Microfluidics-based protocol for differentiation of human ESCs to hepatic cells
Our ability to grow and manipulate human ES (hES) cells has improved markedly in recent years. A number of procedures have been developed to allow differentiation of human ES cells into specific cell types such as insulin-producing cells and liver cells. Despite these tremendous technical advances, several major technical challenges need to be addressed if the intent of such differentiation is cell replacement therapy. To perform any transplantation assay, whether in animal models or in humans, millions to billions of cells must first be induced to transform into a specific cell type to provide adequate starting material. Therefore, a robust procedure for scaling up the culture procedure is the first requirement. At the same time, the transplantable cells must exist in a pure population, devoid of any unwanted, undifferentiated (and therefore potentially tumorigenic) hES cells. In this proposal, we will solve these problems using a new microfluidic platform that delivers growth factors to the cells in a uniform, efficient manner to decrease the heterogeneity and increase the yield of specialized cells from hES cells. Further, we propose to develop prototype bioreactors to produce specialized cells from hES cells in large-scale. During the course of this experiment, we will focus on generating hepatocytes (liver cells) from human ES cells. Availability of a large number of hepatocytes will significantly impact the field of liver diseases for the following reasons. First, stem cell therapy for severe liver failure is particularly attractive as donor livers are rare and intravenous administration of differentiated hepatocytes usually results in their deposition into the liver. Second, there are significant benefits to the field of hepatitis research from the development of a homogeneous and plentiful source of human hepatocyte. Currently there are no effective animal model systems for studying hepatitis virus infection, as the virus infects only human and primate cells. Thus availability of a reliable source of human hepatocytes will accelerate the rate of progress in the field. Third, hES cell lines with genotypes characteristic of various genetic diseases that affect liver function could provide novel insights into mechanisms of the liver disease process and variations in drug response. In summary, this project attempts to accomplish two goals. One is to develop a widely used technique to grow a large population of homogeneous cells. Second is to produce a large numbers of hepatocytes that will be useful in clinical research.
A primary goal of Proposition 71 is to translate basic stem cell research to clinical applications. The disability and loss of earning power and personal freedom resulting from a disease or disorder are devastating and create a financial burden for California in addition to the suffering caused to patients and their families. Therapies using human embryonic stem cells (hES cells) have the potential to change millions of lives. Using hES cells as models of disease will help us understand the underlying causes of disease and likely aid in the development of drugs to treat those diseases.
In this proposal, we will focus to produce a large number of liver cells to cure various liver diseases. It is astonishing that liver diseases such as cirrhosis and hepatitis affect 25 million people in the United States, and that over 27,000 Americans die from cirrhosis annually, making it the country's third leading cause of death for people between the ages of 25 and 59. The 2006 report submitted to the Center for Health Statistic for the State of California indicated that chronic liver disease and cirrhosis age-adjusted death rate was 10.6 deaths per 100,000 population in California. The number is significantly high and this number did not meet the Healthy People 2010 objective of no more than 3 deaths.
Given the fact that often the only way to cure patients with sever liver disease is by transplanting new healthy livers, and that donor livers are rare, a new approach to obtain liver cells for transplantation purpose is urgently needed. Since liver transplantation will require millions to billions of cells, which must first be induced to transform into a specific cell type to provide adequate starting material, a robust procedure for scaling up the culture procedure is the first requirement. At the same time, the transplantable cells must exist in a pure population, devoid of any unwanted, undifferentiated (and therefore potentially tumorigenic) hES cells. Only when these goals are met, we can consider developing clinical trial methodology that will directly impact human testing of stem cell therapies. Our proposed approach can overcome these major issues.
Additional significant benefits obtaining a reliable source of hepatocytes are the following. It would distinctly increase the pace of development of novel therapies for hepatitis patients. Second, it will serve to develop new improved methods for developing and testing drugs for treating liver diseases. It is also anticipated that, in the long term, the return to the State in terms of revenue, health benefits for its Citizens and job creation will be significant. The State of California will also be viewed as the leader in establishing clinical stem cell research programs.