It is expected that research funded by the California Institute of Regenerative Medicine will result in the development of many new human embryonic stem cells. The properties of these cell lines will have to be verified. One of the main attributes of embryonic stem cells is their capacity to differentiate toward an unlimited number of cell fates. This is what will make them a powerful tool in future regenerative medicine. Therefore, we need new methods capable of quickly evaluating the response of many cell lines to many different environments. In this proposal, we will develop and test a new test bed to evaluate the capabilities of potential human embryonic stem cells: the chicken embryo.
Since chicken embryos develop outside the body in an egg, they are highly accessible to experimental manipulations. This enables us both to introduce reagents (ie., human embryonic stem cells) and to visualize the response of those reagents to their local environments as they happen. We propose to introduce human embryonic stem cells to six different organ systems during several stages of development. This will test the ability of these cells to respond to a large number of different environmental stimuli. Since different embryonic cell lines may have different capacities, we will test the abilities of seven different human embryonic stem cells. We will compare their response with that of partially differentiated cells that should have more limited differentiation capabilities. Transplanted cells will be fluorescently tagged so their migration can be traced by fluorescence microscopy. Antibodies and probes of molecular expression will be used to assess the response of these cell lines to different environments. Their origin (human or chicken) can also be confirmed with these methods using different antibodies and probes. This will help us to develop a set of formal criteria that to assess the response capability of hESC as they progressively become more differentiated.
To further understand molecular aspects of the cellular response, we will begin to characterize changes in molecular expression that take place as cells progress toward specific cell fates. This profile will enable us to begin to understand molecular factors which regulate cellular differentiation, so they can be harnessed for effective future regenerative medical applications. This last goal will serve to show the power of this technology, but will have to await a later stage of funding to be completed.
Statement of Benefit to California:
Human embryonic stem cells offer tremendous potential toward significant advances in the new age of regenerative medicine. These cells can be induced to differentiate along many different cell fates, providing the promise of tissue and/or organ replacement or supplementation. This approach offers great hope toward improving health care especially where tissues are damaged due to disease or injury. Ultimately, this approach could reduce health care costs and increase the well being of the general population.
We expect that many new embryonic stem cell lines will be derived with support from the California Institute for Regenerative Medicine. We would not be surprised if the ability of these putative stem cells to differentiate toward specific cell fates differed from cell line to cell line. Additionally, some cell lines may lose their differentiative capacity as they are kept in in vitro culture conditions.
Our research proposal aims to provide an easy and effective assay to test the pluripotentiality of these new putative human embryonic stem cell lines. In order for these cell lines to live up to their full potential and be useful in curing human diseases, we must understand their pluripotential properties. To date, assays of pluripotentiality have depended on 1) in vitro assays with limits in ascertaining the true developmental potential and 2) transplantation to mouse embryos which do not facilitate the high throughput analysis, essential to screen the myriad of generated cell lines. The latter assay also involves causing pain in a sentient being (the mother).
Chicken embryos develop in an egg, outside of the body. Hence it is easily accessible to experimentation including the delivery and observation of putative embryonic stem cells. The chicken embryo is a classic model of development and has been very well documented through years of research. It offers a myriad of developmental microenvironments which can be utilized to test the responsiveness of these new cell lines.
This research would contribute to the progress of stem cell research which ultimately could improve health care for everyone, worldwide. Since California is one of the first states to implement support for human embryonic stem cell research, our findings could also contribute to major economic advantages to the citizens of the state.