3D printing of customized bioceramics for promoting bone tissue regeneration by regulating sympathetic nerve behavior.

Numerous studies have shown that there are multiple neural activities involved in the process of bone resorption and bone regeneration, and promoting osteogenesis by promoting neural network reconstruction is an effective strategy for repairing critical size bone defects. However, traumatic bone defects often cause activation of the sympathetic nervous system (SNS) in the damaged area, releasing excess catecholamines (CAs), resulting in a decrease in the rate of bone formation. Herein, a 3D-printed scaffold loaded with propranolol (PRN) is proposed to reduce CA concentrations in bone defect areas and promote bone regeneration through drug release. For this purpose, PRN-loaded methacrylated gelatin (GelMA) microspheres were mixed with low-concentration GelMA and perfused into a 3D-printed porous hydroxyapatite (HAp) scaffold. By releasing PRN, which can block ß-adrenergic receptors, it hinders the activation of sympathetic nerves and inhibits the release of excess CA by the SNS. At the same time, the composite scaffold recruits bone marrow mesenchymal stem cells (BMSCs) and promotes the differentiation of BMSCs in the direction of osteoblasts, which effectively promotes bone regeneration in the rabbit femoral condyle defect model. The results of the study showed that the release of PRN from the composite scaffold could effectively hinder the activation of sympathetic nerves and promote bone regeneration, providing a new strategy for the treatment of bone defects.

DLP Fabrication of Multiple Hierarchical Biomimetic GelMA/SilMA/HAp Scaffolds for Enhancing Bone Regeneration.

Severe bone defects resulting from trauma and diseases remain a persistent clinical challenge. In this study, a hierarchical biomimetic microporous hydrogel composite scaffold was constructed by mimicking the hierarchical structure of bone. Initially, gelatin methacrylamide (GelMA) and methacrylic anhydride silk fibroin (SilMA) were synthesized, and GelMA/SilMA inks with suitable rheological and mechanical properties were prepared. Biomimetic micropores were then generated by using an aqueous two-phase emulsification method. Subsequently, biomimetic microporous GelMA/SilMA was mixed with hydroxyapatite (HAp) to prepare biomimetic microporous GelMA/SilMA/HAp ink. Hierarchical biomimetic microporous GelMA/SilMA/HAp (M-GSH) scaffolds were then fabricated through digital light processing (DLP) 3D printing. Finally, in vitro experiments were conducted to investigate cell adhesion, proliferation, and inward migration as well as osteogenic differentiation and vascular regeneration effects. In vivo experiments indicated that the biomimetic microporous scaffold significantly promoted tissue integration and bone regeneration after 12 weeks of implantation, achieving 42.39% bone volume fraction regeneration. In summary, this hierarchical biomimetic microporous scaffold provides a promising strategy for the repair and treatment of bone defects.

Fused Deposition Modeling Printed PLA/Nano ß-TCP Composite Bone Tissue Engineering Scaffolds for Promoting Osteogenic Induction Function.

Purpose: Large bone defects caused by congenital defects, infections, degenerative diseases, trauma, and tumors often require personalized shapes and rapid reconstruction of the bone tissue. Three-dimensional (3D)-printed bone tissue engineering scaffolds exhibit promising application potential. Fused deposition modeling (FDM) technology can flexibly select and prepare printed biomaterials and design and fabricate bionic microstructures to promote personalized large bone defect repair. FDM-3D printing technology was used to prepare polylactic acid (PLA)/nano ß-tricalcium phosphate (TCP) composite bone tissue engineering scaffolds in this study. The ability of the bone-tissue-engineered scaffold to repair bone defects was evaluated in vivo and in vitro. Methods: PLA/nano-TCP composite bone tissue engineering scaffolds were prepared using FDM-3D printing technology. The characterization data of the scaffolds were obtained using relevant detection methods. The physical and chemical properties, biocompatibility, and in vitro osteogenic capacity of the scaffolds were investigated, and their bone repair capacity was evaluated using an in vivo animal model of rabbit femur bone defects. Results: The FDM-printed PLA/nano ß-TCP composite scaffolds exhibited good personalized porosity and shape, and their osteogenic ability, biocompatibility, and bone repair ability in vivo were superior to those of pure PLA. The merits of biodegradable PLA and bioactive nano ß-TCP ceramics were combined to improve the overall biological performance of the composites. Conclusion: The FDM-printed PLA/nano-ß-TCP composite scaffold with a ratio of 7:3 exhibited good personalized porosity and shape, as well as good osteogenic ability, biocompatibility, and bone repair ability. This study provides a promising strategy for treating large bone defects.

DLP 3D printing of high-resolution root scaffold with bionic bioactivity and biomechanics for personalized bio-root regeneration.

Digital light projection (DLP) printing of hydroxyapatite (HAp) bioceramic provides a promising strategy for fabrication of complex personalized bio-tooth root scaffold with high-resolution. However, it is still a challenge to fabricate bionic bio-tooth root with satisfied bioactivity and biomechanics. This research studied the HAp-based bioceramic scaffold with bionic bioactivity and biomechanics for personalized bio-root regeneration. Compared to natural decellularized dentine (NDD) scaffolds with unitary shape and restricted mechanical properties, those DLP printing bio-tooth roots with natural size, high precision appearance, excellent structure, and a smooth surface were successfully manufactured, which met various shape and structure requirements for personalized bio-tooth regeneration. Moreover, the bioceramic sintering at 1250 °C enhanced the physicochemical properties of HAp and exhibited good elastic modulus (11.72 ± 0.53 GPa), which was almost twice of early NDD (4.76 ± 0.75 GPa). To further improve the surface activity of sintered biomimetic, the nano-HAw (nano-hydroxyapatite whiskers) coating deposited by hydrothermal treatment increased the mechanical properties and surface hydrophilicity, which indicated positive effects on dental follicle stem cells (DFSCs)' proliferation and enhanced the DFSCs osteoblastic differentiation in vitro. Subcutaneous transplantation in nude mice and in-situ transplantation in rat alveolar fossa proved that the nano-HAw-containing scaffold could promote the DFSCs differentiate into periodontal ligament-like enthesis formation. In conclusion, by combining the optimized sintering temperature and modified nano-HAw interface through hydrothermal treatment, the DLP-printing of HAp-based bioceramic with favorable bioactivity and biomechanics is a promising candidate for personalized bio-root regeneration.

In situ photo-crosslinked adhesive hydrogel loaded with mesenchymal stem cell-derived extracellular vesicles promotes diabetic wound healing.

The delayed healing of diabetic wounds is directly affected by the disturbance of wound microenvironment, resulting from persistent inflammation, insufficient angiogenesis, and impaired cell functions. Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) showed considerable therapeutic potential in diabetic wound healing. However, the low retention rate of MSC-EVs at wound sites hampers their efficacy. For skin wounds exposed to the outer environment, using a hydrogel with tissue adhesiveness under a moist wound condition is a promising strategy for wound healing. In this study, we modified methacryloyl-modified gelatin (GelMA) hydrogel with catechol motifs of dopamine to fabricate a GelMA-dopamine hydrogel. EVs isolated from MSCs were applied in the synthesized GelMA-dopamine hydrogel to prepare a GelMA-dopamine-EV hydrogel. The results demonstrated that the newly formed GelMA-dopamine hydrogel possessed improved properties of softness, adhesiveness, and absorptive capacity, as well as high biocompatibility in the working concentration (15% w/v). In addition, MSC-EVs were verified to promote cell migration and angiogenesis in vitro. In the skin wound model of diabetic rats, the GelMA-dopamine-EV hydrogel exerted prominent wound healing efficacy estimated by collagen deposition, skin appendage regeneration, and the expression of IL-6, CD31, and TGF-ß. In conclusion, this combination of MSC-EVs and the modified hydrogel not only accelerates wound closure but also promotes skin structure normalization by rescuing the homeostasis of the healing microenvironment of diabetic wounds, which provides a potential approach for the treatment of diabetic wounds.

DLP fabrication of customized porous bioceramics with osteoinduction ability for remote isolation bone regeneration.

Currently, various bioceramics have been widely used in bone regeneration. However, it remains a huge challenge to remote isolation bone regeneration, such as severed finger regeneration. The remote isolation bone tissue has a poor regenerative microenvironment that lacks enough blood and nutrition supply. It is very difficult to repair and regenerate. In this study, well-controlled multi-level porous 3D-printed calcium phosphate (CaP) bioceramic scaffolds with precision customized structures were fabricated by high-resolution digital light projection (DLP) printing technology for remote isolation bone regeneration. In vitro results demonstrated that optimizing material processing procedures could achieve multi-level control of 3D-printed CaP bioceramic scaffolds and enhance the osteoinduction ability of bioceramics effectively. In vivo results indicated that 3D-printed CaP bioceramic scaffolds constructed by optimized processing procedure exhibited a promising ability of bone regeneration and osteoinduction in ectopic osteogenesis and in situ caudal vertebrae regeneration in beagles. This study provided a promising strategy based on 3D-printed CaP bioceramic scaffolds constructed by optimized processing procedures for remote isolation bone regeneration, such as severed finger regeneration.

Preparation and characterization of biomimetic gradient multi-layer cell-laden scaffolds for osteochondral integrated repair.

A cell-laden tissue engineering scaffold for osteochondral integrated repair is one of the ideal strategies for osteochondral lesions. In this study, we fabricated cell-laden porous hydrogel scaffolds with gradient nano-hydroxyapatite using methacrylic anhydride gelatin (GelMA), nano-hydroxyapatite (nHA), and polyethylene oxide (PEO) solution for osteochondral tissue regeneration. The scaffold possessed interconnected pores and a nano-hydroxyapatite gradient in the vertical direction. The chemical, physical, mechanical, and biological properties of the hydrogel solutions and scaffolds were characterized. In vitro experiments confirmed that cells were distributed homogeneously and that different pore structures could affect the proliferation and differentiation of BMSCs. The Nonporous hydrogel was beneficial for the chondrogenic differentiation of BMSCs and interconnected pores were conducive to BMSC proliferation and osteogenic differentiation. The osteochondral integrative repair capacity of the scaffold was assessed by implanting the scaffolds into the intercondylar defect of the rabbit femur. By constructing pore structures in different layers, the cells in different layers of the hydrogels were in an intrinsic environment for survival and differentiation. Animal experiments confirmed that tissue engineering scaffolds for osteochondral lesions require different pore structures in different layers, and gradient structure facilitated integrated repair.

Three-Dimensional Printing of Large-Scale, High-Resolution Bioceramics with Micronano Inner Porosity and Customized Surface Characterization Design for Bone Regeneration.

Three-dimensional printing technologies have opened up new possibilities for manufacturing bioceramics with complex shapes in a completely digital fabrication process. Some bioceramics have demonstrated elaborate design and high resolution in their small parts through digital light projection (DLP) printing. However, it is still a challenge to prepare large-scale, high-precision ceramics that can effectively regulate the bioactivity of materials. In this study, we fabricated a large-scale hydroxyapatite porous bioceramic (length >150 mm) using DLP. This bioceramic had highly micronanoporous surface structures (printing resolution <65 µm), which could be controlled by adjusting the solid content and sintering process. Both in vitro and in vivo results indicated that the designed bioceramic had promising bone regeneration ability. This study provides significant evidence for exploring the effects of microenvironments on bone tissue regeneration. These results indicated that DLP technology has the potential to produce large-scale bone tissue engineering scaffolds with accurate porosity.

Biomimetic Methacrylated Gelatin Hydrogel Loaded With Bone Marrow Mesenchymal Stem Cells for Bone Tissue Regeneration.

Large-segment bone defect caused by trauma or tumor is one of the most challenging problems in orthopedic clinics. Biomimetic materials for bone tissue engineering have developed dramatically in the past few decades. The organic combination of biomimetic materials and stem cells offers new strategies for tissue repair, and the fate of stem cells is closely related to their extracellular matrix (ECM) properties. In this study, a photocrosslinked biomimetic methacrylated gelatin (Bio-GelMA) hydrogel scaffold was prepared to simulate the physical structure and chemical composition of the natural bone extracellular matrix, providing a three-dimensional (3D) template and extracellular matrix microenvironment. Bone marrow mesenchymal stem cells (BMSCS) were encapsulated in Bio-GelMA scaffolds to examine the therapeutic effects of ECM-loaded cells in a 3D environment simulated for segmental bone defects. In vitro results showed that Bio-GelMA had good biocompatibility and sufficient mechanical properties (14.22kPa). A rat segmental bone defect model was constructed in vivo. The GelMA-BMSC suspension was added into the PDMS mold with the size of the bone defect and photocured as a scaffold. BMSC-loaded Bio-GelMA resulted in maximum and robust new bone formation compared with hydrogels alone and stem cell group. In conclusion, the bio-GelMA scaffold can be used as a cell carrier of BMSC to promote the repair of segmental bone defects and has great potential in future clinical applications.

Chondrocyte-laden GelMA hydrogel combined with 3D printed PLA scaffolds for auricle regeneration.

The current gold standard for auricular reconstruction after microtia or ear trauma is the autologous cartilage graft with an autologous skin flap overlay. Harvesting autologous cartilage requires an additional surgery that may result in donor area complications. In addition, autologous cartilage is limited and the auricular reconstruction requires complex sculpting, which requires excellent clinical skill and is very time consuming. This work explores the use of 3D printing technology to fabricate bioactive artificial auricular cartilage using chondrocyte-laden gelatin methacrylate (GelMA) and polylactic acid (PLA) for auricle reconstruction. In this study, chondrocytes were loaded within GelMA hydrogel and combined with the 3D-printed PLA scaffolds to biomimetic the biological mechanical properties and personalized shape. The printing accuracy personalized scaffolds, biomechanics and chondrocyte viability and biofunction of artificial auricle have been studied. It was found that chondrocytes were fixed in the PLA auricle scaffolds via GelMA hydrogels and exhibited good proliferative properties and cellular activity. In addition, new chondrocytes and chondrogenic matrix, as well as type II collagen were observed after 8 weeks of implantation. At the same time, the transplanted auricle complex kept full and delicate auricle shape. This study demonstrates the potential of using 3D printing technology to construct in vitro living auricle tissue. It shows a great prospect in the clinical application of auricle regeneration.

Fabrication of customized Ti6AI4V heterogeneous scaffolds with selective laser melting: Optimization of the architecture for orthopedic implant applications.

Orthopedic implants with heterogeneous porous structures were known as ideal bone osteointegration. This research introduced the selective laser melting (SLM), finite element analysis (FEA), and a hydrothermal process (HT) for manufacturing a three-level heterogeneous porous structure. The macroporous structure was designed via CAD and micropores were tuned via laser power regulation. A nano-size layer of hydroxyapatite crystals was coated by an HT process. The mechanical properties were reinforced via a core-shell structure with core reinforcement. The existence of micropores and nano-hydroxyapatite coating enhanced the in vitro proliferation of preosteoblasts and osteogenic cellular behaviors of rBMSCs. Thus, the three-level heterogeneous porous titanium implants could inspire researchers with potential clue of cyto-implant interaction mechanism, therefore building ideal orthopedic implants with accelerated osteointegration. STATEMENT OF SIGNIFICANCE: Porous structures of titanium implants play an important role in bone tissue regeneration; The geometrical environment influence cell behaviour and bone tissue ingrowth in all macro-/micro-/nanoscale. In this study, a novel method to fabricate heterogeneous scaffolds and its macro-/micro-/nanoscopic structures were studied. A CAD model was used to obtain the macroscopic structure and the insufficient laser power was introduced for porous microstructure. Therefore, a layer of nano hydroxyapatite was coated via hydrothermal process. Cytoproliferation and cytodifferentiation results indicated that a integrity of regular/irregular, macro-/micro-/nanoscale porous structure had advance in recruiting stem cells and promoting differentiation. This research is beneficial to the development of bone implants with better bone regeneration ability.