This application seeks to improve our ability to use neural stem cells for brain and spinal cord repair. Human embryonic stem cells (hES) have the ability to produce many kinds of cells, including neural stem cells —cells that form the components of the brain and spinal cord. A fundamental problem is how hES cells maintain the ability to self-renew—to make more copies of themselves—while still retaining the capacity to become other cell types. Analogous to hES, an important question in utilizing hES-derived NSC is to determine how to maintain them in a state in which they can produce more stem cells while retaining the capacity to become (or differentiate to) the cell types needed for neural repair. For example, in stroke, one might want to use stem cells to replace lost neurons. However, for reasons that are unclear, neural stem cells transplanted into the brain often lose their ability to produce neurons. If we have a means to enhance this ability, we could ultimately benefit stroke patients. In our previous work, we have shown that mouse embryonic stem cells missing the gene, PTEN, have a much greater capacity to produce additional embryonic stem cells. Additionally, we have shown that mouse neural stem cells missing the PTEN gene also retain a greater self-renewal capacity as well as the ability to produce neurons than do normal neural stem cells, both in tissue culture as well as in the living animal. The protein that is made by the gene, PTEN, acts by inhibiting a molecular pathway, the PI3K/Akt pathway. In this proposal, we will discover whether this pathway is important for the self-renewal of hES as well as neural stem cells derived from hES. It is our belief that inactivating this pathway will inhibit the production of hES cells and neural stem cells, and that activating this pathway will enhance this production. We will then test whether promoting the pathway in hES-derived neural stem cells has meaningful consequences when we transplant the cells into the living (in vivo) mouse brain. First, we will test whether PTEN-deficient neural stem cells are better at replacing neurons than are normal neural stem cells following transplantation into the brains of mice that do not have the capacity to produce new neurons on their own. Next, we will determine if loss of PTEN function also enhances the ability of hES-derived neural stem cells to repair the brain following a stroke. In addition to the studies that specifically assess the PTEN/PI3K/Akt pathway, we will discover other genes and pathways that regulate the process of hES-derived neural stem cell self-renewal. High priority candidates from these studies will then be tested in our in vivo models. We believe the studies proposed here will have important implications on the ways that we use both transplanted as well as endogenous (the patients’ own) human neural stem cells in stroke and other disorders where replacement of brain or spinal cord cells is a therapeutic option.
Neurological disorders are the leading source of disability, and are not only devastating for individuals, but are an enormous social and financial burden that costs the State of California many billions of dollars per year. Currently there are no treatments that promote repair and recovery in the brain after any major neurological disorder, including stroke, degenerative disease and trauma. Research with stem cells derived from animals indicates that transplantation of stem cells can promote recovery of function in the brain after stroke and models of degenerative disease in experimental animals. This research suggests that human embryonic stem cell therapy may provide a way to promote repair and recovery in patients with brain disorders. Nevertheless, although the animal results look promising thus far, these animal stem cell studies have identified limitations in the procedure because most of the transplanted cells do not survive and integrate into the brain, and there is little understanding of the fate of the transplanted cells and the mechanisms that control this fate. The goals of this grant are: 1) to determine the precise molecular pathways that identify human embryonic stem cells at different stages of differentiation as they turn into neurons, or as they divide in a process of self-renewal; and 2) to identify the molecules that promote human embryonic stem cell survival, differentiation into different cell types including specific types of neurons, and integration into brain circuits to promote functional recovery after neurological conditions such as stroke or degenerative disease. The proposed studies will build on and draw from our work in mouse embryonic stems in which we have identified a key molecular pathway that promotes embryonic stem cell survival, differentiation and repair of the brain after stroke and other forms of cell death. This pathway is known as the “PI3K/Akt pathway”. The results from the studies proposed in this grant will identify molecules that promote human embryonic stem cell growth and survival and that may lead to repair of the human brain after various neurological disorders. If successful, the State of California and its citizens will benefit not only from the improvement of individual health and lifestyle afforded by the development of treatments for currently untreatable neurological conditions, but will benefit also financially as a whole from the economic impact of reduced costs for caring for disabled individuals, as well as from the development of novel medical technologies that will place California at the forefront of a new medical field with a global market.