The VASP-profilin1 (Pfn1) interaction is critical for efficient cell migration and is regulated by cell-substrate adhesion in a PKA-dependent manner.

Dynamic regulation of the actin cytoskeleton is an essential feature of cell motility. Action of Enabled (Ena)/vasodilator-stimulated phosphoprotein (VASP), a family of conserved actin-elongating proteins, is an important aspect of regulation of the actin cytoskeletal architecture at the leading edge that controls membrane protrusion and cell motility. In this study, we performed mutagenesis experiments in overexpression and knockdown-rescue settings to provide, for the first time, direct evidence of the role of the actin-binding protein profilin1 (Pfn1) in VASP-mediated regulation of cell motility. We found that VASP's interaction with Pfn1 is promoted by cell-substrate adhesion and requires down-regulation of PKA activity. Our experimental data further suggest that PKA-mediated Ser137 phosphorylation of Pfn1 potentially negatively regulates the Pfn1-VASP interaction. Finally, Pfn1's ability to be phosphorylated on Ser137 was partly responsible for the anti-migratory action elicited by exposing cells to a cAMP/PKA agonist. On the basis of these findings, we propose a mechanism of adhesion-protrusion coupling in cell motility that involves dynamic regulation of Pfn1 by PKA activity.

Pharmacological intervention of MKL/SRF signaling by CCG-1423 impedes endothelial cell migration and angiogenesis.

De novo synthesis of cytoskeleton-regulatory proteins triggered by the megakaryoblastic leukemia (MKL)/serum response factor (SRF) transcriptional system in response to pro-angiogenic growth factors lies at the heart of endothelial cell (EC) migration (a critical element of angiogenesis) and neovascularization. This study explores whether pharmacological intervention of MKL/SRF signaling axis by CCG-1423 is able to suppress angiogenesis. Our studies show that CCG-1423 inhibits migration and cord morphogenesis of EC in vitro and sprouting angiogenesis ex vivo and in vivo, suggesting CCG-1423 could be a novel anti-angiogenic agent. Kymography analyses of membrane dynamics of EC revealed that CCG-1423 treatment causes a major defect in membrane protrusion. CCG-1423 treatment led to attenuated expression of several actin-binding proteins that are important for driving membrane protrusion including ArpC2, VASP, and profilin1 (Pfn1) with the most drastic effect seen on the expression of Pfn1. Finally, depletion of Pfn1 alone is also sufficient for a dramatic decrease in sprouting angiogenesis of EC in vitro and ex vivo, further suggesting that Pfn1 depletion may be one of the mechanisms of the anti-angiogenic action of CCG-1423.

Histone Deacetylase 3 (HDAC3)-dependent Reversible Lysine Acetylation of Cardiac Myosin Heavy Chain Isoforms Modulates Their Enzymatic and Motor Activity.

Reversible lysine acetylation is a widespread post-translational modification controlling the activity of proteins in different subcellular compartments. We previously demonstrated that a class II histone deacetylase (HDAC), HDAC4, and a histone acetyltransferase, p300/CREB-binding protein-associated factor, associate with cardiac sarcomeres and that a class I and II HDAC inhibitor, trichostatin A, enhances contractile activity of myofilaments. In this study we show that a class I HDAC, HDAC3, is also present at cardiac sarcomeres. By immunohistochemical and electron microscopic analyses, we found that HDAC3 was localized to A-band of sarcomeres and capable of deacetylating myosin heavy chain (MHC) isoforms. The motor domains of both cardiac α- and ß-MHC isoforms were found to be reversibly acetylated. Biomechanical studies revealed that lysine acetylation significantly decreased the Km for the actin-activated ATPase activity of MHC isoforms. By in vitro motility assay, we found that lysine acetylation increased the actin-sliding velocity of α-myosin by 20% and ß-myosin by 36% compared with their respective non-acetylated isoforms. Moreover, myosin acetylation was found to be sensitive to cardiac stress. During induction of hypertrophy, myosin isoform acetylation increased progressively with duration of stress stimuli independently of isoform shift, suggesting that lysine acetylation of myosin could be an early response of myofilaments to increase contractile performance of the heart. These studies provide the first evidence for localization of HDAC3 at myofilaments and uncover a novel mechanism modulating the motor activity of cardiac MHC isoforms.

Differential Effects of Left Ventricular Pacing Sites on Regional Contraction Patterns and Global Performance.

OBJECTIVE: To define the differential effect of site-specific ventricular counterpacing efficacy during cardiac resynchronization therapy (CRT) to identify the most informative imaging views to quantify it. Cross-sectional and long-axis views commonly are used to assess left ventricular (LV) contractility. DESIGN: The effects of LV apical (LVa) and free-wall (LVfw) pacing during CRT on long- and short-axis contraction, cardiac output, and stroke work were assessed in an open-chested acute canine model to determine whether LVa and LVfw would induce earlier apical than basilar LV radial contraction and earlier free-wall than septal contraction, respectively. Apical (CRTa) and free-wall (CRTfw) using right ventricular (RV) pacing-induced dyssynchrony also were examined. SETTING: University large animal research laboratory. PARTICIPANTS: Ten acutely anesthetized and instrumented open-chested purpose-bred dogs. INTERVENTIONS: RV pacing served as the model of cardiac dyssynchrony. Selective LVfw and LVa pacing alone or with RV (CRTfw and CRTa, respectively) were studied relative to right atrial pacing (RA) as the control. MEASUREMENTS AND MAIN RESULTS: Two pairs of 3 ultrasonic crystals were place along the LV longitudinal axis-apex and mid-to-base pairs along septal and free wall lines. Conductance catheter-defined longitudinal LV segmental volumes and pressure-volume data were collected. RV decreased cardiac output and stroke work compared with RA (2.0±0.3 v 1.4±0.1 L/min; 137±22 v 60±14 mJ; p<0.05, respectively). LVfw but not LVa decreased stroke work (130±35 mJ), and CRTa but not CRTfw improved both (2.1±0.2 L/min; 113±13 mJ; p<0.01 v RV pacing). No difference in time to minimal length free wall-to-septal crystal was seen with pacing. Both LVa and CRTa displayed increased apical-to-basilar shortening delay compared with RA, RV, and LVfw (42±47, 9±105, and 1±46 msec, respectively; p<0.05). No matching regional LV volume changes were seen during LVa. CONCLUSIONS: LV functional analysis from only a cross-sectional plane may be insufficient to characterize improved LV contraction synchrony during multisite CRT.

Relaxin suppresses atrial fibrillation by reversing fibrosis and myocyte hypertrophy and increasing conduction velocity and sodium current in spontaneously hypertensive rat hearts.

RATIONALE: Atrial fibrillation (AF) contributes significantly to morbidity and mortality in elderly and hypertensive patients and has been correlated to enhanced atrial fibrosis. Despite a lack of direct evidence that fibrosis causes AF, reversal of fibrosis is considered a plausible therapy. OBJECTIVE: To evaluate the efficacy of the antifibrotic hormone relaxin (RLX) in suppressing AF in spontaneously hypertensive rats (SHR). METHODS AND RESULTS: Normotensive Wistar-Kyoto (WKY) and SHR were treated for 2 weeks with vehicle (WKY+V and SHR+V) or RLX (0.4 mg/kg per day, SHR+RLX) using implantable mini-pumps. Hearts were perfused, mapped optically to analyze action potential durations, intracellular Ca²âº transients, and restitution kinetics, and tested for AF vulnerability. SHR hearts had slower conduction velocity (CV; P<0.01 versus WKY), steeper CV restitution kinetics, greater collagen deposition, higher levels of transcripts for transforming growth factor-ß, metalloproteinase-2, metalloproteinase-9, collagen I/III, and reduced connexin 43 phosphorylation (P<0.05 versus WKY). Programmed stimulation triggered sustained AF in SHR (n=5/5) and SHR+V (n=4/4), but not in WKY (n=0/5) and SHR+RLX (n=1/8; P<0.01). RLX treatment reversed the transcripts for fibrosis, flattened CV restitution kinetics, reduced action potential duration at 90% recovery to baseline, increased CV (P<0.01), and reversed atrial hypertrophy (P<0.05). Independent of antifibrotic actions, RLX (0.1 µmol/L) increased Na⁺ current density, INa (≈2-fold in 48 hours) in human cardiomyocytes derived from inducible pluripotent stem cells (n=18/18; P<0.01). CONCLUSIONS: RLX treatment suppressed AF in SHR hearts by increasing CV from a combination of reversal of fibrosis and hypertrophy and by increasing INa. The study provides compelling evidence that RLX may provide a novel therapy to manage AF in humans by reversing fibrosis and hypertrophy and by modulating cardiac ionic currents.

Myocardial extracellular volume fraction quantified by cardiovascular magnetic resonance is increased in diabetes and associated with mortality and incident heart failure admission.

AIMS: Diabetes may promote myocardial extracellular matrix (ECM) expansion that increases vulnerability. We hypothesized that: (i) type 2 diabetes would be associated with quantitative cardiovascular magnetic resonance (CMR) measures of myocardial ECM expansion, i.e. extracellular volume fraction (ECV); (ii) medications blocking the renin-angiotensin-aldosterone system (RAAS) would be associated with lower ECV; and (iii) ECV in diabetic individuals would be associated with mortality and/or incident hospitalization for heart failure. METHODS AND RESULTS: We enrolled 1176 consecutive patients referred for CMR without amyloidosis and computed ECV from measures of the haematocrit and myocardial and blood T1 pre- and post-contrast. Linear regression modelled ECV; Cox regression modelled mortality and/or hospitalization for heart failure. Diabetic individuals (n = 231) had higher median ECV than those without diabetes (n = 945): 30.2% (IQR: 26.9-32.7) vs. 28.1% (IQR: 25.9-31.0), respectively, P < 0.001). Diabetes remained associated with higher ECV in models adjusting for demographics, comorbidities, and medications (P < 0.001). Renin-angiotensin-aldosterone system blockade was associated with lower ECV (P = 0.028) in multivariable linear models. Over a median of 1.3 years (IQR: 0.8-1.9), 38 diabetic individuals had events (21 incident hospitalizations for heart failure; 24 deaths), and ECV was associated with these events (HR: 1.52, 95% CI: 1.21-1.89 per 3% ECV increase) in multivariable Cox regression models. CONCLUSION: Diabetes is associated with increased ECV. Extracellular volume fraction detects amelioration of ECM expansion associated with RAAS blockade, and is associated with mortality and/or incident hospitalization for heart failure in diabetic individuals. Extracellular matrix expansion may be an important intermediate phenotype in diabetic individuals that is detectable and treatable.

Nonlinear Viscoelastic Modeling of Finger Arteries: Toward Smartphone-Based Blood Pressure Monitoring via the Oscillometric Finger Pressing Method.

OBJECTIVE: Oscillometric finger pressing is a smartphone-based blood pressure (BP) monitoring method. Finger photoplethysmography (PPG) oscillations and pressure are measured during a steady increase in finger pressure, and an algorithm computes systolic BP (SP) and diastolic BP (DP) from the measurements. The objective was to assess the impact of finger artery viscoelasticity on the BP computation. METHODS: Nonlinear viscoelastic models relating transmural pressure (finger BP - applied pressure) to PPG oscillations during finger pressing were developed. The output of each model to a measured transmural pressure input was fitted to measured PPG oscillations from 15 participants. A parametric sensitivity analysis was performed via model simulations to elucidate the viscoelastic effect on the derivative-based BP computation algorithm. RESULTS: A Wiener viscoelastic model comprising a first-order transfer function followed by a static sigmoidal function fitted the measured PPG oscillations better than an elastic model containing only the static function (median (IQR) error of 30.5% (25.6%-34.0%) vs 50.9% (46.7%-53.7%); p<0.01). In Wiener model simulations, the derivative algorithm underestimated SP, especially with high pulse pressure and low transfer function cutoff frequency (i.e., greater viscoelasticity). The mean of the normalized PPG waveform at the maximum oscillation beat was found to correlate with the cutoff frequency (r = -0.8) and could thus possibly be used to compensate for viscoelasticity. CONCLUSION: Finger artery viscoelasticity negatively impacts oscillometric BP computation algorithms but can potentially be compensated for using available measurements. SIGNIFICANCE: These findings may help in converting smartphones into truly cuffless BP monitors for improving hypertension awareness and control.

A smartphone application toward detection of systolic hypertension in underserved populations.

High systolic blood pressure (BP) is the most important modifiable risk factor for cardiovascular disease. Managing systolic hypertension is especially difficult in underserved populations wherein access to cuff BP devices is limited. We showed that ubiquitous smartphones without force sensing can be converted into absolute pulse pressure (PP) monitors. The concept is for the user to perform guided thumb and hand maneuvers with the phone to induce cuff-like actuation and allow built-in sensors to make cuff-like measurements for computing PP. We developed an Android smartphone PP application. The 'app' could be learned by volunteers and yielded PP with total error < 8 mmHg against cuff PP (N = 24). We also analyzed a large population-level database comprising adults less than 65 years old to show that PP plus other basic information can detect systolic hypertension with ROC AUC of 0.9. The smartphone PP app could ultimately help reduce the burden of systolic hypertension in underserved populations and thus health disparities.

Grand Challenges at the Interface of Engineering and Medicine.

Over the past two decades Biomedical Engineering has emerged as a major discipline that bridges societal needs of human health care with the development of novel technologies. Every medical institution is now equipped at varying degrees of sophistication with the ability to monitor human health in both non-invasive and invasive modes. The multiple scales at which human physiology can be interrogated provide a profound perspective on health and disease. We are at the nexus of creating "avatars" (herein defined as an extension of "digital twins") of human patho/physiology to serve as paradigms for interrogation and potential intervention. Motivated by the emergence of these new capabilities, the IEEE Engineering in Medicine and Biology Society, the Departments of Biomedical Engineering at Johns Hopkins University and Bioengineering at University of California at San Diego sponsored an interdisciplinary workshop to define the grand challenges that face biomedical engineering and the mechanisms to address these challenges. The Workshop identified five grand challenges with cross-cutting themes and provided a roadmap for new technologies, identified new training needs, and defined the types of interdisciplinary teams needed for addressing these challenges. The themes presented in this paper include: 1) accumedicine through creation of avatars of cells, tissues, organs and whole human; 2) development of smart and responsive devices for human function augmentation; 3) exocortical technologies to understand brain function and treat neuropathologies; 4) the development of approaches to harness the human immune system for health and wellness; and 5) new strategies to engineer genomes and cells.

Association between extracellular matrix expansion quantified by cardiovascular magnetic resonance and short-term mortality.

BACKGROUND: Extracellular matrix expansion may be a fundamental feature of adverse myocardial remodeling, it appears to be treatable, and its measurement may improve risk stratification. Yet, the relationship between mortality and extracellular matrix is not clear because of difficulties with its measurement. To assess its relationship with outcomes, we used novel, validated cardiovascular magnetic resonance techniques to quantify the full spectrum of extracellular matrix expansion not readily detectable by conventional cardiovascular magnetic resonance. METHODS AND RESULTS: We recruited 793 consecutive patients at the time of cardiovascular magnetic resonance without amyloidosis or hypertrophic cardiomyopathy as well as 9 healthy volunteers (ages 20-50 years). We measured the extracellular volume fraction (ECV) to quantify the extracellular matrix expansion in myocardium without myocardial infarction. ECV uses gadolinium contrast as an extracellular space marker based on T1 measures of blood and myocardium pre- and post-gadolinium contrast and hematocrit measurement. In volunteers, ECV ranged from 21.7% to 26.2%, but in patients it ranged from 21.0% to 45.8%, indicating considerable burden. There were 39 deaths over a median follow-up of 0.8 years (interquartile range 0.5-1.2 years), and 43 individuals who experienced the composite end point of death/cardiac transplant/left ventricular assist device implantation. In Cox regression models, ECV related to all-cause mortality and the composite end point (hazard ratio, 1.55; 95% confidence interval, 1.27-1.88 and hazard ratio, 1.48; 95% confidence interval, 1.23-1.78, respectively, for every 3% increase in ECV), adjusting for age, left ventricular ejection fraction, and myocardial infarction size. CONCLUSIONS: ECV measures of extracellular matrix expansion may predict mortality as well as other composite end points (death/cardiac transplant/left ventricular assist device implantation).

The Microsoft Research Aurora Project: Important Findings on Cuffless Blood Pressure Measurement.

Conventional blood pressure (BP) measurement devices based on an inflatable cuff only provide a narrow view of the continuous BP profile. Cuffless BP measuring technologies could permit numerous BP readings throughout daily life and thereby considerably improve the assessment and management of hypertension. Several wearable cuffless BP devices based on pulse wave analysis (applied to a photoplethysmography or tonometry waveform) with or without use of pulse arrival time are now available on the market. The key question is: Can these devices provide accurate measurement of BP? Microsoft Research recently published a complex article describing perhaps the most important and highest resource project to date (Aurora Project) on assessing the accuracy of several pulse wave analysis and pulse wave analysis-pulse arrival time devices. The overall results from 1125 participants were clear-cut negative. The present article motivates and describes emerging cuffless BP devices and then summarizes the Aurora Project. The study methodology and findings are next discussed in the context of regulatory-cleared devices, physiology, and related studies, and the study strengths and limitations are pinpointed thereafter. Finally, the implications of the Aurora Project are briefly stated and recommendations for future work are offered to finally realize the considerable potential of cuffless BP measurement in health care.

Smartphone-Based Blood Pressure Monitoring via the Oscillometric Finger Pressing Method: Analysis of Oscillation Width Variations Can Improve Diastolic Pressure Computation.

OBJECTIVE: Oscillometric finger pressing is a potential method for absolute blood pressure (BP) monitoring via a smartphone. The user presses their fingertip against a photoplethysmography-force sensor unit on a smartphone to steadily increase the external pressure on the underlying artery. Meanwhile, the phone guides the finger pressing and computes systolic BP (SP) and diastolic BP (DP) from the measured blood volume oscillations and finger pressure. The objective was to develop and evaluate reliable finger oscillometric BP computation algorithms. METHODS: The collapsibility of thin finger arteries was exploited in an oscillometric model to develop simple algorithms for computing BP from the finger pressing measurements. These algorithms extract features from "width" oscillograms (oscillation width versus finger pressure functions) and the conventional "height" oscillogram for markers of DP and SP. Finger pressing measurements were obtained using a custom system along with reference arm cuff BP measurements from 22 subjects. Measurements were also obtained during BP interventions in some subjects for 34 total measurements. RESULTS: An algorithm employing the average of width and height oscillogram features predicted DP with correlation of 0.86 and precision error of 8.6 mmHg with respect to the reference measurements. Analysis of arm oscillometric cuff pressure waveforms from an existing patient database provided evidence that the width oscillogram features are better suited to finger oscillometry. CONCLUSION: Analysis of oscillation width variations during finger pressing can improve DP computation. SIGNIFICANCE: The study findings may help in converting widely available devices into truly cuffless BP monitors for improving hypertension awareness and control.

Subject-specific factors affecting particle residence time distribution of left atrial appendage in atrial fibrillation: A computational model-based study.

Background: Atrial fibrillation (AF) is a prevalent arrhythmia, that causes thrombus formation, ordinarily in the left atrial appendage (LAA). The conventional metric of stroke risk stratification, CHA2DS2-VASc score, does not account for LAA morphology or hemodynamics. We showed in our previous study that residence time distribution (RTD) of blood-borne particles in the LAA and its associated calculated variables (i.e., mean residence time, tm , and asymptotic concentration, C ∞) have the potential to improve CHA2DS2-VASc score. The purpose of this research was to investigate the effects of the following potential confounding factors on LAA tm and C ∞: (1) pulmonary vein flow waveform pulsatility, (2) non-Newtonian blood rheology and hematocrit level, and (3) length of the simulation. Methods: Subject-Specific data including left atrial (LA) and LAA cardiac computed tomography, cardiac output (CO), heart rate, and hematocrit level were gathered from 25 AF subjects. We calculated LAA tm and C ∞ based on series of computational fluid dynamics (CFD) analyses. Results: Both LAA tm and C ∞ are significantly affected by the CO, but not by temporal pattern of the inlet flow. Both LAA tm and C ∞ increase with increasing hematocrit level and both calculated indices are higher for non-Newtonian blood rheology for a given hematocrit level. Further, at least 20,000 s of CFD simulation is needed to calculate LAA tm and C ∞ values reliably. Conclusions: Subject-specific LA and LAA geometries, CO, and hematocrit level are essential to quantify the subject-specific proclivity of blood cell tarrying inside LAA in terms of the RTD function.

Bioengineered 3D Skeletal Muscle Model Reveals Complement 4b as a Cell-Autonomous Mechanism of Impaired Regeneration with Aging.

A mechanistic understanding of cell-autonomous skeletal muscle changes after injury can lead to novel interventions to improve functional recovery in an aged population. However, major knowledge gaps persist owing to limitations of traditional biological aging models. 2D cell culture represents an artificial environment, while aging mammalian models are contaminated by influences from non-muscle cells and other organs. Here, a 3D muscle aging system is created to overcome the limitations of these traditional platforms. It is shown that old muscle constructs (OMC) manifest a sarcopenic phenotype, as evidenced by hypotrophic myotubes, reduced contractile function, and decreased regenerative capacity compared to young muscle constructs. OMC also phenocopy the regenerative responses of aged muscle to two interventions, pharmacological and biological. Interrogation of muscle cell-specific mechanisms that contribute to impaired regeneration over time further reveals that an aging-induced increase of complement component 4b (C4b) delays muscle progenitor cell amplification and impairs functional recovery. However, administration of complement factor I, a C4b inactivator, improves muscle regeneration in vitro and in vivo, indicating that C4b inhibition may be a novel approach to enhance aged muscle repair. Collectively, the model herein exhibits capabilities to study cell-autonomous changes in skeletal muscle during aging, regeneration, and intervention.

Mathematical Modeling of Oscillometric Blood Pressure Measurement: A Complete, Reduced Oscillogram Model.

OBJECTIVE: Oscillogram modeling is a powerful tool for understanding and advancing popular oscillometric blood pressure (BP) measurement. A reduced oscillogram model relating cuff pressure oscillation amplitude ( ∆O) to external cuff pressure of the artery ( Pe) is: [Formula: see text], where g(P) is the arterial compliance versus transmural pressure ( P) curve, Ps and Pd are systolic and diastolic BP, and k is the reciprocal of the cuff compliance. The objective was to determine an optimal functional form for the arterial compliance curve. METHODS: Eight prospective, three-parameter functions of the brachial artery compliance curve were compared. The study data included oscillometric arm cuff pressure waveforms and invasive brachial BP from 122 patients covering a 20-120 mmHg pulse pressure range. The oscillogram measurements were constructed from the cuff pressure waveforms. Reduced oscillogram models, inputted with measured systolic and diastolic BP and each parametric brachial artery compliance curve function, were optimally fitted to the oscillogram measurements in the least squares sense. RESULTS: An exponential-linear function yielded as good or better model fits compared to the other functions, with errors of 7.9±0.3 and 5.1±0.2% for tail-trimmed and lower half-trimmed oscillogram measurements. Importantly, this function was also the most tractable mathematically. CONCLUSION: A three-parameter exponential-linear function is an optimal form for the arterial compliance curve in the reduced oscillogram model and may thus serve as the standard function for this model henceforth. SIGNIFICANCE: The complete, reduced oscillogram model determined herein can potentially improve oscillometric BP measurement accuracy while advancing foundational knowledge.

European Society of Hypertension recommendations for the validation of cuffless blood pressure measuring devices: European Society of Hypertension Working Group on Blood Pressure Monitoring and Cardiovascular Variability.

BACKGROUND: There is intense effort to develop cuffless blood pressure (BP) measuring devices, and several are already on the market claiming that they provide accurate measurements. These devices are heterogeneous in measurement principle, intended use, functions, and calibration, and have special accuracy issues requiring different validation than classic cuff BP monitors. To date, there are no generally accepted protocols for their validation to ensure adequate accuracy for clinical use. OBJECTIVE: This statement by the European Society of Hypertension (ESH) Working Group on BP Monitoring and Cardiovascular Variability recommends procedures for validating intermittent cuffless BP devices (providing measurements every >30 sec and usually 30-60 min, or upon user initiation), which are most common. VALIDATION PROCEDURES: Six validation tests are defined for evaluating different aspects of intermittent cuffless devices: static test (absolute BP accuracy); device position test (hydrostatic pressure effect robustness); treatment test (BP decrease accuracy); awake/asleep test (BP change accuracy); exercise test (BP increase accuracy); and recalibration test (cuff calibration stability over time). Not all these tests are required for a given device. The necessary tests depend on whether the device requires individual user calibration, measures automatically or manually, and takes measurements in more than one position. CONCLUSION: The validation of cuffless BP devices is complex and needs to be tailored according to their functions and calibration. These ESH recommendations present specific, clinically meaningful, and pragmatic validation procedures for different types of intermittent cuffless devices to ensure that only accurate devices will be used in the evaluation and management of hypertension.

HDAC3-dependent reversible lysine acetylation of cardiac myosin heavy chain isoforms modulates their enzymatic and motor activity.

Reversible lysine acetylation is a widespread post-translational modification controlling the activity of proteins in different subcellular compartments. We previously demonstrated that a class II histone deacetylase (HDAC), HDAC4, and a histone acetyltransferase, PCAF, associate with cardiac sarcomeres, and a class I and II HDAC inhibitor, trichostatin A, enhances contractile activity of myofilaments. In this study, we show that a class I HDAC, HDAC3, is also present at cardiac sarcomeres. By immunohistochemical and electron microscopic analyses, we found that HDAC3 was localized to the A band of sarcomeres and was capable of deacetylating myosin heavy chain (MHC) isoforms. The motor domains of both cardiac α- and ß-MHC isoforms were found to be reversibly acetylated. Biomechanical studies revealed that lysine acetylation significantly decreased the K(m) for the actin-activated ATPase activity of both α- and ß-MHC isoforms. By an in vitro motility assay, we found that lysine acetylation increased the actin sliding velocity of α-myosin by 20% and ß-myosin by 36%, compared to their respective non-acetylated isoforms. Moreover, myosin acetylation was found to be sensitive to cardiac stress. During induction of hypertrophy, myosin isoform acetylation increased progressively with duration of stress stimuli, independent of isoform shift, suggesting that lysine acetylation of myosin could be an early response of myofilaments to increase contractile performance of the heart. These studies provide the first evidence for localization of HDAC3 at myofilaments and uncover a novel mechanism modulating the motor activity of cardiac MHC isoforms.

Physiological relevance of quantifying segmental contraction synchrony.

BACKGROUND: Most current indices of synchrony quantify left ventricular (LV) contraction pattern in terms of a single, global (integrated) measure. We report the development and physiological relevance of a novel method to quantify LV segmental contraction synchrony. METHODS: LV pressure-volume and echocardiographic data were collected in seven anesthetized, opened-chest dogs under several pacing modes: right atrial (RA) (control), right ventricular (RV) (dyssynchrony), and additional LV pacing at either apex (CRTa) or free wall (CRTf). Cross-correlation-based integrated (CCSI(int) ) and segmental (CCSI(seg) ) measures of synchrony were calculated from speckle-tracking derived radial strain, along with a commonly used index (maximum time delay). LV contractility was quantified using either E(es) (ESPVR slope) or ESPVR(area) (defined in the manuscript). RESULTS: RV pacing decreased CCSI(int) at LV base (0.95 ± 0.02 [RA] vs 0.64 ± 0.14 [RV]; P < 0.05) and only CRTa improved it (0.93 ± 0.03; P < 0.05 vs RV). The CCSI(seg) analysis identified anteroseptal and septal segments as being responsible for the low CCSI(int) during RV pacing and inferior segment for poor resynchronization with CRTf. Changes in ESPVR(area) , and not in E(es) , indicated depressed LV contractility with RV pacing, an observation consistent with significantly decreased global LV performance (stroke work [SW]: 252 ± 23 [RA] vs 151 ± 24 [RV] mJ; P < 0.05). Only CRTa improved SW and contractility (SW: 240 ± 19 mJ; ESPVR(area) : 545 ± 175 mmHg•mL; both P < 0.01 vs RV). Only changes in CCSI(seg) and global LV contractility were strongly correlated (R(2) = 0.698, P = 0.005). CONCLUSION: CCSI(seg) provided insights into the changes in LV integrated contraction pattern and a better link to global LV contractility changes.

Abdominal aortic aneurysm monitoring via arterial waveform analysis: towards a convenient point-of-care device.

Abdominal aortic aneurysms (AAAs) are lethal but treatable yet substantially under-diagnosed and under-monitored. Hence, new AAA monitoring devices that are convenient in use and cost are needed. Our hypothesis is that analysis of arterial waveforms, which could be obtained with such a device, can provide information about AAA size. We aim to initially test this hypothesis via tonometric waveforms. We study noninvasive carotid and femoral blood pressure (BP) waveforms and reference image-based maximal aortic diameter measurements from 50 AAA patients as well as the two noninvasive BP waveforms from these patients after endovascular repair (EVAR) and from 50 comparable control patients. We develop linear regression models for predicting the maximal aortic diameter from waveform or non-waveform features. We evaluate the models in out-of-training data in terms of predicting the maximal aortic diameter value and changes induced by EVAR. The best model includes the carotid area ratio (diastolic area divided by systolic area) and normalized carotid-femoral pulse transit time ((age·diastolic BP)/(height/PTT)) as input features with positive model coefficients. This model is explainable based on the early, negative wave reflection in AAA and the Moens-Korteweg equation for relating PTT to vessel diameter. The predicted maximal aortic diameters yield receiver operating characteristic area under the curves of 0.83 ± 0.04 in classifying AAA versus control patients and 0.72 ± 0.04 in classifying AAA patients before versus after EVAR. These results are significantly better than a baseline model excluding waveform features as input. Our findings could potentially translate to convenient devices that serve as an adjunct to imaging.

Left ventricular and myocardial function in mice expressing constitutively pseudophosphorylated cardiac troponin I.

RATIONALE: Protein kinase (PK)C-induced phosphorylation of cardiac troponin (cTn)I has been shown to regulate cardiac contraction. OBJECTIVE: Characterize functional effects of increased PKC-induced cTnI phosphorylation and identify underlying mechanisms using a transgenic mouse model (cTnI(PKC-P)) expressing mutant cTnI (S43E, S45E, T144E). METHODS AND RESULTS: Two-dimensional gel analysis showed 7.2+/-0.5% replacement of endogenous cTnI with the mutant form. Experiments included: mechanical measurements (perfused isolated hearts, isolated papillary muscles, and skinned fiber preparations), biochemical and molecular biological measurements, and a mathematical model-based analysis for integrative interpretation. Compared to wild-type mice, cTnI(PKC-P) mice exhibited negative inotropy in isolated hearts (14% decrease in peak developed pressure), papillary muscles (53% decrease in maximum developed force), and skinned fibers (14% decrease in maximally activated force, F(max)). Additionally, cTnI(PKC-P) mice exhibited slowed relaxation in both isolated hearts and intact papillary muscles. The cTnI(PKC-P) mice showed no differences in calcium sensitivity, cooperativity, steady-state force-MgATPase relationship, calcium transient (amplitude and relaxation), or baseline phosphorylation of other myofilamental proteins. The model-based analysis revealed that experimental observations in cTnI(PKC-P) mice could be reproduced by 2 simultaneous perturbations: a decrease in the rate of cross-bridge formation and an increase in calcium-independent persistence of the myofilament active state. CONCLUSIONS: A modest increase in PKC-induced cTnI phosphorylation ( approximately 7%) can significantly alter cardiac muscle contraction: negative inotropy via decreased cross-bridge formation and negative lusitropy via persistence of myofilament active state. Based on our data and data from the literature we speculate that effects of PKC-mediated cTnI phosphorylation are site-specific (S43/S45 versus T144).