PHD Student University of California, Irvine, United States
Abstract: Early-onset myopathy, areflexia, respiratory distress and dysphagia (EMARDD) is a highly fatal skeletal muscle disorder that does not have available cures. EMARDD is a poorly understood disease that is the result of a loss of function mutation in the MEGF10 gene, a membrane-bound protein linked to muscle stem cell support. In EMARDD patients, muscle stem cells (SCs) have impaired myogenic capacities, and thus skeletal muscle myogenesis is stunted. To combat EMARDD, SC transplantation therapies offer an appealing avenue, and induced pluripotent stem cells can be differentiated to skeletal muscle before transplantation to ensure a patient-specific treatment. However, pluripotent derived skeletal muscle offers a relatively weak treatment since less than one percent of transplanted cells persist as PAX7+ stem cells after engraftment. Helping support the retention of transplanted SCs as a quiescent stem cell pool is paramount to addressing EMARDD and other skeletal muscle disorders. We set out to overexpress MEGF10 to improve the retention of quiescent transplanted SCs. We incorporated a Tet-On system of doxycycline-driven inducible overexpression into pluripotent stem cells using CRISPR gene editing. Tet-On pluripotent derived PAX7 cells were transplanted in vivo, and MEGF10 was overexpressed at different timepoints (Day0-10, Day10-20, Day20-30, or Day0-30), to determine the kinetics of MEGF10 expression for PAX7 cell support. We found that after MEGF10 induction for the first 10 days following transplantation resulted in more human myofibers and better SC retention and support. Our work indicates that support of transplanted stem cells after engraftment leads to their long-term stability. To showcase this improved PAX7 support in MEGF10 treated groups, we plan to conduct reinjury assays that demonstrate their predicted improved regenerative abilities. We have collected spatial transcriptomic data on engrafted muscle stem cells with and without MEGF10 treatment at late regenerative timepoints to compare the PAX7 and myofiber transcriptomic landscapes. With this spatial RNA-seq data, we hope to elucidate the dynamic mechanism of MEGF10-mediated muscle stem cell support in vivo.
Funding Source: UCI MAXIMUS T32 Pre-Doctoral Training Fellowship
