Defining Heterogeneity of Human Embryonic Stem Cells

Funding Type: 
SEED Grant
Grant Number: 
RS1-00333
ICOC Funds Committed: 
$0
Disease Focus: 
Amyotrophic Lateral Sclerosis
Neurological Disorders
Spinal Cord Injury
Neurological Disorders
Stem Cell Use: 
Embryonic Stem Cell
Public Abstract: 
Being pluripotent, human embryonic stem cells (hESCs) have the potential to act as a source of cells for regenerative therapies. But before these potential applications of hESCs can be effectively pursued, an understanding of the complex cellular relationships existing within hESC cultures and factors involved in the maintenance of hESC properties is required. hESC lines are known to be morphologically and phenotypically heterogeneous. We propose to select an antibody diversity library directly on live hESCs to identify a panel of monoclonal antibodies that recognize distinct hESC subpopulations, and to further characterize these subpopulations with regards to pluripotency and capacity for self-renewal. These antibodies will be of broad interest to laboratories that are either utilizing and optimizing the existing hESC lines or developing new hECS lines. These antibodies can be used in the future to ensure quality of hECS cultures, develop sublines that are more suitable for regenerative therapy, and identify corresponding antigens for functional studies of hESCs.
Statement of Benefit to California: 
This study aims to develop antibodies that recognize distinct subpopulations of human embryonic stem cells (hESCs), and to further characterize these subpopulations with regards to pluripotency and capacity for self-renewal. These antibodies will be of broad interest to laboratories that are utilizing or optimizing existing hESC lines, or developing new hESC lines. This study benefits California and its citizens because the knowledge and reagents generated can be used in the future to ensure quality of hESC cultures, and develop sublines that are more suitable for regenerative therapy.
Progress Report: 
  • A main goal of research in our laboratory is to identify strategies to promote neural repair in spinal cord injury and related neurological conditions. On the one hand, we have been using mouse models of spinal cord injury to study a long-standing puzzle in the field, namely, why axons, the fibers that connect nerve cells, do not regenerate after injury to the brain and the spinal cord. On the other hand, relevant to this CIRM SEED grant, we have started to explore the developmental and therapeutic potential of human embryonic stem cells (hESCs) for neural repair. We do this by first developing a method to genetically manipulate a HUES line of hESCs. The advent of hESCs has offered enormous potential for regenerative medicine and for basic understanding of human biology. To attain the full potential of hESCs as a tool both for therapeutic development and for basic research, we need to greatly enhance and expand our ability to genetically manipulate hESCs. A major goal for our SEED grant-sponsored research is to establish methods to genetically manipulate the HUES series of hESC lines, which are gaining wide utility in the research community due to the advantages on their growth characteristics over previously developed hESC lines. The first gene that we targeted in HUES cells, Fezf2, is critical for the development of the corticospinal tract, which plays important roles in fine motor control in humans and hence represents an important target for recovery and repair after spinal cord injury. By introducing a fluorescent reporter to the Fezf2 locus, we are now able to monitor the differentiation of hESCs into Fezf2-expressing neuronal lineages. This work has been published. A second goal is to start to explore the developmental and therapeutic potential of these cells and cells that derived from these cells in the brain and spinal cord. We are currently utilizing the cell line genetically engineered above to develop an efficient method to differentiate HUES cells into subcerebral neurons. Results so far have been encouraging. Efforts are also underway to overexpress Fezf2 as a complementary approach to drive the differentiation of HUES cells into specific neuronal types. Together, these studies will lay down the foundation for therapeutic development with HUES cells and their more differentiated derivatives for neurological disorders including spinal cord injury where neural regeneration can be beneficial. The CIRM SEED grant has allowed us to pursue a new, exciting path of research that we would have not pursued had we not been awarded the grant. Furthermore, the CIRM funded research has opened a new window of opportunity for us to explore genetic engineering of hESCs to model human neurological conditions in future.

© 2013 California Institute for Regenerative Medicine