Bone Tissue Engineering with Adipose-Derived Stem Cells in Polycaprolactone/Graphene Oxide/Dexamethasone 3D-Printed Scaffolds.

We fabricated three-dimensional (3D)-printed polycaprolactone (PCL) and PCL/graphene oxide (GO) (PGO) scaffolds for bone tissue engineering. An anti-inflammatory and pro-osteogenesis drug dexamethasone (DEX) was adsorbed onto GO and a 3D-printed PGO/DEX (PGOD) scaffold successfully improved drug delivery with a sustained release of DEX from the scaffold up to 1 month. The physicochemical properties of the PCL, PGO, and PGOD scaffolds were characterized by various analytical techniques. The biological response of these scaffolds was studied for adherence, proliferation, and osteogenic differentiation of seeded rabbit adipose-derived stem cells (ASCs) from DNA assays, alkaline phosphatase (ALP) production, calcium quantification, osteogenic gene expression, and immunofluorescence staining of osteogenic marker proteins. The PGOD scaffold was demonstrated to be the best scaffold for maintaining cell viability, cell proliferation, and osteogenic differentiation of ASCs in vitro. In vivo biocompatibility of PGOD was confirmed from subcutaneous implantation in nude mice where ASC-seeded PGOD can form ectopic bones, demonstrated by microcomputed tomography (micro-CT) analysis and immunofluorescence staining. Furthermore, implantation of PGOD/ASCs constructs into critical-sized cranial bone defects in rabbits form tissue-engineered bones at the defect site, observed using micro-CT and histological analysis.

Astragalus polysaccharide-containing 3D-printed scaffold for traumatized skin repair and proteomic study.

Astragalus polysaccharide-containing 3D-printed scaffold shows great potential in traumatic skin repair. This study aimed to investigate its repairing effect and to combine it with proteomic technology to deeply resolve the related protein expression changes. Thirty SD rats were divided randomly into three groups (n = 10 per group): the sham-operated group, the model group and the scaffold group. Subsequently, we conducted a comparative analysis on trauma blood perfusion, trauma healing rate, histological changes, the expression of the YAP/TAZ signalling pathway and angiogenesis-related factors. Additionally, neonatal skin tissues were collected for proteomic analysis. The blood perfusion volume and wound healing recovery in the scaffold group were better than that in the model group (p < 0.05). The protein expression of STAT3, YAP, TAZ and expression of vascular-related factor A (VEGFA) in the scaffold group was higher than that in the model group (p < 0.05). Proteomic analysis showed that there were 207 differential proteins common to the three groups. Mitochondrial function, immune response, redox response, extracellular gap and ATP metabolic process were the main groups of differential protein changes. Oxidative phosphorylation, metabolic pathway, carbon metabolism, calcium signalling pathway, etc. were the main differential metabolic pathway change groups. Astragalus polysaccharide-containing 3D-printed scaffold had certain reversals of protein disorder. The Astragalus polysaccharide-containing 3D-printed scaffold may promote the VEGFs by activating the YAP/TAZ signalling pathway with the help of STAT3 into the nucleus, accelerating early angiogenesis of the trauma, correcting the protein disorder of the trauma and ultimately realizing the repair of the wound.

Targeting Bacteria-Induced Ferroptosis of Bone Marrow Mesenchymal Stem Cells to Promote the Repair of Infected Bone Defects.

The specific mechanisms underlying bacteria-triggered cell death and osteogenic dysfunction in host bone marrow mesenchymal stem cells (BMSCs) remain unclear, posing a significant challenge to the repair of infected bone defects. This study identifies ferroptosis as the predominant cause of BMSCs death in the infected bone microenvironment. Mechanistically, the bacteria-induced activation of the innate immune response in BMSCs leads to upregulation and phosphorylation of interferon regulatory factor 7 (IRF7), thus facilitating IRF7-dependent ferroptosis of BMSCs through the transcriptional upregulation of acyl-coenzyme A synthetase long-chain family member 4 (ACSL4). Moreover, it is found that intervening in ferroptosis can partially rescue cell injuries and osteogenic dysfunction. Based on these findings, a hydrogel composite 3D-printed scaffold is designed with reactive oxygen species (ROS)-responsive release of antibacterial quaternized chitosan and sustained delivery of the ferroptosis inhibitor Ferrostatin-1 (Fer-1), capable of eradicating pathogens and promoting bone regeneration in a rat model of infected bone defects. Together, this study suggests that ferroptosis of BMSCs is a promising therapeutic target for infected bone defect repair.

Bioinspired soft-hard combined system with mild photothermal therapeutic activity promotes diabetic bone defect healing via synergetic effects of immune activation and angiogenesis.

Background: The comprehensive management of diabetic bone defects remains a substantial clinical challenge due to the hostile regenerative microenvironment characterized by aggravated inflammation, excessive reactive oxygen species (ROS), bacterial infection, impaired angiogenesis, and unbalanced bone homeostasis. Thus, an advanced multifunctional therapeutic platform capable of simultaneously achieving immune regulation, bacterial elimination, and tissue regeneration is urgently designed for augmented bone regeneration under diabetic pathological milieu. Methods and Results: Herein, a photoactivated soft-hard combined scaffold system (PGCZ) was engineered by introducing polydopamine-modified zeolitic imidazolate framework-8-loaded double-network hydrogel (soft matrix component) into 3D-printed poly(ε-caprolactone) (PCL) scaffold (hard matrix component). The versatile PGCZ scaffold based on double-network hydrogel and 3D-printed PCL was thus prepared and features highly extracellular matrix-mimicking microstructure, suitable biodegradability and mechanical properties, and excellent photothermal performance, allowing long-term structural stability and mechanical support for bone regeneration. Under periodic near-infrared (NIR) irradiation, the localized photothermal effect of PGCZ triggers the on-demand release of Zn2+, which, together with repeated mild hyperthermia, collectively accelerates the proliferation and osteogenic differentiation of preosteoblasts and potently inhibits bacterial growth and biofilm formation. Additionally, the photoactivated PGCZ system also presents outstanding immunomodulatory and ROS scavenging capacities, which regulate M2 polarization of macrophages and drive functional cytokine secretion, thus leading to a pro-regenerative microenvironment in situ with enhanced vascularization. In vivo experiments further demonstrated that the PGCZ platform in conjunction with mild photothermal therapeutic activity remarkably attenuated the local inflammatory cascade, initiated endogenous stem cell recruitment and neovascularization, and orchestrated the osteoblast/osteoclast balance, ultimately accelerating diabetic bone regeneration. Conclusions: This work highlights the potential application of a photoactivated soft-hard combined system that provides long-term biophysical (mild photothermal stimulation) and biochemical (on-demand ion delivery) cues for accelerated healing of diabetic bone defects.

3D printed scaffolds with quercetin and vitamin D3 nanocarriers: In vitro cellular evaluation.

Increasing bone diseases and anomalies significantly challenge bone regeneration, necessitating the development of innovative implantable devices for effective healing. This study explores the potential of 3D-printed calcium phosphate (CaP) scaffolds functionalized with natural medicine to address this issue. Specifically, quercetin and vitamin D3 (QVD) encapsulated solid lipid nanoparticles (QVD-SLNs) are incorporated into the scaffold to enhance bone regeneration. The melt emulsification method is utilized to achieve high drug encapsulation efficiency (~98%) and controlled biphasic release kinetics. The process-structure-property performance of these systems allows more controlled release while maintaining healthy cell-material interactions. The functionalized scaffolds show ~1.3- and ~-1.6-fold increase in osteoblast cell proliferation and differentiation, respectively, as compared with the control. The treated scaffold demonstrates a reduction in osteoclastic activity as compared with the control. The QVD-SLN-loaded scaffolds show ~4.2-fold in vitro chemopreventive potential against osteosarcoma cells. Bacterial assessment with both Staphylococcus aureus and Pseudomonas aeruginosa shows a significant reduction in bacterial colony growth over the treated scaffold. These findings summarize that the release of QVD-SLNs through a 3D-printed CaP scaffold can treat various bone-related disorders for low or non-load-bearing applications.

Biomimetic Mineralized 3D-Printed Polycaprolactone Scaffold Induced by Self-Adaptive Nanotopology to Accelerate Bone Regeneration.

Three-dimensional (3D)-printed biodegradable polymer scaffolds are at the forefront of personalized constructs for bone tissue engineering. However, it remains challenging to create a biological microenvironment for bone growth. Herein, we developed a novel yet feasible approach to facilitate biomimetic mineralization via self-adaptive nanotopography, which overcomes difficulties in the surface biofunctionalization of 3D-printed polycaprolactone (PCL) scaffolds. The building blocks of self-adaptive nanotopography were PCL lamellae that formed on the 3D-printed PCL scaffold via surface-directed epitaxial crystallization and acted as a linker to nucleate and generate hydroxyapatite crystals. Accordingly, a uniform and robust mineralized layer was immobilized throughout the scaffolds, which strongly bound to the strands and had no effect on the mechanical properties of the scaffolds. In vitro cell culture experiments revealed that the resulting scaffold was biocompatible and enhanced the proliferation and osteogenic differentiation of mouse embryolous osteoblast cells. Furthermore, we demonstrated that the resulting scaffold showed a strong capability to accelerate in vivo bone regeneration using a rabbit bone defect model. This study provides valuable opportunities to enhance the application of 3D-printed scaffolds in bone repair, paving the way for translation to other orthopedic implants.

A 3D-Printed Scaffold for Repairing Bone Defects.

The treatment of bone defects has always posed challenges in the field of orthopedics. Scaffolds, as a vital component of bone tissue engineering, offer significant advantages in the research and treatment of clinical bone defects. This study aims to provide an overview of how 3D printing technology is applied in the production of bone repair scaffolds. Depending on the materials used, the 3D-printed scaffolds can be classified into two types: single-component scaffolds and composite scaffolds. We have conducted a comprehensive analysis of material composition, the characteristics of 3D printing, performance, advantages, disadvantages, and applications for each scaffold type. Furthermore, based on the current research status and progress, we offer suggestions for future research in this area. In conclusion, this review acts as a valuable reference for advancing the research in the field of bone repair scaffolds.

Enhanced osteochondral regeneration with a 3D-Printed biomimetic scaffold featuring a calcified interfacial layer.

The integrative regeneration of both articular cartilage and subchondral bone remains an unmet clinical need due to the difficulties of mimicking spatial complexity in native osteochondral tissues for artificial implants. Layer-by-layer fabrication strategies, such as 3D printing, have emerged as a promising technology replicating the stratified zonal architecture and varying microstructures and mechanical properties. However, the dynamic and circulating physiological environments, such as mass transportation or cell migration, usually distort the pre-confined biological properties in the layered implants, leading to undistinguished spatial variations and subsequently inefficient regenerations. This study introduced a biomimetic calcified interfacial layer into the scaffold as a compact barrier between a cartilage layer and a subchondral bone layer to facilitate osteogenic-chondrogenic repair. The calcified interfacial layer consisting of compact polycaprolactone (PCL), nano-hydroxyapatite, and tasquinimod (TA) can physically and biologically separate the cartilage layer (TA-mixed, chondrocytes-load gelatin methacrylate) from the subchondral bond layer (porous PCL). This introduction preserved the as-designed independent biological environment in each layer for both cartilage and bone regeneration, successfully inhibiting vascular invasion into the cartilage layer and preventing hyaluronic cartilage calcification owing to devascularization of TA. The improved integrative regeneration of cartilage and subchondral bone was validated through gross examination, micro-computed tomography (micro-CT), and histological and immunohistochemical analyses based on an in vivo rat model. Moreover, gene and protein expression studies identified a key role of Caveolin (CAV-1) in promoting angiogenesis through the Wnt/ß-catenin pathway and indicated that TA in the calcified layer blocked angiogenesis by inhibiting CAV-1.

3D-printed PCL@BG scaffold integrated with SDF-1α-loaded hydrogel for enhancing local treatment of bone defects.

BACKGROUND: The long-term nonunion of bone defects is always a difficult problem in orthopaedics treatment. Artificial bone implants made of polymeric materials are expected to solve this problem due to their suitable degradation rate and good biocompatibility. However, the lack of mechanical strength, low osteogenic induction ability and poor hydrophilicity of these synthetic polymeric materials limit their large-scale clinical application. RESULTS: In this study, we used bioactive glass (BG) (20%, W/W) and polycaprolactone (PCL, 80%, W/W) as raw materials to prepare a bone repair scaffold (PCL@BG20) using fused deposition modelling (FDM) three-dimensional (3D) printing technology. Subsequently, stromal cell-derived factor-1α (SDF-1α) chemokines were loaded into the PCL@BG20 scaffold pores with gelatine methacryloyl (GelMA) hydrogel. The experimental results showed that the prepared scaffold had a porous biomimetic structure mimicking that of cancellous bone, and the compressive strength (44.89 ± 3.45 MPa) of the scaffold was similar to that of cancellous bone. Transwell experiments showed that scaffolds loaded with SDF-1α could promote the recruitment of bone marrow stromal cells (BMSCs). In vivo data showed that treatment with scaffolds containing SDF-1α and BG (PCL@BG-GelMA/SDF-1α) had the best effect on bone defect repair compared to the other groups, with a large amount of new bone and mature collagen forming at the bone defect site. No significant organ toxicity or inflammatory reactions were observed in any of the experimental groups. CONCLUSIONS: The results show that this kind of scaffold containing BG and SDF-1α serves the dual functions of recruiting stem cell migration in vivo and promoting bone repair in situ. We envision that this scaffold may become a new strategy for the clinical treatment of bone defects.

Three Birds, One Stone: An Osteo-Microenvironment Stage-Regulative Scaffold for Bone Defect Repair through Modulating Early Osteo-Immunomodulation, Middle Neovascularization, and Later Osteogenesis.

In order to repair critical-sized bone defects, various polylactic acid-glycolic acid (PLGA)-based hybrid scaffolds are successfully developed as bone substitutes. However, the byproducts of these PLGA-based scaffolds are known to acidify the implanted site, inducing tiresome acidic inflammation. Moreover, these degradation productions cannot offer an osteo-friendly microenvironment at the implanted site, matching natural bone healing. Herein, inspired by bone microenvironment atlas of natural bone-healing process, an osteo-microenvironment stage-regulative scaffold (P80/D10/M10) is fabricated by incorporating self-developed decellularized bone matrix microparticles (DBM-MPs) and multifunctional magnesium hydroxide nanoparticles (MH-NPs) into PLGA with an optimized proportion using low-temperature rapid prototyping (LT-RP) 3D-printing technology. The cell experiments show that this P80/D10/M10 exhibits excellent properties in mechanics, biocompatibility, and biodegradability, meanwhile superior stimulations in osteo-immunomodulation, angiogenesis, and osteogenesis. Additionally, the animal experiments determined that this P80/D10/M10 can offer an osteo-friendly microenvironment in a stage-matched pattern for enhanced bone regeneration, namely, optimization of early inflammation, middle neovascularization, and later bone formation. Furthermore, transcriptomic analysis suggested that the in vivo performance of P80/D10/M10 on bone defect repair is mostly attributed to regulating artery development, bone development, and bone remodeling. Overall, this study reveals that the osteo-microenvironment stage-regulative scaffold provides a promising treatment for bone defect repair.

Collagen-Coated Hyperelastic Bone Promotes Osteoblast Adhesion and Proliferation.

Successfully reconstructing bone and restoring its dynamic function represents a significant challenge for medicine. Critical size defects (CSDs), resulting from trauma, tumor removal, or degenerative conditions, do not naturally heal and often require complex bone grafting. However, these grafts carry risks, such as tissue rejection, infections, and surgical site damage, necessitating the development of alternative treatments. Three-dimensional and four-dimensional printed synthetic biomaterials represent a viable alternative, as they carry low production costs and are highly reproducible. Hyperelastic bone (HB), a biocompatible synthetic polymer consisting of 90% hydroxyapatite and 10% poly(lactic-co-glycolic acid, PLGA), was examined for its potential to support cell adhesion, migration, and proliferation. Specifically, we seeded collagen-coated HB with MG-63 human osteosarcoma cells. Our analysis revealed robust cell adhesion and proliferation over 7 days in vitro, with cells forming uniform monolayers on the external surface of the scaffold. However, no cells were present on the core of the fibers. The cells expressed bone differentiation markers on days 3 and 5. By day 7, the scaffold began to degrade, developing microscopic fissures and fragmentation. In summary, collagen-coated HB scaffolds support cell adhesion and proliferation but exhibit reduced structural support after 7 days in culture. Nevertheless, the intricate 3D architecture holds promise for cellular migration, vascularization, and early osteogenesis.

A tissue engineered 3D printed calcium alkali phosphate bioceramic bone graft enables vascularization and regeneration of critical-size discontinuity bony defects in vivo.

Introduction: Recently, efforts towards the development of patient-specific 3D printed scaffolds for bone tissue engineering from bioactive ceramics have continuously intensified. For reconstruction of segmental defects after subtotal mandibulectomy a suitable tissue engineered bioceramic bone graft needs to be endowed with homogenously distributed osteoblasts in order to mimic the advantageous features of vascularized autologous fibula grafts, which represent the standard of care, contain osteogenic cells and are transplanted with the respective blood vessel. Consequently, inducing vascularization early on is pivotal for bone tissue engineering. The current study explored an advanced bone tissue engineering approach combining an advanced 3D printing technique for bioactive resorbable ceramic scaffolds with a perfusion cell culture technique for pre-colonization with mesenchymal stem cells, and with an intrinsic angiogenesis technique for regenerating critical size, segmental discontinuity defects in vivo applying a rat model. To this end, the effect of differing Si-CAOP (silica containing calcium alkali orthophosphate) scaffold microarchitecture arising from 3D powder bed printing (RP) or the Schwarzwalder Somers (SSM) replica fabrication technique on vascularization and bone regeneration was analyzed in vivo. In 80 rats 6-mm segmental discontinuity defects were created in the left femur. Methods: Embryonic mesenchymal stem cells were cultured on RP and SSM scaffolds for 7d under perfusion to create Si-CAOP grafts with terminally differentiated osteoblasts and mineralizing bone matrix. These scaffolds were implanted into the segmental defects in combination with an arteriovenous bundle (AVB). Native scaffolds without cells or AVB served as controls. After 3 and 6 months, femurs were processed for angio-µCT or hard tissue histology, histomorphometric and immunohistochemical analysis of angiogenic and osteogenic marker expression. Results: At 3 and 6 months, defects reconstructed with RP scaffolds, cells and AVB displayed a statistically significant higher bone area fraction, blood vessel volume%, blood vessel surface/volume, blood vessel thickness, density and linear density than defects treated with the other scaffold configurations. Discussion: Taken together, this study demonstrated that the AVB technique is well suited for inducing adequate vascularization of the tissue engineered scaffold graft in segmental defects after 3 and 6 months, and that our tissue engineering approach employing 3D powder bed printed scaffolds facilitated segmental defect repair.

3D-printed placental-derived bioinks for skin tissue regeneration with improved angiogenesis and wound healing properties.

Extracellular matrix (ECM)-based bioinks has attracted much attention in recent years for 3D printing of native-like tissue constructs. Due to organ unavailability, human placental ECM can be an alternative source for the construction of 3D print composite scaffolds for the treatment of deep wounds. In this study, we use different concentrations (1.5%, 3% and 5%w/v) of ECM derived from the placenta, sodium-alginate and gelatin to prepare a printable bioink biomimicking natural skin. The printed hydrogels' morphology, physical structure, mechanical behavior, biocompatibility, and angiogenic property are investigated. The optimized ECM (5%w/v) 3D printed scaffold is applied on full-thickness wounds created in a mouse model. Due to their unique native-like structure, the ECM-based scaffolds provide a non-cytotoxic microenvironment for cell adhesion, infiltration, angiogenesis, and proliferation. In contrast, they do not show any sign of immune response to the host. Notably, the biodegradation, swelling rate, mechanical property, cell adhesion and angiogenesis properties increase with the increase of ECM concentrations in the construct. The ECM 3D printed scaffold implanted into deep wounds increases granulation tissue formation, angiogenesis, and re-epithelialization due to the presence of ECM components in the construct, when compared with printed scaffold with no ECM and no treatment wound. Overall, our findings demonstrate that the 5% ECM 3D scaffold supports the best deep wound regeneration in vivo, produces a skin replacement with a cellular structure comparable to native skin.

3D-Printed Bioactive Scaffold Loaded with GW9508 Promotes Critical-Size Bone Defect Repair by Regulating Intracellular Metabolism.

The process of bone regeneration is complicated, and it is still a major clinical challenge to regenerate critical-size bone defects caused by severe trauma, infection, and tumor resection. Intracellular metabolism has been found to play an important role in the cell fate decision of skeletal progenitor cells. GW9508, a potent agonist of the free fatty acid receptors GPR40 and GPR120, appears to have a dual effect of inhibiting osteoclastogenesis and promoting osteogenesis by regulating intracellular metabolism. Hence, in this study, GW9508 was loaded on a scaffold based on biomimetic construction principles to facilitate the bone regeneration process. Through 3D printing and ion crosslinking, hybrid inorganic-organic implantation scaffolds were obtained after integrating 3D-printed ß-TCP/CaSiO3 scaffolds with a Col/Alg/HA hydrogel. The 3D-printed ß-TCP/CaSiO3 scaffolds had an interconnected porous structure that simulated the porous structure and mineral microenvironment of bone, and the hydrogel network shared similar physicochemical properties with the extracellular matrix. The final osteogenic complex was obtained after GW9508 was loaded into the hybrid inorganic-organic scaffold. To investigate the biological effects of the obtained osteogenic complex, in vitro studies and a rat cranial critical-size bone defect model were utilized. Metabolomics analysis was conducted to explore the preliminary mechanism. The results showed that 50 µM GW9508 facilitated osteogenic differentiation by upregulating osteogenic genes, including Alp, Runx2, Osterix, and Spp1 in vitro. The GW9508-loaded osteogenic complex enhanced osteogenic protein secretion and facilitated new bone formation in vivo. Finally, the results from metabolomics analysis suggested that GW9508 promoted stem cell differentiation and bone formation through multiple intracellular metabolism pathways, including purine and pyrimidine metabolism, amino acid metabolism, glutathione metabolism, and taurine and hypotaurine metabolism. This study provides a new approach to address the challenge of critical-size bone defects.

3D printed pore morphology mediates bone marrow stem cell behaviors via RhoA/ROCK2 signaling pathway for accelerating bone regeneration.

Bone bionics and structural engineering have sparked a broad interest in optimizing artificial scaffolds for better bone regeneration. However, the mechanism behind scaffold pore morphology-regulated bone regeneration remains unclear, making the structure design of scaffolds for bone repair challenging. To address this issue, we have carefully assessed diverse cell behaviors of bone mesenchymal stem cells (BMSCs) on the ß-tricalcium phosphate (ß-TCP) scaffolds with three representative pore morphologies (i.e., cross column, diamond, and gyroid pore unit, respectively). Among the scaffolds, BMSCs on the ß-TCP scaffold with diamond pore unit (designated as D-scaffold) demonstrated enhanced cytoskeletal forces, elongated nucleus, faster cell mobility, and better osteogenic differentiation potential (for example, the alkaline phosphatase expression level in D-scaffold were 1.5-2 times higher than other groups). RNA-sequencing analysis and signaling pathway intervention revealed that Ras homolog gene family A (RhoA)/Rho-associated kinase-2 (ROCK2) has in-depth participated in the pore morphology-mediated BMSCs behaviors, indicating an important role of mechanical signaling transduction in scaffold-cell interactions. Finally, femoral condyle defect repair results showed that D-scaffold could effectively promote endogenous bone regeneration, of which the osteogenesis rate was 1.2-1.8 times higher than the other groups. Overall, this work provides insights into pore morphology-mediated bone regeneration mechanisms for developing novel bioadaptive scaffold designs.

Black phosphorus nanosheets-enabled DNA hydrogel integrating 3D-printed scaffold for promoting vascularized bone regeneration.

The classical 3D-printed scaffolds have attracted enormous interests in bone regeneration due to the customized structural and mechanical adaptability to bone defects. However, the pristine scaffolds still suffer from the absence of dynamic and bioactive microenvironment that is analogous to natural extracellular matrix (ECM) to regulate cell behaviour and promote tissue regeneration. To address this challenge, we develop a black phosphorus nanosheets-enabled dynamic DNA hydrogel to integrate with 3D-printed scaffold to build a bioactive gel-scaffold construct to achieve enhanced angiogenesis and bone regeneration. The black phosphorus nanosheets reinforce the mechanical strength of dynamic self-healable hydrogel and endow the gel-scaffold construct with preserved protein binding to achieve sustainable delivery of growth factor. We further explore the effects of this activated construct on both human umbilical vein endothelial cells (HUVECs) and mesenchymal stem cells (MSCs) as well as in a critical-sized rat cranial defect model. The results confirm that the gel-scaffold construct is able to promote the growth of mature blood vessels as well as induce osteogenesis to promote new bone formation, indicating that the strategy of nano-enabled dynamic hydrogel integrated with 3D-printed scaffold holds great promise for bone tissue engineering.

Production of 3D Printed Bi-Layer and Tri-Layer Sandwich Scaffolds with Polycaprolactone and Poly (vinyl alcohol)-Metformin towards Diabetic Wound Healing.

Type 2 diabetes mellitus (T2DM) is a chronic disease characterized by impaired insulin secretion, sensitivity, and hyperglycemia. Diabetic wounds are one of the significant complications of T2DM owing to its difficulty in normal healing, resulting in chronic wounds. In the present work, PCL/PVA, PCL/PVA/PCL, and metformin-loaded, PCL/PVA-Met and PCL/PVA-Met/PCL hybrid scaffolds with different designs were fabricated using 3D printing. The porosity and morphological analysis of 3D-printed scaffolds were performed using scanning electron microscopy (SEM). The scaffolds' average pore sizes were between 63.6 ± 4.0 and 112.9 ± 3.0 µm. Molecular and chemical interactions between polymers and the drug were investigated with Fourier transform infrared spectroscopy (FT-IR) and X-ray diffraction (XRD). Mechanical, thermal, and degradation analysis of the scaffolds were undertaken to investigate the physico-chemical characteristics of the scaffolds. Owing to the structure, PCL/PVA/PCL sandwich scaffolds had lower degradation rates than the bi-layer scaffolds. The drug release of the metformin-loaded scaffolds was evaluated with UV spectrometry, and the biocompatibility of the scaffolds on fibroblast cells was determined by cell culture analysis. The drug release in the PCL/PVA-Met scaffold was sustained till six days, whereas in the PCL/PVA-Met/PCL, it continued for 31 days. In the study of drug release kinetics, PCL/PVA-Met and PCL/PVA-Met/PCL scaffolds showed the highest correlation coefficients (R2) values for the first-order release model at 0.8735 and 0.889, respectively. Since the layered structures in the literature are mainly obtained with the electrospun fiber structures, these biocompatible sandwich scaffolds, produced for the first time with 3D-printing technology, may offer an alternative to existing drug delivery systems and may be a promising candidate for enhancing diabetic wound healing.

PCL strut-like scaffolds appear superior to gyroid in terms of bone regeneration within a long bone large defect: An in silico study.

The treatment of large bone defects represents a major clinical challenge. 3D printed scaffolds appear as a promising strategy to support bone defect regeneration. The 3D design of such scaffolds impacts the healing path and thus defect regeneration potential. Among others, scaffold architecture has been shown to influence the healing outcome. Gyroid architecture, characterized by a zero mean surface curvature, has been discussed as a promising scaffold design for bone regeneration. However, whether gyroid scaffolds are favourable for bone regeneration in large bone defects over traditional strut-like architecture scaffolds remains unknown. Therefore, the aim of this study was to investigate whether gyroid scaffolds present advantages over more traditional strut-like scaffolds in terms of their bone regeneration potential. Validated bone defect regeneration principles were applied in an in silico modeling approach that allows to predict bone formation in defect regeneration. Towards this aim, the mechano-biological bone regeneration principles were adapted to allow simulating bone regeneration within both gyroid and strut-like scaffolds. We found that the large surface curvatures of the gyroid scaffold led to a slower tissue formation dynamic and conclusively reduced bone regeneration. The initial claim, that an overall reduced zero mean surface curvature would enhance bone formation, could not be confirmed. The here presented approach illustrates the potential of in silico tools to evaluate in pre-clinical studies scaffold designs and eventually lead to optimized architectures of 3D printed implants for bone regeneration.

Sulfated carboxymethyl cellulose and carboxymethyl κ-carrageenan immobilization on 3D-printed poly-ε-caprolactone scaffolds differentially promote pre-osteoblast proliferation and osteogenic activity.

The lack of bioactivity in three-dimensional (3D)-printing of poly-є-caprolactone (PCL) scaffolds limits cell-material interactions in bone tissue engineering. This constraint can be overcome by surface-functionalization using glycosaminoglycan-like anionic polysaccharides, e.g., carboxymethyl cellulose (CMC), a plant-based carboxymethylated, unsulfated polysaccharide, and κ-carrageenan, a seaweed-derived sulfated, non-carboxymethylated polysaccharide. The sulfation of CMC and carboxymethylation of κ-carrageenan critically improve their bioactivity. However, whether sulfated carboxymethyl cellulose (SCMC) and carboxymethyl κ-carrageenan (CM-κ-Car) affect the osteogenic differentiation potential of pre-osteoblasts on 3D-scaffolds is still unknown. Here, we aimed to assess the effects of surface-functionalization by SCMC or CM-κ-Car on the physicochemical and mechanical properties of 3D-printed PCL scaffolds, as well as the osteogenic response of pre-osteoblasts. MC3T3-E1 pre-osteoblasts were seeded on 3D-printed PCL scaffolds that were functionalized by CM-κ-Car (PCL/CM-κ-Car) or SCMC (PCL/SCMC), cultured up to 28 days. The scaffolds' physicochemical and mechanical properties and pre-osteoblast function were assessed experimentally and by finite element (FE) modeling. We found that the surface-functionalization by SCMC and CM-κ-Car did not change the scaffold geometry and structure but decreased the elastic modulus. Furthermore, the scaffold surface roughness and hardness increased and the scaffold became more hydrophilic. The FE modeling results implied resilience up to 2% compression strain, which was below the yield stress for all scaffolds. Surface-functionalization by SCMC decreased Runx2 and Dmp1 expression, while surface-functionalization by CM-κ-Car increased Cox2 expression at day 1. Surface-functionalization by SCMC most strongly enhanced pre-osteoblast proliferation and collagen production, while CM-κ-Car most significantly increased alkaline phosphatase activity and mineralization after 28 days. In conclusion, surface-functionalization by SCMC or CM-κ-Car of 3D-printed PCL-scaffolds enhanced pre-osteoblast proliferation and osteogenic activity, likely due to increased surface roughness and hydrophilicity. Surface-functionalization by SCMC most strongly enhanced cell proliferation, while CM-κ-Car most significantly promoted osteogenic activity, suggesting that surface-functionalization by CM-κ-Car may be more promising, especially in the short-term, for in vivo bone formation.

Repair of Large-Scale Rib Defects Based on Steel-Reinforced Concrete-Designed Biomimetic 3D-Printed Scaffolds with Bone-Mineralized Microenvironments.

Tissue engineering technology provides a promising approach for large-scale bone reconstruction in cases of extensive chest wall defects. However, previous studies did not consider meticulous scaffold design specific to large-scale rib regeneration in terms of three-dimensional (3D) shape, proper porous structures, enough mechanical strength, and osteogenic microenvironments. Thus, there is an urgent need to develop an appropriate bone biomimetic scaffold (BBS) to address this problem. In this study, a BBS with controllable 3D morphology, appropriate mechanical properties, good biocompatibility and biodegradability, porous structure suitable for cell loading, and a biomimetic osteogenic inorganic salt (OIS) microenvironment was successfully prepared by integrating computer-aided design, 3D-printing, cast-molding, and freeze-drying technologies. The addition of the OIS in the scaffold substantially promoted ectopic bone regeneration in vivo, which might be attributed to the activation of osteogenic and angiogenic signaling pathways as well as upregulated expression of osteogenic genes. More importantly, dual long rib defects could be successfully repaired and medullary cavity recanalized by the rib-shaped mature cortical bone, which might be mediated by the activation of osteoclast signaling pathways. Thus, this paper presents a reliable BBS and proposes a new strategy for the repair of large-scale bone defects.