Gene silencing of endothelial von Willebrand Factor attenuates angiotensin II-induced endothelin-1 expression in porcine aortic endothelial cells.

Expression of endothelin (ET)-1 is increased in endothelial cells exposed to angiotensin II (Ang II), leading to endothelial dysfunction and cardiovascular disorders. Since von Willebrand Factor (vWF) blockade improves endothelial function in coronary patients, we hypothesized that targeting endothelial vWF with short interference RNA (siRNA) prevents Ang II-induced ET-1 upregulation. Nearly 65 ± 2% silencing of vWF in porcine aortic endothelial cells (PAOECs) was achieved with vWF-specific siRNA without affecting cell viability and growth. While showing ET-1 similar to wild type cells at rest, vWF-silenced cells did not present ET-1 upregulation during exposure to Ang II (100 nM/24 h), preserving levels of endothelial nitric oxide synthase activity similar to wild type. vWF silencing prevented AngII-induced increase in nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activity and superoxide anion (O2-) levels, known triggers of ET-1 expression. Moreover, no increase in O2- or ET-1 levels was found in silenced cells treated with AngII or NOX-agonist phorbol ester (PMA 5 nM/48 h). Finally, vWF was required for overexpression of NOX4 and NOX2 in response to AngII and PMA. In conclusion, endothelial vWF knockdown prevented Ang II-induced ET-1 upregulation through attenuation of NOX-mediated O2- production. Our findings reveal a new role of vWF in preventing of Ang II-induced endothelial dysfunction

Late plasma exosome microRNA-21-5p depicts magnitude of reverse ventricular remodeling after early surgical repair of primary mitral valve regurgitation

Introduction: Primary mitral valve regurgitation (MR) results from degeneration of mitral valve apparatus. Mechanisms leading to incomplete postoperative left ventricular (LV) reverse remodeling (Rev–Rem) despite timely and successful surgical mitral valve repair (MVR) remain unknown. Plasma exosomes (pEXOs) are smallest nanovesicles exerting early postoperative cardioprotection. We hypothesized that late plasma exosomal microRNAs (miRs) contribute to Rev–Rem during the late postoperative period. Methods: Primary MR patients (n = 19; age, 45–71 years) underwent cardiac magnetic resonance imaging and blood sampling before (T0) and 6 months after (T1) MVR. The postoperative LV Rev–Rem was assessed in terms of a decrease in LV end-diastolic volume and patients were stratified into high (HiR-REM) and low (LoR-REM) LV Rev–Rem subgroups. Isolated pEXOs were quantified by nanoparticle tracking analysis. Exosomal microRNA (miR)-1, –21–5p, –133a, and –208a levels were measured by RT-qPCR. Anti-hypertrophic effects of pEXOs were tested in HL-1 cardiomyocytes cultured with angiotensin II (AngII, 1 μM for 48 h). Results: Surgery zeroed out volume regurgitation in all patients. Although preoperative pEXOs were similar in both groups, pEXO levels increased after MVR in HiR-REM patients (+0.75-fold, p = 0.016), who showed lower cardiac mass index (–11%, p = 0.032). Postoperative exosomal miR-21-5p values of HiR-REM patients were higher than other groups (p < 0.05). In vitro, T1-pEXOs isolated from LoR-REM patients boosted the AngII-induced cardiomyocyte hypertrophy, but not postoperative exosomes of HiR-REM. This adaptive effect was counteracted by miR-21-5p inhibition. Summary/Conclusion: High levels of miR-21-5p-enriched pEXOs during the late postoperative period depict higher LV Rev–Rem after MVR. miR-21-5p-enriched pEXOs may be helpful to predict and to treat incomplete LV Rev–Rem after successful early surgical MVR

EPIGENETIC MODULATION OF MYOCARDIAL ANGIOGENIC BALANCE: AN EMERGING THERAPEUTIC PERSPECTIVE FOR ADULT FAILING HEART

The term "Epigenetics" refers to chromatin-based pathways involved in the regulation of gene expression without altering DNA sequence. Suppression of angiogenesis may be related to cardiac dysfunction following alterations in capillary microvasculature. An understanding of the relationship between angiogenesis and cardiac remodeling remains the major limitation in order to address the weak self-renewal ability of the adult failing heart. In physiological conditions, genes responsible for sustaining the angiogenic ability of the mature endothelial cells can be categorized into pro- and anti-angiogenic genes. The balanced expression of angiogenic genes maintains a state of equilibrium in capillary density of the adult myocardium. In the context of post-ischemic myocardial dysfunction, transcriptional and posttranscriptional modifications of the gene pool are involved in alterations of angiogenic balance in capillary microvasculature, angiogenesis, myocardial perfusion/contractility match and contractile function. The regulation of angiogenic gene expression by different epigenetic modifications may induce the formation of new vessels from coronary mature endothelial cells rather than endothelial progenitor cells. The role of proangiogenic epigenetic activators is emerging as a new actor in the modulation of the angiogenic balance of failing heart. In fact, the development of an ideal method to promote myocardial revascularization, while attenuating cardiac remodeling, is still a challenging issue. The present review will examine emerging studies on the epigenetic mechanisms involved in the regulation of function and angiogenic ability of mature endothelial cells residing in the adult myocardium

Long-term Intake of Pasta Containing Barley (1-3)Beta-D-Glucan Increases Neovascularization-mediated Cardioprotection through Endothelial Upregulation of Vascular Endothelial Growth Factor and Parkin.

Barley (1-3)β-D-Glucan (BBG) enhances angiogenesis. Since pasta is very effective in providing a BBG-enriched diet, we hypothesized that the intake of pasta containing 3% BBG (P-BBG) induces neovascularization-mediated cardioprotection. Healthy adult male C57BL/6 mice fed P-BBG (n = 15) or wheat pasta (Control, n = 15) for five-weeks showed normal glucose tolerance and cardiac function. With a food intake similar to the Control, P-BBG mice showed a 109% survival rate (P < 0.01 vs. Control) after cardiac ischemia (30 min)/reperfusion (60 min) injury. Left ventricular (LV) anion superoxide production and infarct size in P-BBG mice were reduced by 62 and 35% (P < 0.0001 vs. Control), respectively. The capillary and arteriolar density of P-BBG hearts were respectively increased by 12 and 18% (P < 0.05 vs. Control). Compared to the Control group, the VEGF expression in P-BBG hearts was increased by 87.7% (P < 0.05); while, the p53 and Parkin expression was significantly increased by 125% and cleaved caspase-3 levels were reduced by 33% in P-BBG mice. In vitro, BBG was required to induce VEGF, p53 and Parkin expression in human umbelical vascular endothelial cells. Moreover, the BBG-induced Parkin expression was not affected by pifithrin-α (10 uM/7days), a p53 inhibitor. In conclusion, long-term dietary supplementation with P-BBG confers post-ischemic cardioprotection through endothelial upregulation of VEGF and Parkin

Plasma Exosomes in Cardiac Surgery: Opportunityfor Precision Perioperative Cardioprotection?

Perioperative cardioprotection aims to timely minimize the consequences of myocardial ischemia-reperfusion injury. Despite elective cardiac surgery is safer today than in the past, improvement of perioperative outcome in older patients with co-morbidities and co-medications remains a debated issue.1 Exosomes, smallest extracellular nanovesicles (30-150 nm in diameter), are increasingly being exploited for their unique cardioprotective role mediated by cargo components (i.e.: microRNAs, proteins, metabolites) and surface pro-survival proteins (i.e. heat shock protein 70, pregnancy-associated plasma protein- A). Previous studies have demonstrated that plasma exosomes during cardiac surgery play anti-inflammatory role by specific microRNAs. Moreover, we found that plasma exosomes change protein cargo in older patients undergoing different types of on-pump cardiac surgery.2 Our data ara in accord with recent findings demonstrating change of exosomal protein cargo by external factors (i.e. drugs, hypoxia, free radicals).3 In particular, 22 and 12 anti-apoptotic exosomal proteins were detected early after aortic unclamping during heart valve surgery and coronary artery bypass graft (CABG), respectively. Interestingly, in CABG patients anti-apoptotic plasma exosomes further increased at 72 h after aortic unclamping (P &lt; 0.01) whereas troponin levels decreased.2 In conclusion, plasma exosomes represent tantalizing theranostic tool for prompt clinical applications in critical scenario for inducing and monitoring effective perioperative cardioprotection early after cardiac surgery

Long-term dietary intake of pasta enriched with barley (1–3) beta-d-glucan induces neovascularization-mediated cardioprotection against ischemia/reperfusion injury in mice

Background: The modulation of angiogenesis is known to be a strategy to increase heart resistance to ischemia/reperfusion (I/R). Whereas the angiogenic activity of barley (1- 3)beta-D-Glucan (BG) have been characterized in vitro, there are no clear experimental data demonstrating that chronic dietary intake and intestinal absorption of BG actually protects the heart against I/R. We tested whether long-term consumption of 3% w/v BG included in normal pasta would increase myocardial angiogenesis and render the heart of mice more resistant to I/R. Materials and methods: Healthy adult male C57BL/6 mice were fed for 5 weeks with a low-fat diet supplemented with pasta enriched with BG (3g/100mg) (BG, n=15) or regular pasta (control, n=15). Food intake, glucose tolerance test and cardiac function were weekly assessed. At fifth week of diet, anesthetized mice underwent to 30 min of cardiac ischemia followed by 60 min of reperfusion. The myocardial infarction was induced by ligation of the left anterior descending artery. The myocardial capillary density and infarct size/area at risk were assessed ex vivo. Myocardial Vascular Endothelial Growth Factor 165 (VEGF165) expression and anion superoxide (O2-) levels were measured respectively by western blot and dihydroethidium staining. To evaluate the paracrine activation of protective angiogenesis by BG, we measured the expression of VEGF165, a well known pro-angiogenic growth factor, in endothelial cells exposed for 7 days to 3% BG. Finally, we examined the expression of dectin-1, a receptor of BG, in both endothelial cells and cardiomyocytes. Results: At similar food intake, 3% w/v BG did not affect body weight, glucose metabolism and cardiac function of mice. At the end of reperfusion, BG mice survived 50±2% more than control (P<0.01). Infarct size/area at risk and myocardial O2- load were reduced respectively by 62±5% (P<0.001) and 35±4% (P<0.0001) in β-D-glucan mice. Myocardial capillary density and VEGF165 expression were increased respectively by 12±0.6% (P<0.05) and 47.6±1% (P<0.001) in BG group compared to control mice. In vitro, VEGF165 expression was significantly increased in BG-treated endothelial cells (P<0.001). Dectin-1 was expressed in endothelial cells, but not in cardiomyocytes. Conclusions: Long-term intake of BG-rich diet augments myocardial capillary density and reduces infarct size by sustained expression of VEGF165 in endothelial cells, which express dectin-1. Our findings suggest important potential health benefits of pasta-rich in BG and emphasize the need to develop BG-rich functional foods with protective activities for increasing heart resistance to I/R injury

Firing-Induced Microstructural Properties of Quasi-Diamagnetic Carbonate-Based Porous Ceramics : a 1H NMR Relaxation Correlation Study

This study deals with the application of two-dimensional proton nuclear magnetic resonance relaxometry (2D 1H NMR-R) to the characterization of porous ceramics nearly free of magnetic compounds. Different microstructural properties were obtained by firing a diamagnetic mixture of kaolin, calcium, and magnesium carbonate over a wide range of maximum temperatures (600–1100 °C) and firing times at the maximum temperature (soaking times) (0–10 h). The 2D 1H NMR-R method relies on the correlated measurement of 1H longitudinal (T1) and transverse (T2) relaxation times of pore-filling water by which the properties of the interconnected pore space may be investigated. In the absence of significant magnetic susceptibility effect due to para- and ferro-magnetic compounds, the 2D 1H NMR-R maps allow studying the conjoint effects on pore size distribution and inter-pore coupling due to the variations in both time and temperature of firing. The NMR experiments were performed with a low-field 1H NMR sensor, which allows non-destructive and in situ analysis. For ceramic specimens fired at 600 and 700 °C, the fraction of smallest pores increases with firing time at the expenses of those with intermediate size. The pore shrinkage occurring at this stage, and likely associated with the transformation of kaolinite in metakaolinite, is affected in a similar way by soaking time and firing temperature, in line with the concept of equivalent firing temperature. At temperatures from 800 to 1100 °C, the structural modifications involving interconnectivity and average pore size are driven primarily by firing temperature and, secondarily, by soaking time. The 2D 1H NMR-R results are confirmed by more traditional, but destructive, mineralogical, and structural analyses like X-ray powder diffraction, helium pycnometry, mercury intrusion porosimetry, and nitrogen adsorption/desorption method

Effects of time and temperature of firing on Fe-rich ceramics studied by Mössbauer spectroscopy and two-dimensional 1H-nuclear magnetic resonance relaxometry

The combined effects of firing temperature and soaking time on the microstructure of iron-rich porous ceramics have been studied by 57Fe-Mössbauer spectroscopy and 2D 1H nuclear magnetic resonance (NMR) relaxometry using a single-sided probe. Examining water-saturated ceramics using the relaxation correlation method, where longitudinal (T 1) and transverse (T 2) relaxation times are measured concurrently, provides information about firing-induced changes in both porosity (related to T 1) and magnetic properties (related to T 2). Comparing the information obtained from 1H-NMR analyses with that obtained from Mössbauer spectroscopy (which characterizes changes in iron-bearing species) shows that the T 1-T 2 NMR correlation technique is very sensitive to even subtle modifications in the magnetic behavior of Fe-bearing species. Moreover, the single-sided NMR approach allows us to perform millimeter-scale depth-resolved measurements, which can be used to non-invasively study the microstructural heterogeneities associated with non-uniform firing effects inside ceramics. This is in contrast to Mössbauer spectroscopy, which requires that the ceramic samples be ground. © 2012 American Institute of Physics