Microfluidics-assisted fabrication of natural killer cell-laden microgel enhances the therapeutic efficacy for tumor immunotherapy.

Recently, interest in cancer immunotherapy has increased over traditional anti-cancer therapies such as chemotherapy or targeted therapy. Natural killer (NK) cells are part of the immune cell family and essential to tumor immunotherapy as they detect and kill cancer cells. However, the disadvantage of NK cells is that cell culture is difficult. In this study, porous microgels have been fabricated using microfluidic channels to effectively culture NK cells. Microgel fabrication using microfluidics can be mass-produced in a short time and can be made in a uniform size. Microgels consist of photo cross-linkable polymers such as methacrylic gelatin (GelMa) and can be regulated via controlled GelMa concentrations. NK92 cell-laden three-dimensional (3D) microgels increase mRNA expression levels, NK92 cell proliferation, cytokine release, and anti-tumor efficacy, compared with two-dimensional (2D) cultures. In addition, the study confirms that 3D-cultured NK92 cells enhance anti-tumor effects compared with enhancement by 2D-cultured NK92 cells in the K562 leukemia mouse model. Microgels containing healthy NK cells are designed to completely degrade after 5 days allowing NK cells to be released to achieve cell-to-cell interaction with cancer cells. Overall, this microgel system provides a new cell culture platform for the effective culturing of NK cells and a new strategy for developing immune cell therapy.

3D-printed versatile biliary stents with nanoengineered surface for anti-hyperplasia and antibiofilm formation.

Biliary strictures are characterized by the narrowing of the bile duct lumen, usually caused by surgical biliary injury, cancer, inflammation, and scarring from gallstones. Endoscopic stent placement is a well-established method for the management of biliary strictures. However, maintaining optimal mechanical properties of stents and designing surfaces that can prevent stent-induced tissue hyperplasia and biofilm formation are challenges in the fabrication of biodegradable biliary stents (BBSs) for customized treatment. This study proposes a novel approach to fabricating functionalized polymer BBSs with nanoengineered surfaces using 3D printing. The 3D printed stents, fabricated from bioactive silica poly(ε-carprolactone) (PCL) via a sol-gel method, exhibited tunable mechanical properties suitable for supporting the bile duct while ensuring biocompatibility. Furthermore, a nanoengineered surface layer was successfully created on a sirolimus (SRL)-coated functionalized PCL (fPCL) stent using Zn ion sputtering-based plasma immersion ion implantation (S-PIII) treatment to enhance the performance of the stent. The nanoengineered surface of the SRL-coated fPCL stent effectively reduced bacterial responses and remarkably inhibited fibroblast proliferation and initial burst release of SRL in vitro systems. The physicochemical properties and biological behaviors, including in vitro biocompatibility and in vivo therapeutic efficacy in the rabbit bile duct, of the Zn-SRL@fPCL stent demonstrated its potential as a versatile platform for clinical applications in bile duct tissue engineering.

Longitudinal long term follow up investigation on the carcinogenic impact of polyhexamethylene guanidine phosphate in rat models.

Polyhexamethylene guanidine phosphate (PHMG-p) is a major component in humidifier disinfectants, which cause life-threatening lung injuries. However, to our knowledge, no published studies have investigated associations between PHMG-p dose and lung damage severity with long-term follow-up. Therefore, we evaluated longitudinal dose-dependent changes in lung injuries using repeated chest computed tomography (CT). Rats were exposed to low (0.2 mg/kg, n = 10), intermediate (1.0 mg/kg, n = 10), and high (5.0 mg/kg, n = 10) doses of PHMG-p. All rats underwent repeated CT scans after 10 and 40 weeks following the first exposure. All CT images were quantitatively analyzed using commercial software. Inflammation/fibrosis and tumor counts underwent histopathological evaluation. In both radiological and histopathologic results, the lung damage severity increased as the PHMG-p dose increased. Moreover, the number, size, and malignancy of the lung tumors increased as the dose increased. Bronchiolar-alveolar hyperplasia developed in all groups. During follow-up, there was intergroup variation in bronchiolar-alveolar hyperplasia progression, although bronchiolar-alveolar adenomas or carcinomas usually increase in size over time. Thirty-three carcinomas were detected in the high-dose group in two rats. Overall, lung damage from PHMG-p and the number and malignancy of lung tumors were shown to be dose-dependent in a rat model using repeated chest CT scans during a long-term follow-up.

Hyaluronic acid/polyphenol sunscreens with broad-spectrum UV protection properties from tannic acid and quercetin.

Currently, commercial sunscreens cause a number of biotoxicity and environmental issues, making it imperative to develop biocompatible alternatives. In this study, we aimed to develop an alternative sunscreen from two ecofriendly and biocompatible natural polyphenolic compounds, tannic acid (TA) and quercetin (Que). The sunscreen was prepared through a simple process using an oil-in-water emulsion as the medium and hyaluronic acid (HA) as the base polymer to improve biocompatibility. The HA/TA/Que. sunscreen prepared in this study exhibits 0 % transmittance in the UVB region and <15 % transmittance in the UVA region, resulting in excellent sun-protection properties (SPF 30). Remarkably, the as-prepared HA/TA/Que. sunscreen has a suitable viscosity and similar UV protection properties to those of commercial sunscreens. The HA/TA/Que. sunscreen also exhibits 90.4 % antioxidant activity against 2,2-diphenyl-1-picrylhydrazyl, demonstrating an ability to effectively capture reactive oxygen species that directly affect the skin. In addition, the cell viability was >90 % at a concentration of 50 µg/mL after 7 days, indicating excellent cytocompatibility. Owing to its various advantageous features, the HA/TA/Que. sunscreen with excellent sun protection properties and multiple functionalities is expected to resolve many environmental and biological issues caused by commercial sunscreens.

Peptide derived from stromal cell-derived factor 1δ enhances the in vitro expression of osteogenic proteins via bone marrow stromal cell differentiation and promotes bone formation in in vivo models.

Mesenchymal stem cells (MSCs) rely on chemokines and chemokine receptors to execute their biological and physiological functions. Stromal cell-derived factor-1 (SDF-1) is upregulated in injury sites, where it acts as a chemotactic agent, attracting CXCR4-expressing MSCs, which play a pivotal role in the healing and regeneration of tissue throughout the body. Furthermore, SDF-1 expression has been observed in regions experiencing inflammation-induced bone destruction and fracture sites. In this study, we identified a novel peptide called bone-forming peptide-5 (BFP-5), derived from SDF-1δ, which can promote the osteogenesis of MSCs as well as bone formation and healing. Multipotent bone marrow stromal cells treated with BFP-5 showed enhanced alizarin red S staining and higher alkaline phosphatase (ALP) activity. Moreover, ALP and osterix proteins were more abundantly expressed when cells were treated with BFP-5 than SDF-1α. Histology and microcomputed tomography data at 12 weeks demonstrated that both rabbit and goat models transplanted with polycaprolactone (PCL) scaffolds coated with BFP-5 showed significantly greater bone formation than animals transplanted with PCL scaffolds alone. These findings suggest that BFP-5 could be useful in the development of related therapies for conditions associated with bones.

A wearable colorimetric sweat pH sensor-based smart textile for health state diagnosis.

Sweat pH is an important indicator for diagnosing disease states, such as cystic fibrosis. However, conventional pH sensors are composed of large brittle mechanical parts and need additional instruments to read signals. These pH sensors have limitations for practical wearable applications. In this study, we propose wearable colorimetric sweat pH sensors based on curcumin and thermoplastic-polyurethane (C-TPU) electrospun-fibers to diagnose disease states by sweat pH monitoring. This sensor aids in pH monitoring by changing color in response to chemical structure variation from enol to di-keto form via H-atom separation. Its chemical structure variation changes the visible color due to light absorbance and reflectance changes. Furthermore, it can rapidly and sensitively detect sweat pH due to its superior permeability and wettability. By O2 plasma activation and thermal pressing, this colorimetric pH sensor can be easily attached to various fabric substrates such as swaddling and patient clothing via surface modification and mechanical interlocking of C-TPU. Furthermore, the diagnosable clothing is durable and reusable enough to neutral washing conditions due to the reversible pH colorimetric sensing performance by restoring the enol form of curcumin. This study contributes to the development of smart diagnostic clothing for cystic fibrosis patients who require continuous sweat pH monitoring.

NK cells encapsulated in micro/macropore-forming hydrogels via 3D bioprinting for tumor immunotherapy.

BACKGROUND: Patients face a serious threat if a solid tumor leaves behind partial residuals or cannot be completely removed after surgical resection. Immunotherapy has attracted attention as a method to prevent this condition. However, the conventional immunotherapy method targeting solid tumors, that is, intravenous injection, has limitations in homing in on the tumor and in vivo expansion and has not shown effective clinical results. METHOD: To overcome these limitations, NK cells (Natural killer cells) were encapsulated in micro/macropore-forming hydrogels using 3D bioprinting to target solid tumors. Sodium alginate and gelatin were used to prepare micro-macroporous hydrogels. The gelatin contained in the alginate hydrogel was removed because of the thermal sensitivity of the gelatin, which can generate interconnected micropores where the gelatin was released. Therefore, macropores can be formed through bioprinting and micropores can be formed using thermally sensitive gelatin to make macroporous hydrogels. RESULTS: It was confirmed that intentionally formed micropores could help NK cells to aggregate easily, which enhances cell viability, lysis activity, and cytokine release. Macropores can be formed using 3D bioprinting, which enables NK cells to receive the essential elements. We also characterized the functionality of NK 92 and zEGFR-CAR-NK cells in the pore-forming hydrogel. The antitumor effects on leukemia and solid tumors were investigated using an in vitro model. CONCLUSION: We demonstrated that the hydrogel encapsulating NK cells created an appropriate micro-macro environment for clinical applications of NK cell therapy for both leukemia and solid tumors via 3D bioprinting. 3D bioprinting makes macro-scale clinical applications possible, and the automatic process shows potential for development as an off-the-shelf immunotherapy product. This immunotherapy system could provide a clinical option for preventing tumor relapse and metastasis after tumor resection. Micro/macropore-forming hydrogel with NK cells fabricated by 3D bioprinting and implanted into the tumor site.

Assessment of Esophageal Reconstruction via Bioreactor Cultivation of a Synthetic Scaffold in a Canine Model.

OBJECTIVES: Using tissue-engineered materials for esophageal reconstruction is a technically challenging task in animals that requires bioreactor training to enhance cellular reactivity. There have been many attempts at esophageal tissue engineering, but the success rate has been limited due to difficulty in initial epithelialization in the special environment of peristalsis. The purpose of this study was to evaluate the potential of an artificial esophagus that can enhance the regeneration of esophageal mucosa and muscle through the optimal combination of a double-layered polymeric scaffold and a custom-designed mesenchymal stem cell-based bioreactor system in a canine model. METHODS: We fabricated a novel double-layered scaffold as a tissue-engineered esophagus using an electrospinning technique. Prior to transplantation, human-derived mesenchymal stem cells were seeded into the lumen of the scaffold, and bioreactor cultivation was performed to enhance cellular reactivity. After 3 days of cultivation using the bioreactor system, tissue-engineered artificial esophagus was transplanted into a partial esophageal defect (5×3 cm-long resection) in a canine model. RESULTS: Scanning electron microscopy (SEM) showed that the electrospun fibers in a tubular scaffold were randomly and circumferentially located toward the inner and outer surfaces. Complete recovery of the esophageal mucosa was confirmed by endoscopic analysis and SEM. Esophagogastroduodenoscopy and computed tomography also showed that there were no signs of leakage or stricture and that there was a normal lumen with complete epithelialization. Significant regeneration of the mucosal layer was observed by keratin-5 immunostaining. Alpha-smooth muscle actin immunostaining showed significantly greater esophageal muscle regeneration at 12 months than at 6 months. CONCLUSION: Custom-designed bioreactor cultured electrospun polyurethane scaffolds can be a promising approach for esophageal tissue engineering.

Three-dimensional bioprinting of mesenchymal stem cells using an osteoinductive bioink containing alginate and BMP-2-loaded PLGA nanoparticles for bone tissue engineering.

Hydrogels mimicking the physicochemical properties of the native extracellular matrix have attracted great attention as bioinks for three-dimensional (3D) bioprinting in tissue engineering applications. Alginate is a widely used bioink with beneficial properties of fast gelation and biocompatibility; however, bioprinting using alginate-based bioinks has several limitations, such as poor printability, structural instability, and limited biological activities. To address these issues, we formulated various bioinks using bone morphogenetic protein-2 (BMP-2)-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles and alginate for mesenchymal stem cell (MSC) printing and induction of osteogenic differentiation. Incorporation of PLGA nanoparticles into alginate could enhance the mechanical properties and printability of the bioink. In particular, Alg/NPN30 (30 mg/mL PLGA nanoparticles and 3% w/v alginate) was most suitable for 3D printing with respect to printability and stability. BMP-2-loaded PLGA nanoparticles (NPBMP-2) displayed sustained in vitro release of BMP-2 for up to two weeks. Further in vitro studies indicated that bioinks composed of alginate and NPBMP-2 significantly induced osteogenesis of the MSCs compared with other controls, evidenced by enhanced calcium deposition, alkaline phosphatase activity, and gene expression of osteogenic markers. Our novel bioink consisting of widely used biocompatible components displays good printability, stability, and osteogenic inductivity, and holds strong potential for cell printing and bone tissue engineering applications.

Surface engineering of 3D-printed scaffolds with minerals and a pro-angiogenic factor for vascularized bone regeneration.

Scaffolds functionalized with biomolecules have been developed for bone regeneration but inducing the regeneration of complex structured bone with neovessels remains a challenge. For this study, we developed three-dimensional printed scaffolds with bioactive surfaces coated with minerals and platelet-derived growth factor. The minerals were homogeneously deposited on the surface of the scaffold using 0.01 M NaHCO3 with epigallocatechin gallate in simulated body fluid solution (M2). The M2 scaffold demonstrated enhanced mineral coating amount per scaffold with a greater compressive modulus than the others which used different concentration of NaHCO3. Then, we immobilized PDGF on the mineralized scaffold (M2/P), which enhanced the osteogenic differentiation of human adipose derived stem cells in vitro and promoted the secretion of pro-angiogenic factors. Cells cultured in M2/P showed remarkable ratio of osteocalcin- and osteopontin-positive nuclei, and M2/P-derived medium induced endothelial cells to form tubule structures. Finally, the implanted M2/P scaffolds onto mouse calvarial defects had regenerated bone in 80.8 ± 9.8% of the defect area with the arterioles were formed, after 8 weeks. In summary, our scaffold, which composed of minerals and pro-angiogenic growth factor, could be used therapeutically to improve the regeneration of bone with a highly vascularized structure. STATEMENT OF SIGNIFICANCE: Surface engineered scaffolds have been developed for bone regeneration but inducing the volumetric regeneration of bone with neovessels remains a challenge. In here, we developed 3D printed scaffolds with bioactive surfaces coated with bio-minerals and platelet-derived growth factors. We proved that the 0.01 M NaHCO3 with polyphenol in simulated body fluid solution enhanced the deposition of bio-minerals and even distribution on the surface of scaffold. The in vitro studies demonstrated that the attached cells on the bioactive surface showed the enhanced osteogenic differentiation and secretion of pro-angiogenic factors. Finally, the scaffold with bioactive surface not only improved the regenerated volume of bone tissues but also increased neovessel formation after in vivo implantation onto mouse calvarial defect.

Hyaluronic Acid Coating on Hydrophobic Tracheal Scaffold Enhances Mesenchymal Stem Cell Adhesion and Tracheal Regeneration.

BACKGROUND: Long segmental tracheal repair is challenging in regenerative medicine due to low adhesion of stem cells to tracheal scaffolds. Optimal transplantation of stem cells for tracheal defects has not been established. We evaluated the role of hyaluronic acid (HA) coating of tracheal scaffolds in mesenchymal stem cell (MSC) adhesion and tracheal regeneration in a rabbit model. METHODS: A three-dimensionally printed tubular tracheal prosthesis was incubated with dopa-HA-fluorescein isothiocyanate in phosphate-buffered saline for 2 days. MSCs were incubated with an HA-coated scaffold, and their adhesion was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. HA coated scaffolds with or without MSC seeding were transplanted at the circumferential tracheal defect in rabbits, and survival, rigid bronchoscopy, radiologic findings, and histologic findings were compared between the two groups. RESULTS: HA-coated scaffolds showed better MSC adhesion than non-coated scaffolds. The HA-coated scaffolds with MSC group showed a wider airway and greater mucosal regeneration compared to the HA-coated scaffolds without MSC group. CONCLUSION: HA coating of scaffolds can promote MSC adhesion and tracheal regeneration.

Transplantation of a 3D-printed tracheal graft combined with iPS cell-derived MSCs and chondrocytes.

For successful tracheal reconstruction, tissue-engineered artificial trachea should meet several requirements, such as biocompatible constructs comparable to natural trachea, coverage with ciliated respiratory mucosa, and adequate cartilage remodeling to support a cylindrical structure. Here, we designed an artificial trachea with mechanical properties similar to the native trachea that can enhance the regeneration of tracheal mucosa and cartilage through the optimal combination of a two-layered tubular scaffold and human induced pluripotent stem cell (iPSC)-derived cells. The framework of the artificial trachea was fabricated with electrospun polycaprolactone (PCL) nanofibers (inner) and 3D-printed PCL microfibers (outer). Also, human bronchial epithelial cells (hBECs), iPSC-derived mesenchymal stem cells (iPSC-MSCs), and iPSC-derived chondrocytes (iPSC-Chds) were used to maximize the regeneration of tracheal mucosa and cartilage in vivo. After 2 days of cultivation using a bioreactor system, tissue-engineered artificial tracheas were transplanted into a segmental trachea defect (1.5-cm length) rabbit model. Endoscopy did not reveal granulation ingrowth into tracheal lumen. Alcian blue staining clearly showed the formation of ciliated columnar epithelium in iPSC-MSC groups. In addition, micro-CT analysis showed that iPSC-Chd groups were effective in forming neocartilage at defect sites. Therefore, this study describes a promising approach for long-term functional reconstruction of a segmental tracheal defect.

Tissue-Engineered Graft for Circumferential Esophageal Reconstruction in Rats.

The use of biocompatible materials for circumferential esophageal reconstruction is a technically challenging task in rats and requires an optimal implant technique with nutritional support. Recently, there have been many attempts at esophageal tissue engineering, but the success rate has been limited due to difficulty in early epithelization in the special environment of peristalsis. Here, we developed an artificial esophagus that can improve the regeneration of the esophageal mucosa and muscle layers through a two-layered tubular scaffold, a mesenchymal stem cell-based bioreactor system, and a bypass feeding technique with modified gastrostomy. The scaffold is made of polyurethane (PU) nanofibers in a cylindrical shape with a three-dimensional (3D) printed polycaprolactone strand wrapped around the outer wall. Prior to transplantation, human-derived mesenchymal stem cells were seeded into the lumen of the scaffold, and bioreactor cultivation was performed to enhance cellular reactivity. We improved the graft survival rate by applying surgical anastomosis and covering the implanted prosthesis with a thyroid gland flap, followed by temporary nonoral gastrostomy feeding. These grafts were able to recapitulate the findings of initial epithelialization and muscle regeneration around the implanted sites, as demonstrated by histological analysis. In addition, increased elastin fibers and neovascularization were observed in the periphery of the graft. Therefore, this model presents a potential new technique for circumferential esophageal reconstruction.

Fabrication of 3D plotted scaffold with microporous strands for bone tissue engineering.

Scaffold porosity has played a key role in bone tissue engineering aimed at effective tissue regeneration, by promoting cell attachment, proliferation, and osteogenic differentiation for new bone formation. Three-dimensional plotting systems (3DPSs) have been widely used to introduce porosity to the scaffold; however, introducing certain features in the scaffold strands that improve bone tissue regeneration remains a challenge. In this work, we fabricated bone tissue scaffolds with macro- and microporous structural features using a 3DPS and non-solvent-induced phase separation method. This approach allowed both macro- and micropores to be created in the scaffold strands. The surface morphology and mechanical and degradation properties of the perforated scaffolds were characterized carefully. Human marrow stromal cells were cultured on the scaffolds and then analyzed in vitro to assess scaffold bio-function. The highly porous scaffold exhibited mechanical properties similar to those of cancellous bone. Cell attachment, proliferation, and differentiation were significantly higher in porous scaffold compared to its nonporous counterpart. These results suggest that highly porous scaffolds have tremendous potential as a bone tissue regeneration platform.

Graphene oxide/alginate composites as novel bioinks for three-dimensional mesenchymal stem cell printing and bone regeneration applications.

Three-dimensional (3D) cell printing is a versatile technique enabling the creation of 3D constructs containing hydrogel and cells in the desired shape or pattern. Bioinks exhibiting appropriate mechanical properties and biological activities to support cell growth and/or differentiation toward a specific lineage play critical roles in 3D cell printing and tissue engineering applications. Herein, we explored alginate/graphene oxide (GO) composites as bioinks for their potential to improve printability, structural stability, and osteogenic activities for osteogenic tissue engineering applications. The addition of GO (0.05-1.0 mg mL-1) to 3% alginate significantly enhanced the printing performances of the alginate bioink. In addition, mesenchymal stem cells (MSCs) printed with alginate/GO showed good proliferation and higher survival in an oxidative stress environment. The 3D scaffolds printed with MSCs and alginate/GO demonstrated significantly enhanced osteogenic differentiation compared with those printed with MSCs and alginate. Overall, a bioink of 3% alginate and 0.5 mg mL-1 GO showed the most balanced characteristics in terms of printability, structural stability, and osteogenic induction of the printed MSCs. Alginate/GO composite bioinks will be useful for bioprinting research for various tissue engineering applications.

Tissue-Engineered Esophagus via Bioreactor Cultivation for Circumferential Esophageal Reconstruction.

The use of biomaterials for circumferential esophageal repair is technically challenging in a rat model, and an optimal scaffold implantation technique with nutritional support is essential. The purpose of this study was to investigate the effects of three-dimensional printed esophageal grafts and bioreactor cultivation on muscle regeneration and reepithelialization from circumferential esophageal defects in a rat model. Here, we designed an artificial esophagus that can enhance the regeneration of esophageal mucosa and muscle through the optimal combination of a two-layered tubular scaffold and mesenchymal stem cell-based bioreactor system. The graft was verified by the performance comparison with an omentum-cultured esophageal scaffold. We also applied a new surgical anastomosis technique and a thyroid gland flap over the implanted scaffold to improve graft survival. Although no regenerated mucosal layer was observed around the implants of the control group, histological examination of the regenerative esophagi along the scaffold revealed that the bioreactor system and omentum-cultured groups showed more than 80% of the mucosal regeneration without a fistula. The regenerated tissues showed that the integration of the esophageal scaffold and its native esophageal tissue was intact and were covered with layers of stratified squamous epithelium with several newly developed blood vessels. Therefore, this study describes a novel approach for circumferential esophageal reconstruction. Impact Statement In vivo functional esophageal reconstruction remains challenging due to anastomosis site leakage and necrosis of the implanted scaffold in a circumferential esophageal defect. Therefore, it is necessary to develop a tissue-engineered esophagus that enables regeneration of esophageal mucosa and muscle without leakage of the esophageal anastomosis. In this study, we proposed an intriguing strategy that combines a mesenchymal stem cell-seeded tubular scaffold with a bioreactor system for esophageal reconstruction and introduced a new surgical anastomosis technique with the thyroid gland flap over the implanted scaffold to improve graft survival. We believe that this system should be a powerful platform for circumferential replacement of the esophagus in a rat model.

Development of a three-dimensionally printed scaffold grafted with bone forming peptide-1 for enhanced bone regeneration with in vitro and in vivo evaluations.

Defects in bone are some of the most difficult injuries to treat. Biomimetic scaffolds represent a promising approach for successful bone tissue regeneration. In this study, a three-dimensional (3D) scaffold with osteo-inductive functionality was designed and assayed both in-vitro and in-vivo. Bone formation peptide-1 (BFP1), an osteo-promoting specific peptide, was covalently bound to a 3D printed polycaprolactone (PCL) scaffold using polydopamine (DOPA). The amount of BFP1 immobilized on the surface was found to increase depending on the BFP1 concentration of the loading solution. To observe the biological effects of the 3D scaffolds, human tonsil-derived mesenchymal stem cells (hTMSCs) were isolated. The cells were cultured on the scaffolds and observed to rapidly differentiate into osteoblast-like cells with osteo-promoting capabilities. The scaffolds were implanted in a rabbit calvarial defect model for 8 weeks and successfully stimulated both vessel and bone regeneration. Osteo-promoting 3D scaffolds may provide a safer and more efficient approach for bone repair and remodelling in regenerative medicine.

In situ gold nanoparticle growth on polydopamine-coated 3D-printed scaffolds improves osteogenic differentiation for bone tissue engineering applications: in vitro and in vivo studies.

In this study, we designed scaffolds coated with gold nanoparticles (GNPs) grown on a polydopamine (PDA) coating of a three-dimensional (3D) printed polycaprolactone (PCL) scaffold. Our results demonstrated that the scaffolds developed here may represent an innovative paradigm in bone tissue engineering by inducing osteogenesis as a means of remodeling and healing bone defects.

Prolongation of liver-specific function for primary hepatocytes maintenance in 3D printed architectures.

Isolated primary hepatocytes from the liver are very similar to in vivo native liver hepatocytes, but they have the disadvantage of a limited lifespan in 2D culture. Although a sandwich culture and 3D organoids with mesenchymal stem cells (MSCs) as an attractive assistant cell source to extend lifespan can be used, it cannot fully reproduce the in vivo architecture. Moreover, long-term 3D culture leads to cell death because of hypoxic stress. Therefore, to overcome the drawback of 2D and 3D organoids, we try to use a 3D printing technique using alginate hydrogels with primary hepatocytes and MSCs. The viability of isolated hepatocytes was more than 90%, and the cells remained alive for 7 days without morphological changes in the 3D hepatic architecture with MSCs. Compared to a 2D system, the expression level of functional hepatic genes and proteins was higher for up to 7 days in the 3D hepatic architecture. These results suggest that both the 3D bio-printing technique and paracrine molecules secreted by MSCs supported long-term culture of hepatocytes without morphological changes. Thus, this technique allows for widespread expansion of cells while forming multicellular aggregates, may be applied to drug screening and could be an efficient method for developing an artificial liver.

Generation of Multilayered 3D Structures of HepG2 Cells Using a Bio-printing Technique.

BACKGROUND/AIMS: Chronic liver disease is a major widespread cause of death, and whole liver transplantation is the only definitive treatment for patients with end-stage liver diseases. However, many problems, including donor shortage, surgical complications and cost, hinder their usage. Recently, tissue-engineering technology provided a potential breakthrough for solving these problems. Three-dimensional (3D) printing technology has been used to mimic tissues and organs suitable for transplantation, but applications for the liver have been rare. METHODS: A 3D bioprinting system was used to construct 3D printed hepatic structures using alginate. HepG2 cells were cultured on these 3D structures for 3 weeks and examined by fluorescence microscopy, histology and immunohistochemistry. The expression of liverspecific markers was quantified on days 1, 7, 14, and 21. RESULTS: The cells grew well on the alginate scaffold, and liver-specific gene expression increased. The cells grew more extensively in 3D culture than two-dimensional culture and exhibited better structural aspects of the liver, indicating that the 3D bioprinting method recapitulates the liver architecture. CONCLUSIONS: The 3D bioprinting of hepatic structures appears feasible. This technology may become a major tool and provide a bridge between basic science and the clinical challenges for regenerative medicine of the liver.