One-Year Results of Ear Reconstruction with 3D Printed Implants.

PURPOSE: External ear reconstruction has been a challenging subject for plastic surgeons for decades. Popular methods using autologous costal cartilage or polyethylene still have their drawbacks. With the advance of three-dimensional (3D) printing technique, bioscaffold engineering using synthetic polymer draws attention as an alternative. This is a clinical trial of ear reconstruction using 3D printed scaffold, presented with clinical results after 1 year. MATERIALS AND METHODS: From 2021 to 2022, five adult patients with unilateral microtia underwent two-staged total ear reconstruction using 3D printed implants. For each patient, a patient-specific 3D printed scaffold was designed and produced with polycaprolactone (PCL) based on computed tomography images, using fused deposition modeling. Computed tomography scan was obtained preoperatively, within 2 weeks following the surgery and after 1 year, to compare the volume of the normal side and the reconstructed ear. At 1-year visit, clinical photo was taken for scoring by two surgeons and patients themselves. RESULTS: All five patients had completely healed reconstructed ear at 1-year follow-up. On average, the volume of reconstructed ear was 161.54% of that of the normal side ear. In a range of 0 to 10, objective assessors gave scores 3 to 6, whereas patients gave scores 8 to 10. CONCLUSION: External ear reconstruction using 3D printed PCL implant showed durable, safe results reflected by excellent volume restoration and patient satisfaction at 1 year postoperatively. Further clinical follow-up with more cases and refinement of scaffold with advancing bioprinting technique is anticipated. The study's plan and results have been registered with the Clinical Research Information Service (CRIS No. 3-2019-0306) and the Ministry of Food and Drug Safety (MFDS No. 1182).

Fabrication of 3D-Printed Implant for Two-Stage Ear Reconstruction Surgery and Its Clinical Application.

PURPOSE: Ear reconstruction is one of the most difficult areas in the field of reconstructive surgery. Due to limitations of the current practice, a novel method of auricular reconstruction is needed. Major advancements in three-dimensional (3D) printing technique have rendered the process of ear reconstruction more favorable. Herein, we present our experience in designing and clinically using 3D implants in both 1st and 2nd stage ear reconstruction surgery. MATERIALS AND METHODS: After obtaining 3D CT data from each patient, a 3D geometric ear model was created using mirroring and segmentation processes. The 3D-printed implant design resembles but does not exactly match the normal ear shape, and can be inserted in harmony with the currently used surgical technique. The 2nd stage implant was designed to minimize dead space and support the posterior ear helix. The 3D implants were finally fabricated with a 3D printing system and used in ear reconstruction surgery in our institute. RESULTS: The 3D implants were manufactured for application to the currently used two-stage technique while maintaining the shape of the patient's normal ear. The implants were successfully used for ear reconstruction surgery in microtia patients. A few months later, the 2nd stage implant was used in the 2nd stage operation. CONCLUSION: The authors were able to design, fabricate, and apply patient-specific 3D-printed ear implants for 1st and 2nd stage ear reconstruction surgeries. This design, combined with 3D bioprinting technique, may be a future alternative for ear reconstruction.

Zygomatic arch reconstruction with a patient-specific polycaprolactone/beta-tricalcium phosphate scaffold after parosteal osteosarcoma resection in a dog.

OBJECTIVE: To describe the surgical application of a 3D-printing-based, patient-specific, biocompatible polycaprolactone/beta-tricalcium phosphate (PCL/ß-TCP) scaffold to reconstruct the zygomatic arch after tumor resection in a dog. ANIMAL: A 13 year old female spayed Maltese. STUDY DESIGN: Case report METHODS: The dog's presenting complaint was swelling ventral to her right eye. A round mass arising from the caudal aspect of the right zygomatic arch was identified by computed tomography (CT). The histopathologic diagnosis was a low-grade spindle-cell tumor. Surgical resection was planned to achieve 5 mm margins. A patient-specific osteotomy guide and polycaprolactone/beta-tricalcium phosphate (PCL/ß-TCP) scaffold were produced. Osteotomy, including 30% of total zygomatic arch length, was performed using an oscillating saw aligned with the guide. The scaffold was placed in the defect. Parosteal osteosarcoma was diagnosed based on histopathological examination. Excision was complete, with the closest margin measuring 0.3 mm. RESULTS: Mild epiphora, due to surgical site swelling, subsided after 20 days. Tissue formation within and around the porous scaffold was noted on CT 10 months postoperatively, with no evidence of metastasis or local recurrence. Facial conformation appeared symmetrical, and no complications were noted 16 months postoperatively. CONCLUSION: The use of a 3D-printing-based, patient-specific, biocompatible PCL/ß-TCP scaffold successfully restored the structure and function of the zygomatic arch without complications, even following wide zygomectomy for complete tumor removal.

Clinical Application of 3D-Printed Patient-Specific Polycaprolactone/Beta Tricalcium Phosphate Scaffold for Complex Zygomatico-Maxillary Defects.

(1) Background: In the present study, we evaluated the efficacy of a 3D-printed, patient-specific polycaprolactone/beta tricalcium phosphate (PCL/ß-TCP) scaffold in the treatment of complex zygomatico-maxillary defects. (2) Methods: We evaluated eight patients who underwent immediate or delayed maxillary reconstruction with patient-specific PCL implants between December 2019 and June 2021. The efficacy of these techniques was assessed using the volume and density analysis of computed tomography data obtained before surgery and six months after surgery. (3) Results: Patients underwent maxillary reconstruction with the 3D-printed PCL/ß-TCP scaffold based on various reconstructive techniques, including bone graft, fasciocutaneous free flaps, and fat graft. In the volume analysis, satisfactory volume conformity was achieved between the preoperative simulation and actual implant volume with a mean volume conformity of 79.71%, ranging from 70.89% to 86.31%. The ratio of de novo bone formation to total implant volume (bone volume fraction) was satisfactory with a mean bone fraction volume of 23.34%, ranging from 7.81% to 66.21%. Mean tissue density in the region of interest was 188.84 HU, ranging from 151.48 HU to 291.74 HU. (4) Conclusions: The combined use of the PCL/ß-TCP scaffold with virtual surgical simulation and 3D printing techniques may replace traditional non-absorbable implants in the future owing to its accuracy and biocompatible properties.

Long-term efficacy and safety of 3D printed implant in patients with nasal septal deformities.

PURPOSE: To investigate the long-term safety and efficacy of a 3D-printed bioresorbable polycaprolactone (PCL) nasal implant for nasal septal deformity reconstruction. METHODS: Fourteen patients who had undergone nasal septum reconstruction surgery using 3D-printed PCL nasal septal implants were enrolled. The primary outcome was the change in total Nasal Obstruction Symptom Evaluation (NOSE) scale scores between postoperative 3 months and current status (3.59 ± 0.51 years). The secondary outcomes were changes in the minimum cross-sectional area (MCA) and volume of both nasal cavities based on acoustic rhinometry, the cross-sectional area of the ostiomeatal unit, and the nasal septum angle of the paranasal sinus (PNS) in computed tomography (CT) images, and a visual analog scale (VAS) of the patients' subjective satisfaction. RESULTS: The results showed no significant changes in the MCAs (Cohen's d:0.09; p = 0.711) or nasal volume (Cohen's d:0.26; p = 0.356), the area of the ostiomeatal unit (Cohen's d:0.49; p = 0.064), septum angles (Cohen's d:0.18; p = 0.831), the NOSE scale (Cohen's d:0.14; p = 0.621), or patients' subjective satisfaction (Cohen's d:0.52; p = 0.076) during the follow-up period. CONCLUSIONS: This homogeneous composite microporous PCL nasal septal implant demonstrated long-term clinical efficacy and safety in human tissues that required maintenance of mechanical strength. Therefore, the indications for this implant could extend to various other craniofacial reconstructions in the future.

Evaluation of Auricular Cartilage Reconstruction Using a 3-Dimensional Printed Biodegradable Scaffold and Autogenous Minced Auricular Cartilage.

Auricular cartilage reconstruction represents one of the greatest challenges for otolaryngology-head and neck surgery. The native structure and composition of the auricular cartilage can be achieved by combining a suitable chondrogenic cell source with an appropriate scaffold. In reconstructive surgery for cartilage tissue, autogenous cartilage is considered to be the best chondrogenic cell source. Polycaprolactone is mainly used as a tissue-engineered scaffold owing to its mechanical properties, miscibility with a large range of other polymers, and biodegradability. In this study, scaffolds with or without autogenous minced auricular cartilage were implanted bilaterally in rabbits for auricular regeneration. Six weeks (n = 4) and 16 weeks (n = 4) after implantation, real-time quantitative reverse transcription polymerase chain reaction and histology were used to assess the regeneration of the auricular cartilage. Quantitative reverse transcription polymerase chain reaction analysis revealed that the messenger RNA expression of aggrecan, collagen I, and collagen II was higher in scaffolds with 50% minced cartilage than the scaffold-only groups or scaffolds with 30% minced cartilage (P < 0.05). Furthermore, histological analysis demonstrated significantly superior cartilage regeneration in scaffolds with the minced cartilage group compared with the scaffold-only and control groups (P < 0.05). Autogenous cartilage can be easily obtained and loaded onto a scaffold to promote the presence of chondrogenic cells, allowing for an improvement of the reconstruction of auricular cartilage. Here, the regeneration of auricular cartilage was also successful in the 50% minced cartilage group. The results presented in this study could have clinical implications, as they demonstrate the potential of a 1-stage process for auricular reconstruction.

Efficacy of three-dimensionally printed polycaprolactone/beta tricalcium phosphate scaffold on mandibular reconstruction.

It has been demonstrated that development of three-dimensional printing technology has supported the researchers and surgeons to apply the bone tissue engineering to the oromandibular reconstruction. In this study, poly caprolactone/beta tricalcium phosphate (PCL/ß-TCP) scaffolds were fabricated by multi-head deposition system. The feasibility of the three-dimensionally (3D) -printed PCL/ß-TCP scaffolds for mandibular reconstruction was examined on critical-sized defect of canine mandible. The scaffold contained the heterogeneous pore sizes for more effective bone ingrowth and additional wing structures for more stable fixation. They were implanted into the mandibular critical-sized defect of which periosteum was bicortically resected. With eight 1-year-old male beagle dogs, experimental groups were divided into 4 groups (n = 4 defects per group, respectively). (a) no further treatment (control), (b) PCL/ß-TCP scaffold alone (PCL/TCP), (c) PCL/ß-TCP scaffold with recombinant human bone morphogenetic protein-2 (rhBMP-2) (PCL/TCP/BMP2) and (d) PCL/ß-TCP scaffold with autogenous bone particles (PCL/TCP/ABP). In micro-computed tomography, PCL/TCP/BMP2 and PCL/TCP/ ABP groups showed significant higher bone volume in comparison to Control and PCL/TCP groups (P < 0.05). In histomorphometric analysis, a trend towards more bone formation was observed in PCL/TCP/BMP2 and PCL/TCP/ABP groups, but the results lacked statistical significance (P = 0.052). Within the limitations of the present study, 3D-printed PCL/ß-TCP scaffolds showed acceptable potential for oromandibular reconstruction.

New clinical application of three-dimensional-printed polycaprolactone/ß-tricalcium phosphate scaffold as an alternative to allograft bone for limb-sparing surgery in a dog with distal radial osteosarcoma.

Limb-sparing surgery is one of the surgical options for dogs with distal radial osteosarcoma (OSA). This case report highlights the novel application of a three-dimensional (3D)-printed patient-specific polycaprolactone/ß-tricalcium phosphate (PCL/ß-TCP) scaffold in limb-sparing surgery in a dog with distal radial OSA. The outcomes evaluated included postoperative gait analysis, complications, local recurrence of tumor, metastasis, and survival time. Post-operative gait evaluation showed significant improvement in limb function, including increased weight distribution and decreased asymmetry. The implant remained well in place and increased bone opacity was observed between the host bone and the scaffold. There was no complication due to scaffold or surgery. Significant improvement in limb function and quality of life was noted postoperatively. Local recurrence and pulmonary metastasis were identified at 8 weeks postoperatively. The survival time from diagnosis of OSA to death was 190 days. The PCL/ß-TCP scaffold may be an effective alternative to cortical allograft in limb-sparing surgery for bone tumors.

Ideal scaffold design for total ear reconstruction using a three-dimensional printing technique.

Ear reconstruction using three-dimensional (3D) printing technique has been considered as a good substitute for conventional surgery, because it can provide custom-made 3D framework. However, there are difficulties with its application in clinical use. Researchers have reported 3D scaffolds for ear cartilage regeneration, but the designs of the 3D scaffolds were not appropriate to be used in surgery. Hence, we propose the design of an ideal 3D ear scaffold for use in ear reconstruction surgery. Facial computed tomography (CT) images of the unaffected ear were extracted using a "segmentation" procedure. The selected data were converted to a 3D model and mirrored to create a model of the affected side. The design of 3D model was modified to apply to Nagata's two-stage surgery. Based on the 3D reconstructed model, a 3D scaffold was 3D printed using polycaprolactone. The 3D scaffold closely resembled the real cartilage framework used in current operations in terms of ear anatomy. To account for skin thickness, the 3D scaffold was made 4 mm smaller than the real ear. Furthermore, 2 mm pores were included to allow the implantation of diced cartilage to promote regeneration of the cartilage. 3D printing technology can overcome the limitations of previous auricular reconstruction methods. Further studies are required to achieve a functional and stable substitute for auricular cartilage and to extend the clinical use of the 3D-printed construct. Additionally, the ethical and legal issues regarding the transplantation of 3D-printed constructs and cell culture technologies using human stem cells remain to be solved. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1295-1303, 2019.

Effects of three-dimensionally printed polycaprolactone/ß-tricalcium phosphate scaffold on osteogenic differentiation of adipose tissue- and bone marrow-derived stem cells.

BACKGROUND: Autogenous bone grafts have several limitations including donor-site problems and insufficient bone volume. To address these limitations, research on bone regeneration is being conducted actively. In this study, we investigate the effects of a three-dimensionally (3D) printed polycaprolactone (PCL)/tricalcium phosphate (TCP) scaffold on the osteogenic differentiation potential of adipose tissue-derived stem cells (ADSCs) and bone marrow-derived stem cells (BMSCs). METHODS: We investigated the extent of osteogenic differentiation on the first and tenth day and fourth week after cell culture. Cytotoxicity of the 3D printed PCL/ß-TCP scaffold was evaluated by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay, prior to osteogenic differentiation analysis. ADSCs and BMSCs were divided into three groups: C, only cultured cells; M, cells cultured in the 3D printed PCL/ß-TCP scaffold; D, cells cultured in the 3D printed PCL/ß-TCP scaffold with a bone differentiation medium. Alkaline phosphatase (ALP) activity assay, von Kossa staining, reverse transcription-polymerase chain reaction (RT-PCR), and Western blotting were performed for comparative analysis. RESULTS: ALP assay and von Kossa staining revealed that group M had higher levels of osteogenic differentiation compared to group C. RT-PCR showed that gene expression was higher in group M than in group C, indicating that, compared to group C, osteogenic differentiation was more extensive in group M. Expression levels of proteins involved in ossification were higher in group M, as per the Western blotting results. CONCLUSION: Osteogenic differentiation was increased in mesenchymal stromal cells (MSCs) cultured in the 3D printed PCL/TCP scaffold compared to the control group. Osteogenic differentiation activity of MSCs cultured in the 3D printed PCL/TCP scaffold was lower than that of cells cultured on the scaffold in bone differentiation medium. Collectively, these results indicate that the 3D printed PCL/TCP scaffold promoted osteogenic differentiation of MSCs and may be widely used for bone tissue engineering.

Clinical Application of 3-Dimensional Printing Technology for Patients With Nasal Septal Deformities: A Multicenter Study.

Importance: Studies have shown the controllability and porosity of polycaprolactone as well as the use of 3-dimensional (3-D) printing for nasal reconstruction in animal models. The utility of polycaprolactone with 3-D technology in nasal cartilaginous framework reconstruction in humans remains unknown. Objective: To investigate the safety and efficacy of 3-D printed, bioresorbable polycaprolactone nasal implants. Design, Setting, and Participants: This multicenter clinical trial comprised 20 patients with caudal septal deviations who underwent septoplasty, which used a 3-D printed polycaprolactone mesh, at 2 centers in South Korea. Patients were included if they were aged 18 to 74 years and had nasal septal deviations, Nasal Obstruction Symptom Evaluation scores greater than 20, and persistent nasal obstructions. Twenty-two patients met the inclusion criteria, but 2 patients were excluded before the operation. The study was conducted from July 1, 2016, to June 30, 2017. Main Outcomes and Measures: The change in total Nasal Obstruction Symptom Evaluation score between the preoperative examination and the week 12 postoperative examination was the primary outcome. Changes in bilateral nasal cavity minimum cross-sectional area and volume on acoustic rhinometry at weeks 4 and 12 after the operation as well as changes in the nasal cavity cross-sectional area at the osteomeatal unit and nasal septum angle in the paranasal sinus on computed tomography after week 12 were among the secondary outcomes. Results: Of the 20 patients included in the study, 4 (20%) were female, 16 (80%) were male, with a mean (SD) age of 34.95 (11.96) years. The preoperative and week 12 postoperative results revealed significant changes in the minimal cross-sectional areas on acoustic rhinometry (0.41 [SD, 0.39] vs -0.11 [SD, 0.18]; difference, 0.42; 95% CI, 0.23-0.61), nasal septum angles on computed tomography (11.22 [SD, 6.57] vs 2.89 [SD, 3.12]; difference, 8.33; 95% CI, 5.08-11.58), and Nasal Obstruction Symptom Evaluation scores (73.50 [SD, 19.88] vs 3.75 [SD, 6.26]; difference, 69.75; 95% CI, 59.22-80.28). The surgeons' convenience level with the procedure was favorable (visual analog scale score [SD], 90.90 [9.45]), and so were the patients' symptom improvements and satisfaction after 12 weeks (visual analog scale score [SD], 88.30 [9.87]). Conclusions and Relevance: The 3-D printed, homogeneous, composite, microporous polycaprolactone nasal implant demonstrated proper mechanical support and thinness with excellent biocompatibility and surgical manipulability. Polycaprolactone may be a clinically biocompatible material for use in various craniofacial reconstructions in the future.

Cleft Alveolus Reconstruction Using a Three-Dimensional Printed Bioresorbable Scaffold With Human Bone Marrow Cells.

Bone tissue engineering technology based on scaffold has been applied for cleft lip and palate treatment. However, clinical applications of patient-specific three-dimensional (3D) scaffolds have rarely been performed. In this study, a clinical case using patient-specific 3D-printed bioresorbable scaffold with bone marrow stromal cells collected from iliac crest in the operating room has been introduced. At 6-month after transplantation, the bone volume of the newly regenerated bone was approximately 45% of the total defect volume. Bone mineral density of the newly regenerated bone was about 75% compared to the surrounding bone. The Hounsfield unit value was higher than that of cancellous maxillary alveolar bone and lower than that of the cortical maxillary alveolar bone. Bone-marrow-derived mesenchymal stem cells-seeded 3D-printed patient-specific polycaprolactone scaffolds offer a promising alternative for alveolar cleft reconstruction and other bony defects.

Pre-set extrusion bioprinting for multiscale heterogeneous tissue structure fabrication.

Recent advances in three-dimensional bioprinting technology have led to various attempts in fabricating human tissue-like structures. However, current bioprinting technologies have limitations for creating native tissue-like structures. To resolve these issues, we developed a new pre-set extrusion bioprinting technique that can create heterogeneous, multicellular, and multimaterial structures simultaneously. The key to this ability lies in the use of a precursor cartridge that can stably preserve a multimaterial with a pre-defined configuration that can be simply embedded in a syringe-based printer head. The multimaterial can be printed and miniaturized through a micro-nozzle without conspicuous deformation according to the pre-defined configuration of the precursor cartridge. Using this system, we fabricated heterogeneous tissue-like structures such as spinal cords, hepatic lobule, blood vessels, and capillaries. We further obtained a heterogeneous patterned model that embeds HepG2 cells with endothelial cells in a hepatic lobule-like structure. In comparison with homogeneous and heterogeneous cell printing, the heterogeneous patterned model showed a well-organized hepatic lobule structure and higher enzyme activity of CYP3A4. Therefore, this pre-set extrusion bioprinting method could be widely used in the fabrication of a variety of artificial and functional tissues or organs.

Reconstruction of Complex Maxillary Defects Using Patient-specific 3D-printed Biodegradable Scaffolds.

Reconstruction of maxilla defects has remained one of the most challenging problems in craniomaxillofacial reconstruction because it typically requires harvesting and grafting of autologous bone, which poses limitations related to the difficulties in accurately reconstructing the defected bone and the highly prolonged duration of surgery. We employed tissue-engineered, patient-specific, 3-dimensional (3D)-printed biodegradable scaffolds for maxillofacial bone reconstruction in patients with complex maxillary defects after surgical removal of cancer. A customized polycaprolactone (PCL) scaffold was designed and fabricated for each patient. For this purpose, we used computer-aided design and manufacturing combined with 3D printing technology. The patients implanted with the PCL scaffolds were followed up for up to 2 years with careful evaluation of morphological changes in the face. We confirmed that the patient-specific 3D-printed PCL scaffold effectively filled the maxillary defect and promoted regeneration of the deficient tissue while remaining stable in the body for a relatively long period. Employing customized tissue-engineered scaffolds built using the patient's computed tomography data and an extrusion-based 3D printing system is safe and clinically feasible, helping create and maintain improved morphological features of the face, which represents the most important aspect from the perspective of the patients.

Three-Dimensional Bio-Printed Scaffold Sleeves With Mesenchymal Stem Cells for Enhancement of Tendon-to-Bone Healing in Anterior Cruciate Ligament Reconstruction Using Soft-Tissue Tendon Graft.

PURPOSE: To investigate the efficacy of the insertion of 3-dimensional (3D) bio-printed scaffold sleeves seeded with mesenchymal stem cells (MSCs) to enhance osteointegration between the tendon and tunnel bone in anterior cruciate ligament (ACL) reconstruction in a rabbit model. METHODS: Scaffold sleeves were fabricated by 3D bio-printing. Before ACL reconstruction, MSCs were seeded into the scaffold sleeves. ACL reconstruction with hamstring tendon was performed on both legs of 15 adult rabbits (aged 12 weeks). We implanted 15 bone tunnels with scaffold sleeves with MSCs and implanted another 15 bone tunnels with scaffold sleeves without MSCs before passing the graft. The specimens were harvested at 4, 8, and 12 weeks. H&E staining, immunohistochemical staining of type II collagen, and micro-computed tomography of the tunnel cross-sectional area were evaluated. Histologic assessment was conducted with a histologic scoring system. RESULTS: In the histologic assessment, a smooth bone-to-tendon transition through broad fibrocartilage formation was identified in the treatment group, and the interface zone showed abundant type II collagen production on immunohistochemical staining. Bone-tendon healing histologic scores were significantly higher in the treatment group than in the control group at all time points. Micro-computed tomography at 12 weeks showed smaller tibial (control, 9.4 ± 0.9 mm2; treatment, 5.8 ± 2.9 mm2; P = .044) and femoral (control, 9.6 ± 2.9 mm2; treatment, 6.0 ± 1.0 mm2; P = .03) bone-tunnel areas in the treated group than in the control group. CONCLUSIONS: The 3D bio-printed scaffold sleeve with MSCs exhibited excellent results in osteointegration enhancement between the tendon and tunnel bone in ACL reconstruction in a rabbit model. CLINICAL RELEVANCE: If secure biological healing between the tendon graft and tunnel bone can be induced in the early postoperative period, earlier, more successful rehabilitation may be facilitated. Three-dimensional bio-printed scaffold sleeves with MSCs have the potential to accelerate bone-tendon healing in ACL reconstruction.

Three-Dimensional Printing-based Reconstruction of a Maxillary Bone Defect in a Dog Following Tumor Removal.

Three-dimensional (3D) printing has been applied extensively not only in human, but also veterinary medicine. However, the technique is still used in the clinical area for a surgical plan or education prior to surgery. Thus, we report a case of reconstruction after tumor removal surgery with the use of a 3D-printed scaffold. A 12-year-old female mixed dog had a left caudal maxillary mass. Based on computed tomography images, a defect was confirmed on the maxillary bone due to the oral mass, and a surgical plan was designed to remove the oral mass and graft the 3D printed scaffold. Customized polycaprolactone/ beta-tracalciumphosphate (PCL/ß-TCP) scaffold was fabricated using the micro-extrusion-based 3D printer. In the operation, after the removal of the oral mass, the scaffold was grafted onto the defect site. At follow-up, 8 months after surgery, the result was successful without any special problems in the periodic CT scans and oral examinations. This case is believed to be the first case of reconstruction by using a 3D printed scaffold in the maxillary bone defect, and this 3D printing technique is thought to be very helpful for veterinary patients with bone defects and several other diseases.

Fabrication of In Vitro Cancer Microtissue Array on Fibroblast-Layered Nanofibrous Membrane by Inkjet Printing.

In general, a drug candidate is evaluated using 2D-cultured cancer cells followed by an animal model. Despite successful preclinical testing, however, most drugs that enter human clinical trials fail. The high failure rates are mainly caused by incompatibility between the responses of the current models and humans. Here, we fabricated a cancer microtissue array in a multi-well format that exhibits heterogeneous and batch-to-batch structure by continuous deposition of collagen-suspended Hela cells on a fibroblast-layered nanofibrous membrane via inkjet printing. Expression of both Matrix Metalloproteinase 2 (MMP2) and Matrix Metalloproteinase 9 (MMP9) was higher in cancer microtissues than in fibroblast-free microtissues. The fabricated microtissues were treated with an anticancer drug, and high drug resistance to doxorubicin occurred in cancer microtissues but not in fibroblast-free microtissues. These results introduce an inkjet printing fabrication method for cancer microtissue arrays, which can be used for various applications such as early drug screening and gradual 3D cancer studies.

Effects of 3D-Printed Polycaprolactone/ß-Tricalcium Phosphate Membranes on Guided Bone Regeneration.

This study was conducted to compare 3D-printed polycaprolactone (PCL) and polycaprolactone/ß-tricalcium phosphate (PCL/ß-TCP) membranes with a conventional commercial collagen membrane in terms of their abilities to facilitate guided bone regeneration (GBR). Fabricated membranes were tested for dry and wet mechanical properties. Fibroblasts and preosteoblasts were seeded into the membranes and rates and patterns of proliferation were analyzed using a kit-8 assay and by scanning electron microscopy. Osteogenic differentiation was verified by alizarin red S and alkaline phosphatase (ALP) staining. An in vivo experiment was performed using an alveolar bone defect beagle model, in which defects in three dogs were covered with different membranes. CT and histological analyses at eight weeks after surgery revealed that 3D-printed PCL/ß-TCP membranes were more effective than 3D-printed PCL, and substantially better than conventional collagen membranes in terms of biocompatibility and bone regeneration and, thus, at facilitating GBR.

Osteogenesis of Adipose-Derived and Bone Marrow Stem Cells with Polycaprolactone/Tricalcium Phosphate and Three-Dimensional Printing Technology in a Dog Model of Maxillary Bone Defects.

Bone graft material should possess sufficient porosity and permeability to allow integration with native tissue and vascular invasion, and must satisfy oxygen and nutrient transport demands. In this study, we have examined the use of three-dimensional (3D)-printed polycaprolactone/tricalcium phosphate (PCL/TCP) composite material in bone grafting, to estimate the scope of its potential application in bone surgery. Adipose-derived stem cells (ADSCs) and bone marrow stem cells (BMSCs) are known to enhance osteointegration. We hypothesized that a patient-specific 3D-printed solid scaffold could help preserve seeded ADSCs and BMSCs and enhance osteointegration. Diffuse osteogenic tissue formation was observed by micro-computed tomography with both stem cell types, and the ADSC group displayed similar osteogenesis compared to the BMSC group. In histological assessment, the scaffold pores showed abundant ossification in both groups. Reverse transcription polymerase chain reaction (RT-PCR) showed that the BMSC group had higher expression of genes associated with ossification, and this was confirmed by Western blot analysis. The ADSC- and BMSC-seeded 3D-printed PCL/TCP scaffolds displayed promising enhancement of osteogenesis in a dog model of maxillary bone defects.

Three-dimensional bioprinting of multilayered constructs containing human mesenchymal stromal cells for osteochondral tissue regeneration in the rabbit knee joint.

The use of cell-rich hydrogels for three-dimensional (3D) cell culture has shown great potential for a variety of biomedical applications. However, the fabrication of appropriate constructs has been challenging. In this study, we describe a 3D printing process for the preparation of a multilayered 3D construct containing human mesenchymal stromal cells with a hydrogel comprised of atelocollagen and supramolecular hyaluronic acid (HA). This construct showed outstanding regenerative ability for the reconstruction of an osteochondral tissue in the knee joints of rabbits. We found that the use of a mechanically stable, host-guest chemistry-based hydrogel was essential and allowed two different types of extracellular matrix (ECM) hydrogels to be easily printed and stacked into one multilayered construct without requiring the use of potentially harmful chemical reagents or physical stimuli for post-crosslinking. To the best of our knowledge, this is the first study to validate the potential of a 3D printed multilayered construct consisting of two different ECM materials (atelocollagen and HA) for heterogeneous tissue regeneration using an in vivo animal model. We believe that this 3D printing-based platform technology can be effectively exploited for regeneration of various heterogeneous tissues as well as osteochondral tissue.