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Skeletal muscle is an electrically and mechanically active tissue that contains highly oriented, densely packed myofibrils. The tissue has self-regeneration capacity upon injury, which is limited in the cases of volumetric muscle loss. Several regenerative therapies have been developed in order to enhance this capacity, as well as to structurally and mechanically support the defect site during regeneration. Among them, biomimetic approaches that recapitulate the native microenvironment of the tissue in terms of parallel-aligned structure and biophysical signals were shown to be effective. In this study, we have developed 3D printed aligned and electrically active scaffolds in which the electrical conductivity was provided by carbonaceous material (CM) derived from algae-based biomass. The synthesis of this conductive and functional CM consisted of eco-friendly synthesis procedure such as pre-carbonization and multi-walled carbon nanotube (MWCNT) catalysis. CM obtained from biomass via hydrothermal carbonization (CM-03) and its ash form (CM-03K) were doped within poly(ɛ-caprolactone) (PCL) matrix and 3D printed to form scaffolds with aligned fibers for structural biomimicry. Scaffolds were seeded with C2C12 mouse myoblasts and subjected to electrical stimulation during the in vitro culture. Enhanced myotube formation was observed in electroactive groups compared to their non-conductive counterparts and it was observed that myotube formation and myotube maturity were significantly increased for CM-03 group after electrical stimulation. The results have therefore showed that the CM obtained from macroalgae biomass is a promising novel source for the production of the electrically conductive scaffolds for skeletal muscle tissue engineering.

Synthetic polymers remain to be a major choice for scaffold fabrication due to their structural stability and mechanical strength. However, the lack of functional moieties limits their application for cell-based therapies which necessitate modification and functionalization. Blending synthetic polymers with natural components is a simple and effective way to achieve the desired biological properties for a scaffold. Herein, nanofibrous mats made of polycaprolactone (PCL) and egg white protein (EWP) blend were developed and further evaluated for use as a scaffold for tissue engineering applications. Homogeneous distribution of EWP was achieved throughout the nanofibrous mats, as shown by immunohistochemistry. ATR-FTIR analysis and contact angle measurements have further confirmed the presence of EWP on the surface of the samples. The swelling test showed that PCL/EWP nanofibers have higher water uptake than PCL nanofibrous mats. Also, EWP addition on the nanofibrous mats resulted in an increase in the tensile strength and Young's modulus of the mats, indicating that the presence of protein can greatly enhance the mechanical properties of the mats. A significantly higher, more uniform, and dispersed cell spreading was observed on days 7 and 14 than that on neat PCL mats, demonstrating the importance of providing the required cues for cell homing by the availability of EWP. Hence, EWP is shown to be a simple and low-cost source for the functionalization of PCL nanofibrous mats. EWP is, therefore, a facile candidate to enhance cellular interactions of synthetic polymers for a wide range of tissue engineering applications.

A key challenge in skeletal muscle tissue engineering is the choice of a proper scaffolding material as it should demonstrate elastic behavior to withstand and support the dynamic loading of the tissue microenvironment while being biodegradable and biocompatible. In this study, we tested the applicability of a novel biodegradable polyurethane (PU) elastomer chain extended with fibrinogen (Fib) to fulfill these requirements. Biodegradable polyurethane-fibrinogen (PU-Fib) elastomers were synthesized by step-wise condensation polymerization. Firstly, PU prepolymer was synthesized and then Fib was integrated into PU prepolymer during the second step of polymerization. The chemical, thermal, viscoelastic, mechanical and biodegradation properties of PU-Fib were characterized. FTIR-ATR spectrum showed amide bands specific to PU and Fib, DSC thermograms showed the suitable integration between the components. Dynamic mechanical analysis revealed Tg and Tα* transitions at 64.5 °C and 38.4 °C, respectively. PU and Fib had shown chemically compatible interactions and when compared to PCL, PU-Fib possessed viscoelastic properties more suitable to the native tissue. PU-Fib films were produced and seeded with C2C12 mouse myoblasts. Uniaxial cyclic stretch was applied to the cell seeded films for 21 d to mimic the native dynamic tissue microenvironment. Cell proliferation, viability and the expression of muscle-specific markers (immunofluorescent staining for myogenin and myosin heavy chain) were assessed. Myoblasts proliferated well on PU-Fib films; aligned parallel along their long edge, and express myogenic markers under biomimetic dynamic culture. It was possible to culture myoblasts with high viability on PU-Fib elastomeric films mimicking native muscle microenvironment.