Reconstruction of Pathways involved in cardiomyocyte differentiation from embryonic stem cells

Funding Type: 
SEED Grant
Grant Number: 
RS1-00333
ICOC Funds Committed: 
$0
Disease Focus: 
Amyotrophic Lateral Sclerosis
Neurological Disorders
Spinal Cord Injury
Neurological Disorders
Stem Cell Use: 
Embryonic Stem Cell
Public Abstract: 
Heart disease is the leading cause of mortality and decline in the quality of life in the world. Current therapies are unable to restore function to damaged heart tissue. Recent scientific developments demonstrate the ability of human embryonic stem cells to form cardiomyocytes and this gives rise to the hope that we will be able to use these cells to differentiate and replace damaged myocardium. Despite this preliminary evidence and promise, we are far from being able to successfully accomplish regeneration of mature myocardium. This is largely due to our lack of understanding of all the factors that are involved in the differentiation process and our ability to manipulate the regeneration process. Our current understanding of the differentiation process indicates that there are at least four distinct stages of differentiation leading to cardiomyocytes, namely: (1) establishment of organizing centers; (2) mesendoderm induction; (3) establishment of cardiac precursors; and (4) terminal differentiation of beating cardiomyocytes. Our first quest is to identify what are the factors associated with each stage of differentiation leading to cardiomyocytes. We will carry out measurements of several biological macromolecules that are anticipated to play a role in sending signals for differentiation. This will be followed by systematic reconstruction of biochemical pathways so that we get a systems-level perspective on the differentiation process. In addition, we plan to embark on quantitative modeling of cardiomyogenesis to provide a firm basis for stem cell-based therapy. Quantitative modeling of cardiomyogenesis is certain to provide insights into the etiology of complex congenital heart defects (CHDs). A comprehensive quantitative approach will provide explanations for the malformations beyond that which can be obtained by single gene manipulations in typical experimental systems because of the ability to take into account the interconnected networks of signal transduction pathways. As important as this research is for pediatric cardiology, the significance for adult cardiology is perhaps more profound because it offers the promise of improving prospects for therapeutic intervention. In support of this view, adult cardiac disease, such as ventricular hypertrophy, reiterates molecular genetic pathways that are characteristic of embryonic development, and thus the predictions based on quantitative modeling and in vitro testing might define targets for pharmaceutical intervention.
Statement of Benefit to California: 
In general, cardiovascular disease typically refers to a wide variety of heart and blood vessel diseases, including coronary heart disease, hypertension, stroke, and rheumatic heart disease. In 2000 nearly 200,000 Californians were hospitalized on account of heart disease. The statewide hospitalization rate due to heart disease in California is 6.4 per thousand in 2000. In addition to heart related illnesses, Stroke, hypercholesterolemia, high blood pressure and stroke accounted for another 30 percent of residents in California. Nearly 100,000 deaths resulted from these pathologies. In almost every case damage to the heart preceded death. Several risk factors besides genetics and heredity play a role in heart diseases. While prevention by healthy life styles and avoidance of risk factors is the first choice, this may not be feasible for all the residents of California. Ability of embryonic stem cells to differentiate into cardiac cells provides the hope that therapy by regeneration of damaged heart is a definitely possibility for the future. However, our understanding of the processes that lead a stem cell to differentiate into a cardiac lineage, is very primitive and any efforts at stem cell therapy mandate that we have a molecular and systemic understanding of cardiomyogenesis. This is the major objective of our proposal. Based on obtaining a detailed molecular picture of pathways that lead to cardiac differentiation from stem cells, we will be able to design small molecules/drugs that would trigger such differentiation in pathology. Further, such understanding will also aid preventive measures that will lead to reduction of the number of heart-related fatalities in California.
Progress Report: 
  • A main goal of research in our laboratory is to identify strategies to promote neural repair in spinal cord injury and related neurological conditions. On the one hand, we have been using mouse models of spinal cord injury to study a long-standing puzzle in the field, namely, why axons, the fibers that connect nerve cells, do not regenerate after injury to the brain and the spinal cord. On the other hand, relevant to this CIRM SEED grant, we have started to explore the developmental and therapeutic potential of human embryonic stem cells (hESCs) for neural repair. We do this by first developing a method to genetically manipulate a HUES line of hESCs. The advent of hESCs has offered enormous potential for regenerative medicine and for basic understanding of human biology. To attain the full potential of hESCs as a tool both for therapeutic development and for basic research, we need to greatly enhance and expand our ability to genetically manipulate hESCs. A major goal for our SEED grant-sponsored research is to establish methods to genetically manipulate the HUES series of hESC lines, which are gaining wide utility in the research community due to the advantages on their growth characteristics over previously developed hESC lines. The first gene that we targeted in HUES cells, Fezf2, is critical for the development of the corticospinal tract, which plays important roles in fine motor control in humans and hence represents an important target for recovery and repair after spinal cord injury. By introducing a fluorescent reporter to the Fezf2 locus, we are now able to monitor the differentiation of hESCs into Fezf2-expressing neuronal lineages. This work has been published. A second goal is to start to explore the developmental and therapeutic potential of these cells and cells that derived from these cells in the brain and spinal cord. We are currently utilizing the cell line genetically engineered above to develop an efficient method to differentiate HUES cells into subcerebral neurons. Results so far have been encouraging. Efforts are also underway to overexpress Fezf2 as a complementary approach to drive the differentiation of HUES cells into specific neuronal types. Together, these studies will lay down the foundation for therapeutic development with HUES cells and their more differentiated derivatives for neurological disorders including spinal cord injury where neural regeneration can be beneficial. The CIRM SEED grant has allowed us to pursue a new, exciting path of research that we would have not pursued had we not been awarded the grant. Furthermore, the CIRM funded research has opened a new window of opportunity for us to explore genetic engineering of hESCs to model human neurological conditions in future.

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