Towards the Clinical Translation of 3D PLGA/ß-TCP/Mg Composite Scaffold for Cranial Bone Regeneration.

Recent years have witnessed the rapid development of 3D porous scaffolds with excellent biocompatibility, tunable porosity, and pore interconnectivity, sufficient mechanical strength, controlled biodegradability, and favorable osteogenesis for improved results in cranioplasty. However, clinical translation of these scaffolds has lagged far behind, mainly because of the absence of a series of biological evaluations. Herein, we designed and fabricated a composite 3D porous scaffold composed of poly (lactic-co-glycolic) acid (PLGA), ß-tricalcium phosphate (ß-TCP), and Mg using the low-temperature deposition manufacturing (LDM) technique. The LDM-engineered scaffolds possessed highly porous and interconnected microstructures with a porosity of 63%. Meanwhile, the scaffolds exhibited mechanical properties close to that of cancellous bone, as confirmed by the compression tests. It was also found that the original composition of scaffolds could be maintained throughout the fabrication process. Particularly, two important biologic evaluations designed for non-active medical devices, i.e., local effects after implantation and subchronic systemic toxicity tests, were conducted to evaluate the local and systemic toxicity of the scaffolds. Additionally, the scaffolds exhibited significant higher mRNA levels of osteogenic genes compared to control scaffolds, as confirmed by an in vitro osteogenic differentiation test of MC3T3-E1 cells. Finally, we demonstrated the improved cranial bone regeneration performance of the scaffolds in a rabbit model. We envision that our investigation could pave the way for translating the LDM-engineered composite scaffolds into clinical products for cranial bone regeneration.

Divalent Anion-Induced Biohydrogels with High Strength, Anti-swelling, and Bioactive Capability for Enhanced Skull Bone Regeneration.

Increased intracranial pressure after traumatic brain injury (TBI) is an urgent problem in clinical practice. A pliable hydrogel is preferred for cranioplasty applications after TBI since it can protect brain tissue and promote bone healing. Nevertheless, biohydrogels for cranial bone regeneration still face challenges of poor mechanical properties, large swelling ratios, and low osteogenesis activity. Herein, inspired by Hofmeister effects, biopolymer hydrogels composed of protein and polysaccharides were treated with a Hofmeister series including a series of monovalent and divalent anions. Our results reveal that the divalent anion-cross-linked biohydrogels exhibit stronger mechanical properties and lower swelling ratios compared with monovalent-anion treated gels. Intriguingly, the divalent HPO42- anion induced biohybrid hydrogels with excellent mechanical behaviors (3.7 ± 0.58 MPa, 484 ± 76.7 kPa, and 148.3 ± 6.85 kJ/m3), anti-swelling capability (16.7%), and gradual degradation ability, significantly stimulating osteogenic differentiation and in vivo cranial bone regeneration. Overall, this study may provide new insights into the design of biomimetic hydrogels for treating cranial bone defects after TBI.

Nanoclay Promotes Mouse Cranial Bone Regeneration Mainly through Modulating Drug Binding and Sustained Release.

Nanoclay (Nanosilicates, NS) is appearing as an intriguing 2D nanomaterial for bone tissue engineering with multiple proposed functions, e.g., intrinsic osteoinductivity, improving mechanical properties, and drug release capacity. However, the mechanism of NS for in vivo bone regeneration has been hardly defined so far. This knowledge gap will significantly affect the design/application of NS-based biomaterials. To determine the role of NS in osteoblastic differentiation and bone formation, we used the mouse calvarial-derived pre-osteoblasts (MC3T3-E1) and a clinically-relevant mouse cranial bone defect model. Instead of a hydrogel, we prepared biomimetic 3D gelatin nanofibrous scaffolds (GF) and NS-blended composite scaffolds (GF/NS) to determine the essential role of NS in critical low-dose (0.5 µg per scaffold) of BMP2-induced cranial bone regeneration. In contrast to "osteoinductivity", our data indicated that NS could enable single-dose of BMP2, promoting significant osteoblastic differentiation while multiple-dose of BMP2 (without NS) was required to achieve similar efficacy. Moreover, our release study revealed that direct binding to NS in GF scaffolds provided stronger protection to BMP2 and sustained release compared to GF/NS composite scaffolds. Consistently, our in vivo data indicated that only BMP2/NS direct binding treatment was able to repair the large mouse cranial bone defects after 6 weeks of transplantation while neither BMP2, NS alone, nor BMP2 released from GF/NS scaffolds was sufficient to induce significant cranial bone defect repair. Therefore, we concluded that direct nanoclay-drug binding enabled sustained release is the most critical contribution to the significantly improved bone regeneration compared to other possible mechanisms based on our study.

Tuning pore features of mineralized collagen/PCL scaffolds for cranial bone regeneration in a rat model.

Porosity is indispensable for a bone tissue-engineered scaffold for facilitating endogenous cell migration and nascent bone ingrowth. In large-sized cranial bone defect repair, porous scaffolds meet great challenges to match cranial bone regeneration and provide sufficient protection with structural integrity. Therefore, the pore features of the scaffolds for cranial bone regeneration should differ from those typical porous scaffolds used in tubular bone repair and be finely tuned. In this study, a series of porous mineralized collagen/PCL scaffolds with different pore features were fabricated via freeze-drying and applied in a Sprague Dawley rat cranial bone calvarial defect model. The pore size for four groups increased from 10-45 µm to 40-130 µm. As scaffold porosity increased, the compressive strength decreased from 2.09 ±â€¯0.12 MPa to 0.51 ±â€¯0.04 MPa. The micro-computed tomography three-dimensional reconstruction images showed that as pore size and porosity increased, the amount of new bone formation had a maximum in group 3 (pore size: 20-100 µm, compressive strength: 1.06 ±â€¯0.03 MPa). In addition, the histological and histomorphometric analyses showed a consistent tendency which confirmed the Micro-CT results. Meanwhile, histological findings including bony bridging, tissue response at the bone-implant interface and fibrous capsule thickness indicated that the dura mater pathway played the most important role in the regenerative process of this calvarial defect model.

Role of HA and BG in engineering poly(ε-caprolactone) porous scaffolds for accelerating cranial bone regeneration.

Effects of varied bioactive fillers on the biological behavior of porous polymer/inorganic composite scaffolds are lack of comprehensive comparison and remain elusive. Moreover, composite scaffolds with high porosity suffer from inferior mechanical performance. Herein, high-pressure molding and salt leaching were employed to prepare poly(ε-caprolactone) (PCL) composite porous scaffolds loaded with hydroxyapatite (HA) and bioactive glass (BG), respectively. Structural analysis indicated all the porous scaffolds presented interconnected open-pore structure with the porosity of ~87% and pore size of ~180 µm, hinging on the amounts and size of porogen. Compared to PCL/HA scaffolds, PCL/BG scaffolds showed ~2.3-fold augment in the water absorption. Attributing to the compact framework, the PCL/HA and PCL/BG porous scaffolds exhibited outstanding compressive modulus, which was notably higher than other PCL composite porous scaffolds reported in literatures. Cells culture results demonstrated that PCL/BG scaffolds displayed higher expression of osteogenic differentiation than PCL and PCL/HA scaffolds. Furthermore, in vivo results showed that more mature bone was formed within PCL/BG scaffolds than PCL/HA scaffolds, manifesting that the introduction of BG accelerated cranial bone regeneration to obtain complete bone healing within a short time. Therefore, these data indicate that PCL/BG scaffolds are more competitive for bone tissue engineering application. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 654-662, 2019.

Polydopamine-Assisted Anchor of Chitosan onto Porous Composite Scaffolds for Accelerating Bone Regeneration.

Surface function has an importance for the bioactivity of porous polymeric scaffolds. The goal of the present study is to immobilize highly bioactive chitosan (CS) onto the surface of porous composite scaffolds to accelerate bone regeneration. Porous poly(ε-caprolactone) (PCL)/bioactive glass (BG) composite scaffolds with strong anchor of CS were fabricated via mussel-inspired polydopamine (PDA) coating as a bridging layer. In vitro cell culture showed that firm immobilization of CS onto the composite scaffolds significantly enhanced protein adsorption, cell adhesion, and osteogenic differentiation compared to CS-decorated scaffolds via physical adsorption. In vivo assessments demonstrated that covalent immobilization of CS onto the surface of scaffolds obviously promoted cranial bone regeneration in comparison with the counterparts with physical adsorption of CS. The proposed method offers a feasible and effective means to fabricate artificial bioactive scaffolds for bone tissue engineering application.

Novel 3D Hybrid Nanofiber Aerogels Coupled with BMP-2 Peptides for Cranial Bone Regeneration.

An ideal synthetic bone graft is a combination of the porous and nanofibrous structure presented by natural bone tissue as well as osteoinductive biochemical factors such as bone morphogenetic protein 2 (BMP-2). In this work, ultralight 3D hybrid nanofiber aerogels composed of electrospun PLGA-collagen-gelatin and Sr-Cu codoped bioactive glass fibers with incorporation of heptaglutamate E7 domain specific BMP-2 peptides have been developed and evaluated for their potential in cranial bone defect healing. The nanofiber aerogels are surgically implanted into 8 mm × 1 mm (diameter × thickness) critical-sized defects created in rat calvariae. A sustained release of E7-BMP-2 peptide from the degradable hybrid aerogels significantly enhances bone healing and defect closure over 8 weeks in comparison to unfilled defects. Histomorphometry and X-ray microcomputed tomography (µ-CT) analysis reveal greater bone volume and bone formation area in case of the E7-BMP-2 peptide loaded hybrid nanofiber aerogels. Further, histopathology data divulged a near complete nanofiber aerogel degradation along with enhanced vascularization of the regenerated tissue. Together, this study for the first time demonstrates the fabrication of 3D hybrid nanofiber aerogels from 2D electrospun fibers and their loading with therapeutic osteoinductive BMP-2 mimicking peptide for cranial bone tissue regeneration.

Cranial bone regeneration via BMP-2 encoding mesenchymal stem cells.

Cranial bone repair and regeneration via tissue engineering principles has attracted a great deal of interest from researchers during last decade. Here, within this study, 6 mm critical-sized bone defect regeneration via genetically modified mesenchymal stem cells (MSC) were monitored up to 4 months. Cranial bone repair and new bone formations were evaluated by histological staining and real time PCR analysis in five different groups including autograft and bone morphogenetic protein-2 (BMP-2) transfected MSC groups. Results presented here indicate a proper cranial regeneration in autograft groups and a prospering regeneration for hBMP-2 encoding mesenchymal stem cells.

High-Pressure Compression-Molded Porous Resorbable Polymer/Hydroxyapatite Composite Scaffold for Cranial Bone Regeneration.

Fabricating porous scaffolds with sufficient mechanical properties is a challenge for healing bone defects. High-pressure compression-molded (HPCM) porous composite scaffold comprising poly(l-lactide) (PLLA), poly(lactide-co-glycolide) (PLGA), and hydroxyapatite (HA) was prepared and showed upregulated mechanical properties due to a solid network structure and a highly ordered crystalline architecture. The compressive yield strength and modulus of the HPCM scaffold molded at 1000 MPa and 180 °C were 0.91 and 6.84 MPa, respectively. The HPCM scaffold also exhibited an interconnected porous architecture with porosity greater than 80%, an appropriate degradation rate, and enhanced cell proliferation. Moreover, the HPCM scaffold supported the healing of a rat calvarial defect in vivo.