Micropatterning on PDMS by UV photolithography of an organic-inorganic hybrid polymer and its effect on human primary myoblasts growth - 20/03/24

Muscle modeling is getting more and more attention for a better understanding of numerous pathologies. In vivo models are the most accurate but due to financial and ethical considerations, ex vivo and in vitro models tend to draw more attention. In vitro models usually focus on specific caracteristics like reproducing the Extra Cellular Matrix (ECM), inducing the alignement of the myotubes and co-culturing with other cell types. Our aim was to propose an in vitro model of differentiated and aligned human skeletal muscle fibers.

To this end, we describe the micropatterning on a PDMS [poly(dimethylsiloxane)] substrate of EETMOS [2-(3.4 epoxycyclohexylethyltrimethoxysilane)] an organic-inorganic hybrid polymer, to produce a network of parallel lines that can be stretchable to optimise différenciation. EETMOS-based resin was synthesized by sol-gel process and polymerized using UV-photolithography. Human primary myoblasts were seeded onto the microstructurated substrate to be, after proliferation, stretched to differentiate into aligned myotubes. The effect of the spacing between the parallel lines was assessed by immunofluorescence.

After an optimization of the model, we ended up with 30μm large myotubes reached with a 75μm spacing. Then we functionalized the silicone with the use of silylated peptide ligands derived from extracellular matrix adhesion proteins to avoid detachment of the fibers from their support, during the stretching protocol (10% from L0). An improvement in the expression of sarcomere proteins was observed with stretching in relation to better differentiation of the myogenic progeniteur.

We successfully produced aligned myotubes which afterwards, reacted to mechanical stretching. The in vitro model that we propose would be a very useful tool to evaluate in a patient, from a microbiopsy, his muscular responses to mechanical stress.