Role of p120-catenin in restricting reprogramming of human keratinocytes

Funding Type: 
Basic Biology II
Grant Number: 
RB2-01637
ICOC Funds Committed: 
$0
Disease Focus: 
Alzheimer's Disease
Neurological Disorders
Stem Cell Use: 
Embryonic Stem Cell
iPS Cell
Public Abstract: 
Adult human cells such as skin cells can be turn into “reprogrammed” stem cells that acquire properties of pluripotency - the ability to become multiple types of specialized cells of the body. These reprogrammed stem cells are called induced pluripotent stem cells (iPSCs). Generation of iPSCs may enable the development of new therapies for many human diseases such as Alzheimer's diseases, spinal cord injury, stroke, burns, heart disease, type I diabetes, osteoarthritis, and rheumatoid arthritis. For example, type I diabetes occurs when the pancreatic beta cells that produce insulin are damaged or die. The only current cure is a pancreatic transplant from a recently deceased donor, but the demand for transplants far outweighs the supply. If reprogramming becomes possible, skin cells can be easily taken from a patient with type I diabetes and converted to iPSCs which can then be grown into the insulin-producing beta cells. These beta cells genetically match the patient’s cells and can be transplanted back into the patient’s body without any risk of rejection. This would potentially cure the disease. Therefore, the reprogramming technology has enormous therapeutic potential. The most widely used method for reprogramming is delivering certain stem cell-associated genes or proteins called reprogramming factors (RFs) into adult human cells. The major drawback of this type of approach is the low efficiency of reprogramming, which compromises the clinical use of this powerful approach. Recently, it has been found that stimulation of cells with a protein called Wnt3a increases efficiency of the conversion of adult cells to iPSCs, but the efficiency is still too low for clinical use. However, adult cells containing less amount of a protein called p53 has higher efficiency of reprogramming. These observations suggest existence of barriers in the cell to restrict reprogramming. Our proposed study is to determine whether p120-catenin serves as a barrier to the reprogramming. To achieve this goal, we will use skin cells (keratinocytes) in which p120-catenin expression is deleted or reduced. We will then reprogram these cells by introducing RFs and/or Wnt, and examine the reprogramming efficiency. If the cells lacking p120-catenin or with low level of p120-catenin are more easily reprogrammed than are cells with normal level of p120-catenin, is would suggest that p120-catenin serves as barrier to the reprogramming. We will further determine how p120-catenin functions as a barrier by examining the interaction between p120-catenin and Wnt regulated events. Skin cells which are relatively easy to obtain and reprogram would be an ideal source of iPSCs. Successful completion of this study will provide information that helps develop new strategies such as inhibition of p120-catenin to improve the reprogramming efficiency. This would overcome existing hurdles that hamper the application of the technology useful in transplantation therapy of many human diseases.
Statement of Benefit to California: 
Chronic degenerative diseases, such as Alzheimer's diseases, type I diabetes, heart disease, osteoarthritis, rheumatoid arthritis, etc., afflict a significant number of California’s population. Today, donated organs or tissues which match host’s immune system are often used to replace ailing or destroyed tissue of the patient, but the need for transplantable tissues and organs far outweighs the available supply. Right now, over 21,000 patients in Californian are waiting for life-saving transplants. That's 21 percent of the more than 100,000 people waiting across our country. One third of them will die before receiving a transplant. Stem cells can give rise to a variety of cell types, which offer the possibility of a renewable source of replacement cells and tissues to treat degenerative diseases. Adult somatic cells can be converted to induced pluripotent stem cells (iPSCs). This process is called reprogramming. The reprogrammed iPSCs have the potential to become any type of cells in the body. The iPSCs are remarkably similar to embryonic stem cells and would offer all the benefits of embryonic stem cells without the controversial use of embryos. For example, it may become possible to generate healthy heart muscle cells from patient’s own tissue in the laboratory and then transplant those cells into patients with chronic heart disease without risk of immune rejection. However, the low efficiency of the reprogramming limited clinical application of this powerful method. Our study is aimed at determining whether p120-catenin serves as barrier to prevent somatic cells from dedifferentiation (reprogramming). This study will uncover the role for p120-catenin in regulating somatic reprogramming and provide information for developing strategy about how to make reprogramming more efficient so that iPSCs can be generated on a large scale for transplantation, which is beneficial to the patients with degenerative diseases in California. The improved reprogramming technology can be commercialized by the biotech industry in California to generate revenue and create new job opportunities.
Progress Report: 
  • We are interested in identifying soluble protein factors in blood which can either promote or inhibit stem cell activity in the brain. Through a previous aging study and the transfer of blood from young to old mice and vice versa we had identified several proteins which correlated with reduced stem cell function and neurogenesis in young mice exposed to old blood. Over the past year we studied two factors, CCL11/eotaxin and beta2-microglobulin in more detail in tissue culture and in mice. We could demonstrate that both factors administered into the systemic environment of mice reduce neurogenesis in a brain region involved in learning and memory. We have also begun to test the effect of these factors on human neural stem cells and we started experiments to try to identify protein factors which can enhance neurogenesis.
  • While age-related cognitive dysfunction and dementia in humans are clearly distinct entities and affect different brain regions, the aging brain shows the telltale molecular and cellular changes that characterize most neurodegenerative diseases. Remarkably, the aging brain remains plastic and exercise or dietary changes can increase cognitive function in humans and animals, with animal brains showing a reversal of some of the aforementioned biological changes associated with aging. We showed recently that blood-borne factors coming outside the brain can inhibit or promote adult neurogenesis in an age-dependent fashion in mice. Accordingly, exposing an old mouse to a young systemic environment or to plasma from young mice increased neurogenesis, synaptic plasticity, and improved contextual fear conditioning and spatial learning and memory. Preliminary proteomic studies show several proteins with stem cell activity increase in old “rejuvenated” mice supporting the notion that young blood may contain increased levels of beneficial factors with regenerative capacity. We believe we have identified some of these factors now and tested them on cultured mouse and human neural stem cell derived cells. Preliminary data suggest that these factors have beneficial effects and we will test whether these effects hold true in living mice.
  • Cognitive function in humans declines in essentially all domains starting around age 50-60 and neurodegeneration and Alzheimer’s disease seems to be inevitable in all but a few who survive to very old age. Mice with a fraction of the human lifespan show similar cognitive deterioration indicating that specific biological processes rather than time alone are responsible for brain aging. While age-related cognitive dysfunction and dementia in humans are clearly distinct entities the aging brain shows the telltale molecular and cellular changes that characterize most neurodegenerative diseases including synaptic loss, dysfunctional autophagy, increased inflammation, and protein aggregation. Remarkably, the aging brain remains plastic and exercise or dietary changes can increase cognitive function in humans and animals. Using heterochronic parabiosis or systemic application of plasma we showed recently that blood-borne factors present in the systemic milieu can rejuvenate brains of old mice. Accordingly, exposing an old mouse to a young systemic environment or to plasma from young mice increased neurogenesis, synaptic plasticity, and improved contextual fear conditioning and spatial learning and memory. Unbiased genome-wide transcriptome studies from our lab show that hippocampi from old “rejuvenated” mice display increased expression of a synaptic plasticity network which includes increases in c-fos, egr-1, and several ion channels. In our most recent studies, plasma from young but not old humans reduced neuroinflammation in brains of immunodeficient mice (these mice allow us to avoid an immune response against human plasma). Together, these studies lend strong support to the existence of factors with beneficial, “rejuvenating” activity in young plasma and they offer the opportunity to try to identify such factors.
  • Cognitive function in humans declines in essentially all domains starting around age 50-60 and neurodegeneration and dementia seem to be inevitable in all but a few who survive to very old age. Mice with a fraction of the human lifespan show similar cognitive deterioration indicating that specific biological processes rather than time alone are responsible for brain aging. While age-related cognitive dysfunction and dementia in humans are clearly distinct entities and affect different brain regions the aging brain shows the telltale molecular and cellular changes that characterize most neurodegenerative diseases including synaptic loss, dysfunctional autophagy, increased inflammation, and protein aggregation. Remarkably, the aging brain remains plastic and exercise or dietary changes can increase cognitive function in humans and animals, with animal brains showing a reversal of some of the aforementioned biological changes associated with aging. Using heterochronic parabiosis we showed recently that blood-borne factors present in the systemic milieu can inhibit or promote adult neurogenesis in an age-dependent fashion in mice. Accordingly, exposing an old mouse to a young systemic environment or to plasma from young mice increased neurogenesis, synaptic plasticity, and improved contextual fear conditioning and spatial learning and memory. Over the past three years we discovered that factors in blood can actively change the number of new neurons that are being generated in the brain and that local cells in areas were neurons are generated respond to cues from the blood. We have started to identify some of these factors and hope they will allow us to regulate the activity of neural stem cells in the brain and hopefully improve cognition in diseases such as Alzheimer's.

© 2013 California Institute for Regenerative Medicine