Sacrificial biomaterials in 3D fabrication of scaffolds for tissue engineering applications.

Three-dimensional (3D) printing technology has progressed exceedingly in the area of tissue engineering. Despite the tremendous potential of 3D printing, building scaffolds with complex 3D structure, especially with soft materials, still exist as a challenge due to the low mechanical strength of the materials. Recently, sacrificial materials have emerged as a possible solution to address this issue, as they could serve as temporary support or templates to fabricate scaffolds with intricate geometries, porous structures, and interconnected channels without deformation or collapse. Here, we outline the various types of scaffold biomaterials with sacrificial materials, their pros and cons, and mechanisms behind the sacrificial material removal, compare the manufacturing methods such as salt leaching, electrospinning, injection-molding, bioprinting with advantages and disadvantages, and discuss how sacrificial materials could be applied in tissue-specific applications to achieve desired structures. We finally conclude with future challenges and potential research directions.

Microtube embedded hydrogel bioprinting for vascularization of tissue-engineered scaffolds.

Vascular tissue engineering has been considered promising as one of the alternatives for viable artificial tissues and organs. Macro- and microscale hollow tubes fabricated with various techniques have been widely studied to mimic blood vessels. To date, the fabrication of biomimetic capillary vessels with sizes ranging from 1 to 10 µm is still challenging. In this paper, core-sheath microtubes were electrospun to mimic capillary vessels and were embedded in carboxymethyl cellulose/sodium alginate hydrogel for bioprinting. The results showed improved printing fidelity and promoted cell attachment. The tube concentration and tube length both had significant influences on filament size and merging area. Printed groups with higher microtube concentration showed higher microtube density, with filament/nozzle size ratio, and printed/designed grid area ratio closer to 100%. In the in vitro experiments, microtubes were not only compatible with human umbilical vein endothelial cells but also provided microtopographical cues to promote cell proliferation and morphogenesis in three-dimensional space. In summary, the microtubes fabricated by our groups have the potential for the bioprinting of vascularized soft tissue scaffolds.

Effects of Viscosities and Solution Composition on Core-Sheath Electrospun Polycaprolactone(PCL) Nanoporous Microtubes.

Vascularization for tissue engineering applications has been challenging over the past decades. Numerous efforts have been made to fabricate artificial arteries and veins, while few focused on capillary vascularization. In this paper, core-sheath electrospinning was adopted to fabricate nanoporous microtubes that mimic the native capillaries. The results showed that both solution viscosity and polyethylene oxide (PEO) ratio in polycaprolactone (PCL) sheath solution had significant effects on microtube diameter. Adding PEO into PCL sheath solution is also beneficial to surface pore formation, although the effects of further increasing PEO showed mixed results in different viscosity groups. Our study showed that the high viscosity group with a PCL/PEO ratio of 3:1 resulted in the highest average microtube diameter (2.14 µm) and pore size (250 nm), which mimics the native human capillary size of 1-10 µm. Therefore, our microtubes show high potential in tissue vascularization of engineered scaffolds.

Fabrication of Nanopores Polylactic Acid Microtubes by Core-Sheath Electrospinning for Capillary Vascularization.

There has been substantial progress in tissue engineering of biological substitutes for medical applications. One of the major challenges in development of complex tissues is the difficulty of creating vascular networks for engineered constructs. The diameter of current artificial vascular channels is usually at millimeter or submillimeter level, while human capillaries are about 5 to 10 µm in diameter. In this paper, a novel core-sheath electrospinning process was adopted to fabricate nanoporous microtubes to mimic the structure of fenestrated capillary vessels. A mixture of polylactic acid (PLA) and polyethylene glycol (PEO) was used as the sheath solution and PEO was used as the core solution. The microtubes were observed under a scanning electron microscope and the images were analyzed by ImageJ. The diameter of the microtubes ranged from 1-8 microns. The diameter of the nanopores ranged from 100 to 800 nm. The statistical analysis showed that the microtube diameter was significantly influenced by the PEO ratio in the sheath solution, pump rate, and the viscosity gradient between the sheath and the core solution. The electrospun microtubes with nanoscale pores highly resemble human fenestrated capillaries. Therefore, the nanoporous microtubes have great potential to support vascularization in engineered tissues.

Musculoskeletal Tissue Engineering Using Fibrous Biomaterials.

In tissue engineering, scaffolds should provide the topological and physical cues as native tissues to guide cell adhesion, growth, migration, and differentiation. Fibrous structure is commonly present in human musculoskeletal tissues, including muscles, tendons, ligaments, and cartilage. Biomimetic fibrous scaffolds are thus critical for musculoskeletal tissue engineering. Electrospinning is a versatile technique for fabricating nanofibers from a variety of biomaterials. However, conventional electrospinning can only generate 2D nanofiber mats. Postprocessing methods are often needed to create 3D electrospun nanofiber scaffolds. In this chapter, we present two novel electrospinning-based scaffold fabrication techniques, which can generate 3D nanofiber scaffolds in one-station process: divergence electrospinning and hybrid 3D printing with parallel electrospinning. These techniques can be applied for engineering tissues with aligned fiber structures.

Tunable 3D Nanofiber Architecture of Polycaprolactone by Divergence Electrospinning for Potential Tissue Engineering Applications.

The creation of biomimetic cell environments with micro and nanoscale topographical features resembling native tissues is critical for tissue engineering. To address this challenge, this study focuses on an innovative electrospinning strategy that adopts a symmetrically divergent electric field to induce rapid self-assembly of aligned polycaprolactone (PCL) nanofibers into a centimeter-scale architecture between separately grounded bevels. The 3D microstructures of the nanofiber scaffolds were characterized through a series of sectioning in both vertical and horizontal directions. PCL/collagen (type I) nanofiber scaffolds with different density gradients were incorporated in sodium alginate hydrogels and subjected to elemental analysis. Human fibroblasts were seeded onto the scaffolds and cultured for 7 days. Our studies showed that the inclination angle of the collector had significant effects on nanofiber attributes, including the mean diameter, density gradient, and alignment gradient. The fiber density and alignment at the peripheral area of the 45°-collector decreased by 21% and 55%, respectively, along the z-axis, while those of the 60°-collector decreased by 71% and 60%, respectively. By altering the geometry of the conductive areas on the collecting bevels, polyhedral and cylindrical scaffolds composed of aligned fibers were directly fabricated. By using a four-bevel collector, the nanofibers formed a matrix of microgrids with a density of 11%. The gradient of nitrogen-to-carbon ratio in the scaffold-incorporated hydrogel was consistent with the nanofiber density gradient. The scaffolds provided biophysical stimuli to facilitate cell adhesion, proliferation, and morphogenesis in 3D.

Engineering the hard-soft tissue interface with random-to-aligned nanofiber scaffolds.

Tendon injuries can be difficult to heal and have high rates of relapse due to stress concentrations caused by scar formation and the sutures used in surgical repair. Regeneration of the tendon/ligament-to-bone interface is critical to provide functional graft integration after injury. The objective of this study is to recreate the tendon-to-bone interface using a gradient scaffold which is fabricated by a one-station electrospinning process. Two cell phenotypes were grown on a poly-ε-caprolactone nanofiber scaffold which possesses a gradual transition from random to aligned nanofiber patterns. We assessed the effects of the polymer concentration, tip-to-collector distance, and electrospinning time on the microfiber diameter and density. Osteosarcoma and fibroblast cells were seeded on the random and aligned sections of scaffolds, respectively. A random-to-aligned cocultured tissue interface which mimicked the native transition in composition of enthesis was created after 96 h culturing. The results showed that the microstructure gradient influenced the cell morphology, tissue topology, and promoted enthesis formation. This study demonstrates a heterogeneous nanofiber scaffold strategy for interfacial tissue regeneration. It provides a potential solution for mimicking transitional interface between distinct tissues, and can be further developed as a heterogeneous cellular composition platform to facilitate the formation of multi-tissue complex systems.

Recent Progress of Fabrication of Cell Scaffold by Electrospinning Technique for Articular Cartilage Tissue Engineering.

As a versatile nanofiber manufacturing technique, electrospinning has been widely employed for the fabrication of tissue engineering scaffolds. Since the structure of natural extracellular matrices varies substantially in different tissues, there has been growing awareness of the fact that the hierarchical 3D structure of scaffolds may affect intercellular interactions, material transportation, fluid flow, environmental stimulation, and so forth. Physical blending of the synthetic and natural polymers to form composite materials better mimics the composition and mechanical properties of natural tissues. Scaffolds with element gradient, such as growth factor gradient, have demonstrated good potentials to promote heterogeneous cell growth and differentiation. Compared to 2D scaffolds with limited thicknesses, 3D scaffolds have superior cell differentiation and development rate. The objective of this review paper is to review and discuss the recent trends of electrospinning strategies for cartilage tissue engineering, particularly the biomimetic, gradient, and 3D scaffolds, along with future prospects of potential clinical applications.