Title : The impact of decellularization methods on human muscle extracellular matrix
Abstract:
Extracellular matrix (ECM) obtained after cell depletion (decellularization process) has been recently appointed as the best system to recapitulate fundamental aspects of the native tissue structure and soluble factors. In this work we propose the decellularization method as tool to study ECM composition of human skeletal muscle tissue. In order to balance both cell removal and the maintenance of the tissue structure and biochemical composition, we compared the efficiency of two decellularization methods. The detergent-enzymatic protocol involved cycles of sodium deoxycholate (SDC), the enzyme DNaseI and water. The non-detergent protocol expected the use of the chemical latrunculin B and water. DNA and ECM components quantification, electron microscopy imaging, immunohistochemistry and immunofluorescence analysis were performed. The use of SDC ensured the desired cell removal, the maintenance of the extracellular matrix (ECM) structural proteins such as laminin and fibronectin, but the sulfated glycosaminoglycans (GAGs) strongly decreased. On the contrary, Latrunculin B treatment allows a much better
maintenance of GAGs in respect to the shape of the muscle fibers. GAGs interact with ECM constituents, adhesion molecules and growth factors that play a crucial role in muscle development and maintenance. These results demonstrated that SDC and Latruculin B treatment can provide a good tridimensional mold for ex vivo modeling of healthy and pathological muscle. Duchenne muscular dystrophy is an example of muscle pathology in which GAGs are dysregulated and they could represent a new therapeutic target.
