Vasodilator reactive oxygen species ameliorate perturbed myocardial oxygen delivery in exercising swine with multiple comorbidities.

Multiple common cardiovascular comorbidities produce coronary microvascular dysfunction. We previously observed in swine that a combination of diabetes mellitus (DM), high fat diet (HFD) and chronic kidney disease (CKD) induced systemic inflammation, increased oxidative stress and produced coronary endothelial dysfunction, altering control of coronary microvascular tone via loss of NO bioavailability, which was associated with an increase in circulating endothelin (ET). In the present study, we tested the hypotheses that (1) ROS scavenging and (2) ETA+B-receptor blockade improve myocardial oxygen delivery in the same female swine model. Healthy female swine on normal pig chow served as controls (Normal). Five months after induction of DM (streptozotocin, 3 × 50 mg kg-1 i.v.), hypercholesterolemia (HFD) and CKD (renal embolization), swine were chronically instrumented and studied at rest and during exercise. Sustained hyperglycemia, hypercholesterolemia and renal dysfunction were accompanied by systemic inflammation and oxidative stress. In vivo ROS scavenging (TEMPOL + MPG) reduced myocardial oxygen delivery in DM + HFD + CKD swine, suggestive of a vasodilator influence of endogenous ROS, while it had no effect in Normal swine. In vitro wire myography revealed a vasodilator role for hydrogen peroxide (H2O2) in isolated small coronary artery segments from DM + HFD + CKD, but not Normal swine. Increased catalase activity and ceramide production in left ventricular myocardial tissue of DM + HFD + CKD swine further suggest that increased H2O2 acts as vasodilator ROS in the coronary microvasculature. Despite elevated ET-1 plasma levels in DM + HFD + CKD swine, ETA+B blockade did not affect myocardial oxygen delivery in Normal or DM + HFD + CKD swine. In conclusion, loss of NO bioavailability due to 5 months exposure to multiple comorbidities is partially compensated by increased H2O2-mediated coronary vasodilation.

Questionnaire survey on cardiologists' view and management of coronary microvascular disease in clinical practice.

OBJECTIVE: We aimed to assess the opinion of Dutch cardiologists on coronary microvascular disease (CMD) and its management in clinical practice, and to assess the need for a CMD guideline among Dutch cardiologists. METHODS: We developed an online questionnaire including different aspects of CMD which was reviewed by an expert panel. The questionnaire was distributed by e­mail among all members of the Dutch Society of Cardiology. RESULTS: A total of 103 cardiologists (70% male) completed the questionnaire (response rate: 10%). Median age and years of experience as a cardiologist were 49 ± 15 and 12 ± 12 years, respectively. Overall, 93% of the cardiologists had considered the CMD diagnosis, 85% had ever made such a diagnosis, 90% had treated a patient with CMD, and 61% had referred patients to tertiary care. The median (interquartile range) self-rated knowledge level was 7.0 (2.0) (scale of 0-10). 84% rated their knowledge as sufficient (>5.5) and 58% viewed CMD as a disease entity. Overall, 61% and 17%, respectively, agreed that evidence-based diagnostic and treatment modalities for CMD do not exist, while 56% believed that CMD patients have a higher risk for cardiovascular disease and mortality. Finally, 82% of the responders stated that a CMD guideline is needed, and 91% wanted to receive the guideline once developed. DISCUSSION: Fifty-eight per cent of the responders recognise CMD as a separate disease entity. Our study underscores the need for a dedicated CMD guideline for Dutch cardiology practice. However, the response rate was low (10%), and it is likely that mainly cardiologists interested in CMD have participated in our study.

CECR1-mediated cross talk between macrophages and vascular mural cells promotes neovascularization in malignant glioma.

Glioblastomas (glioblastoma multiforme, GBM) are most malignant brain tumors characterized by profound vascularization. The activation of macrophages strongly contributes to tumor angiogenesis during GBM development. Previously, we showed that extracellular adenosine deaminase protein Cat Eye Syndrome Critical Region Protein 1 (CECR1) is highly expressed by M2-like macrophages in GBM where it defines macrophage M2 polarization and contributes to tumor expansion. In this study, the effect of CECR1 in macrophages on tumor angiogenesis was investigated. Immunohistochemical evaluation of GBM tissue samples showed that the expression of CECR1 correlates with microvascular density in the tumors, confirming data from the TCGA set. In a three-dimensional co-culture system consisting of human pericytes, human umbilical vein endothelial cells and THP1-derived macrophages, CECR1 knockdown by siRNA and CECR1 stimulation of macrophages inhibited and promoted new vessel formation, respectively. Loss and gain of function studies demonstrated that PDGFB mRNA and protein levels in macrophages are modulated by CECR1. The proangiogenic properties of CECR1 in macrophages were partially mediated via paracrine activation of pericytes by PDGFB-PDGFRß signaling. CECR1-PDGFB-PDGFRß cross-activation between macrophages and pericytes promoted pericyte migration, shown by transwell migration assay, and enhanced expression and deposition of periostin, a matrix component with proangiogenic properties. CECR1 function in (M2-like) macrophages mediates cross talk between macrophages and pericytes in GBM via paracrine PDGFB-PDGFRß signaling, promoting pericyte recruitment and migration, and tumor angiogenesis. Therefore, CECR1 offers a new portent target for anti-angiogenic therapy in GBM via immune modulation.

Animal models of heart failure with preserved ejection fraction.

Heart failure with preserved ejection fraction (HFpEF) constitutes a clinical syndrome in which the diagnostic criteria of heart failure are not accompanied by gross disturbances of systolic function, as assessed by ejection fraction. In turn, under most circumstances, diastolic function is impaired. Although it now represents over 50 % of all patients with heart failure, the mechanisms of HFpEF remain understood, precluding effective therapy. Understanding the pathophysiology of HFpEF has been restricted by both limited access to human myocardial biopsies and by the lack of animal models that fully mimic human pathology. Animal models are valuable research tools to clarify subcellular and molecular mechanisms under conditions where the comorbidities and other confounding factors can be precisely controlled. Although most of the heart failure animal models currently available represent heart failure with reduced ejection fraction, several HFpEF animal models have been proposed. However, few of these fulfil all the features present in human disease. In this review we will provide an overview of the currently available models to study HFpEF from rodents to large animals as well as present advantages and disadvantages of these models.

Peripheral arterial tonometry cannot detect patients at low risk of coronary artery disease.

BACKGROUND: Endothelial dysfunction precedes coronary artery disease (CAD) and can be measured by peripheral arterial tonometry (PAT). We examined the applicability of PAT to detect a low risk of CAD in a chest pain clinic. METHODS: In 93 patients, PAT was performed resulting in reactive hyperaemia (RHI) and augmentation (AIx) indices. Patients were risk classified according to HeartScore, Diamond and Forrester pretest probability (DF), exercise testing (X-ECG), and computed tomography calcium scoring (CCS) and angiography (CTA). Correlations, risk group differences and prediction of revascularisation within 1 year were calculated. RESULTS: RHI correlated with HeartScore (r = - 0.21, p = 0.05), AIx with DF (r = 0.26, p = 0.01). However, both were not significantly different between normal and ischaemic X-ECG groups. In addition RHI and AIx were similar between low risk as compared with intermediate-to-high risk, based on risk algorithms (RHI: 1.98 (0.67) vs 1.94 (0.78); AIx: 0.0 (21) vs 5.0 (25); p = NS), or CCS and CTA (RHI: 1.99 (0.58) vs 1.89 (0.82); AIx: - 2.0 (24) vs 4.0 (25); p = NS). Finally, RHI and AIx failed to predict revascularisation (RHI: OR 1.42, CI 0.65-3.1; AIx: OR 1.02, CI 0.98-1.05). CONCLUSIONS: PAT cannot detect a low risk of CAD, possibly because RHI and AIx versus X-ECG, CCS and CTA represent independent processes.

The impact of Fourier-Domain optical coherence tomography catheter induced motion artefacts on quantitative measurements of a PLLA-based bioresorbable scaffold.

Intracoronary Fourier-Domain optical coherence tomography (FD-OCT) enables imaging of the coronary artery within 2-4 seconds, a so far unparalleled speed. Despite such fast data acquisition, cardiac and respiratory motion can cause artefacts due to longitudinal displacement of the catheter within the artery. We studied the influence of longitudinal FD-OCT catheter displacement on serial global lumen and scaffold area measurements in coronary arteries of swine that received PLLA-based bioresorbable scaffolds. In 10 swine, 20 scaffolds (18 × 3.0 mm) were randomly implanted in two epicardial coronary arteries. Serial FD-OCT imaging was performed immediately after implantation (T1) and at 3 (T2) and 6 months (T3) follow-up. Two methods for the selection of OCT cross-sections were compared. Method A did not take into account longitudinal displacement of the FD-OCT catheter. Method B accounted for longitudinal displacement of the FD-OCT catheter. Fifty-one OCT pullbacks of 17 scaffolds were serially analyzed. The measured scaffold length differed between time points, up to one fourth of the total scaffold length, indicating the presence of longitudinal catheter displacement. Between method A and B, low error was demonstrated for mean area measurements. Correlations between measurements were high: R2 ranged from 0.91 to 0.99 for all mean area measurements at all time points. Considerable longitudinal displacement of the FD-OCT catheter was observed, diminishing the number of truly anatomically matching cross-sections in serial investigations. Global OCT dimensions such as mean lumen and scaffold area were not significantly affected by this displacement. Accurate co-registration of cross-sections, however, is mandatory when specific regions, e.g. jailed side branch ostia, are analyzed.

Cardiovascular performance of adult breeding sows fails to obey allometric scaling laws.

In view of the remarkable decrease of the relative heart weight (HW) and the relative blood volume in growing pigs, we investigated whether HW, cardiac output (CO), and stroke volume (SV) of modern growing pigs are proportional to BW, as predicted by allometric scaling laws: HW (or CO or SV) = a·BW(b), in which a and b are constants, and constant b is a multiple of 0.25 (quarter-power scaling law). Specifically, we tested the hypothesis that both HW and CO scale with BW to the power of 0.75 (HW or CO = a·BW(0.75)) and SV scales with BW to the power of 1.00 (SV = a·BW(1.0)). For this purpose, 2 groups of pigs (group 1, consisting of 157 pigs of 50 ± 1 kg; group 2, consisting of 45 pigs of 268 ± 18 kg) were surgically instrumented with a flow probe or a thermodilution dilution catheter, under open-chest anesthetized conditions to measure CO and SV, after which HW was determined. The 95% confidence intervals of power-coefficient b for HW were 0.74 to 0.80, encompassing the predicted value of 0.75, suggesting that HW increased proportionally with BW, as predicted by the allometric scaling laws. In contrast, the 95% confidence intervals of power-coefficient b for CO and SV as measured with flow probes were 0.40 to 0.56 and 0.39 to 0.61, respectively, and values obtained with the thermodilution technique were 0.34 to 0.53 and 0.40 to 0.62, respectively. Thus, the 95% confidence limits failed to encompass the predicted values of b for CO and SV of 0.75 and 1.0, respectively. In conclusion, although adult breeding sows display normal heart growth, cardiac performance appears to be disproportionately low for BW. This raises concern regarding the health status of adult breeding sows.

Postconditioning against ischaemia-reperfusion injury: ready for wide application in patients?

Ischaemic postconditioning (IPOC) is an intervention in which brief, intermittent periods of reocclusion at the onset of reperfusion (i.e. stuttering reperfusion) protect myocardium from lethal reperfusion injury. The mechanism underlying the cardioprotective effects of IPOC is incompletely understood. However, it is perceived that IPOC begins with specific cell-surface receptors responsible for activating a number of signalling pathways, many of which appear to converge at the mitochondrial level. IPOC has been demonstrated both in animal models and in patients with acute myocardial infarction (AMI) in small proof-of-concept trials. This intervention offers the possibility of further limiting infarct size in patients undergoing primary percutaneous coronary intervention (PCI). Here, we provide a brief overview of the concept of IPOC and the mechanisms underlying this phenomenon. (Neth Heart J 2010;18:389-93.).

Does cardiovascular performance of modern fattening pigs obey allometric scaling laws?

In view of the remarkable decrease of the relative heart weight and the relative blood volume in growing pigs, we investigated whether cardiac output (CO) and stroke volume (SV) of modern growing pigs are proportional to body mass (M), as predicted by allometric scaling laws: CO (or SV) = a.M(b), in which b is a multitude of 0.25 (quarter power scaling law). Specifically, we tested the hypothesis that CO scales with M to the power of 0.75 (CO = a.M(0.75)) and SV scales with M to the power of 1.00 (SV = a.M(1.0)) and investigated whether these relations persisted during increased cardiac stress. For this purpose, 2 groups of pigs (group 1 of 57 +/- 3 kg in Lelystad, and group 2 of 28 +/- 1 kg in Rotterdam) were chronically instrumented with a flow probe to measure CO and SV; instrumented pigs were studied at rest and during strenuous exercise (at approximately 85% of maximum heart rate). Analysis of both groups of pigs (analyzed separately or combined) under resting conditions demonstrated that the 95% confidence intervals of power-coefficient b for CO encompassed 0.75 and for SV encompassed 1.0. During exercise, similar results were obtained, except for SV in group 2, in which the 95% confidence limits remained below 1.0, which may have been due to the relatively small range of BW in group 2. These observations indicate that CO and SV of growing pigs with M less than 75 kg are still proportional to M, even during strenuous exercise, and that CO and SV scale with M according to the quarter power scaling laws. In conclusion, the concerns about disproportional growth and development of modern growing pigs with BW up to 75 kg were not confirmed by the present study.

Cell transplantation for cardiac regeneration: where do we stand?

During the last decade transplantation of cells into the heart has emerged as a novel therapy for the prevention and treatment of heart failure. Although various cell types have been used, most experience has been obtained with the progenitor cells of skeletal muscle, also called myoblasts, and a wide array of bone marrow-derived cell types. The first preclinical studies demonstrated an improvement in global and regional heart function that was attributed mainly to a direct contractile effect of the transplanted cells. Furthermore, it was suggested that multiple cell types are able to form true cardiomyocytes and truly 'regenerate' the myocardium. More recent studies have questioned these early findings. Other mechanisms such as paracrine effects on the infarct and remote myocardium, a reduction in adverse remodelling and improvement of mechanical properties of the infarct tissue likely play a more important role. On the basis of encouraging preclinical studies, multiple early-phase clinical trials and several randomised controlled trials have been conducted that have demonstrated the feasibility, safety and potential efficacy of this novel therapy in humans. This review summarises the available evidence on cardiac cell transplantation and provides an outlook on future preclinical and clinical research that has to fill in the remaining gaps. (Neth Heart J 2008;16:88-95.).

Contrast enhancement of coronary arteries in cardiac surgery: a new multispectral stereoscopic camera technique.

OBJECTIVE: During open heart surgery, the myocardium usually provides sufficient visual contrast with both epicardial veins and arteries. However, visibility of coronary arteries may occasionally be impaired due to, e.g., intra-myocardial course of coronary arteries, increased epicardial fat, epicardial post-surgical adhesions, or pericarditis. Seen within the near infra-red range, coronary arteries show higher contrasts in relation to the myocardium than coronary veins. Hence, we developed a non-contact stereo-optical camera to selectively enhance coronary arteries by combining visible and near infra-red images. In this paper we present our first results on porcine and human hearts. MATERIALS AND METHODS: Two CMOS-cameras, with apochromatic lenses and dual-band LED-arrays, -captured visible colour (visible range, or VIS, 400-780nm) and near infra-red grey-scale (near infra-red range, or NIR, 910-920nm) images by sequentially switching between LED-array emission bands. Data was recorded by computer and processed off-line. Arterial NIR contrasts were algorithmically distinguished from shadows and specular reflections. Detected arteries were selectively enhanced and back-projected into the stereoscopic VIS-colour-image using either a 3D-display or conventional shutter glasses. RESULTS: Our technique visualised coronary vasculature and allowed to identify concealed parts of coronary arteries using off-line processing. Raw VIS & NIR images were real-time, processing took < 15s after filming. CONCLUSION: The applied principle works, but needs further development.

In vitro imaging of single living human umbilical vein endothelial cells with a clinical 3.0-T MRI scanner.

Iron oxide-labelled, single, living human umbilical vein endothelial cells (HUVECs) were imaged over time in vitro using a clinical 3.0-T magnetic resonance (MR) microscopy system. Labelling efficiency, toxicity, cell viability, proliferation and differentiation were assessed using flow cytometry, magnetic cell sorting and a phenanthroline assay. MR images were compared with normal light and fluorescence microscopy. Efficient uptake of iron oxide into HUVECs was shown, although with higher label uptake dose-dependent cytotoxic effects were observed, affecting cell viability. For MR imaging, a T2* weighted three-dimensional protocol was used with in-plane resolution of 39 x 48 microm2 and 100-microm slices with a scan time of 13 min. MRI could detect living cells in standard culture dishes at single-cell resolution, although label loss was observed that corresponded with the intracellular iron measurements. MR microscopy using iron oxide labels is a promising tool for studying HUVEC migration and cell biology in vitro and in vivo, but possible toxic effects of label uptake and loss of label over time should be taken into account.

Alterations in myofilament function contribute to left ventricular dysfunction in pigs early after myocardial infarction.

Myocardial infarction (MI) initiates cardiac remodeling, depresses pump function, and predisposes to heart failure. This study was designed to identify early alterations in Ca2+ handling and myofilament proteins, which may contribute to contractile dysfunction and reduced beta-adrenergic responsiveness in postinfarct remodeled myocardium. Protein composition and contractile function of skinned cardiomyocytes were studied in remote, noninfarcted left ventricular (LV) subendocardium from pigs 3 weeks after MI caused by permanent left circumflex artery (LCx) ligation and in sham-operated pigs. LCx ligation induced a 19% increase in LV weight, a 69% increase in LV end-diastolic area, and a decrease in ejection fraction from 54+/-5% to 35+/-4% (all P<0.05), whereas cardiac responsiveness to exercise-induced increases in circulating noradrenaline levels was blunted. Endogenous protein kinase A (PKA) was significantly reduced in remote myocardium of MI animals, and a negative correlation (R=0.62; P<0.05) was found between cAMP levels and LV weight-to-body weight ratio. Furthermore, SERCA2a expression was 23% lower after MI compared with sham. Maximal isometric force generated by isolated skinned myocytes was significantly lower after MI than in sham (15.4+/-1.5 versus 19.2+/-0.9 kN/m2; P<0.05), which might be attributable to a small degree of troponin I (TnI) degradation observed in remodeled postinfarct myocardium. An increase in Ca2+ sensitivity of force (pCa50) was observed after MI compared with sham (DeltapCa50=0.17), which was abolished by incubating myocytes with exogenous PKA, indicating that the increased Ca2+ sensitivity resulted from reduced TnI phosphorylation. In conclusion, remodeling of noninfarcted pig myocardium is associated with decreased SERCA2a and myofilament function, which may contribute to depressed LV function. The full text of this article is available online at http://circres.ahajournals.org.

High-resolution magnetic resonance imaging of iron-labeled myoblasts using a standard 1.5-T clinical scanner.

Myoblast transplantation is a promising means of restoring cardiac function in infarcted areas. For optimization of transplant protocols, tracking the location and fate of the injected cells is necessary. An attractive imaging modality for this is magnetic resonance imaging (MRI) as it is noninvasive and as iron-labeled myoblasts provide a signal attenuation in T2*-weighted protocols. The aim of this study was to develop an efficient iron-labeling protocol for myoblasts and to visualize single-labeled cells using a clinical 1.5-T scanner. Pig myoblasts were labeled with a superparamagnetic iron oxide (SPIO) agent using a liposome transfection agent. Labeling efficiency, toxicity, cell viability, and proliferative capacity were measured for 10 days. Magnetic resonance (MR) of myoblast cultures used a T2*-weighted three-dimensional protocol with a maximum in-plane resolution of 19.5 x 26.0 microm2 and 50 microm slices. Use of liposomes improved SPIO labeling efficiency. Labeling did not induce toxicity or affect cell viability or proliferation. The cell distribution as observed with light and fluorescence microscopy matched the signal voids observed in the MRI datasets. Liposomes promote fast, nontoxic and efficient SPIO labeling of myoblasts that can be tracked by MRI microscopy in clinical scanners using susceptibility-weighted protocols.

Regulation of coronary blood flow. Effect of coronary artery stenosis.

The consistently high level of myocardial oxygen extraction requires tight control of coronary blood flow, because an increase in myocardial oxygen demand, as occurs during exercise, cannot be obtained by a further increase in oxygen extraction. Consequently, adequate control of coronary vascular resistance is critical. Coronary resistance vessel tone is the result of a myriad of vasodilator and vasoconstrictor influences, which are exerted by the myocardium, endothelium and neurohumoral status. Unraveling of the integrative mechanisms controlling metabolic vasodilation has been difficult, more than likely due to the redundancy design of vasomotor control. In contrast to the traditional view that myocardial ischemia produced by a coronary artery stenosis causes maximal microvascular dilation, more recent studies have shown that the coronary microvessels retain some degree of vasodilator reserve during ischemia and remain responsive to vasoconstrictor stimuli. These observations raise the question of whether pharmacologic vasodilators acting at the microvascular level might be therapeutically useful. The critical property of effective vasodilator therapy requires selective dilation of small arteries, while avoiding coronary steal by not interfering with metabolic vasoregulation at the level of the arterioles.