Muscle Microbiopsy to Delineate Stem Cell Involvement in Young Patients: A Novel Approach for Children With Cerebral Palsy

Abstract

Cerebral palsy (CP), the single largest cause of childhood physical disability, is characterized firstly by a lesion in the immature brain, and secondly by musculoskeletal problems that progress with age. Previous research reported altered muscle properties, such as reduced volume and satellite cell (SC) numbers and hypertrophic extracellular matrix compared to Typically Developing (TD) children (>10 years). Unfortunately, data on younger CP patients are scarce and studies on SCs and other muscle stem cells in CP are insufficient or lacking. Therefore, it remains difficult to understand the early onset and trajectory of altered muscle properties in growing CP children. Because muscle stem cells are responsible for postnatal growth, repair and remodeling, multiple adult stem cell populations from young CP children could play a role in altered muscle development. To this end, new methods for studying muscle samples of young children, valid to delineate the features and to elucidate the regenerative potential of muscle tissue, are necessary. Using minimal invasive muscle microbiopsy, which was applied in young subjects under general anaesthesia for the first time, we aimed to isolate and characterize muscle stem cell-derived progenitors of TD children and patients with CP. Data of 15 CP patients, 3-9 years old, and 5 aged-matched TD children were reported. The muscle microbiopsy technique was tolerated well in all participants. Through the explant technique, we provided muscle stem cell-derived progenitors from the Medial Gastrocnemius. Via fluorescent activated cell sorting, using surface markers CD56, ALP and PDGFRa, we obtained SC-derived progenitors, mesoangioblasts and fibro-adipogenic progenitors, respectively. Adipogenic, skeletal and smooth muscle differentiation assays confirmed the cell identity and ability to give rise to different cell types after appropriate stimuli. Myogenic differentiation in CP SC-derived progenitors showed enhanced fusion index and altered myotube formation based on MYOSIN HEAVY CHAIN expression, as well as disorganization of nuclear spreading, which were not observed in TD myotubes. In conclusion, the microbiopsy technique allows more focused muscle research in young CP patients. Current results show altered differentiation abilities of muscle stem cell-derived progenitors and support the hypothesis of their involvement in CP-altered muscle growth.