Increasing the number and survival of hESC and the derived lineages in tissue culture.

Funding Type: 
SEED Grant
Grant Number: 
RB2-01645
ICOC Funds Committed: 
$0
Stem Cell Use: 
Embryonic Stem Cell
iPS Cell
Public Abstract: 
Our research for methods to protect the cells during aging revealed that N-t-butyl hydroxylamine (NtBHA, active form of a drug in clinical trial for stroke) and methylene blue (MB, a well known drug) significantly extend the vigor and lifetime of cells. MB or NtBHA at very low concentrations delay senescence of the cells; increase the number of the cells by 6-7 million cells/week; enhance cellular resistance to oxidative stress and toxic agents; and improve energy production. When fed to old rats MB and NtBHA enhance cognitive function and improve muscle strength. Preliminary observations suggest that NtBHA and MB, accomplish this, in part, by improving the quality of the mitochodnria, which are the cellular source for energy. MB and NtBHA also lowered the levels of oxidative damage to the cell compnents. We will use MB and NtBHA to improve the maintenance of hESC and the derived neuronal lineages in culture. We are expecting MB and NtBHA to improve the culturing of hESC and derived cells when added to the established growth media. Thus, we hypothesize that maintaining hESC and derived neuronal lineages with MB or NtBHA considerably increases the total number of cells, efficiency of derivation, and survival in the tissue culture conditions. If the goals of this proposal are successfully accomplished, then the quality and the number of the available stem cell units for cell therapy should improve. Additionally, the storage of stem cells at –80 °C will be made safer. A future longer-term objective of our proposed research is to test whether MB and NtBHA promote the function of endogenous stem cells when administered in experimental animal models. The basis for this hypothesis is that MB and NtBHA enhanced the cognitive function and improved muscle strength in rats. A one possible explanation for this effect is that MB and NtBHA, in part, improve the function of endogenous stem cells in the rat’s tissues (e.g. increased the secretion of specific hormones). Additionally, MB and NtBHA may enhance the metabolic activity of the somatic cells adjacent to the stem cells, which, based on recent research, should preserve the function of the endogenous stem cells in the rat’s tissues. Promoting the function of endogenous stem cells has been proposed as a therapeutic strategy to prevent and cure several diseases in human. A pharmacological approach to enhancing the function of native stem cells in tissue (e.g. the use of MB and NtBHA) to prevent disease (e.g. Alzheimer disease) may complement stem-cell transplantation therapy.
Statement of Benefit to California: 
In order for the objectives of Proposition 71 to be accomplished and patients in Californian to benefit from the use of human embryonic stem cells (hESC) to regenerate tissues to replace tissues damaged by disease and injury, researchers must first be able to grow and maintain sufficient numbers of healthy and pure hESC and their derived lineages. Current hESC research is impeded by the technical difficulties and safety concerns in getting hESC, especially the derived lineages, to grow in adequate numbers under laboratory conditions and being able to maintain the cells in a healthy state. We are proposing to test the effect of two compounds, which have been shown to improve the health of aging cells from old rats, without harmful side effects. These two compounds appear to accomplish this by preventing the dysfunction of key organelles known as mitochondria, which produce energy that cells use for all biological processes, including cell division, cell repair, and the production of enzymes and hormones. If our research is successful, it will help speed up basic hESC research, use of hESC in the private biotechnology sector, and cell replacement therapy in California. This research, in conjunction with the ongoing research at many of California’s institutes, should help position California at the forefront of stem cells research.
Progress Report: 
  • The ultimate goal of the proposed study is to identify approaches to increase the production of therapeutically useful blood cells from human ESCs and patient-specific iPSCs. Currently, bone marrow transplantation is the best way to cure many blood-related disorders, such as sickle cell anemia, thalassemia, and blood cancers like leukemia. Furthermore, blood transfusion is an effective way to rapidly counteract blood cell loss due to ablative treatments, such as chemotherapy and radiation therapy. Unfortunately, the limiting factor in transplantation and transfusion treatments is the lack of matched donors. The ability to producing unlimited numbers of blood stem cells and/or functioning differentiated blood cells from human ESCs and patient-derived iPSCs will greatly improve the opportunity of such treatments. Transcription factors play important roles in regulating cell proliferation and differentiation. RUNX1 is a transcription factor that is expressed in blood cells and regulates the expression of many blood cell related genes. Therefore, the specific aim of our studies is to examine the effect of RUNX1 on blood cell formation, expansion, and differentiation from human embryonic stem cells and induced pluripotent stem cells. During the first year of funding period, we have established the cell culture and differentiation systems in our laboratory, generated necessary DNA constructs for the proposed studies, and produced transcription factors for testing the effect.
  • The ultimate goal of the proposed study is to identify approaches to increase the production of therapeutically useful blood cells from human ESCs and patient-specific iPSCs. Currently, bone marrow transplantation is the best way to cure many blood-related disorders, such as sickle cell anemia, thalassemia, and blood cancers like leukemia. Furthermore, blood transfusion is an effective way to rapidly counteract blood cell loss due to ablative treatments, such as chemotherapy and radiation therapy. Unfortunately, the limiting factor in transplantation and transfusion treatments is the lack of matched donors. The ability to producing unlimited numbers of blood stem cells and/or functioning differentiated blood cells from human ESCs and patient-derived iPSCs will greatly improve the opportunity of such treatments. Transcription factors play important roles in regulating cell proliferation and differentiation. RUNX1 is a transcription factor that is expressed in blood cells and regulates the expression of many blood cell related genes. Therefore, the specific aim of our studies is to examine the effect of RUNX1 on blood cell formation, expansion, and differentiation from human embryonic stem cells and induced pluripotent stem cells. During the second year of funding period, we have performed hematopoietic cell differentiation using both human ESCs and iPSCs in the presence and absence of this transcription factor. Our results indicate that this factor promotes the production of blood stem cells and progenitors. We will make additional amount of this factor and further confirm our initial finding.
  • The ultimate goal of the proposed study is to identify approaches to increase the production of therapeutically useful blood cells from human ESCs and patient-specific iPSCs. Currently, bone marrow transplantation is the best way to cure many blood-related disorders, such as sickle cell anemia, thalassemia, and blood cancers like leukemia. Furthermore, blood transfusion is an effective way to rapidly counteract blood cell loss due to ablative treatments, such as chemotherapy and radiation therapy. Unfortunately, the limiting factor in transplantation and transfusion treatments is the lack of matched donors. The ability to producing unlimited numbers of blood stem cells and/or functioning differentiated blood cells from human ESCs and patient-derived iPSCs will greatly improve the opportunity of such treatments. Transcription factors play important roles in regulating cell proliferation and differentiation. RUNX1 is a transcription factor that is expressed in blood cells and regulates the expression of many blood cell related genes. Therefore, the specific aim of our studies is to examine the effect of RUNX1 on blood cell formation, expansion, and differentiation from human embryonic stem cells and induced pluripotent stem cells. During the second year of funding period, we have performed hematopoietic cell differentiation using both human ESCs and iPSCs in the presence and absence of this transcription factor. Our results indicate that this factor promotes the production of blood stem cells and progenitors. We will make additional amount of this factor and further confirm our initial finding.

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