Adult stem cells function throughout our life to replenish dying cells and regenerate damaged tissues. This unique ability of adult stem cells originates from their capacity to divide indefinitely and to give rise to specialized cells, processes known as stem cell self-renewal and differentiation. Understanding how stem cells self-renew and differentiate will have broad implications in regenerative medicine and cancer treatment. For example, decreased self-renewal or derailed differentiation of normal stem cells leads to tissue degeneration, while increased self-renewal of cancer stem cells results in tumorigenesis. We aim to identify the molecular regulators that control the self-renewal and differentiation of adult stem cells. As mammals age, the capacity of adult stem cells to self-renew declines and stem cell differentiation is dysregulated. Thus, we propose to use aging as a platform to understand the molecular network regulating stem cell self-renewal and differentiation. Identification of these molecular regulators holds promise for new avenues of regenerative medicine and cancer therapy.
Most studies on adult stem cells take advantage of the power of mouse genetics. However, one limitation of these studies is that it is not clear whether human adult stem cells act the same way as mouse stem cells. We will take advantage of a culture condition of human adult stem cells that mimics their living condition within the tissue and a system that permits functional evaluation of human stem cells under physiological conditions. These studies directly dissect the molecular basis underlying the self-renewal and differentiation of human adult stem cells, bringing us one step closer to therapeutic treatments in humans.
Statement of Benefit to California:
This proposal addresses the major gaps in our knowledge of adult stem cells, which form several research focuses of this RFA: human stem cell aging, molecular basis of human pluripotent stem cell (hPSC) self-renewal, and molecular determinants of stem cell fate decisions during hPSC differentiation. These knowledge are critical to our understanding of many devastating human diseases associated with aging. With CIRM's mission in mind, the proposed studies will be conducted in human adult stem cells. Therefore, the output of these studies can be directly translated into practice in human. This project will provide mechanistic insight into adult stem cell biology and significantly enhance the development of stem cell-based therapeutics for human diseases in California, such as cancer and tissue degenerative diseases.
The aged community in California will benefit most from the proposed research. The size of this community is increasing, as the baby boomers begin to enter their 60’s. The younger community will also benefit from these studies through prevention of the
diseases of aging. Developing treatments or preventions of diseases of aging will greatly improve the quality of life of aged people, allow younger people to age gracefully, and significantly lessen the healthcare burden in California. In addition, many students will be trained in this cutting-edge research and drive the progress of proposed studies. This research activity will provide an excellent training ground for the next generation of scientists in California.
This proposal is focused on the identification of genetic regulators of hematopoietic stem cell (HSC) aging. While it is well documented that the self-renewal and differentiation properties of HSCs change with age, the molecular mechanisms remain to be determined. In Aim 1, the applicant proposes to examine the role of a mitochondrial protein in regulating the self-renewal of human HSCs and their capacity for engraftment in immunodeficient mice. In Aim 2, the applicant will investigate the role of this protein in the HSC niche by testing the effects of its overexpression in stromal cells on co-cultured HSC self-renewal. Finally, in Aim 3, the applicant proposes a genome-wide screen for genes up- or downregulated as a result of aging or caloric restriction (a known lifespan enhancer), followed by testing of candidate genes for effects on human HSC self-renewal.
Reviewers agreed that this proposal addresses a major unsolved problem but felt that its potential impact is limited by flaws in its experimental design. For example, the overall goal is to understand human HSC aging but no experiments actually involve aged cells. A reviewer cautioned that cell proliferation, the main readout proposed, doesn’t necessarily share the same mechanisms as cell aging and recommended evaluating longer-term outcomes. Reviewers did not find the proposal particularly innovative or creative. One commented that the experimental approaches would provide data that is largely descriptive and lacking focus on underlying mechanisms.
Reviewers raised a number of concerns about the research plan. One reviewer found the proposal too simplistic, noting that it doesn’t consider other potentially important factors in HSC aging, such as oxidative stress, telomerase activity and the other subtypes of the mitochondrial protein under investigation. This reviewer was particularly concerned that the presence of two of these subtypes in the mitochondria of HSCs could complicate the interpretation of results. Another reviewer questioned the feasibility of Aim 1, given that the lentiviral transduction efficiency of HSCs is not provided and could be quite low. One reviewer noted that the in vitro co-culture studies with human HSC overexpressing the mitochondrial protein would yield data that only measures effects on the relatively more mature progenitor cells rather than on HSC and questioned the value of this approach. In addition, this aim lacks experimental details for HSC transplant into immunodeficient mice. Reviewers questioned the approach of Aim 2, calling the hypothesis speculative and the rationale unclear. Another reviewer suggested measuring oxidative stress in HSCs in Aim 2, rather than just proliferation, as this would more directly address the hypothesis. Reviewers found Aim 3 to be overambitious and lacking detail. One noted that the applicant describes array data in the preliminary results and was unclear how the strategy for Aim 3 would differ. Two other reviewers felt that the applicant significantly underestimated the resources and time required to complete the studies proposed in Aim 3.
Reviewers did not find the preliminary data compelling. One was surprised at the omission of data demonstrating expression of the mitochondrial protein in HSCs. Another reviewer would have appreciated more detail about the phenotype of the mitochondrial protein knock out mouse. This reviewer noted that the effect on hematopoiesis in this knock out mouse appears to be quite minimal and thus the overall hypothesis that the mitochondrial protein keeps HSCs in a youthful state is not convincing. Finally, the reviewer cautioned that the osteoblast might not be the key component of the hematopoietic niche, in which case the irradiation data would not be relevant to the proposal.
Reviewers praised the applicant as an exceptional young investigator with an impressive publication record. However, they noted an absence of collaborators and suggested that additional expertise in the areas of hematological assays, HSC transplant and high-throughput screening might the aid the proposal.
Overall, reviewers found this proposal to be overly simplistic and raised a number of concerns about the research plan that lead them to question its feasibility and potential impact.