In vitro evaluation of electrospun silk fibroin/nano-hydroxyapatite/BMP-2 scaffolds for bone regeneration

Bone tissue engineering by using osteoinductive scaffolds seeded with stem cells to promote bone extracellular matrix (ECM) production and remodeling has evolved into a promising approach for bone repair and regeneration. In order to mimic the ECM of bone tissue structurally and compositionally, nanofibrous silk fibroin (SF) scaffolds containing hydroxyapatite (HAP) nanoparticles and bone morphogenetic protein 2 (BMP-2) were fabricated in this study using electrospinning technique. The microstructure, mechanical property, biocompatibility, and osteogenic characteristics were examined. It was found that the HAP nanoparticles were successfully incorporated in the SF nanofibers (diameter, 200–500 nm). The mechanical properties of SF/HAP/BMP-2 composite scaffolds increased with HAP content when it was less than 20 wt%, after which the mechanical properties dropped as HAP content increased. Cell culture tests using bone marrow mesenchymal stem cells (BMSCs) showed that the scaffolds had good biocompatibility and promoted the osteogenic differentiation of BMSCs. Therefore, the electrospun SF/HAP/BMP-2 scaffolds may serve as a promising biomaterial for bone tissue engineering.     

Collagenous matrix supported by a 3D-printed scaffold for osteogenic differentiation of dental pulp cells

Objective

A systematic characterization of hybrid scaffolds, fabricated based on combinatorial additive manufacturing technique and freeze-drying method, is presented as a new platform for osteoblastic differentiation of dental pulp cells (DPCs).

Present and future of tissue engineering scaffolds for dentin‐pulp complex regeneration

More than two thirds of the global population suffers from tooth decay, which results in cavities with various levels of lesion severity. Clinical interventions to treat tooth decay range from simple coronal fillings to invasive root canal treatment. Pulp capping is the only available clinical option to maintain the pulp vitality in deep lesions, but irreversible pulp inflammation and reinfection are frequent outcomes for this treatment. When affected pulp involvement is beyond repair, the dentist has to perform endodontic therapy leaving the tooth non‐vital and brittle. On‐going research strategies have failed to overcome the limitations of existing pulp capping materials so that healthy and progressive regeneration of the injured tissues is attained. Preserving pulp vitality is crucial for tooth homeostasis and durability, and thus, there is a critical need for clinical interventions that enable regeneration of the dentin‐pulp complex to rescue millions of teeth annually. The identification and development of appropriate biomaterials for dentin‐pulp scaffolds are necessary to optimize clinical approaches to regenerate these hybrid dental tissues. Likewise, a deep understanding of the interactions between the micro‐environment, growth factors, and progenitor cells will provide design basis for the most fitting scaffolds for this purpose. In this review, we first introduce the long‐lasting clinical dental problem of rescuing diseased tooth vitality, the limitations of current clinical therapies and interventions to restore the damaged tissues, and the need for new strategies to fully revitalize the tooth. Then, we comprehensively report on the characteristics of the main materials of naturally‐derived and synthetically‐engineered polymers, ceramics, and composite scaffolds as well as their use in dentin‐pulp complex regeneration strategies. Finally, we present a series of innovative smart polymeric biomaterials with potential to overcome dentin‐pulp complex regeneration challenges.     

Silk fibroin scaffolds loaded with angiogenic genes in adenovirus vectors for tissue regeneration

Vascularization remains a critical challenge in dermal tissue regeneration. In this study, a vascular endothelial growth factor (VEGF165) and angiopoietin‐1 (Ang‐1) dual gene coexpression vector that encoded green fluorescent protein (GFP) was constructed from an arginine–glycine–aspartic acid‐modified adenovirus. Silk fibroin (SF) scaffolds loaded with adenovirus vectors were fabricated by freeze‐drying method. In vitro, the human endothelial‐derived cell line EA.hy926 was infected with adenovirus vectors and then expressed GFP, secreted VEGF165 and Ang‐1, and promoted cell proliferation effectively. The VEGF165 and Ang‐1 genes loaded in the SF scaffolds significantly promoted the formation of abundant microvascular networks in the chick embryo chorioallantoic membrane. In vivo, angiogenic genes loaded in the scaffolds promoted vascularization and collagen deposition in scaffolds, thus effectively accelerating dermal tissue regeneration in a dorsal full‐thickness skin defect wound model in Sprague–Dawley rats. In conclusion, SF scaffolds loaded with arginine–glycine–aspartic acid‐modified adenovirus vectors encoding VEGF165 and Ang‐1 could stimulate the formation of vascular networks through the effective expression of target genes in vascular endothelial cells, thereby accelerating the regeneration of dermal tissue.     

Cytomodulin‐functionalized porous PLGA particulate scaffolds respond better to cell migration,actin production and wound healing in rodent model

In the present study, porous PLGA microparticulate scaffolds (PMS_P), surface‐hydrolysed scaffolds (PMS_Hyd) and cytomodulin‐coupled scaffolds (PMS_CM) were prepared and characterized. After coupling the particles with cytomodulin, the size was reduced from 334 µm (span 0.53) to 278 µm due to hydrolysis, and contact angle also decreased from 70.87 ± 8.56 to 31.43 ± 7.43, indicating an increase in hydrophilicity. Surface roughness and pore density increased, along with an increase in surface area from 9.59 ± 0.36 to 16.82 ± 0.064 m²/g after attaching the biomolecule CM onto the PLGA particles. In vitro cell culture experiments on human dermal fibroblasts (HDFs) were performed for 21 days, in which MTT assay indicated two‐fold higher cell proliferation on PMS_Hyd than on PMS_CM; however, cell distribution, cell spreading and actin production were significantly higher on PMS_CM than on other scaffolds. Migration of cells from PMS_CM to a 2D plate was gradual but the migrated cells attained early confluence, indicating the preservation of normal cellular functions. In a full‐thickness wound mouse model, PMS_CM exhibited 80% wound closure within 2 weeks. Further, at the end of week 3, the inflammatory cell count in the PMS_CM group was reduced to one‐third of the control group, while in PMS_P and PMS_Hyd the extent of inflammation was much higher and more severe. In the case of PMS_CM, abundant fibroblast proliferation, early formation of the scar tissue, eschar formation and inward movement of the wound margins (a zipper‐like movement) towards the deeper layers of the skin suggested advanced wound healing. Cytomodulin‐coupled scaffolds ensured better cell spreading and migration and thus enabled rapid wound healing (see Supporting information, Figure S1). Copyright © 2012 John Wiley & Sons, Ltd.     

Bone Conduction Capacity of Highly Porous 3D-Printed Titanium Scaffolds Based on Different Pore Designs

In porous titanium scaffolds manufactured via 3D printing, the differences in bone formation according to pore design and implantation period were studied. Titanium scaffolds with three types of different pore structures (Octadense, Gyroid, and Dode) were fabricated via 3D printing using the selective laser melting method. Mechanical properties of scaffolds were investigated. Prepared specimens were inserted into both femurs of nine rabbits and their clinical characteristics were observed. Three animals were sacrificed at the 2nd, 4th, and 6th weeks, and the differences in bone formation were radiologically and histologically analyzed. The percentage of new bone and surface density in the pore structure were observed to be approximately 25% and 8 mm²/mm³, respectively. There was no difference in the amount of newly formed bone according to the pore design at 2, 4, and 6 weeks. In addition, no differences in the amount of newly formed bone were observed with increasing time within the same pore design for all three designs. During the 6-week observation period, the proportion of new bones in the 3D-printed titanium scaffold was approximately 25%. Differences in bone formation according to the pore design or implantation period were not observed.     

低热高压法制作PLGA支架的三维结构研究

目的制作不含有机溶剂、三维结构良好的聚丙交酯-乙交酯共聚物（PLGA）支架,使之符合组织工程骨修复的需要,探讨一种新型聚合物支架制作方法。方法将聚合物与氯化钠粉碎后,采用低热高压法制作PLGA泡沫结构支架,经密度法、氯化钠法测定其空隙率、开孔率;扫描电镜观察表面和内部结构、测定孔径。结果利用此种方法制作的PLGA支架,空隙率达到90.0％和92.5％、孔径在200-250μm之间、开孔率为98.0％以上（P<0.01）,平均氯化钠沥净时间为12～13h。结论使用低热高压法制作的组织工程支架,三维结构稳定,各项参数可控制;根据模具的大小可以制作不同体积的支架;依据盐的颗粒粒度与数量控制支架的孔径和空隙率,在制作过程中不使用有机溶剂,减少了有机溶剂残留可能引起的对细胞的毒性。使用这种方法要对聚合物与氯化钠颗粒进行充分混合。     

Evaluation of methods for the construction of collagenous scaffolds with a radial pore structure for tissue engineering

Type I collagen is used widely as a biomaterial. The structure of collagenous biomaterials, including pore sizes and general architecture, can be varied by a number of techniques. In this study, we developed a method to construct flat fibrillar type I collagen scaffolds, 6 cm in diameter and with a radially orientated pore structure, by the use of directional freezing. Different methodologies were tested, the optimal one being freezing of a collagen suspension inside-out, using a centrally positioned liquid nitrogen-cooled tube. Pore sizes could be varied by the use of different tube materials. Use of aluminium tubes resulted in radial scaffolds with a pore size of 20-30 μm, whereas use of stainless steel produced radial scaffolds with 70-100 μm pore sizes. Brass- and copper-based tubes produced scaffolds with less homogeneous radial pores, pore sizes being 90-100 and 50-80 μm, respectively. Fibreglass tubes gave even less uniformity (pore size 100-150 μm). Scaffolds were free of cracks, except in case of aluminium. Scaffolds with a radial inner structure may be especially suitable for tissue engineering of organs with a radial scaffold structure, such as the diaphragm.