Abstract
Duchenne Muscular Dystrophy (DMD) is caused by mutations in the gene coding for dystrophin, which functions
to maintain muscle fiber structure and function in the whole body by preventing it from being damaged during
muscle contraction. Presently, there is no definitive treatment for DMD patients, and current therapies focus on
prolonging survival and improving quality of life. Definitive treatment will require that functional dystrophin protein
is restored in all affected muscle groups. Possible approaches to restoring dystrophin expression in muscle fibers
include cell therapy.We hope that transplantation of a particular type of cell capable of regenerating muscle will
help us to develop new therapeutic approaches to this disease. Muscle stem cells, termed satellite cells, isolated
from healthy donors or patients should be able to provide dystrophin and repair muscle damage in DMD patients.
For efficient DMD therapy, satellite cells which maintain the ability to self-renew are also necessary. However,
satellite cells are rare (2-7% of all muscle nuclei) and often difficult to isolate. In addition, efficient myogenic
differentiation and the scale-up of myogenic differentiation remain elusive and must be developed further in order
to generate effective cell-based therapy for chronic muscle diseases. Patient-derived induced pluripotent stem
cells (iPSCs) are the ideal cell source to obtain an unlimited number of myogenic cells that escape immune
rejection after engraftment. However, the failure of systemic delivery of the injected cells has hindered practical
application in patients. Our long-term goals are to develop an effective supply of satellite cells for cell therapy, to
find an efficient systemic delivery method that can treat all affected muscles, and to enable the injected cells to
self-renew and reach an effective mass. The specific aims of this application: 1. To determine possible
interactions between CD24-expressing satellite cells and capillary endothelial cells via real-time imaging and 3-
D imaging after our recently established whole muscle tissue clearing protocol. 2. To develop systemic delivery
methods for satellite cells and hiPSC-derived myogenic progenitor cells in combination with genetic modification,
including transduction of an extravasation factor, CD24, and Mannitol, an agent for enhancing vascular
permeabilization. 3. To develop systemic delivery methods for satellite cells via Extracellular Vesicle (EV)-
mediated CD24 transfer. In combination with gene and protein transduction, this concerted approach will help
us to make satellite cells that can be systemically transplanted into patients for a definitive cure of DMD. The
proposed specific aims will test whether systemically-injected satellite cells can be delivered to a target injured
muscle and contribute to muscle regeneration as well as to the satellite cell pool. The anticipated outcome of the
proposed specific aims will provide valuable insights for satellite cell-based cell therapies in muscular dystrophies
and other muscle diseases. Eventually, our experimental scheme will be directly applied to human satellite cells
isolated from DMD patients. The DMD-specific satellite cells stably transfected with CD24 and microdystrophin
will be able to produce enough satellite cells for autologous and systemic cell transplantation to DMD patients.