Keywords: Cerebral Palsy, Skeletal Muscle, Mechanical Properties  

Background: Cerebral Palsy (CP) is caused by hypoxic-ischemic brain injury in utero. Spastic CP is the common subtype, and is characterized by passive resistance of limb motion. Current literature suggests that spasticity results from extensive fibrofatty deposits in the endomysial extracellular matrix (ECM). However, there are no studies that have explored the impact of fibrosis on the mechanical properties of CP myocytes and matrix. 

Objective: To address this gap in literature, we used correlated imaging to characterize the differences in mechanical properties present in muscle tissue from patients with CP compared to typically developing (TD) children. 

Methods: Quadriceps and hamstring muscle biopsies (1mm^3) were acquired from TD children (n = 6) and children with CP (n = 18). Samples were snap-frozen and stored at -80ºC. Select samples were chosen for sectioning using a Leica Cryostat at 50um and placed onto a microscope slide. The Leica SP8 DIVE Deep In Vivo Explorer 2-Photon microscope was used to image tissue sections. 2-Photon images were used as correlation maps for atomic force microscopy (AFM) experiments, which were conducted using Bruker BioAFM. Data analysis was performed using JPK NanoWizard®. 

Results: A total of 3 TD samples and 4 CP samples were included in the final analysis. 2-Photon images showed increased quantity and thickness of endomysium and perimysium in CP compared to TD. The overall stiffness of the myocytes and endomysium was decreased in CP compared to TD.  

Conclusion: While our results align with the current literature about increased collagen deposition in CP skeletal muscles, our study is the first to suggest that mechanical properties of this tissue have significant impairments. Future studies will explore proteomic changes in skeletal muscle as well as utilize single harmonic generation imaging (SHG) to assess structural changes of collagen bundles that may decrease overall stiffness.