Jeong Hun Park

A novel tissue-engineered trachea with a mechanical behavior similar to native trachea

A novel tissue-engineered trachea was developed with appropriate mechanical behavior and substantial regeneration of tracheal cartilage. We designed hollow bellows scaffold as a framework of a tissue-engineered trachea and demonstrated a reliable method for three-dimensional (3D) printing of monolithic bellows scaffold. We also functionalized gelatin sponge to allow sustained release of TGF-β1 for stimulating tracheal cartilage regeneration and confirmed that functionalized gelatin sponge induces cartilaginous tissue formation in vitro. A tissue-engineered trachea was then created by assembling chondrocytes-seeded functionalized gelatin sponges into the grooves of bellows scaffold and it showed very similar mechanical behavior to that of native trachea along with substantial regeneration of tracheal cartilage in vivo. The tissue-engineered trachea developed here represents a novel concept of tracheal substitute with appropriate mechanical behavior similar to native trachea for use in reconstruction of tracheal stenosis.

Unit cell-based computer-aided manufacturing system for tissue engineering

Scaffolds play an important role in the regeneration of artificial tissues or organs. A scaffold is a porous structure with a micro-scale inner architecture in the range of several to several hundreds of micrometers. Therefore, computer-aided construction of scaffolds should provide sophisticated functionality for porous structure design and a tool path generation strategy that can achieve micro-scale architecture. In this study, a new unit cell-based computer-aided manufacturing (CAM) system was developed for the automated design and fabrication of a porous structure with micro-scale inner architecture that can be applied to composite tissue regeneration. The CAM system was developed by first defining a data structure for the computing process of a unit cell representing a single pore structure. Next, an algorithm and software were developed and applied to construct porous structures with a single or multiple pore design using solid freeform fabrication technology and a 3D tooth/spine computer-aided design model. We showed that this system is quite feasible for the design and fabrication of a scaffold for tissue engineering.

Indirect three-dimensional printing of synthetic polymer scaffold based on thermal molding process.

One of the major issues in tissue engineering has been the development of three-dimensional (3D) scaffolds, which serve as a structural template for cell growth and extracellular matrix formation. In scaffold-based tissue engineering, 3D printing (3DP) technology has been successfully applied for the fabrication of complex 3D scaffolds by using both direct and indirect techniques. In principle, direct 3DP techniques rely on the straightforward utilization of the final scaffold materials during the actual scaffold fabrication process. In contrast, indirect 3DP techniques use a negative mold based on a scaffold design, to which the desired biomaterial is cast and then sacrificed to obtain the final scaffold. Such indirect 3DP techniques generally impose a solvent-based process for scaffold fabrication, resulting in a considerable increase in the fabrication time and poor mechanical properties. In addition, the internal architecture of the resulting scaffold is affected by the properties of the biomaterial solution. In this study, we propose an advanced indirect 3DP technique using projection-based micro-stereolithography and an injection molding system (IMS) in order to address these challenges. The scaffold was fabricated by a thermal molding process using IMS to overcome the limitation of the solvent-based molding process in indirect 3DP techniques. The results indicate that the thermal molding process using an IMS has achieved a substantial reduction in scaffold fabrication time and has also provided the scaffold with higher mechanical modulus and strength. In addition, cell adhesion and proliferation studies have indicated no significant difference in cell activity between the scaffolds prepared by solvent-based and thermal molding processes.

Development of a 3D bellows tracheal graft: mechanical behavior analysis, fabrication and an in vivo feasibility study

Artificial tracheal grafts should have not only enough compressive strength to maintain an open tracheal lumen, but also sufficient flexibility for stable mechanical behavior, similar to the native trachea at the implant site. In this study, we developed a new 3D artificial tracheal graft using a bellows design for considering its mechanical behavior. To investigate the mechanical behavior of the bellows structure, finite element method (FEM) analysis in terms of longitudinal tension/compression, bending and radial compression was conducted. The bellows structure was then compared with the cylinder structure generally used for artificial tracheal grafts. The FEM analysis showed that the bellows had outstanding flexibility in longitudinal tension/compression and bending. Moreover, the bellows kept the lumen open without severe luminal deformation in comparison with the cylinder structure. A three-dimensional artificial tracheal graft with a bellows design was fabricated using indirect solid freeform fabrication technology, and the actual mechanical test was conducted to investigate the actual mechanical behavior of the bellows graft. The fabricated bellows graft was then applied to segmental tracheal reconstruction in a rabbit model to assess its applicability. The bellows graft was completely incorporated into newly regenerated connective tissue and no obstruction at the implanted site was observed for up to 8 weeks after implantation. The data suggested that the developed bellows tracheal graft could be a promising alternative for tracheal reconstruction.

Human Inferior Turbinate: An Alternative Tissue Source of Multipotent Mesenchymal Stromal Cells

Objective Mesenchymal stromal cells (MSCs) are multipotent progenitor cells in adult tissues. Current challenges for the clinical application of MSCs include donor site morbidity, which underscores the need to identify alternative sources of MSCs. This study aimed to explore potential new sources of multipotent MSCs for use in tissue regeneration and the functional restoration of organs. Study Design Mixed methods research. Setting Tertiary care center. Subjects and Methods The authors isolated MSCs from human inferior turbinate tissues discarded during turbinate surgery of 10 patients for nasal obstruction. The expression of surface markers for MSCs was assessed by fluorescence-activated cell sorting. The differentiation potential of human turbinate mesenchymal stromal cells (hTMSCs) was analyzed by immunohistochemistry, reverse transcriptase–polymerase chain reaction, and Western blot analysis. Results Surface epitope analysis revealed that hTMSCs were negative for CD14, CD19, CD34, and HLA-DR and positive for CD29, CD73, and CD90, representing a characteristic phenotype of MSCs. Extracellular matrices with characteristics of cartilage, bone, and adipose tissue were produced by inducing the chondrogenic, osteogenic, and adipogenic differentiation of hTMSCs, respectively. The expression of neuron-specific markers in hTMSCs was confirmed immunocytochemically. Conclusion The hTMSCs represent a new source of multipotent MSCs that are potentially applicable to tissue engineering and regenerative medicine. The availability of differentiated adult cells will allow the development of an effective tissue regeneration method.

Effect of Pore Architecture on Oxygen Diffusion in 3D Scaffolds for Tissue Engineering

The aim of this study was to maximize oxygen diffusion within a three-dimensional scaffold in order to improve cell viability and proliferation. To evaluate the effect of pore architecture on oxygen diffusion, we designed a regular channel shape with uniform diameter, referred to as cylinder shaped, and a new channel shape with a channel diameter gradient, referred to as cone shaped. A numerical analysis predicted higher oxygen concentration in the cone-shaped channels than in the cylinder-shaped channels, throughout the scaffold. To confirm these numerical results, we examined cell proliferation and viability in 2D constructs and 3D scaffolds. Cell culture experiments revealed that cell proliferation and viability were superior in the constructs and scaffolds with cone-shaped channels.

Three-Dimensional Printing of Tissue/Organ Analogues Containing Living Cells

The technical advances of three-dimensional (3D) printing in the field of tissue engineering have enabled the creation of 3D living tissue/organ analogues. Diverse 3D tissue/organ printing techniques with computer-aided systems have been developed and used to dispose living cells together with biomaterials and supporting biochemicals as pre-designed 3D tissue/organ models. Furthermore, recent advances in bio-inks, which are printable hydrogels with living cell encapsulation, have greatly enhanced the versatility of 3D tissue/organ printing. Here, we introduce 3D tissue/organ printing techniques and biomaterials that have been developed and widely used thus far. We also review a variety of applications in an attempt to repair or replace the damaged or defective tissue/organ, and develop the in vitro tissue/organ models. The potential challenges are finally discussed from the technical perspective of 3D tissue/organ printing.

Human turbinate mesenchymal stromal cell sheets with bellows graft for rapid tracheal epithelial regeneration.

Rapid functional epithelial regeneration on the luminal surface is essential when using artificial tracheal grafts to repair tracheal defects. In this study, we imposed human turbinate mesenchymal stromal cell (hTMSC) sheets for tracheal epithelial regeneration, and then assessed their potential as a new clinical cell source. In vitro, hTMSCs sheets showed high capacity to differentiate into tracheal epithelium. We fabricated a poly(ε-caprolactone) (PCL) tracheal graft by indirect three-dimensional (3D) printing technique and created a composite construct by transplanting the hTMSC sheets to its luminal surface of the tracheal graft, then applied this tissue-engineered tracheal graft to non-circumferential tracheal reconstruction in a rabbit model. 4 weeks after implantation, the luminal surface of tissue-engineered tracheal graft was covered by a mature and highly-ciliated epithelium, whereas tracheal grafts without hTMSC sheets were covered by only a thin, immature epithelium. Therefore, hTMSC sheets on the luminal surface of a tissue-engineered tracheal graft can accelerate the tracheal epithelial regeneration, and the tissue-engineered tracheal graft with hTMSC sheets provides a useful clinical alternative for tracheal epithelial regeneration.

Development of a 3D cell printed structure as an alternative to autologs cartilage for auricular reconstruction.

Surgical technique using autologs cartilage is considered as the best treatment for cartilage tissue reconstruction, although the burdens of donor site morbidity and surgical complications still remain. The purpose of this study is to apply three-dimensional (3D) cell printing to fabricate a tissue-engineered graft, and evaluate its effects on cartilage reconstruction. A multihead tissue/organ building system is used to print cell-printed scaffold (CPS), then assessed the effect of the CPS on cartilage regeneration in a rabbit ear. The cell viability and functionality of chondrocytes were significantly higher in CPS than in cell-seeded scaffold (CSS) and cell-seeded hybrid scaffold (CSHS) in vitro. CPS was then implanted into a rabbit ear that had an 8 mm-diameter cartilage defect; at 3 months after implantation the CPS had fostered complete cartilage regeneration whereas CSS and autologs cartilage (AC) fostered only incomplete healing. This result demonstrates that cell printing technology can provide an appropriate environment in which encapsulated chondrocytes can survive and differentiate into cartilage tissue in vivo. Moreover, the effects of CPS on cartilage regeneration were even better than those of AC. Therefore, we confirmed the feasibility of CPS as an alternative to AC for auricular reconstruction.

A new method of fabricating a blend scaffold using an indirect three-dimensional printing technique

Due to its simplicity and effectiveness, the physical blending of polymers is considered to be a practical strategy for developing a versatile scaffold having desirable mechanical and biochemical properties. In the present work, an indirect three-dimensional (i3D) printing technique was proposed to fabricate a 3D free-form scaffold using a blend of immiscible materials, such as polycaprolactone (PCL) and gelatin. The i3D printing technique includes 3D printing of a mold and a sacrificial molding process. PCL/chloroform and gelatin/water were physically mixed to prepare the blend solution, which was subsequently injected into the cavity of a 3D printed mold. After solvent removal and gelatin cross-linking, the mold was dissolved to obtain a PCL-gelatin (PG) scaffold, with a specific 3D structure. Scanning electron microscopy and Fourier transform infrared spectroscopy analysis indicated that PCL masses and gelatin fibers in the PG scaffold homogenously coexisted without chemical bonding. Compression tests confirmed that gelatin incorporation into the PCL enhanced its mechanical flexibility and softness, to the point of being suitable for soft-tissue engineering, as opposed to pure PCL. Human adipose-derived stem cells, cultured on a PG scaffold, exhibited enhanced in vitro chondrogenic differentiation and tissue formation, compared with those on a PCL scaffold. The i3D printing technique can be used to blend a variety of materials, facilitating 3D scaffold fabrication for specific tissue regeneration. Furthermore, this convenient and versatile technique may lead to wider application of 3D printing in tissue engineering.