Oleg F. Sharifov

Intramural Virtual Electrodes During Defibrillation Shocks in Left Ventricular Wall Assessed by Optical Mapping of Membrane Potential

Background—It is believed that defibrillation is due to shock-induced changes of transmembrane potential (&Dgr;Vm) in the bulk of ventricular myocardium (so-called virtual electrodes), but experimental proof of this hypothesis is absent. Here, intramural shock-induced &Dgr;Vm were measured for the first time in isolated preparations of left ventricle (LV) by an optical mapping technique. Methods and Results—LV preparations were excised from porcine hearts (n=9) and perfused through a coronary artery. Rectangular shocks (duration 10 ms, field strength E ≈2 to 50 V/cm) were applied across the wall during the action potential plateau by 2 large electrodes. Shock-induced &Dgr;Vm were measured on the transmural wall surface with a 16×16 photodiode array (resolution 1.2 mm/diode). Whereas weak shocks (E≈2 V/cm) induced negligible &Dgr;Vm in the wall middle, stronger shocks produced intramural &Dgr;Vm of 2 types. (1) Shocks with E>4 V/cm produced both positive and negative intramural &Dgr;Vm that changed their sign on changing shock polarity, possibly reflecting large-scale nonuniformities in the tissue structure; the &Dgr;Vm patterns were asymmetrical, with &Dgr;V−m>&Dgr;V+m. (2) Shocks with E>34 V/cm produced predominantly negative &Dgr;Vm across the whole transmural surface, independent of the shock polarity. These relatively uniform polarizations could be a result of microscopic discontinuities in tissue structure. Conclusions—Strong defibrillation shocks induce &Dgr;Vm in the intramural layers of LV. During action potential plateau, intramural &Dgr;Vm are typically asymmetrical (&Dgr;V−m>&Dgr;V+m) and become globally negative during very strong shocks.

Optical Mapping of Impulse Propagation in Engineered Cardiac Tissue

Cardiac tissue engineering has a potential to provide functional, synchronously contractile tissue constructs for heart repair, and for studies of development and disease using in vivo-like yet controllable in vitro settings. In both cases, the utilization of bioreactors capable of providing biomimetic culture environments is instrumental for supporting cell differentiation and functional assembly. In the present study, neonatal rat heart cells were cultured on highly porous collagen scaffolds in bioreactors with electrical field stimulation. A hallmark of excitable tissues such as myocardium is the ability to propagate electrical impulses. We utilized the method of optical mapping to measure the electrical impulse propagation. The average conduction velocity recorded for the stimulated constructs (14.4 +/- 4.1 cm/s) was significantly higher than that of the nonstimulated constructs (8.6 +/- 2.3 cm/s, p = 0.003). The measured electrical propagation properties correlated to the contractile behavior and the compositions of tissue constructs. Electrical stimulation during culture significantly improved amplitude of contractions, tissue morphology, and connexin-43 expression compared to the nonsimulated controls. These data provide evidence that electrical stimulation during bioreactor cultivation can improve electrical signal propagation in engineered cardiac constructs.

Optical mapping of transmural activation induced by electrical Shocks in isolated left ventricular wall wedge preparations

Introduction: It is believed that electrical shocks interrupt fibrillation by directly stimulating the bulk of ventricular myocardium in excitable states, but how shocks activate intramural tissue layers is not known. In this study, Vm responses and transmural activation patterns induced by shocks during diastole were measured in isolated coronary perfused preparations of porcine left ventricle.

Diagnostic Accuracy of Tissue Doppler Index E/è for Evaluating Left Ventricular Filling Pressure and Diastolic Dysfunction/Heart Failure With Preserved Ejection Fraction: A Systematic Review and Meta‐Analysis

Background Tissue Doppler index E/è is used clinically and in multidisciplinary research for estimation of left ventricular filling pressure (LVFP) and diastolic dysfunction (DD)/heart failure with preserved ejection fraction (HFpEF). Its diagnostic accuracy is not well studied. Methods and Results From the PubMed, Scopus, Embase, and Cochrane databases, we identified 24 studies reporting E/è and invasive LVFP in preserved EF (≥50%). In random‐effects models, E/è had poor to mediocre linear correlation with LVFP. Summary sensitivity and specificity (with 95% CIs) for the American Society of Echocardiography–recommended E/è cutoffs (lateral, mean, and septal, respectively) to identify elevated LVFP was estimated by using hierarchical summary receiver operating characteristic analysis. Summary sensitivity was 30% (9–48%), 37% (13–61%), and 24% (6–46%), and summary specificity was 92% (82–100%), 91% (80–99%), and 98% (92–100%). Positive likelihood ratio (LR+) was <5 for lateral and mean E/è. LR+ was slightly >10 for septal E/è obtained from 4 studies (cumulative sample size <220). For excluding elevated LVFP, summary sensitivity for E/è (lateral, mean, and septal, respectively) was 64% (38–86%), 36% (3–74%), and 50% (14–81%), while summary specificity was 73% (54–89%), 83% (49–100%), and 89% (66–100%). Because of data set limitations, meaningful inference for identifying HFpEF by using E/è could not be drawn. With the use of quality assessment tool for diagnostic accuracy studies (Quality Assessment of Diagnostic Accuracy Studies questionnaire), we found substantial risks of bias and/or applicability. Conclusions There is insufficient evidence to support that E/è can reliably estimate LVFP in preserved EF. The diagnostic accuracy of E/è to identify/exclude elevated LVFP and DD/HFpEF is limited and requires further validation in a well‐designed prospective clinical trial.

Apolipoprotein E Mimetics and Cholesterol-Lowering Properties

Apolipoprotein E (apoE) is a ligand for clearance of lipoprotein remnants such as chylomicrons and very low-density lipoproteins. It has anti-atherogenic and anti-inflammatory properties. Therefore, there is extensive ongoing research to create peptides that can mimic properties of apoE. A number of synthetic peptides that encompass different regions of apoE have been studied for inhibiting inflammatory states, including Alzheimer disease. However, peptides that clear atherogenic lipoproteins, analogous to apoE, via enhanced hepatic uptake have not been previously reviewed. Toward this end, we describe the design and studies of a dual-domain apoE mimetic peptide, Ac-hE18A-NH2. This peptide consists of residues 141–150, the putative receptor-binding region of human apoE, covalently linked to a well characterized class A amphipathic helix, 18A, which has no sequence homology to any other exchangeable apolipoprotein sequences. It demonstrates dramatic effects in reducing plasma cholesterol levels in dyslipidemic mouse and rabbit models. We discuss the scientific rationale and review the literature for the design and efficacy of the peptide. Analogous to apoE, this peptide bypasses the low-density lipoprotein receptor for the hepatic uptake of atherogenic lipoproteins via heparan sulfate proteoglycan (HSPG). ApoE mimetics such as Ac-hE18A-NH2 may therefore restore or replace ligands in genetically induced hyperlipidemias to enable reduction in atherogenic lipoproteins via HSPG even in the absence of functional low-density lipoprotein receptors. Therefore, this and similar peptides may be useful in the treatment of dyslipidemic disorders such as familial hyperlipidemia and atherosclerosis.

Anti-Inflammatory Mechanisms of Apolipoprotein A-I Mimetic Peptide in Acute Respiratory Distress Syndrome Secondary to Sepsis

Acute respiratory distress syndrome (ARDS) due to sepsis has a high mortality rate with limited treatment options. High density lipoprotein (HDL) exerts innate protective effects in systemic inflammation. However, its role in ARDS has not been well studied. Peptides such as L-4F mimic the secondary structural features and functions of apolipoprotein (apo)A-I, the major protein component of HDL. We set out to measure changes in HDL in sepsis-mediated ARDS patients, and to study the potential of L-4F to prevent sepsis-mediated ARDS in a rodent model of lipopolysaccharide (LPS)-mediated acute lung injury, and a combination of primary human leukocytes and human ARDS serum. We also analyzed serum from non-lung disease intubated patients (controls) and sepsis-mediated ARDS patients. Compared to controls, ARDS demonstrates increased serum endotoxin and IL-6 levels, and decreased HDL, apoA-I and activity of anti-oxidant HDL-associated paraoxanase-1. L-4F inhibits the activation of isolated human leukocytes and neutrophils by ARDS serum and LPS in vitro. Further, L-4F decreased endotoxin activity and preserved anti-oxidant properties of HDL both in vitro and in vivo. In a rat model of severe endotoxemia, L-4F significantly decreased mortality and reduces lung and liver injury, even when administered 1 hour post LPS. Our study suggests the protective role of the apoA-I mimetic peptide L-4F in ARDS and gram-negative endotoxemia and warrant further clinical evaluation. The main protective mechanisms of L-4F are due to direct inhibition of endotoxin activity and preservation of HDL anti-oxidant activity.

Intramural Virtual Electrodes in Ventricular Wall Effects on Epicardial Polarizations

Background—Intramural virtual electrodes (IVEs) are believed to play an important role in defibrillation, but their existence in intact myocardium remains unproven. Here, IVEs were detected by use of optical recordings of shock-induced transmembrane potential (Vm) changes (ΔVm) measured from the intact epicardial heart surface. Methods and Results—To detect IVEs, isolated porcine left ventricles were sequentially stained with a Vm-sensitive dye by 2 methods: (1) surface staining (SS) and (2) global staining (GS) via coronary perfusion. Shocks (2 to 50 V/cm) were applied across the ventricular wall in an epicardial-to-endocardial direction during the action potential plateau via transparent mesh electrodes, and shock-induced ΔVm were measured optically from the same epicardial locations after SS and GS. Optical recordings revealed significant differences between ΔVm of 2 types that became more prominent with increasing shock strength: (1) for weak shocks, SS-ΔVm were larger and faster than GS-ΔVm; (2) for intermediate shocks, cathodal GS-ΔVm became multiphasic, whereas SS-ΔVm remained monophasic; and (3) for strong shocks, cathodal GS-ΔVm became uniformly negative, whereas SS-ΔVm typically remained positive. The radical differences in the shape and polarity of SS and GS polarizations can be explained by the contribution of subepicardial IVEs to optical signals. Histological examination revealed a dense network of collagen septa in the subepicardium, which could form the IVE substrate. Conclusions—Intramural virtual electrodes are reflected in optical measurements of shock-induced ΔVm on the intact epicardial surface. These IVEs could be a result of microscopic resistive discontinuities formed by collagen septa.

Pharmacological evidence for Orai channel activation as a source of cardiac abnormal automaticity.

Calcium transport through plasma membrane voltage-independent calcium channels is vital for signaling events in non-excitable and excitable cells. Following up on our earlier work, we tested the hypothesis that this type of calcium transport can disrupt myocardial electromechanical stability. Our Western and immunofluorescence analyses show that left atrial and ventricular myocytes express the Orai1 and the Orai3 calcium channels. Adding the Orai activator 2-aminoethoxydiphenyl borate (2-APB) to the superfusate of rat left atria causes these non-automatic muscles to contract spontaneously and persistently at rates of up to 10 Hz, and to produce normal action potentials from normal resting potentials, all in the absence of external stimulation. 2-APB likewise induces such automatic activity in superfused rat left ventricular papillary muscles, and the EC(50)s at which 2-APB induces this activity in both muscles are similar to the concentrations which activate Orais. Importantly, the voltage-independent calcium channel inhibitor 1-[2-(4-methoxyphenyl)-2-[3-(4-methoxyphenyl) propoxy]ethyl-1H-imidazole (SKF-96365) suppresses this automaticity with an IC(50) of 11 ± 0.6 μM in left atria and 6 ± 1.6 μM in papillary muscles. 1-(5-Iodonaphthalene-1-sulfonyl)-hexahydro-1,4-diazepine (ML-7), a second voltage-independent calcium channel inhibitor, and two calmodulin inhibitors also prevent 2-APB automaticity while two calmodulin-dependent protein kinase II inhibitors do not. Thus an activator of the Orai calcium channels provokes a novel type of high frequency automaticity in non-automatic heart muscle.

Evidence that 2-aminoethoxydiphenyl borate provokes fibrillation in perfused rat hearts via voltage-independent calcium channels.

We tested whether 2-aminoethoxydiphenyl borate (2-APB) induces arrhythmia in perfused rat hearts and whether this arrhythmia might result from the activation of voltage-independent calcium channels. Rat hearts were Langendorff perfused and beat under sinus rhythm. An isovolumic balloon inserted into the left ventricle was used to record mechanical function while bipolar electrograms were recorded from electrodes sutured to the base and the apex of hearts. Western and immunofluorescence analyses were performed on rat left ventricular protein extracts and left ventricular frozen sections, respectively. Rat ventricular myocytes express Orai 1 and Orai 3, and ventricle also contains the Orai regulator Stim1. Rat hearts (n=5) perfused with Krebs-Henseleit (KH) alone maintained sinus rhythm at 4.8 ± 0.1 Hz and stable mechanical function. By contrast, perfusing hearts (n=5) with (KH+22 μM 2-APB) provoked a period of tachycardic ectopy at rates of up to 10.8 ± 0.2 Hz. As perfusion with (KH+22 μM 2-APB) continued, the rate of spontaneous ventricular depolarization increased to 21.8 ± 1.2 Hz and became disorganized. Heart mechanical function collapsed as developed pressure decreased from 87 ± 8.8 to 3.5 ± 1.9 mm Hg. Flow rate did not change between normal (16.6 ± 0.9 ml/min) and fibrillating (17.4 ± 0.8 ml/min) hearts. The addition of 20 μM 1-[2-(4-methoxyphenyl)-2-[3-(4-methoxyphenyl) propoxy]ethyl-1H-imidazole (SKF-96365) to (KH+22 μM 2-APB) perfusates (n=4) restored sinus rhythm and heart mechanical output. These data indicate that activating myocardial voltage-independent calcium channels, possibly the Orais, may be a novel cause of ventricular arrhythmia.

What Is the Evidence That the Tissue Doppler Index E/e′ Reflects Left Ventricular Filling Pressure Changes After Exercise or Pharmacological Intervention for Evaluating Diastolic Function? A Systematic Review

Background Noninvasive echocardiographic tissue Doppler assessment (E/e′) in response to exercise or pharmacological intervention has been proposed as a useful parameter to assess left ventricular (LV) filling pressure (LVFP) and LV diastolic dysfunction. However, the evidence for it is not well summarized. Methods and Results Clinical studies that evaluated invasive LVFP changes in response to exercise/other interventions and echocardiographic E/e′ were identified from PubMed, Scopus, Embase, and Cochrane Library databases. We grouped and evaluated studies that included patients with preserved LV ejection fraction (LVEF), patients with mixed/reduced LVEF, and patients with specific cardiac conditions. Overall, we found 28 studies with 9 studies for preserved LVEF, which was our primary interest. Studies had differing methodologies with limited data sets, which precluded quantitative meta‐analysis. We therefore descriptively summarized our findings. Only 2 small studies (N=12 and 10) directly or indirectly support use of E/e′ for assessing LVFP changes in preserved LVEF. In 7 other studies (cumulative N=429) of preserved LVEF, E/e′ was not useful for assessing LVFP changes. For mixed/reduced LVEF groups or specific cardiac conditions, results similar to preserved LVEF were found. Conclusions We find that there is insufficient evidence that E/e′ can reliably assess LVFP changes in response to exercise or other interventions. We suggest that well‐designed prospective studies should be conducted for further evaluation.