Hydrogel-assisted electrospinning for fabrication of 3D compartmentalized nanofibrous scaffolds toward muscle tissue reconstruction

Abstract

The present thesis is about the fabrication process of various 3D compartmentalized, tailored nanofibrous scaffold for the reconstruction of muscle-tissue interface from in vitro to in vivo. Given the huge demand for the organ transplants and the limited number of donors, significant efforts have been devoted to tissue engineering and regenerative medicine, which provides the potential to artificially construct implantable tissues and organs. With the significant advances of 3D biofabrication processes, more complex-structured tissue constructs with heterogeneous architectures are realized, in comparison with the simple tissue-engineered substitutes (e.g. artificial hip joint). Among various engineered tissue constructs, the 3D muscle construct holds great potential in regenerating muscle tissues that comprise 45% of human body weight. Despite its importance in reconstructing muscle tissues, the development of 3D compartmentalized scaffolds for 3D muscle construct which enable physiological functions of muscle has still remained unrealized. In this thesis, a 3D compartmentalized nanofibrous scaffold that realizes both structural configuration and physical/mechanical functions was fabricated by developing hydrogel-assisted electrospinning (GelES) process. Unlike the conventional electrospinning that fabricates 2D flat nanofiber mats, the GelES process enables the production of 3D compartmentalized nanofibrous scaffolds having tailorable 3D macroscopic configurations. The increased degree of freedom in fabricating 3D compartmentalized nanofibrous scaffold by GelES is stemmed from the employment of 3D hydrogel structure as a grounded collector for electrospinning. In preparing the 3D hydrogel structure, we utilized the fused deposition modeling-based 3D printing for fabricating the mold for the 3D hydrogel structure. Thus, the high degree of freedom of 3D printing embodied the fabrication of a 3D compartmentalized nanofibrous scaffold by GelES. To profoundly understand the GelES process, the electrical properties of the hydrogel were investigated whether it can concentrate electric field developed during electrospinning, and thereby depositing the as-spun nanofibers on its exterior surface. Interestingly, the electrical properties of hydrogel were effective to concentrate the electric field, owing to the presence of mobile ions inside the hydrogel, and the considerable dielectric constant of the hydrogel. The numerical simulations modeling the mobile ions and dielectric constant presented the capability of hydrogel in concentrating electric field, which is 98 % similar to that of the conventional metal collector. Furthermore, the actual deposition of as-spun nanofibers on the 3D hydrogel collector was proven through experimental validations. One of the challenging problems of the GelES was the different transcription quality of the 3D compartmentalized nanofibrous scaffold depending on the 3D surfaces furnished in the 3D hydrogel collector. Herein, an effort to enhance the transcription quality of GelES on 3D compartmentalized surfaces is applied by conducting a parametric sensitivity analysis. With the categorization of 3D surfaces based on Gaussian curvature (K), we identified the difficulty in ensuring transcription quality is in applying GelES on 3D surfaces with K = 0 and K < 0. Then, we quantitatively evaluated the limited transcription quality on the 3D surfaces, with newly-defined measures, coverage ratio (CR), and corner profile fidelity (FC), respectively. Notably, the strategies to increase the transcription quality on 3D surfaces with K = 0 and K < 0 provided the same optimal condition. Therefore, a strategy to fabricate a 3D compartmentalized nanofibrous scaffold that delicately transcript any kinds of 3D surfaces by GelES was developed. At last, the two different types of 3D muscle constructs were recapitulated based on the GelES process. Firstly, the cardiac tissue interface was reconstructed in vitro, by employing the opposite strategy to increase the transcription quality on the 3D surface with K < 0. Considering that the aligned nanofibers are fabricated when the GelES failed to ensure transcription quality on the 3D surface with K < 0, we utilized the strategy to fabricate an aligned-random, heterogeneous NF mats, and subsequently stacking them to recapitulate the 3D cardiac anisotropy. Secondly, the skeletal muscle-nerve interface was reproduced by enveloping the 3D compartmentalized nanofibrous scaffold on the 3D bioprinted muscle construct and connecting it with in vivo tibial nerve of the rat. The 3D PCL nanofiber macrostructure enabled the robust innervation of nerve, similar to the fascia in the human body while providing permeable and flexible characteristics for the viability of 3D bioprinted muscle construct. In conclusion, hydrogel-assisted electrospinning (GelES) to fabricate a 3D compartmentalized nanofibrous scaffold in this thesis are expected to provide a versatile and powerful tool to reproduce physiologically relevant 3D muscle constructs. Furthermore, the practical applications on in vitro cardiac tissue engineering and in vivo innervated skeletal muscle tissue engineering demonstrated novel strategies for the development of regenerative medicine. From the abovementioned promising results, it is believed that the GelES to realize muscle-tissue interface has raised the level of completion of related research fields.