3D_SpineTracker - Automated remyelination detection and classification of axons for sub-acutely injured spinal cord section images for temporal tracking of remyelination after stem cell treatment

Funding Type: 
SEED Grant
Grant Number: 
RS1-00416
ICOC Funds Committed: 
$0
Stem Cell Use: 
Embryonic Stem Cell
Public Abstract: 
3D_SpineTracker Spinal cord injury, or myelopathy, is a disturbance of the spinal cord that results in loss of sensation and mobility. It can be caused by trauma or disease, and it can be either complete or incomplete. Symptoms include loss of sensor or motor function, loss of reflexes, and loss of bladder or bowel control. A traumatic injury site is typically characterized by a loss of the myelin layer, which serves as an insulator for the axon and as a track along which regrowth can occur. Unmyelinated axon fibers do not regenerate. Preliminary studies in rats and mice have shown that human embryonic stem cells have the capability to differentiate into various cell types, aiding in the remyelination of axons. Animals undergoing this treatment have been shown to exhibit myelin sheaths restoration, which ultimately lead to regaining mobility. In order to quantify and control this process, it is crucial to develop methods for precise temporal tracking of remyelinated axons at the injury site. This can be done by distinguishing between two cell types, namely oligodendrocytes and Schwann cells, and by accurately computing the cell count for each type. In order to monitor cell development at the repair site, histological sections are taken at increasing time intervals, and the cell count is determined for each time step. The results of this quantitative analysis can be visualized in an animated, 3D rendering of the spinal cord injury site. One of the challenges for a statistical analysis is the large amount of histological sections and the large number of cells in each section. The development of a growth model as a quantitative analysis tool is a critical step toward a timely deployment of the treatment method in a human clinical trial.
Statement of Benefit to California: 
A stem cell is a type of cell found in both animals and humans that has the potential to develop into many different types of specialized cells in the body. Scientists found that stem cells have the potential to migrate toward injury sites and perform repair functions by replacing damaged cells with healthy cells. The SEED Grant will allow scientists to conduct research toward a better understanding of how stem cells differentiate into certain cell types, thus offering new treatment options ranging from tissue replacement to restoration of mobility functions after spinal cord injury. The citizens of the State of California will be among the first to benefit from this kind of research. Clinical trials on animals and humans are currently underway and are expected to lead to better insight into the processes that cause a stem cell to develop into a new cell type. More research is needed to fully understand the mechanisms of stem cell migration and differentiation. Preliminary results from animal studies indicate that new treatment options could be developed for spinal cord injury patients, in particular for those with recent post-traumatic conditions. It may be a long way until full recovery from spinal cord injury is possible, and even partial restoration of some mobility or other body functions, such as bowel or bladder control, would be considered a tremendous success. Computational methods will be developed to aid in this endeavor. The quantitative analysis and 3D visualization software to be developed under this grant will lead to an improved understanding of how stem cell derived progenitor cells migrate into the injury site to perform repair on the myelin sheaths that are critical for restoring function in axons.
Progress Report: 
  • The original goal of this project was to generate oocytes (eggs) from human embryonic stem (hES) cells in cell culture dishes in the laboratory. Such oocytes could be of use as vehicles to reprogram the DNA from cells of patients with life-threatening or debilitating conditions, thereby allowing generation of new lines of hES cells that are immune matched to the patient. The paucity of donated human oocytes precludes research using such material, and production of human oocytes from hES cells in the laboratory would in theory provide a limitless source of material.
  • Since our last progress report another CIRM-funded group, Dr. Renee Reijo Pera’s lab at Stanford University, has published exciting results demonstrating successful production of primordial oocytes from mouse ES (mES) and human ES (hES) cells. Consequently, during the remaining period of the award we propose to use the Reijo Pera methods for production of female germ line cells in our lab using H9 and HUES-9 female hES cells. After accomplishing this, we will introduce human mtDNA containing mutations that cause either a severe or mild reduction in oxidative phosphorylation (energy) production into H9 and HUES-9 hES cells and investigate the impact of the different mtDNA mutations on the ability of hES cells to form cells with characteristics of PGCs, then primordial oocytes in vitro and in vivo. An important related goal of this research is to investigate whether development of oocytes from ES cells could be used as a method to remove deleterious mtDNA mutations from the hES cell population, thereby improving the utility and possibly safety of derived cell types for therapeutic purposes.
  • During this reporting period we have focused on our approved revised research plan. We continued our efforts to generate embryonic female germ cells from human embryonic stem (hES) cells in vitro using methods reported in the literature at the end of 2009. Our revised plan included the new goal of using in vitro developed female embryonic germ cells (oocytes) as a resource to investigate how mitochondrial genomic DNA containing deleterious mutations is segregated during female germ cell development. As well as providing novel information about the biology of germ cell development, this research may provide important information relevant to development of safe methods for therapeutic cloning. We began by using two different female hES cell lines (HUES-6 and H9) to investigate whether we could use the reported methods to develop female germ cells from hES in our lab. A third female hES cell line (HUES-9) we originally intended to use was found to have a high propensity to gain an extra chromosome (become aneuploid) and was therefore not used. Despite following the reported methods that demonstrated in vitro differentiation of hES cells into female germ cells, we were unable to reproduce the previously reported results in our own lab. The reasons for this are currently unclear but may involve subtle, but important, differences in the methodology or materials we used.

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