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.

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.

Bone Fracture-Treatment Method: Fixing 3D-Printed Polycaprolactone Scaffolds with Hydrogel Type Bone-Derived Extracellular Matrix and ß-Tricalcium Phosphate as an Osteogenic Promoter.

Bone formation and growth are crucial for treating bone fractures. Improving bone-reconstruction methods using autologous bone and synthetic implants can reduce the recovery time. Here, we investigated three treatments using two different materials, a bone-derived decellularized extracellular matrix (bdECM) and ß-tricalcium phosphate (ß-TCP), individually and in combination, as osteogenic promoter between bone and 3D-printed polycaprolactone scaffold (6-mm diameter) in rat calvarial defects (8-mm critical diameter). The materials were tested with a human pre-osteoblast cell line (MG63) to determine the effects of the osteogenic promoter on bone formation in vitro. A polycaprolactone (PCL) scaffold with a porous structure was placed at the center of the in vivo rat calvarial defects. The gap between the defective bone and PCL scaffold was filled with each material. Animals were sacrificed four weeks post-implantation, and skull samples were preserved for analysis. The preserved samples were scanned by micro-computed tomography and analyzed histologically to examine the clinical benefits of the materials. The bdECM-ß-TCP mixture showed faster bone formation and a lower inflammatory response in the rats. Therefore, our results imply that a bdECM-ß-TCP mixture is an ideal osteogenic promoter for treating fractures.

Osteogenesis of 3D-Printed PCL/TCP/bdECM Scaffold Using Adipose-Derived Stem Cells Aggregates; An Experimental Study in the Canine Mandible.

Three-dimensional (3D) printing is perceived as an innovative tool for change in tissue engineering and regenerative medicine based on research outcomes on the development of artificial organs and tissues. With advances in such technology, research is underway into 3D-printed artificial scaffolds for tissue recovery and regeneration. In this study, we fabricated artificial scaffolds by coating bone demineralized and decellularized extracellular matrix (bdECM) onto existing 3D-printed polycaprolactone/tricalcium phosphate (PCL/TCP) to enhance osteoconductivity and osteoinductivity. After injecting adipose-derived stem cells (ADSCs) in an aggregate form found to be effective in previous studies, we examined the effects of the scaffold on ossification during mandibular reconstruction in beagle dogs. Ten beagles were divided into two groups: group A (PCL/TCP/bdECM + ADSC injection; n = 5) and group B (PCL/TCP/bdECM; n = 5). The results were analyzed four and eight weeks after intervention. Computed tomography (CT) findings showed that group A had more diffuse osteoblast tissue than group B. Evidence of infection or immune rejection was not detected following histological examination. Goldner trichrome (G/T) staining revealed rich ossification in scaffold pores. ColI, Osteocalcin, and Runx2 gene expressions were determined using real-time polymerase chain reaction. Group A showed greater expression of these genes. Through Western blotting, group A showed a greater expression of genes that encode ColI, Osteocalcin, and Runx2 proteins. In conclusion, intervention group A, in which the beagles received the additional ADSC injection together with the 3D-printed PCL/TCP coated with bdECM, showed improved mandibular ossification in and around the pores of the scaffold.

Bioprinting of Multiscaled Hepatic Lobules within a Highly Vascularized Construct.

Highly vascularized complex liver tissue is generally divided into lobes, lobules, hepatocytes, and sinusoids, which can be viewed under different types of lens from the micro- to macro-scale. To engineer multiscaled heterogeneous tissues, a sophisticated and rapid tissue engineering approach is required, such as advanced 3D bioprinting. In this study, a preset extrusion bioprinting technique, which can create heterogeneous, multicellular, and multimaterial structures simultaneously, is utilized for creating a hepatic lobule (≈1 mm) array. The fabricated hepatic lobules include hepatic cells, endothelial cells, and a lumen. The endothelial cells surround the hepatic cells, the exterior of the lobules, the lumen, and finally, become interconnected with each other. Compared to hepatic cell/endothelial cell mixtures, the fabricated hepatic lobule shows higher albumin secretion, urea production, and albumin, MRP2, and CD31 protein levels, as well as, cytochrome P450 enzyme activity. It is found that each cell type with spatial cell patterning in bioink accelerates cellular organization, which could preserve structural integrity and improve cellular functions. In conclusion, preset extruded hepatic lobules within a highly vascularized construct are successfully constructed, enabling both micro- and macro-scale tissue fabrication, which can support the creation of large 3D tissue constructs for multiscale tissue engineering.

Restoration of the inferomedial orbital strut using a standardized three-dimensional printing implant.

The inferomedial orbital strut (IOS) is the thin bony junction of the orbital medial wall and floor. Its fracture is common and leads to serious complications, including enophthalmos, globe dystopia and diplopia. However, anatomical restoration of the IOS is challenging owing to reduced structural support; sound anatomical background and accurate implants are therefore essential. The aim of the present study was to incorporate data from cadaveric orbit anatomy into three-dimensional (3D) printing technology and to reconstruct the complex orbital fracture elaborately. After averaging the data from computed tomography (CT) images of 100 adult cadavers, the dimensions of the IOS were extracted, and a tangent sphere was created using a computer-aided design program. The curves were compared with the CT data of 10 adult patients from the simulation test. Based on these data, a standardized 3D implant, 1.15 mm thick, was designed using polycaprolactone. The implant was placed in five patients with complex orbital fractures. The radius of the sphere in contact with the orbit, measuring 33.54 mm, was confirmed to be appropriate. A comparison between the normal side volume (V0) and the postoperative volume (Vpost ) showed that they were statistically similar. Furthermore, a comparison between V0 and the preoperative volume (Vpre ), and Vpost compared with Vpre also showed a statistically significant difference (P < 0.05). On follow-up, the preoperative ocular symptoms were resolved. The orbital data obtained from 100 cadavers provided standardized orbital anatomy, and 3D printed implants were created. The implants were anatomically accurate with regard to the orbital cavity and adequately covered the simulation model. The implant also showed satisfactory results when applied clinically in actual patients.

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.

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.

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.

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.

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).

Effect of solid freeform fabrication-based polycaprolactone/poly(lactic-co-glycolic acid)/collagen scaffolds on cellular activities of human adipose-derived stem cells and rat primary hepatocytes.

Highly biocompatible polycaprolactone (PCL)/poly(lactic-co-glycolic acid) (PLGA)/collagen scaffolds in which the PCL/PLGA collagen solution was selectively dispensed into every other space between the struts were fabricated using solid freeform fabrication (SFF) technology, as we described previously. The objective of this study was to evaluate and compare the PCL/PLGA/collagen scaffolds (group 3) with PCL/PLGA-only scaffolds (group 1) and PCL/PLGA scaffolds with collagen by the dip-coating method (group 2) using human adipose-derived stem cells (hASCs) and rat primary hepatocytes. The selectively dispensed collagen formed a three-dimensional (3D) network of nanofibers in group 3, as observed by scanning electron microscopy. The compressive strength and modulus of group 3 were approximately 140 and 510 times higher, respectively, than those of a sponge-type collagen scaffold whose weak mechanical properties were regarded as a critical drawback. Proliferation and osteogenic differentiation of hASCs were promoted significantly in group 3 compared to groups 1 and 2. In addition, we found that the viability and albumin secretion ability of rat primary hepatocytes were highly retained for 10 days in group 3 but not group 1. Interestingly, hepatocyte aggregation, which enhances hepatic function through cell-cell interactions, was observed particularly in group 3. In conclusion, group 3, in which the collagen was selectively dispensed in the 3D space of the porous PCL/PLGA framework, will be a promising 3D scaffold for culturing various cell types.

Multi-scale vascularization strategy for 3D-bioprinted tissue using coaxial core-shell pre-set extrusion bioprinting and biochemical factors.

Three-dimensional bioprinting is a key technology in bioartificial organ production. However, production of bioartificial organs has significant limitations because it is hard to build vascular structures, especially capillaries, in printed tissue owing to its low resolution. As the vascular structure plays a critical role in delivering oxygen and nutrients to cells and removing metabolic waste, building vascular channels in bioprinted tissue is essential for bioartificial organ production. In this study, we demonstrated an advanced strategy for fabricating multi-scale vascularized tissue using a pre-set extrusion bioprinting technique and endothelial sprouting. Using a coaxial precursor cartridge, mid-scale vasculature-embedded tissue was successfully fabricated. Furthermore, upon generating a biochemical gradient environment in the bioprinted tissue, capillaries were formed in this tissue. In conclusion, this strategy for multi-scale vascularization in bioprinted tissue is a promising technology for bioartificial organ production.

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.

Stimulation of healing within a rabbit calvarial defect by a PCL/PLGA scaffold blended with TCP using solid freeform fabrication technology.

The purpose of this study was to investigate the healing capacity within an 8-mm rabbit calvarial defect using a polycaprolactone (PCL)/poly(lactic-co-glycolic acid) (PLGA) scaffold blended with tri-calcium phosphate (TCP) that was constructed using solid freeform fabrication (SFF) technology. The PCL/PLGA/TCP scaffold showed a 37 % higher compressive strength and rougher surface than the PCL/PLGA scaffold. In animal experiments, new bone formation was analyzed using microcomputed tomography (micro-CT) and histological and histometric analyses. The PCL/PLGA/TCP groups had significantly greater neo-tissue areas as compared with the control groups at 4 and 8 weeks (P < 0.05). The PCL/PLGA/TCP group had significantly greater bone density as compared with the control and PCL/PLGA groups at 4 and 8 weeks (P < 0.005). The results of this study suggest that the PCL/PLGA/TCP scaffold fabricated using SFF technology is useful for recovering and enhancing new bone formation in bony defects in rabbits.

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.

Impact of microstructure on cell behavior and tissue mechanics in collagen and dermal decellularized extra-cellular matrices.

Skin models are used for many applications such as research and development or grafting. Unfortunately, most lack a proper microenvironment producing poor mechanical properties and inaccurate extra-cellular matrix composition and organization. In this report we focused on mechanical properties, extra-cellular matrix organization and cell interactions in human skin samples reconstructed with pure collagen or dermal decellularized extra-cellular matrices (S-dECM) and compared them to native human skin. We found that Full-thickness S-dECM samples presented stiffness two times higher than collagen gel and similar to ex vivo human skin, and proved for the first time that keratinocytes also impact dermal mechanical properties. This was correlated with larger fibers in S-dECM matrices compared to collagen samples and with a differential expression of F-actin, vinculin and tenascin C between S-dECM and collagen samples. This is clear proof of the microenvironment's impact on cell behaviors and mechanical properties. STATEMENT OF SIGNIFICANCE: In vitro skin models have been used for a long time for clinical applications or in vitro knowledge and evaluation studies. However, most lack a proper microenvironment producing a poor combination of mechanical properties and appropriate biological outcomes, partly due to inaccurate extra-cellular matrix (ECM) composition and organization. This can lead to limited predictivity and weakness of skin substitutes after grafting. This study shows, for the first time, the importance of a complex and rich microenvironment on cell behaviors, matrix macro- and micro-organization and mechanical properties. The increased composition and organization complexity of dermal skin decellularized extra-cellular matrix populated with differentiated cells produces in vitro skin models closer to native human skin physiology.

Bioprinting on 3D Printed Titanium Scaffolds for Periodontal Ligament Regeneration.

The three-dimensional (3D) cell-printing technique has been identified as a new biofabrication platform because of its ability to locate living cells in pre-defined spatial locations with scaffolds and various growth factors. Osseointegrated dental implants have been regarded as very reliable and have long-term reliability. However, host defense mechanisms against infections and micro-movements have been known to be impaired around a dental implant because of the lack of a periodontal ligament. In this study, we fabricated a hybrid artificial organ with a periodontal ligament on the surface of titanium using 3D printing technology. CEMP-1, a known cementogenic factor, was enhanced in vitro. In animal experiments, when the hybrid artificial organ was transplanted to the calvarial defect model, it was observed that the amount of connective tissue increased. 3D-printed hybrid artificial organs can be used with dental implants, establishing physiological tooth functions, including the ability to react to mechanical stimuli and the ability to resist infections.

Production of Multiple Cell-Laden Microtissue Spheroids with a Biomimetic Hepatic-Lobule-Like Structure.

The construction of an in vitro 3D cellular model to mimic the human liver is highly desired for drug discovery and clinical applications, such as patient-specific treatment and cell-based therapy in regenerative medicine. However, current bioprinting strategies are limited in their ability to generate multiple cell-laden microtissues with biomimetic structures. This study presents a method for producing hepatic-lobule-like microtissue spheroids using a bioprinting system incorporating a precursor cartridge and microfluidic emulsification system. The multiple cell-laden microtissue spheroids can be successfully generated at a speed of approximately 45 spheroids min-1 and with a uniform diameter. Hepatic and endothelial cells are patterned in a microtissue spheroid with the biomimetic structure of a liver lobule. The spheroids allow long-term culture with high cell viability, and the structural integrity is maintained longer than that of non-structured spheroids. Furthermore, structured spheroids show high MRP2, albumin, and CD31 expression levels. In addition, the in vivo study reveals that structured microtissue spheroids are stably engrafted. These results demonstrate that the method provides a valuable 3D structured microtissue spheroid model with lobule-like constructs and liver functions.