Optimization of Skeletal Muscle Differentiation with the Incorporation of Biological Ligands

  • Khaled Abdel-Raouf Ahmed

Student thesis: Master's Thesis


Skeletal muscle tissue engineering has greatly flourished in the past few years due to advancements in extracting matrix compositions and engineering scaffolds. In addition the derivation of multiple viable cell sources has paved the way for greater translational applications. One particular aspect has yet to be adequately fulfilled, the creation of a skeletal muscle tissue model that is identical to human physiology and preserves the same architecture that can be used for pharmacological or genetic testing. In our goal to reaching such a model, skeletal muscle differentiation must be optimized for long term culture and sustained function. With this objective in mind, we identified eleven biological ligands that are regulators of myogenesis, and tested the impact on skeletal muscle differentiation of three cell types, mouse skeletal myoblasts (C2C12s), Human skeletal myoblasts, and skeletal myotubes derived from transdifferentiated fibroblasts at two concentrations 1ng/ml and 10ng/ml. Differentiation was evaluated based on immunofluorescent imaging of sarcomeric aActinin and MF20 to determine skeletal muscle area percent of images covered in myotubes and extent of multinucleation. A combination experiment, where cells were exposed to ligand combinations from four optimal ligands (10 ng/ml, each), determined from screening experiment, to test the additive effects of these proteins on C2C12 and transdifferentiated fibroblasts differentiation. Cells were quantified for myotubes elongation (area percent) and multinucleation and stained for anti-MyoD, anti-ki67, anti-mTOR and anti-CASQ1 as additional differentiators of maturation. We also created skeletal muscle tissues derived from transdifferentiated fibroblasts and C2C12s. These tissues gave us insight into alignment, compactness and extent of maturations skeletal myotubes reached.
Date of AwardApr 2017
Original languageAmerican English
SupervisorNicolas Christoforou (Supervisor)


  • Skeletal Muscle; Tissue Engineering; Biological Ligands.

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