The Bernstein laboratory studies cardiac and skeletal muscle development and regeneration. Our longterm goal is to exploit cellular processes to achieve tissue repair.
Perspective. Myocardial disease. Each year, over 900,000 people in the United States experience an acute myocardial infarction, and ~500,000 succumb to the sequelae of myocardial insufficiency. The already significant economic impact of this disease is likely to escalate as our ability to save people from the acute event improves. At the cellular level, myocardial insufficiency results from the cumulative death of cardiac myocytes and the inability of remaining cells to proliferate. Indeed, prognosis directly correlates with viable tissue mass after infarct.
Muscular dystrophy. Muscular dystrophies are profoundly debilitating disorders that affect more than 1 in 3,500 male births. They comprise a group of genetic diseases that cause progressive weakness and degeneration of skeletal muscle resulting from defective proteins critical to muscle integrity. These defective proteins are thought to predispose muscle to damage from normal activity, leading to premature exhaustion of the muscle stem cell reservoir that maintains muscle integrity during normal use. While satellite cells are thought to comprise the principal source of muscle regeneration in adults, several other types of stem-like cells have been shown to have regenerative properties, including muscle side population (SP) cells, Sca-1-expressing cells, CD133+ muscle progenitors, and myo-endothelial cells. However, the position of these variously identified cells within the myogenic lineage, and how they may be related to satellite cells and each other, remains unclear.
Elucidating the myogenic progenitor pool in both the developing heart and adult skeletal muscle could lead to molecular and cellular strategies for regenerating these tissues, and would be important contributions to therapy for these diseases.
Overview. Striated muscle cells cease division within weeks of birth. While skeletal muscle retains limited capacity for regeneration through recruitment of satellite cells, resident populations of adult myocardial stem cells have only recently been identified. Both tissues have limited innate ability to regenerate. Our laboratory’s goal is to understand mechanisms regulating cell division, and how such processes play a role in muscle biology and disease.
Our work focuses on three main areas of basic investigation: 1) mechanisms of cell cycle withdrawal during muscle differentiation;2) cardiomyocyte and skeletal myoblast fate determination and subspecialization, and; 3) the role of cell cycle machinery in cellular hypertrophy. In addition, we recently have initiated two areas of translational and clinical research that apply our understanding of how muscle cells behave to the development of new diagnostic and therapeutic approaches to heart failure and muscular dystrophy: 4) human embryonic and induced pluripotent stem cell-based therapies for heart failure and muscular dystrophies; and 5) identification of biomarkers of heart failure in patients with congenital heart disease.
(Beating cardiac muscle cells derived from human embryonic stem cells)
Mechanisms of cell cycle withdrawal during differentiation.
To study the cell cycle withdrawal program in skeletal and cardiac myocytes, we have developed methods for differentiating rodent and avian myoblasts into myocytes with spontaneous contractile activity, and for establishing and manipulating primary myoblast cultures. Using microarrays, we have identified several regulatory proteins that are differentially expressed in proliferating myoblasts versus post-mitotic myotubes. We have demonstrated that one of these proteins, Stem cell antigen-1 (Sca-1/Ly6A), specifically regulates the proliferation/differentiation transition during skeletal myogenesis. Current efforts in primary myoblasts and mouse models are focused on determining the mechanisms by which Sca-1 specifically controls myoblast proliferation and regulates myogenic precursor cell self-renewal.
Cardiomyocyte and skeletal myoblast fate determination and subspecialization.
To examine the regulation of human cardiac and skeletal myoblast fate determination, our lab is identifying human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) that preferentially differentiate into cardiomyocytes and skeletal myocytes, investigating the regulatory mechanisms that govern these cell fate decisions, and developing methods for isolating developmentally-synchronized stem cell-derived myocardial and myoblast precursors. This will facilitate efforts to determine the contributions of genetic programming and environmental stimuli to subspecialization of human cardiomyocytes into atrial and ventricular myocardium, and conduction tissue, and the development of physiologically distinct muscle fiber types.
The role of cell cycle machinery in cellular hypertrophy.
We have demonstrated that hypertrophic stimuli cause a transient burst of Cdk4 activity, remodeling of the retinoblastoma protein complex, and activation of a subset of E2F-1 target genes in murine myoblasts. This has led us to identify a physiological role for both E2F-1-mediated activation and repression of genes involved in cell growth versus division, respectively. Currently, we are investigating the mechanism(s) by which Cip1/Kip1 and INK4 classes of Cdk-inhibitors facilitate this burst of Cdk4 activity, and the role of chromatin remodeling in the hypertrophic response.
Human embryonic and induced pluripotent stem cell-based therapies for heart failure and muscular dystrophies.
Over the past several years, the identification of heterogeneous populations of muscle stem cells in the heart, bone marrow, and adult skeletal muscle has generated great enthusiasm for new approaches to muscle repair and regeneration. However, these studies also have exposed the limitations of current strategies. Taking cues from the highly plastic, developing human heart, our lab is exploring stem cell-based therapies for heart failure and muscular dystrophy. Using mouse models of global and focal myocardial injury, as well as genetic models of muscular dystrophy, we are determining the developmental stages at which embryonic and induced pluripotent stem cell-derived myocardial cells and myoblasts engraft in vivo, and examining the effects of stem cell therapy on muscle physiology and function.
Identification of biomarkers of heart failure in patients with congenital heart disease.
To complement our efforts toward cell-based therapies for heart failure, our group also is investigating better ways to monitor heart failure and the response to therapy in children with single ventricle heart disease, the most difficult congenital heart defect to manage. We have an ongoing human research protocol to measure the levels of proteins and microRNAs found in blood in children with single ventricle compared to children with structurally normal hearts, to determine whether any of these potential biomarkers predict the presence or degree of heart failure in these children. New efforts will be directed toward using proteomics to establish biomarker arrays for pediatric heart failure.
See the Book!
Tissue Engineering in Regenerative Medicine (Stem Cell Biology and Regenerative Medicine Series )
Bernstein, Harold S. (Ed.)
1st Edition., 2011, 510 p. 56 illus., 52 in color.
Humana Press (Springer)
Hardcover, ISBN 978-1-61779-321-9
Available: August 31, 2011
Over the past decade, significant advances in the fields of stem cell biology, bioengineering, and animal models have converged on the discipline of regenerative medicine. This volume provides a state-of-the-art report on tissue engineering toward the goals of tissue and organ restoration and regeneration. Examples from different organ systems are used to illustrate progress with growth factors to assist in tissue remodeling; the capacity of stem cells for restoring damaged tissues; novel synthetic biomaterials to facilitate cell therapy; transplantable tissue patches that preserve three-dimensional structure; synthetic organs generated in culture; aspects of the immune response to transplanted cells and materials; and the development of suitable animal models for non-human clinical trials. Throughout the chapters, the reader will observe a common theme of basic discovery informing clinical translation, and clinical studies in animals and humans guiding subsequent experiments at the bench.
Sources of Support.
The Bernstein Lab and its members have been supported by ~
University of California, San Francisco
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