Faculty Bibliography

The Summer Institute Program to Increase Diversity (SIPID) in Health-Related Research is a career advancement opportunity sponsored by the National Heart, Lung, and Blood Institute. Three mentored programs address difficulties experienced by junior investigators in establishing independent research careers and academic advancement. Aims are to increase the number of faculty from under-represented minority groups who successfully compete for external research funding. Data were collected using a centralized data-entry system from three Summer Institutes. Outcomes include mentees' satisfaction rating about the program, grant and publications productivity and specific comments. Fifty-eight junior faculty mentees (38% male) noticeably improved their rates of preparing/submitting grant applications and publications, with a 18-23% increase in confidence levels in planning and conducting research. According to survey comments, the training received in grantsmanship skills and one-on-one mentoring were the most valuable program components. The SIPID mentoring program was highly valued by the junior faculty mentees. The program will continue in 2011-2014 as PRIDE (PRogram to Increase Diversity among individuals Engaged in health-related research). Long-term follow-up of current mentees will be indexed at five years post training (2013). In summary, these mentoring programs hope to continue increasing the diversity of the next generation of scientists in biomedical research.

BACKGROUND: The neuroendocrine Cav1.3 L-type Ca channels have been recently found in the Human fetal heart and shown to play a vital role in Ca entry from the sarcolemma into the cell and in Ca homeostasis. Calreticulin, a Ca binding endoplasmic reticulum (ER) resident protein, has been recently shown to translocate to the cell surface where its role and function are just emerging. Here, we demonstrated a novel mechanism of Cav1.3 and calreticulin interaction resulting in downregulation of Cav1.3 channel densities in native Human fetal cardiac cells and Human Embryonic Kidney cell lines (tsA201). METHODS AND RESULTS: Cell surface and cytoplasmic staining of calreticulin was demonstrated first in cultured human fetal cardiomyocytes (HFC), gestational age 18-24weeks, using confocal microscopy thereby establishing that calreticulin is present at the cell surface in HFC. Co-immunoprecipitation from HFC using anti-Cav1.3 Ca channel antibody, and probing with anti-calreticulin antibody revealed a 46kDa band corresponding to calreticulin suggesting that Cav1.3 Ca channel and calreticulin co-assemble in a macromolecular complex. Co-expression of Cav1.3 and calreticulin in tsA201 cells resulted in a decrease in surface expression of Cav1.3 Ca channels. These findings were consistent with the electrophysiological studies showing that co-transfection of Cav1.3 Ca channel and calreticulin resulted in 55% reduction of Cav1.3 Ca current densities recorded from tsA201 cells. CONCLUSIONS: The results show the first evidence that calreticulin: (1) is localized outside the ER on the cell surface of HFC; (2) coimmunoprecipitates with Cav1.3 L-type Ca channel; (3) negatively regulates Cav1.3 surface expression thus resulting in decreased Cav1.3 Ca current densities. The data demonstrate a novel mechanism of modulation of Cav1.3 Ca channel by calreticulin, which may be involved in pathological settings such as autoimmune associated congenital heart block where Cav1.3 Ca channels are downregulated.

Ghrelin is a peptide hormone produced mainly in the stomach that has widespread tissue distribution and diverse hormonal, metabolic and cardiovascular activities. The circulating ghrelin concentration increases during fasting and decreases after food intake. Ghrelin secretion may thus be initiated by food intake and is possibly controlled by nutritional factors. Lean subjects have increased levels of circulating ghrelin compared with obese subjects. Recent reports show that low plasma ghrelin is associated with elevated fasting insulin levels, insulin resistance and type 2 diabetes mellitus. Factors involved in the regulation of ghrelin secretion have not yet been defined; however, it is assumed that blood glucose levels represent a significant regulator. Recent evidence indicates that ghrelin can increase myocardial contractility, enhance vasodilatation, and has protective effect from myocardial damage. It has been shown that ghrelin may improve cardiac function through growth hormone (GH)-dependent mechanisms but there is also evidence to suggest that ghrelin's cardioprotective activity is independent of GH. Recent data demonstrate that ghrelin can influence key events in atherogenesis. Thus, ghrelin may be a new target for the treatment of some cardiovascular diseases. In this review, we consider the current literature focusing on ghrelin as a potential antiatherogenic agent in the treatment of various pathophysiological conditions.

Long QT syndrome (LQTS) is a congenital abnormality of cardiac repolarization that manifests as a prolonged QT interval on 12-lead electrocardiograms (ECGs). The syndrome may lead to syncope and sudden death from ventricular tachyarrhythmias known as torsades de pointes. An increased persistent Na(+) current is known to cause a Ca(2+) overload in case of ischemia for example. Such increased Na(+) persistent current is also usually associated to the LQT3 syndrome. The purpose of this study was to investigate the pathological consequences of a novel mutation in a family affected by LQTS. The impact of biophysical defects on cellular homeostasis are also investigated. Genomic DNA was extracted from blood samples, and a combination of PCR and DNA sequencing of several LQTS-linked genes was used to identify mutations. The mutation was reproduced in vitro and was characterized using the patch clamp technique and in silico quantitative analysis. A novel mutation (Q1476R) was identified on the SCN5A gene encoding the cardiac Na(+) channel. Cells expressing the Q1476R mutation exhibited biophysical alterations, including a shift of SS inactivation and a significant increase in the persistent Na(+) current. The in silico analysis confirmed the arrhythmogenic character of the Q1476R mutation. It further revealed that the increase in persistent Na(+) current causes a frequency-dependent Na(+) overload in cardiomyocytes co-expressing WT and mutant Nav1.5 channels that, in turn, exerts a moderating effect on the lengthening of the action potential (AP) duration caused by the mutation. The Q1476R mutation in SCN5A results in a three-fold increase in the window current and a persistent inward Na(+) current. These biophysical defects may expose the carrier of the mutation to arrhythmias that occur preferentially in the patient at rest or during tachycardia. However, the Na(+) overload counterbalances the gain-of-function of the mutation and is beneficial in that it prevents severe arrhythmias at intermediate heart rates.

Pervious biochemical and hemodynamic studies have highlighted the important role of epsilonPKC in cardioprotection during ischemic preconditioning. However, little is known about the electrophysiological consequences of epsilonPKC modulation in ischemic hearts. Membrane permeable peptide epsilonPKC selective activator and inhibitor were used to investigate the role of epsilonPKC modulation in reperfusion arrhythmias. Methods: Protein transduction domain from HIV-TAT was used as a carrier for peptide delivery into intact Langendorff perfused guinea pig hearts. Action potentials were imaged and mapped (124 sites) using optical techniques and surface ECG was continuously recorded. Hearts were exposed to 30min stabilization period, 15min of no-flow ischemia, followed by 20min reperfusion. Peptides (0.5muM) were infused as follows: (a) control (vehicle-TAT peptide; TAT-scrambled psiepsilonRACK peptide); (b) epsilonPKC agonist (TAT-psiepsilonRACK); (c) epsilonPKC antagonist (TAT-epsilonV1). Results: Hearts treated with epsilonPKC agonist psiepsilonRACK had reduced incidence of ventricular tachycardia (VT, 64%) and fibrillation (VF, 50%) compared to control (VT, 80%, P<0.05) and (VF, 70%, P<0.05). However, the highest incidence of VT (100%, P<0.05) and VF (80%) occurred in hearts treated with epsilonPKC antagonist peptide epsilonV1 compared to control and to epsilonPKC agonist psiepsilonRACK. Interestingly, at 20min reperfusion, 100% of hearts treated with epsilonPKC agonist psiepsilonRACK exhibited complete recovery of action potentials compared to 40% (P<0.05) of hearts treated with epsilonPKC antagonist peptide, epsilonV1 and 65% (P<0.5) of hearts in control. At 20min reperfusion, maps of action potential duration from epsilonPKC agonist psiepsilonRACK showed minimal dispersion (48.2+/-9ms) compared to exacerbated dispersion (115.4+/-42ms, P<0.05) in epsilonPKC antagonist and control (67+/-20ms, P<0.05). VT/VF and dispersion from hearts treated with scrambled agonist or antagonist peptides were similar to control. Conclusion: The results demonstrate that epsilonPKC activation by psiepsilonRACK peptide protects intact hearts from reperfusion arrhythmias and affords better recovery. On the other hand, inhibition of epsilonPKC increased the incidence of arrhythmias and worsened recovery compared to controls. The results carry significant therapeutic implications for the treatment of acute ischemic heart disease by preconditioning-mimicking agents.

Cardiac excitability and electrical activity are determined by the sum of individual ion channels, gap junctions and exchanger activities. Electrophysiological remodeling during heart disease involves changes in membrane properties of cardiomyocytes and is related to higher prevalence of arrhythmia-associated morbidity and mortality. Pharmacological and genetic manipulation of cardiac cells as well as animal models of cardiovascular diseases are used to identity changes in electrophysiological properties and the molecular mechanisms associated with the disease. Protein kinase C (PKC) and several other kinases play a pivotal role in cardiac electrophysiological remodeling. Therefore, identifying specific therapies that regulate these kinases is the main focus of current research. PKC, a family of serine/threonine kinases, has been implicated as potential signaling nodes associated with biochemical and biophysical stress in cardiovascular diseases. In this review, we describe the role of PKC isozymes that are involved in cardiac excitability and discuss both genetic and pharmacological tools that were used, their attributes and limitations. Selective and effective pharmacological interventions to normalize cardiac electrical activities and correct cardiac arrhythmias will be of great clinical benefit.

Transgenic Murine Models of CHB. Introduction: Congenital heart block (CHB) is a passively acquired autoimmune disease considered to be due to the transfer of maternal autoantibodies, anti-SSA/Ro -SSB/La, to the fetus resulting in atrioventricular (AV) block and sinus bradycardia. We previously established a murine model for CHB where pups born to immunized wild-type (WT) mothers exhibited electrocardiographic abnormalities similar to those seen in CHB and demonstrated inhibition of L-type Ca channels (LTCCs) by maternal antibodies. Here, we hypothesize that overexpression of LTCC should rescue, whereas knockout of LTCC should worsen the electrocardiographic abnormalities in mice. Methods and Results: Transgenic (TG) mice were immunized with SSA/Ro and SSB/La antigens. Pups born to immunized WT mothers had significantly greater sinus bradycardia and AV block compared to pups from nonimmunized WT. TG pups overexpressing LTCC had significantly less sinus bradycardia and AV block compared to their non-TG littermates and to pups born to immunized WT mothers. All LTCC knockout pups born to immunized mothers had sinus bradycardia, advanced degree of AV block, and decreased fetal parity. No sinus bradycardia or AV block were manifested in pups from control nonimmunized WT mothers. IgG from mothers with CHB children, but not normal IgG, completely inhibited intracellular Ca transient ([Ca](i) T) amplitude. Conclusions: Cardiac-specific overexpression of LTCC significantly reduced the incidence of AV block and sinus bradycardia in pups exposed to anti-SSA/Ro -SSB/La autoantibodies, whereas exposure of LTCC knockout pups to these autoantibodies significantly worsened the electrocardiographic abnormalities. These findings support the hypothesis that maternal antibodies inhibit LTCC and [Ca](i) T thus contributing to the development of CHB. Altogether, the results are relevant to the development of novel therapies for CHB. (J Cardiovasc Electrophysiol, Vol. pp. 1-9)

The novel Cav1.3 (alpha1D) L-type Ca channel plays a significant role in sino-atrial, atrioventricular nodes function and in atrial fibrillation. However, the characterization of alpha1D Ca channel during heart development is very limited. We used real-time RT-PCR, Western blotting and indirect immunostaining to characterize the developmental expression and localization of alpha1D Ca channel in rat hearts. Both protein and mRNA levels of alpha1D Ca channel decreased postnatally. Two forms of alpha1D Ca channel protein (250 kD and 190 kD) were observed, with the full length (250kD) channel protein being predominant in the prenatal stages. Both Western blots and confocal imaging demonstrated that alpha1D Ca channel protein was expressed in both atria and ventricles at fetal and neonatal stages but was absent in the adult ventricles. Interestingly, alpha1D Ca channel was also found at the nucleus/perinucleus of immature, but not adult atrial cells. Furthermore, the nuclear staining was reproduced in adult atrial cell line, HL-1 cells, which possess immature properties. The data are first to show that alpha1D Ca channel has unique age-dependent expression profile and subcellular localization in the heart, suggesting a developmental stage dependent specific function. ABBREVIATIONS: