replacement there is a need to develop pharmacological therapies that retard the disease progression. Lipid-lowering therapies have been the focus of research, however, so far with negative results. Future studies, including animal models, shall provide an opportunity to further evaluate the disease mechanisms of CAVD and to discover potential disease biomarkers and pharmacological interventions that can reduce the need for valve replacement.
Aortic valve disease (AVD) is the most common form of valvular heart disease in the western world. The only proven therapy for severe AVD is open aortic valve replacement, with trans-catheter aortic valve implantation emerging as a promising modality to treat severe aortic stenosis in a selected group of patients. AVD has a long asymptomatic phase with symptoms occurring late in the disease and once symptoms develop, prognosis is poor. There is a growing appreciation that aortic valvular heart disease incorporates a disease process that extends beyond the valve itself leading to an aortic valvular 'heart' disease. The renin-angiotensin system is known to modulate adverse left ventricular remodeling and myocardial fibrosis, which could be caused by increased load caused by the AVD. In this review, the authors explore evidence that suggest that drugs that target the renin-angiotensin system may have a potential therapeutic role in AVD.
Aims Aortic valve stenosis (AS) and its precursor, aortic valve sclerosis (ASc), occur frequently in Western populations. Investigations to retard the progression of AS using statins have been unsuccessful. Development of ASc in humans is associated with increased aortic valve backscatter (AVBS) and poor tissue nitric oxide (NO) responsiveness. In an animal model, ramipril retarded AS/ASc development. We have now set out to identify factors associated with the progression of ASc in humans.Methods and resultsAt baseline and after 4 years, 204 randomly selected subjects (age 63 ± 6 years at study entry) underwent echocardiography with the determination of AVBS values, measurements of platelet NO responsiveness, plasma asymmetric dimethylarginine concentrations, lipid profile, high-sensitivity-C-reactive protein, routine biochemistry, and 25-hydroxy-vitamin D levels. During the study period, 68 of subjects had detectable AVBS progression. On multivariate analysis, higher calcium concentrations (β 0.22; P 0.004), poor platelet NO responsiveness (β 0.18; P 0.018), and increased arterial stiffness (β 0.15; P 0.044) were independent predictors of disease progression. The use of angiotensin-converting enzyme-inhibitors/angiotensin II receptor blockers (ACE-I/ARB) predicted the lack of disease progression (assessed categorically) in the overall cohort and in those without ASc at baseline (n 159) (β 0.8; P 0.025 and β 1.3; P 0.001, respectively). No conventional coronary risk factors were associated with disease progression.Conclusion
This study of early aortic valve disease (i) demonstrates that disease progression occurs in the majority of the normal ageing population over a 4-year period; (ii) provides evidence of the importance of the NO signalling cascade in disease development and progression; and (iii) provides additional data linking ACE-I/ARB use with the retardation of ASc.
Aortic valve stenosis (AS) is the most common form of valvular heart disease in the Western world, affecting ~40% of the population over the age of 80; to date the only established treatment is valve replacement. However, AS progression occurs over many years, and is associated from its earliest stages with increased risk of coronary events. Recent insight into the pathophysiology of AS has included central roles for angiotensin II, for diminished nitric oxide effect at the level of valve endothelium and matrix, and for inflammatory activation/redox stress culminating in activation of pro-calcific stimuli. Despite the presence of atheroma within the stenotic valve, hyperlipidemia per se does not play a critic role in the development of obstructive disease. We review emerging options for pharmacotherapy of AS, including in particular retardation of disease progression. The various clinical evaluations of lipid-reducing therapy have been uniformly unsuccessful in slowing AS progression. However, recent studies in animal models and retrospective evaluations in humans suggest that ACE inhibitors and/or angiotensin receptor blockers may be effective in this regard. Furthermore, agents normally utilized to treat osteoporosis also offer promise in retarding AS. Given the considerable morbidity, mortality and health care costs associated with AS, such therapeutic developments should be expedited.
Heart valve disease is a frequently encountered pathology, related to high morbidity and mortality rates in industrialized and developing countries. Animal models are interesting to investigate the causality, but also underlying mechanisms and potential treatments of human valvular diseases. Recently, animal models of heart valve disease have been developed, which allow to investigate the pathophysiology, and to follow the progression and the potential regression of disease with therapeutics over time. The present review provides an overview of animal models of primary, organic heart valve disease: myxoid age-related, infectious, drug-induced, degenerative calcified, and mechanically induced valvular heart disease.
Calcific aortic valve stenosis (CAVS) is a major health problem facing aging societies. The identification of osteoblast-like and osteoclast-like cells in human tissue has led to a major paradigm shift in the field. CAVS was thought to be a passive, degenerative process, whereas now the progression of calcification in CAVS is considered to be actively regulated. Mechanistic studies examining the contributions of true ectopic osteogenesis, nonosseous calcification, and ectopic osteoblast-like cells (that appear to function differently from skeletal osteoblasts) to valvular dysfunction have been facilitated by the development of mouse models of CAVS. Recent studies also suggest that valvular fibrosis, as well as calcification, may play an important role in restricting cusp movement, and CAVS may be more appropriately viewed as a fibrocalcific disease. High-resolution echocardiography and magnetic resonance imaging have emerged as useful tools for testing the efficacy of pharmacological and genetic interventions in vivo. Key studies in humans and animals are reviewed that have shaped current paradigms in the field of CAVS, and suggest promising future areas for research.
Interleukin-1beta (IL-1beta), reactive oxygen species (ROS), and thioredoxin-interacting protein (TXNIP) are all implicated in the pathogenesis of type 2 diabetes mellitus (T2DM). Here we review mechanisms directing IL-1beta production and its pathogenic role in islet dysfunction during chronic hyperglycemia. In doing so, we integrate previously disparate disease-driving mechanisms for IL-1beta, ROS, and TXNIP in T2DM into one unifying model in which the NLRP3 inflammasome plays a central role. The NLRP3 inflammasome also drives IL-1beta maturation and secretion in another disease of metabolic dysregulation, gout. Thus, we propose that the NLRP3 inflammasome contributes to the pathogenesis of T2DM and gout by functioning as a sensor for metabolic stress.
Recent data have shown that aortic valve stenosis (AS) is an active and highly regulated process which shares similarities with atherosclerosis. However, AS cannot be considered as a purely atherosclerotic phenomenon, and a hypercholesterolemic rabbit model might not be fully representative of human AS pathophysiology.
Twenty-eight New Zealand White rabbits were assigned to three groups: group 1 (no dietary supplement for three months); group 2 (0.3% cholesterol-enriched-diet + 50,000 IU/day vitamin D2 for six months); and group 3 (1% cholesterol-enriched-diet + vitamin D2 for three months). The peak aortic gradient and permeability index (outflow tract/aortic velocity-time-integral) were assessed, as well as calcium staining within the aortic valve and ascending aorta.
AS hemodynamic severity was not different among the groups. The peak gradient was 4 +/- 2 mmHg at baseline, 4 +/- 2 mmHg at three months in controls, 4 +/- 1 mmHg at three months and 6 +/-3 mmHg at six months in group 2, and 4 +/- 1 mmHg at three months in group 3 (p = NS). The permeability index was 64 +/- 7 at baseline, 60 +/- 12 at three months in controls, 63 +/- 14 at three months and 58 +/- 12 at six months in group 2, and 60 +/- 5 at three months in group 3 (p = NS). The aortic valve of cholesterol-enriched-diet rabbits was thickened but not calcified, whereas the ascending aorta was both thickened and calcified.
When using a hypercholesterolemic rabbit model plus vitamin D2, no adverse hemodynamic effect or aortic valve calcification was observed, despite a high-level and prolonged cholesterol-regimen supplementation. These results raise questions with regard to the extrapolation of this animal model to humans.
Epinephrine plays a key role in the control of vasomotor tone; however, the participation of the NO/cGMP pathway in response to beta-adrenoceptor activation remains controversial. To evaluate the involvement of the endothelium in the vascular response to epinephrine, we assessed NO production, endothelial NO synthase phosphorylation, and tissue accumulation of cGMP in the perfused arterial mesenteric bed of rat. Epinephrine elicited a concentration-dependent increase in NO (EC(50) of 45.7 pM), which was coupled to cGMP tissue accumulation. Both NO and cGMP production were blocked by either endothelium removal (saponin) or NO synthase inhibition (N(omega)-nitro-L-arginine). Blockade of beta(1)- and beta(2)-adrenoceptors with 1 microM propranolol or beta(3)-adrenoceptor with 10 nM SR 59230A displaced rightward the concentration-NO production curve evoked by epinephrine. Selective stimulation of beta(1)-, beta(2)-, or beta(3)-adrenoceptors also resulted in NO and cGMP production. Propranolol (1 microM) inhibited the rise in NO induced by isoproterenol or the beta(2)-adrenoceptor agonists salbutamol, terbutaline, or fenoterol. Likewise, 10 nM SR 59230A reduced the effects of the beta(3)-adrenoceptor agonists BRL 37344, CGP 12177, SR 595611A, or pindolol. The NO production induced by epinephrine and BRL 37344 was associated with the activation of the phosphatidylinositol 3-kinase/Akt pathway and phosphorylation of eNOS in serine 1177. In addition, in anaesthetized rats, bolus administration of isoproterenol, salbutamol, or BRL 37344 produced NO-dependent reductions in systolic blood pressure. These findings indicate that beta(1)-, beta(2)-, and beta(3)-adrenoceptors are coupled to the NO/cGMP pathway, highlighting the role of the endothelium in the vasomotor action elicited by epinephrine and related beta-adrenoceptor agonists.
We examined whether impaired renal function causes thickening of the aortic valve leaflets in hyperlipidemic apoE-knockout (apoE-/-) mice, and whether the putative effect on the aortic valves could be prevented by inhibiting the angiotensin-converting enzyme (ACE) with enalapril.
Thickening of the aortic valve leaflets in apoE-/- mice was induced by producing mild or moderate chronic renal failure resulting from unilateral nephrectomy (1/2 NX, n = 18) or subtotal nephrectomy (5/6 NX, n = 22), respectively. Additionally, the 5/6 NX mice were randomized to no treatment (n = 8) or enalapril treatment (n = 13). The maximal thickness of each leaflet was measured from histological sections of the aortic roots.
Leaflet thickness was significantly greater in the 5/6 NX mice than in the 1/2 NX mice (P = 0.030) or the unoperated mice (P = 0.003). The 5/6 NX mice treated with enalapril had significantly thinner leaflets than did the untreated 5/6 NX mice (P = 0.014).
Moderate uremia causes thickening of the aortic valves in apoE-/- mice, which can be attenuated by ACE inhibition. The nephrectomized apoE-/- mouse constitutes a new model for investigating the mechanisms of uremia-induced aortic valve disease, and also provides an opportunity to study its pharmacologic prevention.
In type 2 diabetes, glucose toxicity leads to beta cell apoptosis with decreased beta cell mass as a consequence. Thioredoxin-interacting protein (TXNIP) is a critical mediator of glucose-induced beta cell apoptosis. Since hyperglycaemia leads to elevated serum insulin, we hypothesised that insulin is involved in the regulation of TXNIP protein levels in beta cells.
We studied the production of TXNIP in INS-1E beta cells and in islets of Psammomys obesus, an animal model of type 2 diabetes, in response to glucose and different modulators of insulin secretion.
TXNIP production was markedly augmented in islets from diabetic P. obesus and in beta cells exposed to high glucose concentration. In contrast, adding insulin to the culture medium or stimulating insulin secretion with different secretagogues suppressed TXNIP. Inhibition of glucose and fatty acid-stimulated insulin secretion with diazoxide increased TXNIP production in beta cells. Nitric oxide (NO), a repressor of TXNIP, enhanced insulin signal transduction, whereas inhibition of NO synthase abolished its activation, suggesting that TXNIP inhibition by NO is mediated by stimulation of insulin signalling. Treatment of beta cells chronically exposed to high glucose with insulin reduced beta cell apoptosis. Txnip knockdown mimicking the effect of insulin prevented glucose-induced beta cell apoptosis.
Insulin is a potent repressor of TXNIP, operating a negative feedback loop that restrains the stimulation of TXNIP by chronic hyperglycaemia. Repression of TXNIP by insulin is probably an important compensatory mechanism protecting beta cells from oxidative damage and apoptosis in type 2 diabetes.
Background
Hyperlipidemia has been suggested as a risk factor for stenosis of the aortic valve, but lipid-lowering studies have had conflicting results.
Methods
We conducted a randomized, double-blind trial involving 1873 patients with mild-to-moderate, asymptomatic aortic stenosis. The patients received either 40 mg of simvastatin plus 10 mg of ezetimibe or placebo daily. The primary outcome was a composite of major cardiovascular events, including death from cardiovascular causes, aortic-valve replacement, nonfatal myocardial infarction, hospitalization for unstable angina pectoris, heart failure, coronary-artery bypass grafting, percutaneous coronary intervention, and nonhemorrhagic stroke. Secondary outcomes were events related to aortic-valve stenosis and ischemic cardiovascular events.
Results
During a median follow-up of 52.2 months, the primary outcome occurred in 333 patients (35.3%) in the simvastatin–ezetimibe group and in 355 patients (38.2%) in the placebo group (hazard ratio in the simvastatin–ezetimibe group, 0.96; 95% confidence interval [CI], 0.83 to 1.12; P=0.59). Aortic-valve replacement was performed in 267 patients (28.3%) in the simvastatin–ezetimibe group and in 278 patients (29.9%) in the placebo group (hazard ratio, 1.00; 95% CI, 0.84 to 1.18; P=0.97). Fewer patients had ischemic cardiovascular events in the simvastatin–ezetimibe group (148 patients) than in the placebo group (187 patients) (hazard ratio, 0.78; 95% CI, 0.63 to 0.97; P=0.02), mainly because of the smaller number of patients who underwent coronary-artery bypass grafting. Cancer occurred more frequently in the simvastatin–ezetimibe group (105 vs. 70, P=0.01).
Conclusions
Simvastatin and ezetimibe did not reduce the composite outcome of combined aortic-valve events and ischemic events in patients with aortic stenosis. Such therapy reduced the incidence of ischemic cardiovascular events but not events related to aortic-valve stenosis. (ClinicalTrials.gov number, NCT00092677.)
In a prospective study of 1,980 subjects (mean age 81 +/- 8 years) without valvular aortic stenosis, 981 (50%) had valvular aortic sclerosis diagnosed by 2-dimensional and continuous-wave Doppler echocardiography. Independent risk factors for new coronary events were prior coronary artery disease (p = 0.0001, risk ratio 2.8), male gender (p = 0.002, risk ratio 1.3), and valvular aortic sclerosis (p = 0.0001, risk ratio 1.8).
To study in a rabbit model the expression of endothelial nitric oxide synthase (eNOS) in association with the development of calcification of the aortic valve, and to assess the effects of atorvastatin on eNOS expression, nitrite concentration, and aortic valve calcification.
Rabbits (n = 48) were treated for three months: 16, forming a control group, were fed a normal diet; 16 were fed a 0.5% (wt/wt) high cholesterol diet; and 16 were fed a 0.5% (wt/wt) cholesterol diet plus atorvastatin (2.5 mg/kg/day). The aortic valves were examined with eNOS immunostains and western blotting. Cholesterol and high sensitivity C reactive protein (hsCRP) concentrations were determined by standard assays. Serum nitrite concentrations were measured with a nitric oxide analyser. eNOS was localised by electron microscopy and immunogold labelling. Calcification in the aortic valve was evaluated by micro-computed tomography (CT).
Cholesterol, hsCRP, and aortic valve calcification were increased in the cholesterol fed compared with control animals. Atorvastatin inhibited calcification in the aortic valve as assessed by micro-CT. eNOS protein concentrations were unchanged in the control and cholesterol groups but increased in the atorvastatin treated group. Serum nitrite concentrations were decreased in the hypercholesterolaemic animals and increased in the group treated with atorvastatin.
These data provide evidence that chronic experimental hypercholesterolaemia produces bone mineralisation in the aortic valve, which is inhibited by atorvastatin.
Aortic stenosis (AS) is characterized by extensive remodelling of the valves, including infiltration of inflammatory cells, extracellular matrix degradation, and fibrosis. The molecular mechanisms behind this adverse remodelling have remained obscure. In this article, we study whether cathepsin G, an angiotensin II (Ang II)-forming elastolytic enzyme, contributes to progression of AS.
Stenotic aortic valves (n = 86) and control valves (n = 17) were analysed for cathepsin G, transforming growth factor-beta1 (TGF-beta1), and collagens I and III with RT-PCR and immunohistochemistry. Valvular collagen/elastin ratio was quantified by histochemistry. In stenotic valves, cathepsin G was present in mast cells and showed increased expression (P < 0.001), which correlated positively (P < 0.001) with the expression levels of TGF-beta1 and collagens I and III. TGF-beta1 was also present in mast cell-rich areas and cathepsin G induced losartan-sensitive TGF-beta1 expression in cultured fibroblasts. Collagen/elastin ratio was increased in stenotic valves (P < 0.001) and correlated positively with smoking (P = 0.02). Nicotine in cigarette smoke activated mast cells and induced TGF-beta1 expression in cultured fibroblasts. Fragmented elastin was observed in stenotic valves containing activated cathepsin G-secreting mast cells and in normal valves treated with cathepsin G.
In stenotic aortic valves, mast cell-derived cathepsin G may cause adverse valve remodelling and AS progression.
Carboxypeptidase cleavage of the C-terminal Arg of kinins generates specific agonists of the B1 receptor. Activation of B1 receptors produces nitric oxide via eNOS in bovine endothelial cells and iNOS in cytokine-stimulated human endothelial cells. Angiotensin-converting enzyme (ACE) inhibitors are direct agonists of B1 receptors in endothelial cells, although they release NO via a different signaling pathway than peptide ligands in bovine cells. This brief review discusses carboxypeptidase M as a required processing enzyme for generating B1 agonists, how ACE inhibitors and peptide ligands stimulate NO production and the evidence for, as well as some consequences of, the direct activation of B1 receptors by ACE inhibitors.
background Calcific aortic stenosis has many characteristics in common with atherosclerosis, in- cluding hypercholesterolemia. We hypothesized that intensive lipid-lowering therapy would halt the progression of calcific aortic stenosis or induce its regression. methods In this double-blind, placebo-controlled trial, patients with calcific aortic stenosis were randomly assigned to receive either 80 mg of atorvastatin daily or a matched placebo. Aortic-valve stenosis and calcification were assessed with the use of Doppler echocardi- ography and helical computed tomography, respectively. The primary end points were change in aortic-jet velocity and aortic-valve calcium score. results Seventy-seven patients were assigned to atorvastatin and 78 to placebo, with a median follow-up of 25 months (range, 7 to 36). Serum low-density lipoprotein cholesterol concentrations remained at 130±30 mg per deciliter in the placebo group and fell to 63±23 mg per deciliter in the atorvastatin group (P
We sought to examine the effect of angiotensin receptor blocker (ARB) on the formation of lesions in the aortic valves of hypercholesterolemic rabbits.
Recently, atherosclerosis has been recognized as a mechanism that is responsible for calcific aortic stenosis. The effect of ARBs might help to prevent aortic stenosis because they have multiple antiatherosclerotic effects.
Male Japanese white rabbits (n = 36) were separated as follows: control with chow diet (C) and vehicle (V) groups, both of which were fed a 1% cholesterol diet for 8 weeks, and an ARB group (A), which was fed a 1% cholesterol diet for 8 weeks with ARB (olmesartan, 1 mg/kg/day) for the last 4 weeks.
This dose of olmesartan did not affect either blood pressure or cholesterol levels. Dietary cholesterol induced fatty deposition with macrophage accumulation and osteopontin coexpression in valve leaflets, whereas ARB decreased macrophage accumulation (% area: V, 9.3 +/- 0.34; A, 1.4 +/- 0.30; p = 0.003) and osteopontin expression (p = 0.017). Angiotensin-converting enzyme was also up-regulated in V and decreased by olmesartan (p = 0.015). Immunohistochemistry with anti-CD31 antibody revealed that dietary cholesterol disrupted and olmesartan preserved endothelial integrity on the lesion-prone aortic side of the valve (% CD31-positive circumference: V, 30 +/- 3.7; A, 62 +/- 4.8; p = 0.003). Numbers of alpha-smooth muscle actin-positive myofibroblasts were increased in V and decreased by olmesartan (p = 0.003). Real-time polymerase chain reaction revealed that increased amounts of messenger ribonucleic acid for osteoblast-specific transcription factor core binding factor alpha-1 in V were diminished by olmesartan.
Atherosclerotic changes in the aortic valves of rabbits fed with cholesterol were inhibited by ARB, whereas endo-thelial integrity was preserved and transdifferentiation into myofibroblasts and/or osteoblasts in valve leaflets was inhibited.
Calcific aortic valve disease is associated with increased morbidity and mortality, especially in the elderly. To date, pharmacological therapies have not proven as effective as surgical intervention. Here, we used a hyperlipidemic rabbit model to investigate the potential effects of selective aldosterone inhibition on the early stages of aortic valve calcification, a pharmacological strategy that has not yet been tested. Forty New Zealand male rabbits fed a standard diet for 4 weeks were separated into three groups: (1) control (n=10), fed a standard diet; (2) vehicle (n=15), fed a hyperlipidemic diet (cholesterol 1%) plus vehicle; and (3) eplerenone (n=15), fed a hyperlipidemic diet plus 100 mg/kg/d eplerenone (last 4 weeks). After 8 weeks, animals were sacrificed and prepared aortic valve sections were examined with Von Kossa silver stain and by immunostaining for mineralocorticoid receptor, macrophages and angiotensin-converting enzyme. The presence of calcium deposits was confirmed by scanning electron microscopy. Eplerenone increased aldosterone levels but did not affect blood pressure, cholesterol or potassium levels. Hyperlipidemia induced macrophage accumulation and angiotensin-converting enzyme expression, as well as calcium deposition in the leaflets. All markers were decreased by eplerenone treatment. Immunohistochemistry for mineralocorticoid (aldosterone) receptors revealed similar expression in the leaflets of both control and hyperlipidemic groups. Collectively, these results indicate that aldosterone receptors are present in rabbit aortic valve leaflets and their selective blockade with eplerenone inhibits formation of the sclerotic lesions induced by a high fat diet.
Aortic stenosis (AS) is an active process with similarities to atherosclerosis. The objective of this study was to assess the effect of cholesterol lowering with rosuvastatin on the progression of AS.
This was a randomized, double-blind, placebo-controlled trial in asymptomatic patients with mild to moderate AS and no clinical indications for cholesterol lowering. The patients were randomized to receive either placebo or rosuvastatin 40 mg daily. A total of 269 patients were randomized: 134 patients to rosuvastatin 40 mg daily and 135 patients to placebo. Annual echocardiograms were performed to assess AS progression, which was the primary outcome; the median follow-up was 3.5 years. The peak AS gradient increased in patients receiving rosuvastatin from a baseline of 40.8+/-11.1 to 57.8+/-22.7 mm Hg at the end of follow-up and in patients with placebo from 41.6+/-10.9 mm Hg at baseline to 54.8+/-19.8 mm Hg at the end of follow-up. The annualized increase in the peak AS gradient was 6.3+/-6.9 mm Hg in the rosuvastatin group and 6.1+/-8.2 mm Hg in the placebo group (P=0.83). Treatment with rosuvastatin was not associated with a reduction in AS progression in any of the predefined subgroups.
Cholesterol lowering with rosuvastatin 40 mg did not reduce the progression of AS in patients with mild to moderate AS; thus, statins should not be used for the sole purpose of reducing the progression of AS. Clinical Trial Registration Information- URL: http://www.controlled-trials.com/. Clinical trial registration number: ISRCTN 32424163.
The NLRP3 inflammasome has a major role in regulating innate immunity. Deregulated inflammasome activity is associated with several inflammatory diseases, yet little is known about the signaling pathways that lead to its activation. Here we show that NLRP3 interacted with thioredoxin (TRX)-interacting protein (TXNIP), a protein linked to insulin resistance. Inflammasome activators such as uric acid crystals induced the dissociation of TXNIP from thioredoxin in a reactive oxygen species (ROS)-sensitive manner and allowed it to bind NLRP3. TXNIP deficiency impaired activation of the NLRP3 inflammasome and subsequent secretion of interleukin 1beta (IL-1beta). Akin to Txnip(-/-) mice, Nlrp3(-/-) mice showed improved glucose tolerance and insulin sensitivity. The participation of TXNIP in the NLRP3 inflammasome activation may provide a mechanistic link to the observed involvement of IL-1beta in the pathogenesis of type 2 diabetes.
We sought to identify clinical, physiological, and biochemical correlates, including markers of endothelial dysfunction and of tissue nitric oxide (NO) responsiveness, of the presence of aortic sclerosis (ASc) in an aging population.
Aortic sclerosis has been regarded predominantly as a precursor of hemodynamically significant aortic stenosis. However, ASc also represents an independent correlate of increased risk of cardiovascular morbidity and mortality; the basis of this association is incompletely understood. The assumption that the pathogenesis of aortic valve disease is similar to that of atherosclerosis has not been supported by recent studies; rather there has been increasing evidence of a pathogenetic role of inflammation and endothelial dysfunction. Furthermore, we have recently developed methodology for echocardiographic quantitation of early aortic valve disease.
Randomly selected subjects (n = 253) ages 51 to 77 years underwent transthoracic echocardiography; aortic valve ultrasonic backscatter score (AV(BS)) was used to quantitate echogenicity of the aortic valve. Conventional coronary risk factors were identified. Integrity of NO generation/response was assessed via: 1) plasma asymmetric dimethylarginine concentrations, as a marker of endothelial dysfunction; 2) inhibition of platelet aggregation by the NO donor sodium nitroprusside, as a measure of tissue NO responsiveness and also a coronary prognostic marker; and 3) aortic augmentation index, as a measure of arterial stiffness/wave reflection. All putative correlations with AV(BS) were examined by univariate and multiple linear regression analyses.
On the basis of AV(BS) scores, ASc was present in 19.4% of subjects. The AV(BS) directly correlated with patients' age but inversely correlated with high-sensitivity C-reactive protein, creatinine clearance, and platelet NO responsiveness. On multiple linear regression, ASc was associated with impaired platelet NO responsiveness (beta = -0.16, p = 0.02), advancing age (beta = 0.21, p = 0.003), and low body mass index (beta = -0.23, p = 0.001).
Aortic sclerosis is associated with platelet NO resistance rather than conventional coronary risk factors: this might explain the increased thrombotic risk in ASc.
Excessive reactive oxygen species play a key role in the pathogenesis of diabetic nephropathy, but to what extent these result from increased generation, impaired antioxidant systems, or both is incompletely understood. Here, we report the expression, localization, and activity of the antioxidant thioredoxin and its endogenous inhibitor thioredoxin interacting protein (TxnIP) in vivo and in vitro. In normal human and rat kidneys, expression of TxnIP mRNA and protein was most abundant in the glomeruli and distal nephron (distal convoluted tubule and collecting ducts). In contrast, thioredoxin mRNA and protein localized to the renal cortex, particularly within the proximal tubules and to a lesser extent in the distal nephron. Induction of diabetes in rats increased expression of TxnIP but not thioredoxin mRNA. Kidneys from patients with diabetic nephropathy had significantly higher levels of TxnIP than control kidneys, but thioredoxin expression did not differ. In vitro, high glucose increased TxnIP expression in mesangial, NRK (proximal tubule), and MDCK (distal tubule/collecting duct) cells, and decreased the expression of thioredoxin in mesangial and MDCK cells. Knockdown of TxnIP with small interference RNA suggested that TxnIP mediates the glucose-induced impairment of thioredoxin activity. Knockdown of TxnIP also abrogated both glucose-induced 3H-proline incorporation (a marker of collagen production) and oxidative stress. Taken together, these findings suggest that impaired thiol reductive capacity contributes to the generation of reactive oxygen species in diabetes in a site- and cell-specific manner.
Calcific aortic stenosis displays some similarities to atherosclerosis including evidence of endothelial dysfunction. Whether nitric oxide (NO), which is produced by valvular endothelium, has direct protective effects extending to calcification processes in aortic valve cells has not previously been examined. In vitro calcifying nodules in porcine aortic valve interstitial cell cultures, formed in response to transforming growth factor-beta1 (TGF-beta1) 5 ng/ml, were inhibited by NO donors DETA-NONOate 5-100 microM, and sodium nitroprusside (SNP) 3 microM. Raising intracellular cGMP concentrations, via 8-bromo cGMP 1 mM or via brain natiuretic peptide and C-type natiuretic peptide 0.1 microM, inhibited TGF-beta1-induced nodule formation, potentially implicating the cGMP pathway in the NO effect. Stimulation of interstitial cells with substance P or calcium ionophone (A23187) caused NO release and increased intracellular cGMP respectively. However in the presence of TGF-beta1 basal levels of NO production via nitric oxide synthase (NOS) were insufficient to affect nodule formation. Increased dihydroethidium (DHE) fluorescence in response to TGF-beta1, which was inhibited by DETA-NONOate and TEMPOL, suggested a role for intracellular superoxide in TGF-beta1 signalling. Moreover, nodule formation was suppressed by superoxide scavengers TEMPOL, hydralazine and polyethylene glycol-superoxide dismutase (PEG-SOD), but not SOD. In conclusion, NO donors, or agents raising intracellular cGMP levels, may protect aortic valve interstitial cells from early events leading to calcification.
Calcific aortic stenosis is the most common indication for surgical valve replacement in the United States. For years this disease has been described as a passive degenerative process during which serum calcium attaches to the valve surface and binds to the leaflet to form nodules. Therefore, surgical treatment of this disease has been the approach toward relieving outflow obstruction in these patients. Recent studies demonstrate an association between atherosclerosis and its risk factors for aortic valve disease. In 2008, there are increasing number of epidemiology and experimental studies to provide evidence that this disease process is not a passive phenomena. There is an active cellular process that develops within the valve leaflet and causes a regulated bone formation to develop. If the atherosclerotic hypothesis is important in the initiation of aortic stenosis, then treatments used in slowing the progression of atherosclerosis may be effective in patients with aortic valve disease. This review will discuss the pathogenesis and the potential for medical therapy in the management of patients with calcific aortic stenosis by examining the lessons provided from the experimental research.
The aim of this study was to determine whether oxidative stress is increased in calcified, stenotic aortic valves and to examine mechanisms that might contribute to increased oxidative stress.
Oxidative stress is increased in atherosclerotic lesions and might play an important role in plaque progression and calcification. The role of oxidative stress in valve disease is not clear.
Superoxide (dihydroethidium fluorescence and lucigenin-enhanced chemiluminescence), hydrogen peroxide H2O2 (dichlorofluorescein fluorescence), and expression and activity of pro- and anti-oxidant enzymes were measured in normal valves from hearts not suitable for transplantation and stenotic aortic valves that were removed during surgical replacement of the valve.
In normal valves, superoxide levels were relatively low and distributed homogeneously throughout the valve. In stenotic valves, superoxide levels were increased 2-fold near the calcified regions of the valve (p < 0.05); noncalcified regions did not differ significantly from normal valves. Hydrogen peroxide levels were also markedly elevated in calcified regions of stenotic valves. Nicotinamide adenine dinucleotide phosphate oxidase activity was not increased in calcified regions of stenotic valves. Superoxide levels in stenotic valves were significantly reduced by inhibition of nitric oxide synthases (NOS), which suggests uncoupling of the enzyme. Antioxidant mechanisms were reduced in calcified regions of the aortic valve, because total superoxide dismutase (SOD) activity and expression of all 3 SOD isoforms was significantly decreased. Catalase expression also was reduced in pericalcific regions.
This study provides the first evidence that oxidative stress is increased in calcified regions of stenotic aortic valves from humans. Increased oxidative stress is due at least in part to reduction in expression and activity of antioxidant enzymes and perhaps to uncoupled NOS activity. Thus, mechanisms of oxidative stress differ greatly between stenotic aortic valves and atherosclerotic arteries.
This article reports the surface structure of the endothelial cells covering the cardiac valves and their changes in chronic inactive rheumatic valvulitis and bacterial endocarditis. The endocardial surface of the normal cardiac valve was covered by a layer of endothelial cells, each of which was recognized because of the presence of centrally located nuclear bulge and marginal folds at the cell boundaries. In the cardiac valves affected by chronic rheumatic valvulitis, the endothelial layer denuded extensively, even in the regions where only slight thickening was noticed in the intravalvular tissues. As compared to the inflow side of the valve, denudation of the endothelial layer was more conspicuous on the outflow side. There was no morphological difference between the two hemodynamically different conditions, i.e. stenosis and regurgitation. In the cardiac valves affected by bacterial endocarditis, in contrast, the destruction of the endothelial layer was preferentially observed along the closing margins of the valve, and in the region of calcification and vegetation. These findings are compatible to the interpretation that differences in morphological changes of the endothelial layer in these two acquired valvular diseases may reflect the differences of their pathogeneses. The present study thus suggest an importance of endothelial cell in elucidating the pathogenesis of acquired valvular diseases.
Surface features of 19 stenotic aortic valves from patients undergoing valve replacement were investigated by scanning electron microscopy. Villi, prominent on five valves, were distributed either singularly or in clusters and differed in shape. Endothelial cells had microvilli and bulbous surface projections. Endothelial disruption with a focal loss of endothelial cells was uniformly observed. Erythrocytes were found scattered over the exposed subendothelial surface or enmeshed within fibrin networks on 11 of the valves. Activated leukocytes were seen on four valves and showed veil-like projections as well as microvilli. Platelets, observed on three valves, displayed pseudopodial formation and hyalomeric spreading, signifying an increased degree of membrane response. Most platelet aggregates were composed entirely of dendritic forms (reversible aggregates), but a few also contained spread forms (irreversible aggreates). Focal deposits of crystalline material, presumably containing calcium, were observed in areas of endocardial disruption.
Nonrheumatic stenosis of trileaflet aortic valves, often termed senile or calcific valvular aortic stenosis, is considered a "degenerative" process, but little is known about the cellular or molecular factors that mediate its development.
To characterize the developing aortic valvular lesion, we performed histological and immunohistochemical studies on Formalin-fixed and methanol-Carnoy's-fixed paraffin-embedded aortic valve leaflets or on frozen sections obtained at autopsy from 27 adults (age, 46 to 82 years) with normal leaflets (n = 6), mild macroscopic leaflet thickening (n = 15), or clinical aortic stenosis (n = 6). Focal areas of thickening ("early lesions") were characterized by (1) subendothelial thickening on the aortic side of the leaflet, between the basement membrane (PAS-positive) and elastic lamina (Verhoeff-van Gieson), (2) the presence of large amounts of intracellular and extracellular neutral lipids (oil red O) and fine, stippled mineralization (von Kossa), and (3) disruption of the basement membrane overlying the lesion. Regions of the fibrosa adjacent to these lesions were characterized by thickening and by protein, lipid, and calcium accumulation. Control valves showed none of these abnormalities. Immunohistochemical studies were performed using monoclonal antibodies directed against macrophages (anti-CD68 or HAM-56), and contractile proteins of smooth muscle cells or myofibroblasts (anti-alpha-actin and HHF-35) or rabbit polyclonal antiserum against T lymphocytes (anti-CD3). In normal valves, scattered macrophages were present in the fibrosa and ventricularis, and occasional muscle actin-positive cells were detected in the proximal portion of the ventricularis near the leaflet base, but no T lymphocytes were found. In contrast, early lesions were characterized by the presence of an inflammatory infiltrate composed of non-foam cell and foam cell macrophages, occasional T cells, and rare alpha-actin-positive cells. In stenotic aortic valves, a similar but more advanced lesion was seen.
The early lesion of "degenerative" aortic stenosis is an active inflammatory process with some similarities (lipid deposition, macrophage and T-cell infiltration, and basement membrane disruption) and some dissimilarities (presence of prominent mineralization and small numbers of smooth muscle cells) to atherosclerosis.
We investigated whether L-arginine induces regression of preexisting atheromatous lesions and reversal of endothelial dysfunction in hypercholesterolemic rabbits, whether similar effects can be obtained by cholesterol-lowering therapy with lovastatin, and which mechanism leads to these effects.
Rabbits were fed 1% cholesterol for 4 weeks and 0.5% cholesterol for an additional 12 weeks. Two groups of cholesterol-fed rabbits were treated with L-arginine (2.0% in drinking water) or lovastatin (10 mg/d) during weeks 5 through 16. Systemic nitric oxide (NO) formation was assessed as the urinary excretion rates of nitrate and cGMP in weekly intervals. Cholesterol feeding progressively reduced urinary nitrate excretion to approximately 40% of baseline (P<.05) and increased plasma concentrations of asymmetrical dimethylarginine (ADMA), an endogenous NO synthesis inhibitor. Dietary L-arginine reversed the reduction in plasma L-arginine/ADMA ratio and partly restored urinary excretion of nitrate and cGMP (each P<.05 vs cholesterol) but did not change plasma cholesterol levels. L-Arginine completely blocked the progression of carotid intimal plaques, reduced aortic intimal thickening, and preserved endothelium-dependent vasodilator function. Lovastatin treatment reduced plasma cholesterol by 32% but did not improve urinary nitrate or cGMP excretion or endothelium-dependent vasodilation. Lovastatin had a weaker inhibitory effect on carotid plaque formation and aortic intimal thickening than L-arginine. L-Arginine inhibited but lovastatin potentiated superoxide radical generation in the atherosclerotic vascular wall.
Dietary L-arginine improves NO-dependent vasodilator function in cholesterol-fed rabbits and completely blocks the progression of plaques via restoration of NO synthase substrate availability and reduction of vascular oxidative stress. Lovastatin treatment has a weaker inhibitory effect on the progression of atherosclerosis and no effect on vascular NO elaboration, which may be due to its stimulatory effect on vascular superoxide radical generation.
Although aortic-valve stenosis is clearly associated with adverse cardiovascular outcomes, it is unclear whether valve sclerosis increases the risk of cardiovascular events.
We assessed echocardiograms obtained at base line from 5621 men and women 65 years of age or older who were enrolled in a population-based prospective study. On echocardiography, the aortic valve was normal in 70 percent (3919 subjects), sclerotic without outflow obstruction in 29 percent (1610), and stenotic in 2 percent (92). The subjects were followed for a mean of 5.0 years to assess the risk of death from any cause and of death from cardiovascular causes. Cardiovascular morbidity was defined as new episodes of myocardial infarction, angina pectoris, congestive heart failure, or stroke.
There was a stepwise increase in deaths from any cause (P for trend, <0.001) and deaths from cardiovascular causes (P for trend, <0.001) with increasing aortic-valve abnormality; the respective rates were 14.9 and 6.1 percent in the group with normal aortic valves, 21.9 and 10.1 percent in the group with aortic sclerosis, and 41.3 and 19.6 percent in the group with aortic stenosis. The relative risk of death from cardiovascular causes among subjects without coronary heart disease at base line was 1.66 (95 percent confidence interval, 1.23 to 2.23) for those with sclerotic valves as compared with those with normal valves, after adjustment for age and sex. The relative risk remained elevated after further adjustment for clinical factors associated with sclerosis (relative risk, 1.52; 95 percent confidence interval, 1.12 to 2.05). The relative risk of myocardial infarction was 1.40 (95 percent confidence interval, 1.07 to 1.83) among subjects with aortic sclerosis, as compared with those with normal aortic valves.
Aortic sclerosis is common in the elderly and is associated with an increase of approximately 50 percent in the risk of death from cardiovascular causes and the risk of myocardial infarction, even in the absence of hemodynamically significant obstruction of left ventricular outflow.
The guanidino-methylated arginine analogue NG monomethyl-L-arginine (L-NMMA) has been the standard nitric oxide synthase inhibitor used to evaluate the role of the L-arginine:nitric oxide pathway. However, L-NMMA and other methylated arginine residues are also synthesised in vivo by the action of a family of enzymes known as protein arginine methyltransferases. Proteolysis of proteins containing methylated arginine residues releases free methylarginine residues into the cytosol from where they may pass out of the cell into plasma. Of the three known methylarginine residues produced in mammals only asymmetrically methylated forms (L-NMMA and asymmetric dimethylarginine (ADMA)) but not symmetrically methylated arginine (symmetric dimethylarginine (SDMA)) inhibit nitric oxide synthase (NOS). We and others have proposed that endogenously produced asymmetrically methylated arginines may modulate NO production and that the accumulation of these residues in disease states may contribute to pathology. The activity of the enzyme dimethylarginine dimethylaminohydrolase that metabolises asymmetric methylarginines may be of critical importance in affecting NO pathways in health or disease.
Reactive oxygen species (ROS) are known to induce apoptotic cell death in various cell types. In the vessel wall, ROS can be formed by macrophages within the atherosclerotic plaque or can act on the endothelium after adhesion of monocytes or leucocytes. Moreover, ROS are endogenously synthesized by endothelial and vascular smooth muscle cells by NAD(P)H oxidase. Enhanced ROS production is a very early hallmark in the atherogenic process, suggesting a link between ROS and apoptosis. In endothelial cells, the endogenous generation of ROS is induced by different pro-inflammatory and pro-atherosclerotic factors such as Ang II, oxLDL or TNFalpha, which all promote the execution of programmed cell death. ROS synthesis is thereby causally involved in apoptosis induction, because antioxidants prevent endothelial cell death. The pro-apoptotic effects of endogenous ROS in endothelial cells mechanistically seems to involve the disturbance of mitochondrial membrane permeability followed by cytochrome c release, which finally activates the executioner caspases. In contrast to the pro-apoptotic capacity of ROS in endothelial cells, in vascular smooth muscle cells emerging evidence suggests that endogenous ROS synthesis promotes cell proliferation and hypertrophy and does not affect cell survival. However, high concentrations of exogenous ROS can also stimulate smooth muscle cell apoptosis as shown for other cell types probably via activation of p53. Taken together, the double-edged effects of endogenously derived ROS in endothelial cells versus VSMC may provide a mechanistic clue to the anti-atherosclerotic effects of antioxidants shown in experimental studies, given that the promotion of endothelial survival in combination with inhibition of VSMC proliferation blocks two very important steps in the pathogenesis of atherosclerosis.
Accumulating evidence suggests that oxidant stress alters many functions of the endothelium, including modulation of vasomotor tone. Inactivation of nitric oxide (NO(.)) by superoxide and other reactive oxygen species (ROS) seems to occur in conditions such as hypertension, hypercholesterolemia, diabetes, and cigarette smoking. Loss of NO(.) associated with these traditional risk factors may in part explain why they predispose to atherosclerosis. Among many enzymatic systems that are capable of producing ROS, xanthine oxidase, NADH/NADPH oxidase, and uncoupled endothelial nitric oxide synthase have been extensively studied in vascular cells. As the role of these various enzyme sources of ROS become clear, it will perhaps be possible to use more specific therapies to prevent their production and ultimately correct endothelial dysfunction.
Angiotensin-converting enzyme inhibitors improve endothelial function, inhibit experimental atherogenesis, and decrease ischemic events. The Quinapril Ischemic Event Trial was designed to test the hypothesis that quinapril 20 mg/day would reduce ischemic events (the occurrence of cardiac death, resuscitated cardiac arrest, nonfatal myocardial infarction, coronary artery bypass grafting, coronary angioplasty, or hospitalization for angina pectoris) and the angiographic progression of coronary artery disease in patients without systolic left ventricular dysfunction. A total of 1,750 patients were randomized to quinapril 20 mg/day or placebo and followed a mean of 27 +/- 0.3 months. The 38% incidence of ischemic events was similar for both groups (RR 1.04; 95% confidence interval 0.89 to 1.22; p = 0.6). There was also no significant difference in the incidence of patients having angiographic progression of coronary disease (p = 0.71). The rate of development of new coronary lesions was also similar in both groups (p = 0.35). However, there was a difference in the incidence of angioplasty for new (previously unintervened) vessels (p = 0.018). Quinapril was well tolerated in patients after angioplasty with normal left ventricular function. Quinapril 20 mg did not significantly affect the overall frequency of clinical outcomes or the progression of coronary atherosclerosis. However, the absence of the demonstrable effect of quinapril may be due to several limitations in study design.
Endothelium-dependent/nitric oxide (NO)-mediated vasodilation is impaired in hypertensive individuals. Asymmetric dimethylarginine (ADMA), an endogenous inhibitor of NO synthase, is synthesized by many types of cells including vascular endothelial cells. The serum level of ADMA is elevated in patients with essential hypertension, but the mechanism for this increase is unknown. Therefore, the present study examined whether the renin-angiotensin system (RAS) is involved. Patients with essential hypertension [systolic blood pressure (BP) > 160 mmHg and/or diastolic BP > 95 mmHg] were randomized to an angiotensin-converting enzyme (ACE) inhibitor treatment group (perindopril, 4mg/day for 4 weeks, n = 7), an angiotensin II type 1 (AT1) receptor antagonist treatment group (losartan, 50 mg/day for 4 weeks, n = 7) or a beta-blocker treatment group (bisoprolol, 5 mg/day for 4 weeks, n = 7). Before and after the treatment, BP, serum concentration of ADMA and plasma concentration of von Willebrand factor (vWF, a biological marker of endothelial injury) were measured. Perindopril, losartan and bisoprolol decreased BP to a similar extent, and either perindopril or losartan, but not bisoprolol, significantly decreased serum ADMA and plasma vWF. These findings suggest that the RAS may contribute to the mechanism of increased serum ADMA as well as to the endothelial injury observed in hypertensive patients. The vasculoprotective actions of ACE inhibitors or AT1 receptor antagonists may be explained at least in part by amelioration of the endothelial injury through a decrease in the serum ADMA concentration.
It is now well established that vascular inflammation is an independent risk factor for the development of atherosclerosis. In otherwise healthy patients, chronic elevations of circulating interleukin-6 or its biomarkers are predictors for increased risk in the development and progression of ischemic heart disease. Although multifactorial in etiology, vascular inflammation produces atherosclerosis by the continuous recruitment of circulating monocytes into the vessel wall and by contributing to an oxidant-rich inflammatory milieu that induces phenotypic changes in resident (noninflammatory) cells. In addition, the renin-angiotensin system (RAS) has important modulatory activities in the atherogenic process. Recent work has shown that angiotensin II (Ang II) has significant proinflammatory actions in the vascular wall, inducing the production of reactive oxygen species, inflammatory cytokines, and adhesion molecules. These latter effects on gene expression are mediated, at least in part, through the cytoplasmic nuclear factor-kappaB transcription factor. Through these actions, Ang II augments vascular inflammation, induces endothelial dysfunction, and, in so doing, enhances the atherogenic process. Our recent studies have defined a molecular mechanism for a biological positive-feedback loop that explains how vascular inflammation can be self-sustaining through upregulation of the vessel wall Ang II tone. Ang II produced locally by the inflamed vessel induces the synthesis and secretion of interleukin-6, a cytokine that induces synthesis of angiotensinogen in the liver through a janus kinase (JAK)/signal transducer and activator of transcription (STAT)-3 pathway. Enhanced angiotensinogen production, in turn, supplies more substrate to the activated vascular RAS, where locally produced Ang II synergizes with oxidized lipid to perpetuate atherosclerotic vascular inflammation. These observations suggest that one mechanism by which RAS antagonists prevent atherosclerosis is by reducing vascular inflammation. Moreover, antagonizing the vascular nuclear factor-kappaB and/or hepatic JAK/STAT pathways may modulate the atherosclerotic process.
Recent studies have demonstrated that lesions of aortic sclerosis and stenosis share several similarities with lesions of atherosclerosis. In atherosclerosis, angiotensin-converting enzyme (ACE) is expressed by a subset of macrophages. This study was undertaken to determine whether ACE might be present in aortic sclerosis or stenosis lesions.
Immunohistochemistry was performed on 26 paraffin-embedded human aortic valves. Monospecific antibodies were used to identify ACE, macrophages, angiotensin II type 1 receptor (AT-1 receptor), angiotensin II, and apolipoprotein B. Human low-density lipoprotein (LDL) and high-density lipoprotein (HDL) were isolated from plasma of normal volunteers by sequential density-gradient ultracentrifugation. ACE was not present in normal valves but was present in all valves with aortic sclerosis or stenosis lesions. ACE was detected on a subset of lesion macrophages but was present primarily in an extracellular distribution, where it colocalized with apolipoprotein B. ACE was detected by Western blotting on plasma LDL but not on HDL isolated from normal volunteers. Angiotensin II, the enzymatic product of ACE, was colocalized with ACE in valve lesions. ACE also was colocalized with apolipoprotein B in an adjacent coronary atherosclerotic plaque.
ACE is present in aortic sclerosis and stenosis lesions, where it may participate in lesion development, as is evidenced by the presence of its enzymatic product, angiotensin II. The observation of an association of ACE with LDL in both lesions and plasma suggests that LDL may deliver ACE to lesions and has implications for the role of ACE-containing LDL in other diseases, such as atherosclerosis.
Aortic valve stenosis characteristically progresses due to cuspal calcification, often necessitating valve replacement surgery. The present study investigated the hypothesis that TGF-beta1, a cytokine that causes calcification of vascular smooth muscle cells in culture, initiates apoptosis of valvular interstitial cells as a mechanistic event in cuspal calcification.
Noncalcified and calcified human aortic valve cusps were obtained at autopsy or at the time of cardiac surgery. The distributions within cusps of TGF-beta1, latent-TGF-beta1-associated peptide, and TGF-beta receptors were studied using immunohistochemistry. The effects of TGF-beta1 on mechanistic events contributing to aortic valve calcification were also investigated using sheep aortic valve interstitial cell (SAVIC) cultures.
Immunohistochemistry studies revealed that calcific aortic stenosis cusps characteristically contained within the extracellular matrix qualitatively higher levels of TGF-beta1 than noncalcified cusps. Noncalcified normal valves demonstrated only focal intracellular TGF-beta1. Addition of TGF-beta1 to SAVIC cultures led to a cascade of events, including: cellular migration, aggregation, formation of apoptotic-alkaline phosphatase enriched nodules, and calcification of these nodules. The time course of these events in the SAVIC culture system was rapid with nodule formation with apoptosis by 72 hours, and calcification after 7 days. Furthermore, ZVAD-FMK, an antiapoptosis agent (caspase inhibitor), significantly inhibited calcification and apoptosis induced by TGF-beta1, but had no effect on nodule formation. However, cytochalasin D, an actin-depolymerizing agent, inhibited nodule formation, but not calcification.
TGF-beta1 is characteristically present within calcific aortic stenosis cusps, and mediates the calcification of aortic valve interstitial cells in culture through mechanisms involving apoptosis.
We studied a known rabbit model of atherosclerosis to assess the effect of a hypercholesterolemic diet on aortic valve morphology and function. We also evaluated the effects of the combination of this diet with vitamin D supplements on the development of the disease and the occurrence of valve calcification.
Aortic valve stenosis (AVS) is the most common valvular heart disease. Recent observations have suggested a link between atherosclerosis and the development of AVS. However, until now, there has been no solid direct proof of this potential link.
Rabbits were divided in three groups: 1) no treatment; 2) cholesterol-enriched diet (0.5% cholesterol); and 3) cholesterol-enriched diet plus vitamin D(2) (50,000 IU/day). Echocardiographic assessment of the aortic valve was done at baseline and after 12 weeks of treatment. The aortic valve area (AVA) and maximal and mean transvalvular gradients were recorded and compared over time.
Control animals displayed no abnormalities of the aortic valve. Despite important increases in blood total cholesterol levels, animals in group 2 did not develop any significant functional aortic valve abnormality over 12 weeks. However, eight of 10 of the animals in group 3 developed a significant decrease in AVA (p = 0.004) and significant increases in transvalvular gradients (p = 0.003).
This study supports a potential link between atherosclerosis and the development of AVS. The differences noted between hypercholesterolemic animals with or without vitamin D(2) supplementation imply a significant role of calcium in the development of AVS, meriting further attention.
Activation of vascular NAD(P)H oxidases and the production of reactive oxygen species (ROS) by these enzyme systems are common in cardiovascular disease. In the past several years, a new family of NAD(P)H oxidase subunits, known as the non-phagocytic NAD(P)H oxidase (NOX) proteins, have been discovered and shown to play a role in vascular tissues. Recent studies make clearer the mechanisms of activation of the endothelial and vascular smooth muscle NAD(P)H oxidases. ROS produced following angiotensin II-mediated stimulation of NAD(P)H oxidases signal through pathways such as mitogen-activated protein kinases, tyrosine kinases and transcription factors, and lead to events such as inflammation, hypertrophy, remodeling and angiogenesis. Studies in mice that are deficient in p47(phox) and gp91(phox) (also known as NOX2) NAD(P)H oxidase subunits show that ROS produced by these oxidases contribute to cardiovascular diseases including atherosclerosis and hypertension. Recently, efforts have been devoted to developing inhibitors of NAD(P)H oxidases that will provide useful experimental tools and might have therapeutic potential in the treatment of human diseases.
Monocyte extravasation into the vessel wall has been shown to be a critical step in the development of atherosclerosis. Upon activation, monocytes produce a burst of superoxide anion due to activation of the NADPH oxidase enzyme complex. Monocyte-derived superoxide anion contributes to oxidant stress in inflammatory sites, is required for monocyte-mediated LDL oxidation, and alters basic cell functions such as adhesion and proliferation. We hypothesize that monocyte-derived superoxide anion production contributes to atherosclerotic lesion formation. In this brief review, we summarize our current understanding of the signal transduction pathways regulating NADPH oxidase activation and related superoxide anion production in activated human monocytes. Novel pathways are identified that may serve as future targets for therapeutic intervention in this pathogenic process. The contributions of superoxide anion and NADPH oxidase to atherogenesis are discussed. Future experiments are needed to clarify the exact role of NADPH oxidase-derived superoxide anion in atherogenesis, particularly that derived from monocytes.
The thioredoxin (TRX) system (TRX, TRX reductase, and NADPH) is a ubiquitous thiol oxidoreductase system that regulates cellular reduction/oxidation (redox) status. The oxidation mechanism for disease pathogenesis states that an imbalance in cell redox state alters function of multiple cell pathways. In this study, we review the essential role for TRX to limit oxidative stress directly via antioxidant effects and indirectly by protein-protein interaction with key signaling molecules, such as apoptosis signal-regulating kinase 1. We propose that TRX and its endogenous regulators are important future targets to develop clinical therapies for cardiovascular disorders associated with oxidative stress.
A large number of studies have demonstrated the role of angiotensin II in cardiac preconditioning against ischemic reperfusion injury. Generally, angiotensin II is a detrimental factor for the heart, and its inhibition with an ACE inhibitor provides cardioprotection. This review provides an explanation for such paradoxical behavior of angiotensin II. Angiotensin II can potentiate the induction of the expression of a variety of redox-sensitive factors including p38 MAPK, JNK and Akt, IGF-IR, EGF-R, and HO-1 as well as redox-regulated genes and transcription factors such as NFkappaB. It becomes increasingly apparent that during the earlier phase, the heart attempts to adapt itself against the detrimental effects of angiotensin II by upregulating several cardioprotective genes and proteins. These genes and proteins are redox-regulated and the antioxidants or ROS scavengers block their expressions. Interestingly, an identical pattern of cardioprotective proteins and genes are expressed in the preconditioned heart, which are also inhibited with ROS scavengers. It is tempting to speculate that the induction of the expression of the redox-sensitive cardioprotective proteins is the results of adaptation of the heart against the oxidative stress resulting from angiotensin II; and preconditioning is the net result of harnessing its own protection during ischemic and/or oxidative stress through its ability to trigger redox signaling.
A new HPLC assay for the detection of L-arginine, NG, NG-dimethyl-L-arginine (ADMA) and NG, NG' -dimethyl-L-arginine (SDMA) in plasma using the derivatisation reagent AccQ-Fluor (6-aminoquinolyl-N-hydroxysuccinimidyl carbamate) is described. The fluorescent derivatives produced are extremely stable enabling routine processing of large numbers of samples. Arginine and its metabolites are extracted from plasma on strong cation exchange (SCX) cartridges with NG-monomethyl-L-arginine (NMMA) as internal standard, derivatised and separated on a C18 column with acetonitrile in 0.1M sodium acetate buffer pH 6. Separation of the stereoisomers ADMA and SDMA was excellent and improvements to the solid phase extraction (SPE) procedure enabled good recovery (>80%) of arginine, ADMA and SDMA. The utility of the method is exemplified by comparison of plasma concentrations of ADMA, SDMA and arginine in healthy volunteers and diabetic/ischaemic patients.
Recently, statins and angiotensin-converting enzyme inhibitors (ACEIs) have been shown to slow aortic valve calcium accumulation. Although several studies also suggest that statins may reduce the hemodynamic progression of aortic stenosis (AS), no data are available for ACEIs or the combination of both.
A total of 211 consecutive patients (aged 70+/-10 years, 104 females) with native AS, defined by a peak velocity >2.5 m/s (valve area 0.84+/-0.23 cm(2), mean gradient 42+/-19 mm Hg), with normal left ventricular function and no other significant valvular lesion who were examined between 2000 and 2002 and who had 2 echocardiograms separated by at least 6 months were included. Of these, 102 patients were treated with ACEIs, 50 patients received statins, and 32 patients received both. Hemodynamic progression of AS was assessed and related to medical treatment. Annualized increase in peak aortic jet velocity for the entire study group was 0.32+/-0.44 m x s(-1) x y(-1). Progression was significantly lower in patients treated with statins (0.10+/-0.41 m x s(-1) x y(-1)) than in those who were not (0.39+/-0.42 m x s(-1) x y(-1); P<0.0001). This effect was observed both in mild-to-moderate and severe AS. ACEI use, however, did not significantly affect hemodynamic progression (P=0.29). Furthermore, ACEIs had no additional effect on AS progression when given in combination with statins (0.11+/-0.42 versus 0.08+/-0.43 m x s(-1) x y(-1) for combination versus statin only; P=0.81). Cholesterol levels did not correlate with hemodynamic progression either in the group receiving statins or in the group that did not.
ACEIs do not appear to slow AS progression. However, statins significantly reduce the hemodynamic progression of both mild-to-moderate and severe AS, an effect that may not be related to cholesterol lowering.
The development of therapeutic interventions to prevent progressive valve damage is more likely to limit the progression of structural damage to the aortic valve with normal function (aortic sclerosis [ASC]) than clinically apparent aortic stenosis. Currently, the ability to appreciate the progression of ASC is compromised by the subjective and qualitative evaluation of sclerosis severity.
We sought to reveal whether the intensity of ultrasonic backscatter could be used to quantify sclerosis severity in 26 patients with ASC and 23 healthy young adults. Images of the aortic valve were obtained in the parasternal long-axis view and saved in raw data format. Six square-shaped 11 x 11 pixel regions of interest were placed on the anterior and posterior leaflets, and calibrated backscatter values were obtained by subtracting the regions of interest in the blood pool from the averaged backscatter values obtained from the leaflets.
Mean ultrasonic backscatter values for sclerotic valves exceeded the results in normal valve tissue (16.3 +/- 4.4 dB vs 9.8 +/- 3.3 dB, P < .0001). Backscatter values were greater (22.0 +/- 3.5 dB) in a group of 6 patients with aortic stenosis. Within the sclerosis group, the magnitude of backscatter was directly correlated (P < .05) with a subjective sclerosis score, and with transvalvular pressure gradient. Mean reproducibility was 2.4 +/- 1.8 dB (SD) between observers, and 2.3 +/- 1.7 dB (SD) between examinations.
Measurement of backscatter from the valve leaflets of patients with ASC may be a feasible means of following the progression and treatment response of aortic sclerosis.
The purpose of this study was to investigate the expression of angiotensin II (Ang II)-producing enzyme systems in normal and stenotic aortic valves.
Chronic inflammation and fibrosis are involved in the pathogenesis of aortic stenosis (AS), but the detailed molecular mechanisms of this atherosclerosis-like process remain obscure. Angiotensin II, a powerful mediator of inflammation and fibrosis, may participate in AS progression.
Stenotic aortic valves (n = 86) were obtained from patients undergoing valve replacement surgery, and control valves (n = 11) were obtained from patients undergoing cardiac transplantation. Angiotensin-converting enzyme (ACE) and mast cell (MC)-derived chymase were quantified by reverse-transcription polymerase chain reaction, autoradiography, and immunostaining. The MCs, macrophages, and T lymphocytes were detected by immunohistochemistry, and angiotensin II type 1 receptor (AT-1R) by autoradiography.
Compared with control valves, stenotic aortic valves showed a significant increase in both messenger ribonucleic acid (mRNA) (p = 0.001) and protein (p < 0.001) expression of ACE, which colocalized with macrophages. Similarly, the expression of AT-1R protein and chymase mRNA and protein was upregulated (p < 0.001), and the number of MCs was six-fold higher in stenotic than in normal valves. The MCs were associated with the calcified areas, and-in contrast to control valves-showed an increased degree of degranulation, a prerequisite for chymase secretion and action.
Angiotensin-converting enzyme and chymase, two Ang II-forming enzymes, are locally expressed in aortic valves, and owing to infiltration of macrophages and MCs, are further upregulated in stenotic valves. These novel findings, implicating chronic inflammation and an increased expression of local Ang II-forming systems, suggest that therapeutic interventions aiming at inhibiting these processes may slow AS progression.
Long-term administration of angiotensin II causes myocardial loss and cardiac fibrosis. We previously found iron deposition in the heart of the angiotensin II-infused rat, which may promote angiotensin II-induced cardiac damage. In the present study, we have investigated whether an iron chelator (deferoxamine) and a free radical scavenger (T-0970) affect the angiotensin II-induced upregulation of transforming growth factor-beta1 (TGF-beta1). Angiotensin II infusion for 7 days caused a robust increase in TGF-beta1 mRNA expression in vascular smooth muscle cells, myofibroblast-like cells, and migrated monocytes/macrophages. T-0970 and deferoxamine suppressed the upregulation of TGF-beta1 mRNA and reduced the extent of cardiac fibrosis in the heart of rats treated with angiotensin II. These agents blocked the angiotensin II-induced upregulation of heme oxygenase-1, a potent oxidative and cellular stress-responsive gene, but they did not significantly affect systolic blood pressure or plasma levels of aldosterone. In addition, T-0970 and deferoxamine suppressed the angiotensin II-induced upregulation of monocyte chemoattractant protein-1 in the heart. These results collectively suggest that iron and the iron-mediated generation of reactive oxygen species may contribute to angiotensin II-induced upregulation of profibrotic and proinflammatory genes, such as TGF-beta1 and monocyte chemoattractant protein-1.
Calcium accumulation in the aortic valve is a hallmark of aortic sclerosis and aortic stenosis. Because lipoproteins, angiotensin-converting enzyme, and angiotensin II colocalize with calcium in aortic valve lesions, we hypothesized an association between angiotensin-converting enzyme inhibitor (ACEI) use and lowered aortic valve calcium (AVC) accumulation, as measured by electron beam computed tomography.
Rates of change in volumetric AVC scores were determined retrospectively for 123 patients who had undergone 2 serial electron beam computed tomographic scans. The mean (+/-SD) interscan interval was 2.5 (+/-1.7) years; 80 patients did not receive ACEIs and 43 received ACEIs. The relationship of ACEI use to median rates of AVC score change (both unadjusted and adjusted for baseline AVC scores and coronary heart disease risk factors) was determined. We also examined the relationship of ACEI use to the likelihood of and adjusted odds ratio for definite progression (AVC change >2 times the median interscan variability).
Unadjusted and adjusted median rates of AVC score change were significantly higher in the no-ACEI group than in the ACEI group (adjusted median AVC changes [95% confidence interval]: relative, 28.7%/y [18.9%-38.5%/y] vs 11.0%/y [-1.9% to 24.0%/y], P = .04; absolute: 25.1/y [19.7-30.5/y] vs 12.2/y [4.5-19.9/y], P = .02). The adjusted odds ratio (95% confidence interval) for definite AVC progression was significantly lower for patients who received ACEIs (0.29 [0.11-0.75], P = .01).
This retrospective study finds a significant association between ACEI use and a lower rate of AVC accumulation. The results support the need for prospective, randomized trials of ACEIs in calcific aortic valve disease.
Calcific aortic stenosis has many characteristics in common with atherosclerosis, including hypercholesterolemia. We hypothesized that intensive lipid-lowering therapy would halt the progression of calcific aortic stenosis or induce its regression.
In this double-blind, placebo-controlled trial, patients with calcific aortic stenosis were randomly assigned to receive either 80 mg of atorvastatin daily or a matched placebo. Aortic-valve stenosis and calcification were assessed with the use of Doppler echocardiography and helical computed tomography, respectively. The primary end points were change in aortic-jet velocity and aortic-valve calcium score.
Seventy-seven patients were assigned to atorvastatin and 78 to placebo, with a median follow-up of 25 months (range, 7 to 36). Serum low-density lipoprotein cholesterol concentrations remained at 130+/-30 mg per deciliter in the placebo group and fell to 63+/-23 mg per deciliter in the atorvastatin group (P<0.001). Increases in aortic-jet velocity were 0.199+/-0.210 m per second per year in the atorvastatin group and 0.203+/-0.208 m per second per year in the placebo group (P=0.95; adjusted mean difference, 0.002; 95 percent confidence interval, -0.066 to 0.070 m per second per year). Progression in valvular calcification was 22.3+/-21.0 percent per year in the atorvastatin group, and 21.7+/-19.8 percent per year in the placebo group (P=0.93; ratio of post-treatment aortic-valve calcium score, 0.998; 95 percent confidence interval, 0.947 to 1.050).
Intensive lipid-lowering therapy does not halt the progression of calcific aortic stenosis or induce its regression. This study cannot exclude a small reduction in the rate of disease progression or a significant reduction in major clinical end points. Long-term, large-scale, randomized, controlled trials are needed to establish the role of statin therapy in patients with calcific aortic stenosis.
Patients with aortic stenosis (AS) exhibit increased platelet aggregability, and thrombus formation has been documented on calcific and severely stenosed valves. Isolated porcine and canine aortic valves (AV) release nitric oxide (NO) and prostacyclin, which exert local antithrombotic effects; to date, this has not been studied in humans. In the present study the possible interaction of AV tissue with platelet aggregation was examined, using fragments of AV obtained from patients with AS and aortic regurgitation (AR).
Fragments of AV tissue, excised from patients undergoing AV replacement, were co-incubated with blood samples obtained from normal subjects. The direct effects of valve tissue from patients with AS (n = 14) or with predominant AR (n = 13) on ADP-induced platelet aggregation and intraplatelet cGMP and cAMP content were compared.
In whole blood, non-calcified AV fragments from AR patients inhibited platelet aggregation by 57 +/- 6% (p < 0.01); in platelet-rich plasma results were analogous. In order to determine whether this anti-aggregatory effect could be attributed to the valvular release of NO or prostacyclin, intraplatelet cGMP and cAMP formation was assessed, respectively. While there were no significant changes in cGMP content, cAMP increased by 26 +/- 4% (p < 0.02). Both, anti-aggregatory and cAMP-stimulating effects were similar to those produced by 10 nM prostaglandin E1, a prostacyclin mimetic. Fragments from stenotic valves did not inhibit aggregation and did not affect cGMP or cAMP. Furthermore, fragments from heavily calcified regions potentiated aggregation and, in some cases, induced spontaneous aggregation.
Minimally calcified aortic valves (i.e., AR) and, therefore, presumably also normal valves, exert anti-aggregatory effects, most likely via prostacyclin release. AS is associated with a loss of this effect, thus potentially contributing to thrombotic risk.