Endodermal differentiation of human ES cells

Endodermal differentiation of human ES cells

Funding Type: 
SEED Grant
Grant Number: 
RS1-00308
Approved funds: 
$611,027
Disease Focus: 
Diabetes
Stem Cell Use: 
Embryonic Stem Cell
Public Abstract: 
The goals of this proposal are to investigate endodermal differentiation and proliferation in human ES cell cultures. Endodermal cells give rise to the epithelial lining of the respiratory and digestive tract as well as to the liver and pancreas. The future treatment of diseases such as type I diabetes using stem cell therapy relies on our ability to differentiate stem cells into endoderm, a prerequisite step to forming pancreatic beta cells. In 2005, D’Amour et al. reported the efficient differentiation of human ES cells into endoderm. This report provides a potentially effective protocol that needs to be further evaluated (specific aim 1). In addition, given that the success of stem-cell therapy depends on our ability to generate large numbers of differentiated cells (e.g. 200-700 million beta cells per patient are currently being used in the Edmonton protocol), we will investigate the ability of the endodermal generated in specific aim 1 cells to proliferate in culture (specific aim 2).
Statement of Benefit to California: 
Stem cell therapy relies on the development of efficient and reproducible protocols to differentiate stem cells into various cell types such as pancreatic beta cells. The first step to making pancreatic beta cells is the differentiation of stem cells into so-called endodermal cells, one of the 3 basic cell types of the body. In addition, in order to make stem cell therapy a viable option, one needs to be able to generate large numbers of differentiated cells from stem cells. Our proposal aims to establish such protocols. The health of California and its citizens will ultimately benefit from this research as it will help develop stem cell therapies.
Progress Report: 

Year 1

The goals of this proposal are to investigate endodermal differentiation and proliferation of human ES cells in culture. Endodermal differentiation is a necessary step towards making pancreatic beta cells, as well as other endodermal cells such as liver cells. Pancreatic beta cells generated from human ES cells could be used to treat type I diabetics. In the past two years, we have incorporated human ES cell culture technology into our laboratory and have been able to replicate data obtained by other research groups. While several other research groups and companies around the world are focused on making pancreatic beta cells as quickly as possible, we strongly believe that a more detailed understanding of the biology of human ES cell differentiation into endoderm will help the optimization of this protocol. Therefore, we have focused our efforts on testing a number of variables in the initial step of creating definitive endoderm. We have found that different human ES cell lines have very different capacity to differentiate into endoderm under the same culture conditions. In addition, we have recently focused our research effort on the post-translational modifications of key regulators of endoderm differentiation, and found a critical role for a poorly appreciated modification, namely a sugar modification called GlcNAcylation. In summary, developing a reproducible and efficient way to differentiate human ES cells into endoderm, as well as a thorough understanding of this key step, will allow us and others to elucidate the detailed set of molecular and biochemical events underlying this critical differentiation step, and will improve differentiation protocols.

Year 2

The goals of this proposal are to investigate endodermal differentiation and proliferation of human ES cells in culture. Endodermal differentiation is a necessary step towards making pancreatic beta cells, as well as other endodermal cells, such as liver cells. Pancreatic beta cells generated from human ES cells could be used to treat type I diabetes. In the past two years, we have incorporated human ES cell culture technology into our laboratory and have been able to replicate data obtained by other research groups. While several other research groups and companies around the world are focused on making pancreatic beta cells as quickly as possible, we strongly believe that a more detailed understanding of the biology of human eS cell differentiation into endoderm will help the optimization of this protocol. Therefore, we have focused our efforts on testing a number of variables in the initial step of creating definitive endoderm. We have found that different human ES cell lines have very different capacity to differentiate into endoderm under the same culture conditions. IN addition, we have recently focused our research effort on the post-translational modifications of key regulators of endoderm differentiation, and found a critical role for a poorly appreciated modification—namely a sugar modification called GlcNAcylation. In summary, developing a reproducible and efficient way to differentiate human ES cells into endoderm, as well as thorough understanding of this key step, will allow us and others to elucidate the detailed set of molecular and biochemical events underlying this critical differentiation step, and will improve differentiation protocols. We initiated a project on the role of post-translational modifications during hES cell differentiation into endodermal lineages, specifically on the GlcNAcylation sugar modification. We found that this modification appears to be important for endoderm formation in hES cell cultures. Identification of modified proteins is an important next step in understanding the mechanisms of this phenomenon and may ultimately provide a basis to develop assays for screening drugs that enhance endoderm/beta-cell formation.

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