Research Team

Postdoctoral Fellows

Ludovic Benard, PhD

Dr. Benard’s work focuses on the role of stromal interaction molecule 1 (STIM1) in the development of cardiac hypertrophy and heart failure (HF). Cardiac STIM1 expression is increased during HF, but its mechanism is still non-elucidated. The strategy is to shut down STIM1 expression in pathological contexts to see its implication in cardiac dysfunction.

The aim of his work is to find how STIM1 is activated using two approaches:

  1. Decrease STIM1 expression using adeno-associated virus (AAV) vectors at different stages of cardiac dysfunction in an aortic banding induced HF in mice.
  2. Generate a cardiac specific knockout mouse strain in order to explore molecular STIM1 partners.

Antoine Chaanine, MD

Dr. Chaanine is investigating how endoplasmic reticulum via the emission of calcium signals can affect the function of the neighboring intermyofibrillar mitochondria. The endoplasmic reticulum and the intermyofibrillar mitochondria are interconnected with tethering complexes and cross talk with each other by means of calcium signals. In heart failure (HF), there is a mitochondrial calcium overload that leads to mitochondrial dysfunction. Via gene therapy, the laboratory will study if therapeutic targeted transfer of AAV9-SERCA2a can attenuate mitochondrial calcium overload and rescue the mitochondria from initiating autophagic and apoptotic signaling pathways in a rat model of pressure overload induced HF.

He is also investigating the role of BNIP3 in selectively inducing mitochondrial autophagy in the heart. The laboratory attributes the increase in mitochondrial removal by autophagy through BNIP3 signaling, which is maladaptive, and leads to a deterioration of cardiac function in cardiac hypertrophy and in HF. Via gene therapy, the laboratory will study if therapeutic targeted down-regulation of BNIP3, using AAV9-SiBNIP3, can attenuate mitochondrial autophagy and improve cardiac function in a rat model of pressure overload induced hypertrophy and HF.

Elie Chemaly, MD

Dr. Chemaly is working on several projects, one of which has involved the characterization of the in vivo cardiac effects of long-term overexpression of resistin in rats.

Another series of projects involves the effects of left ventricular (LV) wall stress on electrical remodeling, SERCA2a expression and its potential role in the progression to left ventricular dilatation, and reduced ejection fraction in the setting of pressure overloaded left ventricular hypertrophy in rats. His data indicates that stroke volume preservation would be the other component driving the remodeling.

He also optimized a protocol of dobutamine infusion in rats to study the dose-response on contractility increase, afterload decrease, and their interdependence using conductance catheters for volume. In that setting, he is characterizing the response to dobutamine in vivo of different models of increased preload, and increased afterload hypertrophy.

Jiqiu Chen, MD

Dr. Chen is currently working on the role of fibrosis in congestive heart failure (HF) in rats. The cardiac fibrosis was induced by aortic constriction plus coronary arterial ischemia-reperfusion followed by de-aortic banding. The model has been used to test the effects of AAV9.hsp20 gene transferring on HF and cardiovascular fibrosis. His previous data shows that de-overload ischemic left ventricular (LV) could trigger neointimal formation in coronary arteries. He is exploring whether perivascular fibroblasts and iNOS from vessel walls (endothelial dysfunction) are involved in the events. Bundle collagen fibers will be isolated from adult rats and visualized with CNA35 fluorescent probe in vitro. The mechanical character of isolated bundle collagen fibers will be studied if he can arrange a collaboration with another laboratory. The overall aim is to understand CHF mechanisms associated with non-myocardial elements and to explore how to raise the efficiency of non-inotropic medicine in treatment of HF.

Elisa Yaniz-Galende, PhD

The laboratory is interested in understanding how the heart responds to injury after myocardial infarction. From our work and that of many others, it has emerged that molecules such as stem cell factor (SCF) play an important role in cardiac regeneration, regulating the maintenance, function and activation of progenitors and stem cells. Our recent research is based on cell therapy methods, we developed to enhance the regenerative potential of cardiac resident stem cells, driven by SCF adenoviral gene transfer upon myocardial infarction (MI). During this work, we establish that SCF treatment enhances cardiac function and regeneration after MI, as a consequence of cardiac progenitors and stem cells recruitment to the damage area, together with an increase in proliferation. This important result suggests that SCF adenoviral gene transfer may have a positive clinical benefit in patients with MI and heart failure.

Lahouaria Hadri, PhD

Growth and migration of vascular smooth muscle cells (VSMCs) are responses to arterial injury that are highly important to the processes of restenosis after percutaneous transluminal coronary angioplasty and atherosclerosis. Proliferation is associated with alterations in gene expression of the VSMCs that go from a quiescent/differentiated phenotype to a proliferating/dedifferentiated one. Vascular remodeling includes alteration in Ca2+ handling proteins such as replacement of the L-type Ca2+ channels by the T-type {Kuga, 1996 #1; Gollasch, 1998 #2} and increase in the capacitative Ca2+ entry {Golovina, 2001 #3;Golovina, 1999 #4}. The laboratory has shown that SR Ca2+ handling is also altered with a loss of the ryanodine receptor Ca2+ channels and of the sarco(endo)plasmic reticulum Ca2+ ATPase isoform SERCA2a {Lipskaia, 2003 #5;Vallot, 2000 #6}. Ca2+ controls different functions of the VSMCs including gene expression via modulation of Ca2+-regulated transcription factors. Two main Ca2+-regulated transcription factors have been described in VSMCs: CREB (cAMP-responsive element binding protein) and NFAT (nuclear factor of activated T-cells). CREB is activated by Ca2+-dependent phosphorylation by Ca2+ /calmodulin or MAP kinases whereas NFAT is dephosphorylated by calcineurin, a Ca2+-dependent phosphatase. Protein phosphatase 1 (PP1), the main regulator of the transcription factor CREB, is strongly involved in the control of cell proliferation via p53 and p21. PP1 is regulated by phosphatase inhibitor-1 (I-1) which is activated by PKA phosphorylation and inhibited by protein phosphatase PP2A and PP2B (calcineurin) {Hagiwara, 1992 #12} {Alberts, 1994 #11}. I-1 has been postulated as an integrator of multiple neurohormonal pathways associated with Ca2+ homeostasis and proper contractile function. I-1 is expressed VSMCs, but does not appear to play a significant role in contractile or relaxant response {Carr, 2001 #68}. Because calcineurin strongly controls VSMCs proliferation {Lipskaia, 2005 #79}, the protein phosphatase inhibitor-1, by regulating PP1 and thus controlling the CREB phosphorylation, can regulate VSMCs phenotype and control vascular remodeling.

Kiyotake Ishikawa, MD

The laboratory is researching restoration of cardiac function using large animals. Various types of heart failure (HF) models, such as acute myocardial infarction, chronic coronary occlusion, and mitral valve regurgitation, are created in our lab. These models develop HF which closely reflects severe HF in humans. The laboratory tests the efficacy and the safety of newly invented therapies in these models, and work to bridge small animal studies to clinical accommodations. Our work will help many patients who suffer from severe HF that show minimum improvement with available treatments.

Dongtak Jeong, PhD

Dr. Jeong’s focus is on two different projects. The first project aims to figure out the beneficial effect of SUMO-1 in heart failure (HF) models using transgenic mice and AAV injection. Our group has shown that SUMO-1 is one of the key partners in regulating the SERCA2a activity and stability in vitro. The laboratory has also shown that SUMO-1 expression was significantly downregulated in HF in both human and experimental models. It is, however, not known whether these findings can be translated in the genetically modified mice. The laboratory is figuring out how the data can be applied to the in vivo setting. Dr. Jeong is also focusing on the molecular manipulation of AAV vectors in order to find out the optimal gene delivery vector which is minimally affected by its neutralizing antibody in various species.

Ioannis Karakikes, PhD

Direct reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) by ectopic expression of defined transcription factors has opened up the way for potentially new diagnostic tools. The iPSC technology in combination with tissue engineering can provide advanced in vitro models for drug testing and disease modeling. Dr. Karakikes’s current research is focused on the development of 3-dimensional human engineered cardiac tissues (3D-hECT) containing cardiomyocytes derived from human embryonic stem cells (ESCs) and patient-specific iPSCs. These 3-D models display physiological characteristics similar to human myocardium and are useful for in vitro drug screening, toxicology assays, and disease modeling. Additionally, he has established patient-specific iPSC lines and developed a chemically defined, efficient, and reproducible protocol for differentiating human iPSCs toward cardiomyocytes. Using these iPSC-derived cardiomyocytes, He is investigating the molecular and cellular mechanisms underlying the developmental of cardiovascular defects and assessing their capacity to recapitulate key aspects of patient-specific cardiac phenotypes.

Chang Won Kho, PhD

Knowledge of both global alternation of proteins and protein–protein interactions is crucial to understanding how cells function and the general principles that govern this function. This knowledge is important in helping to understand disease processes. Cardiac sarco/endoplasmic reticulum calcium ATPase (SERCA2a) pump is a promising therapeutic target in heart failure (HF). The SERCA2a gene therapy, as a way to repair SERCA2a function in patients with advanced HF, is under evaluation in a human clinical trial. During this study, our group has gained great experience in targeting calcium cycling by AAV mediated gene transfer in pre-clinical swine models of HF. Using these swine models, Dr. Kho has studied the combined different proteomic approaches to investigate global alternations in protein expression and protein–protein interaction following restoration of SERCA2a that more closely mimic human physiology, function, and anatomy. The long-term goals of this study are 1) to explore molecular alternations of cardiac proteins related to HF, and 2) to investigate the cardiac signaling effects of SERCA2a in swine models of HF by establishing a SERCA2a networking map. The most important thing that Dr. Kho hopes to understand is the global networks of failing cardiac proteins in pre-clinical large animal models. This project will provide a valuable resource for the understanding of the changes in molecular mechanisms related to HF and the effect of SERCA2a restoration which may result in the generation of new diagnostic and therapeutic markers.

Erik Kohlbrenner, BS

Gene therapy has proven to be a useful tool for basic biological research and it is hoped that it will ultimately bring about breakthrough medical treatments for a number of chronic diseases. Recombinant adeno-associated virus (rAAV) vectors have proven to be capable of delivering therapeutic genes into animals and in recent clinical trials, humans as well. The laboratory is interested in understanding the molecular basis of heart failure (HF) and rAAV has proven to be a valuable tool.

A number of candidate genes are being investigated in our laboratory. Once a gene is found to have a role in HF by in vitro experiments, an AAV is produced and delivered in vivo in a small animal model setting. If initial studies into physiological effects prove promising, the laboratory will scale up production for large animal use.

The production methods are based on published protocols such as transfection of HEK293T cells and purification by iodixanol gradient ultracentrifugation. Quality control assays are done to characterize purity, titer, in vitro infectivity, and confirm expected gene expression in infected cells. The laboratory is working on developing improved methods for production, purification, and characterization of rAAV.

In addition to production for in-house experiments, we also produce rAAV for collaborations within and outside of Mount Sinai.

Jason Kovacic, MD

Vascular biology is characterized by the interplay of multiple cell types including endothelial cells, vascular smooth muscle cells (VSMCs), platelets and immune cells. Surprisingly, the origins of many of these cell populations are unknown. For example, VSMCs were once thought to arise locally from the vessel wall. However, this was refuted by subsequent studies indicating a bone marrow (BM) origin. In turn, recent studies have argued against a BM origin, leaning again towards a situation in which VSMCs arise locally from the artery itself. As a possible explanation to these controversies, emerging data suggest that ‘cellular phenotypic switching’, in particular of an endothelial to mesenchymal cell or VSMC (endothelial to mesenchymal transition [EndMT]), may play a role in adult vascular biology.

Similarly, the role of endothelial cells in endothelial homeostasis is equally unclear. While some researchers argue for the BM origins of circulating ‘endothelial progenitor cells’ (EPCs), others suggest that non-BM derived circulating progenitors may account for a large proportion of adult neovascularization and endothelial repair. Adding complexity, several studies have now questioned the identity and importance of EPCs,9 with very recent data suggesting that circulating EPCs play no role whatsoever in atherosclerotic endothelial cell homeostasis.

Collectively, these knowledge gaps and uncertainties represent a major ‘road block’ in our understanding of normal arterial biology and atherosclerosis. The proposed research in the laboratory will directly address these deficiencies by characterizing the origins, role and fate of endothelial cells during arterial homeostasis, injury, aging and atherosclerosis. These experiments will expand our understanding of atherosclerosis and endothelial cell functionality with the aim of identifying novel preventive strategies or treatments for vascular disease.

This project will capitalize on unique and purpose-bred murine lines (end.Scl-Cre, R26R-YFP, ApoE-/- and inter-bred combinations of these lines) and will improve scientific knowledge by defining 1) the role of endothelial lineage-derived cells in endothelial homeostasis (as opposed to other sources that might provision cells to the endothelial compartment) , and 2) the role of endothelial phenotypic switching in atherosclerosis. There are numerous potential outcomes of this research that may impact the way clinical medicine is practiced. For example, the finding that endothelial-lineage derived cells are entirely responsible for the maintenance of endothelial integrity and homeostasis would, in conjunction with other emerging studies, dispel the theory that this is the purview of hematopoietic bone marrow-derived progenitor cells, shifting the focus of contemporary research back towards the endothelium. Therefore, this research has the potential to alter the conceptual paradigms with which we view vascular homeostasis, plaque formation, and atherosclerosis.

The proposed research seeks to characterize the potential contribution of endothelial lineage-derived cells to arterial homeostasis, arterial remodeling, and atherosclerosis. Our goal is to improve our understanding of the molecular and cellular events underlying these processes which ultimately may lead to the development of novel therapeutic interventions.

Razmig Garo-Kratlian, MD

The laboratory is studying the transduction efficiency of the different adeno-associated virus (AAV) serotypes 1-9 in vivo, to the arterial vasculature, with its different layers of tissues using the Rat carotid artery model. This work is highly important in order to elucidate which is the best AAV serotype to efficiently deliver a particular therapeutic gene of interest to the vasculature of the rat carotid artery and to study its effects on a particular disease.

The laboratory is also studying the effects of SERCA2a gene on Pulmonary Artery Hypertension (PAH). We shall be using the Monocrotaline (MCT) injury model of PAH in the rats to observe the ability of SERCA2a gene delivery to pulmonary arteries and its overexpression to prevent PAH in rats. We also will be studying the molecular markers and pathways involved in the process of SERC2a gene preventing PAH.

Thomas LaRocca, MD, PhD Student

7-transmembrane receptors (7TMR) are now well known to initiate G-protein independent events and have the ability to selectively activate particular pathways due to unique ligand binding or due to receptor mutation, termed pluripotential efficacy, or biased signaling. B-arrestins were originally described to participate in 7TMR desensitization/internalization upon ligand binding. Recently it has been determined b-arrestins can also initiate signaling events in the absence of G-protein activation. It is the balance between G-protein and b-arrestins that can determine the bias of the receptor on downstream signaling pathways. The Gi coupled chemokine receptor, CXCR4, has been implicated in stem cell recruitment, angiogenesis, and possibly cardiogenesis in the heart post-acute myocardial infarction. However, CXCR4 signaling events in the cardiac myocyte are still relatively unknown. We are particularly interested in the ability of CXCR4 in regulating long-term, hypertrophic responses and how G-protein and b-arrestin dependent events are involved. The laboratory has generated AAV9 vectors encoding the CXCR4 gene and the CXCR4 WHIM mutant which encodes a 19aa c-terminal truncation that fails to effectively recruit b-arrestin 2. The laboratory utilizes the murine TAC model to induce pressure overload in the heart and, through echocardiography and in vivo hemodynamics using a pressure-volume conductance catheter, we are determining the effects of biased CXCR4 signaling on hypertrophy and heart failure progression. These studies will shed further light on biased 7TMR signaling and provide novel insights into the role of CXCR4 on remodeling responses in the cardiac myocyte.

Ahyoung Lee, PhD

Post-translational modification (PTM) of proteins is an important way to change their function, activity, localization, stability, or turnover rate in response to environmental stresses after their synthesis has been completed. The overall goal of the research is to detect effective PTMs of SERCA2a for developing new therapies for heart disease using proteomic approaches. This may be a key step to understanding progressive cardiac pathologies including heart failure, where multiple genes are known to be involved. In particular, SUMOylation has been identified to be a novel PTM of the calcium cycling protein SERCA2a in vitro and in vivo. Dr. Le is researching the molecular effects and physiological consequences of the SERCA2a SUMOylation using a conditional transgenic mouse model and AAV9-mediated animal study in parallel. Ultimately, the laboratory hopes the findings can be used to reverse the calcium cycling abnormalities and contractile dysfunction in the failing heart by using cardiac SUMO technology.

Larissa Lipskaia, PhD

With an extensive background in basic cardiovascular research, Dr. Lipskaia directs a laboratory focusing on vascular muscle and endothelial cells functions, and calcium dynamics. She is focusing on the role of sarco/endoplasmicreticulum calcium ATPase (SERCA) isoforms in regulation of calcium homeostasis and calcium regulated transcription pathways in normal and diseased vessels. Her project centers on the use of gene therapy to prevent the coronary artery disease in patients with heart failure. The working hypothesis is that SERCA2a isoform controlling the frequency dependence of intracellular Ca2+ signaling may prevent the cascade of events that leads ultimately to vascular remodeling and cardiovascular disease.

Lifan Liang, MD

Kleopatra Rapti, PhD

Chronic heart failure is one of the leading causes of morbidity and mortality in western countries. Pharmacological approaches such as beta-adrenergic blockage, delay disease progression, but they do not cure the disease. Gene therapy using adeno-associated virus (AAV) vectors has emerged in the past few years as a promising therapeutic approach, mainly due to 1) the ability of AAV to display broad tissue tropism, 2) the comparatively low immunogenicity of AAV, and 3) its ability to maintain long-term expression. Several AAV serotypes with distinct tissue tropism have been identified so far. Three serotypes in particular display muscle tropism in vivo: AAV1, 6 and 9, the latter being the most cardiotropic, at least in mice and rats. Interestingly, in vitro they display a different transduction profile―AAV6 showing the highest transduction efficiency in rat adult cardiac myocytes (preliminary data). Furthermore, AAV tropism appears to be species specific. For example, in mice, AAV9 transduces liver three-times more efficiently than heart. In rats, this is quite the opposite, heart is transduced two-fold more efficiently than liver. Little is known about the tissue tropism in larger animals.

The hypothesis is that the differences between in vivo tissue tropism and in vitro transduction profiles stem from one or, more likely, a combination of several of the following factors:

  1. Differential receptor binding
  2. Postbinding events, such as viral endocytosis and/or intracellular trafficking
  3. The differential ability of the AAV serotypes to cross the vascular endothelial barrier
  4. Differences in preexisting neutralizing antibody titers to AAVs in animal models

Whereas, the studies are conducted mainly in rats, the expectation is that the basic conclusions hold true in other animal species as well. Overall, the goal is to understand better the contribution of each of these factors in the interspecies diversity of AAV tissue tropism.

Grant Senyei

His current project focuses on the effects of small molecules on the directed differentiation of human embryonic stem cells into mature cardiomyocytes. The improved differentiation efficiency over current standard protocols allows for a renewable and reliable source of cardiac cells for in vitro studies. Additionally, the specific pathways upon which the small molecules impact the cells’ fate provide a direction for studies aimed at better understanding cardiac development. Using this small molecule-mediated differentiation, he is dissecting the role of the cellular signaling pathways and candidate genes that direct the multistep process of embryonic stem cell to cardiomyocyte differentiation. The enhanced efficacy of a small molecule-directed cardiomyocyte differentiation protocol can facilitate studies of cardiac development and, ultimately, of cell replacement therapies.