Projection-Based 3D Printing of Cell Patterning Scaffolds with Multiscale Channels.

PubMed

Xue, Dai; Wang, Yancheng; Zhang, Jiaxin; Mei, Deqing; Wang, Yue; Chen, Shaochen

2018-06-13

To fully actualize artificial, cell-laden biological models in tissue engineering, such as 3D organoids and organs-on-a-chip systems, cells need to be patterned such that they can precisely mimic natural microenvironments in vitro. Despite increasing interest in this area, patterning cells at multiscale (∼10 μm to 10 mm) remains a significant challenge in bioengineering. Here, we report a projection-based 3D printing system that achieves rapid and high-resolution fabrication of hydrogel scaffolds featuring intricate channels for multiscale cell patterning. Using this system, we were able to use biocompatible poly(ethylene glycol)diacrylate in fabricating a variety of scaffold architectures, ranging from regular geometries such as serpentine, spiral, and fractal-like to more irregular/intricate geometries, such as biomimetic arborescent and capillary networks. A red food dye solution was able to freely fill all channels in the scaffolds, from the trunk (>1100 μm in width) to the small branch (∼17 μm in width) without an external pump. The dimensions of the printed scaffolds remained stable over 3 days while being immersed in Dulbecco's phosphate-buffered saline at 37 °C, and a penetration analysis revealed that these scaffolds are suitable for metabolic and nutrient transport. Cell patterning experiments showed that red fluorescent protein-transfected A549 human nonsmall lung cancer cells adhered well in the scaffolds' channels, and showed further attachment and penetration during cell culture proliferation.

3D-printed bioceramic scaffolds with antibacterial and osteogenic activity.

PubMed

Zhang, Yongliang; Zhai, Dong; Xu, Mengchi; Yao, Qingqiang; Zhu, Huiying; Chang, Jiang; Wu, Chengtie

2017-06-20

Bacterial infection poses a significant risk with the wide application of bone graft materials. Designing bone grafts with good antibacterial performance and excellent bone-forming activity is of particular significance for bone tissue engineering. In our study, a 3D printing method was used to prepare β-tricalcium phosphate (β-TCP) bioceramic scaffolds. Silver (Ag) nanoparticles were uniformly dispersed on graphene oxide (GO) to form a homogeneous nanocomposite (named Ag@GO) with different Ag-to-graphene oxide mass ratios, with this being synthesized via the liquid chemical reduction approach. Ag@GO nanocomposites were successfully modified on the β-TCP scaffolds by a simple soaking method to achieve bifunctional biomaterials with antibacterial and osteogenic activity. The prepared scaffolds possessed a connected network with triangle pore morphology and the surfaces of the β-TCP scaffolds were uniformly modified by the Ag@GO nanocomposite layers. The Ag content in the scaffolds was controlled by changing the coating times and concentration of the Ag@GO nanocomposites. The antibacterial activity of the scaffolds was assessed with Gram-negative bacteria (Escherichia coli, E. coli). The results demonstrated that the scaffolds with Ag@GO nanocomposites presented excellent antibacterial activity. In addition, the scaffolds coated with Ag@GO nanocomposites conspicuously accelerated the osteogenic differentiation of rabbit bone marrow stromal cells by improving their alkaline phosphatase activity and bone-related gene expression (osteopontin, runt-related transcription factor 2, osteocalcin and bone sialoprotein). This study demonstrates that bifunctional scaffolds with a combination of antibacterial and osteogenic activity can be achieved for the reconstruction of large-bone defects while preventing or treating infections.

Incorporation of zinc oxide nanoparticles into chitosan-collagen 3D porous scaffolds: Effect on morphology, mechanical properties and cytocompatibility of 3D porous scaffolds.

PubMed

Ullah, Saleem; Zainol, Ismail; Idrus, Ruszymah Hj

2017-11-01

The zinc oxide nanoparticles (particles size <50nm) incorporated into chitosan-collagen 3D porous scaffolds and investigated the effect of zinc oxide nanoparticles incorporation on microstructure, mechanical properties, biodegradation and cytocompatibility of 3D porous scaffolds. The 0.5%, 1.0%, 2.0% and 4.0% zinc oxide nanoparticles chitosan-collagen 3D porous scaffolds were fabricated via freeze-drying technique. The zinc oxide nanoparticles incorporation effects consisting in chitosan-collagen 3D porous scaffolds were investigated by mechanical and swelling tests, and effect on the morphology of scaffolds examined microscopically. The biodegradation and cytocompatibility tests were used to investigate the effects of zinc oxide nanoparticles incorporation on the ability of scaffolds to use for tissue engineering application. The mean pore size and swelling ratio of scaffolds were decreased upon incorporation of zinc oxide nanoparticles however, the porosity, tensile modulus and biodegradation rate were increased upon incorporation of zinc oxide nanoparticles. In vitro culture of human fibroblasts and keratinocytes showed that the zinc oxide nanoparticles facilitated cell adhesion, proliferation and infiltration of chitosan-collagen 3D porous scaffolds. It was found that the zinc oxide nanoparticles incorporation enhanced porosity, tensile modulus and cytocompatibility of chitosan-collagen 3D porous scaffolds. Copyright © 2017 Elsevier B.V. All rights reserved.

Apple Derived Cellulose Scaffolds for 3D Mammalian Cell Culture

PubMed Central

Modulevsky, Daniel J.; Lefebvre, Cory; Haase, Kristina; Al-Rekabi, Zeinab; Pelling, Andrew E.

2014-01-01

There are numerous approaches for producing natural and synthetic 3D scaffolds that support the proliferation of mammalian cells. 3D scaffolds better represent the natural cellular microenvironment and have many potential applications in vitro and in vivo. Here, we demonstrate that 3D cellulose scaffolds produced by decellularizing apple hypanthium tissue can be employed for in vitro 3D culture of NIH3T3 fibroblasts, mouse C2C12 muscle myoblasts and human HeLa epithelial cells. We show that these cells can adhere, invade and proliferate in the cellulose scaffolds. In addition, biochemical functionalization or chemical cross-linking can be employed to control the surface biochemistry and/or mechanical properties of the scaffold. The cells retain high viability even after 12 continuous weeks of culture and can achieve cell densities comparable with other natural and synthetic scaffold materials. Apple derived cellulose scaffolds are easily produced, inexpensive and originate from a renewable source. Taken together, these results demonstrate that naturally derived cellulose scaffolds offer a complementary approach to existing techniques for the in vitro culture of mammalian cells in a 3D environment. PMID:24842603

Apple derived cellulose scaffolds for 3D mammalian cell culture.

PubMed

Modulevsky, Daniel J; Lefebvre, Cory; Haase, Kristina; Al-Rekabi, Zeinab; Pelling, Andrew E

2014-01-01

There are numerous approaches for producing natural and synthetic 3D scaffolds that support the proliferation of mammalian cells. 3D scaffolds better represent the natural cellular microenvironment and have many potential applications in vitro and in vivo. Here, we demonstrate that 3D cellulose scaffolds produced by decellularizing apple hypanthium tissue can be employed for in vitro 3D culture of NIH3T3 fibroblasts, mouse C2C12 muscle myoblasts and human HeLa epithelial cells. We show that these cells can adhere, invade and proliferate in the cellulose scaffolds. In addition, biochemical functionalization or chemical cross-linking can be employed to control the surface biochemistry and/or mechanical properties of the scaffold. The cells retain high viability even after 12 continuous weeks of culture and can achieve cell densities comparable with other natural and synthetic scaffold materials. Apple derived cellulose scaffolds are easily produced, inexpensive and originate from a renewable source. Taken together, these results demonstrate that naturally derived cellulose scaffolds offer a complementary approach to existing techniques for the in vitro culture of mammalian cells in a 3D environment.

Deposition of phosphate coatings on titanium within scaffold structure.

PubMed

Trybuś, Bartłomiej; Zieliński, Andrzej; Beutner, Rene; Seramak, Tomasz; Scharnweber, Dieter

2017-01-01

Existing knowledge about the appearance, thickness, and chemical composition of phosphate coatings on titanium inside porous structures is insufficient. Such knowledge is important for the design and fabrication of porous implants. Metallic scaffolds were fabricated by selective laser melting of 316L stainless steel powder. Phosphate coatings were deposited on Ti sensors placed either outside the scaffolds or in the holes in the scaffolds. The electrochemically-assisted cathodic deposition of phosphate coatings was performed under galvanostatic conditions in an electrolyte containing the calcium and phosphate ions. The phosphate deposits were microscopically investigated; this included the performance of mass weight measurements and chemical analyses of the content of Ca2+ and  24 PO ions after the dissolution of deposits. The thicknesses of the calcium phosphate coatings were about 140 and 200 nm for isolated titanium sensors and 170 and 300 nm for titanium sensors placed inside pores. Deposition of calcium phosphate occurred inside the pores up to 150 mm below the scaffold surface. The deposits were rich in Ca, with a Ca/P ratio ranging from 2 to 2.5. Calcium phosphate coatings can be successfully deposited on a Ti surface inside a model scaffold. An increase in cathodic current results in an increase in coating thickness. Any decrease in the cathodic current inside the porous structure is slight. The calcium phosphate inside the pores has a much higher Ca/P ratio than that of stoichiometric HAp, likely due to a gradual increase in Ca fraction with distance from the surface.

Fabrication of β-tricalcium phosphate composite ceramic sphere-based scaffolds with hierarchical pore structure for bone regeneration.

PubMed

He, Fupo; Qian, Guowen; Ren, Weiwei; Li, Jiyan; Fan, Peirong; Shi, Haishan; Shi, Xuetao; Deng, Xin; Wu, Shanghua; Ye, Jiandong

2017-04-24

Polymer sphere-based scaffolds, which are prepared by bonding the adjacent spheres via sintering the randomly packed spheres, feature uniform pore structure, full three-dimensional (3D) interconnection, and considerable mechanical strength. However, bioceramic sphere-based scaffolds fabricated by this method have never been reported. Due to high melting temperature of bioceramic, only limited diffusion rate can be achieved when sintering the bioceramic spheres, which is far from enough to form robust bonding between spheres. In the present study, for the first time we fabricated 3D interconnected β-tricalcium phosphate ceramic sphere-based (PG/TCP) scaffolds by introducing phosphate-based glass (PG) as sintering additive and placing uniaxial pressure during the sintering process. The sintering mechanism of PG/TCP scaffolds was unveiled. The PG/TCP scaffolds had hierarchical pore structure, which was composed by interconnected macropores (>200 μm) among spheres, pores (20–120 μm) in the interior of spheres, and micropores (1–3 μm) among the grains. During the sintering process, partial PG reacted with β-TCP, forming β-Ca2P2O7; metal ions from PG substituted to Ca2+ sites of β-TCP. The mechanical properties (compressive strength 2.8–10.6 MPa; compressive modulus 190–620 MPa) and porosity (30%–50%) of scaffolds could be tailored by manipulating the sintering temperatures. The introduction of PG accelerated in vitro degradation of scaffolds, and the PG/TCP scaffolds showed good cytocompatibility. This work may offer a new strategy to prepare bioceramic scaffolds with satisfactory physicochemical properties for application in bone regeneration.

Low temperature fabrication of magnesium phosphate cement scaffolds by 3D powder printing.

PubMed

Klammert, Uwe; Vorndran, Elke; Reuther, Tobias; Müller, Frank A; Zorn, Katharina; Gbureck, Uwe

2010-11-01

Synthetic bone replacement materials are of great interest because they offer certain advantages compared with organic bone grafts. Biodegradability and preoperative manufacturing of patient specific implants are further desirable features in various clinical situations. Both can be realised by 3D powder printing. In this study, we introduce powder-printed magnesium ammonium phosphate (struvite) structures, accompanied by a neutral setting reaction by printing farringtonite (Mg(3)(PO(4))(2)) powder with ammonium phosphate solution as binder. Suitable powders were obtained after sintering at 1100°C for 5 h following 20-40 min dry grinding in a ball mill. Depending on the post-treatment of the samples, compressive strengths were found to be in the range 2-7 MPa. Cytocompatibility was demonstrated in vitro using the human osteoblastic cell line MG63.

3D braid scaffolds for regeneration of articular cartilage.

PubMed

Ahn, Hyunchul; Kim, Kyoung Ju; Park, Sook Young; Huh, Jeong Eun; Kim, Hyun Jeong; Yu, Woong-Ryeol

2014-06-01

Regenerating articular cartilage in vivo from cultured chondrocytes requires that the cells be cultured and implanted within a biocompatible, biodegradable scaffold. Such scaffolds must be mechanically stable; otherwise chondrocytes would not be supported and patients would experience severe pain. Here we report a new 3D braid scaffold that matches the anisotropic (gradient) mechanical properties of natural articular cartilage and is permissive to cell cultivation. To design an optimal structure, the scaffold unit cell was mathematically modeled and imported into finite element analysis. Based on this analysis, a 3D braid structure with gradient axial yarn distribution was designed and manufactured using a custom-built braiding machine. The mechanical properties of the 3D braid scaffold were evaluated and compared with simulated results, demonstrating that a multi-scale approach consisting of unit cell modeling and continuum analysis facilitates design of scaffolds that meet the requirements for mechanical compatibility with tissues. Copyright © 2014 Elsevier Ltd. All rights reserved.

Evaluation of PBS Treatment and PEI Coating Effects on Surface Morphology and Cellular Response of 3D-Printed Alginate Scaffolds.

PubMed

Mendoza García, María A; Izadifar, Mohammad; Chen, Xiongbiao

2017-11-01

Three-dimensional (3D) printing is an emerging technology for the fabrication of scaffolds to repair/replace damaged tissue/organs in tissue engineering. This paper presents our study on 3D printed alginate scaffolds treated with phosphate buffered saline (PBS) and polyethyleneimine (PEI) coating and their impacts on the surface morphology and cellular response of the printed scaffolds. In our study, sterile alginate was prepared by means of the freeze-drying method and then, used to prepare the hydrogel for 3D printing into calcium chloride, forming 3D scaffolds. Scaffolds were treated with PBS for a time period of two days and seven days, respectively, and PEI coating; then they were seeded with Schwann cells (RSC96) for the examination of cellular response (proliferation and differentiation). In addition, swelling and stiffness (Young's modulus) of the treated scaffolds was evaluated, while their surface morphology was assessed using scanning electron microscopy (SEM). SEM images revealed significant changes in scaffold surface morphology due to degradation caused by the PBS treatment over time. Our cell proliferation assessment over seven days showed that a two-day PBS treatment could be more effective than seven-day PBS treatment for improving cell attachment and elongation. While PEI coating of alginate scaffolds seemed to contribute to cell growth, Schwann cells stayed round on the surface of alginate over the period of cell culture. In conclusion, PBS-treatment may offer the potential to induce surface physical cues due to degradation of alginate, which could improve cell attachment post cell-seeding of 3D-printed alginate scaffolds.

Mesoporous bioactive glass nanolayer-functionalized 3D-printed scaffolds for accelerating osteogenesis and angiogenesis

NASA Astrophysics Data System (ADS)

Zhang, Yali; Xia, Lunguo; Zhai, Dong; Shi, Mengchao; Luo, Yongxiang; Feng, Chun; Fang, Bing; Yin, Jingbo; Chang, Jiang; Wu, Chengtie

2015-11-01

The hierarchical microstructure, surface and interface of biomaterials are important factors influencing their bioactivity. Porous bioceramic scaffolds have been widely used for bone tissue engineering by optimizing their chemical composition and large-pore structure. However, the surface and interface of struts in bioceramic scaffolds are often ignored. The aim of this study is to incorporate hierarchical pores and bioactive components into the bioceramic scaffolds by constructing nanopores and bioactive elements on the struts of scaffolds and further improve their bone-forming activity. Mesoporous bioactive glass (MBG) modified β-tricalcium phosphate (MBG-β-TCP) scaffolds with a hierarchical pore structure and a functional strut surface (~100 nm of MBG nanolayer) were successfully prepared via 3D printing and spin coating. The compressive strength and apatite-mineralization ability of MBG-β-TCP scaffolds were significantly enhanced as compared to β-TCP scaffolds without the MBG nanolayer. The attachment, viability, alkaline phosphatase (ALP) activity, osteogenic gene expression (Runx2, BMP2, OPN and Col I) and protein expression (OPN, Col I, VEGF, HIF-1α) of rabbit bone marrow stromal cells (rBMSCs) as well as the attachment, viability and angiogenic gene expression (VEGF and HIF-1α) of human umbilical vein endothelial cells (HUVECs) in MBG-β-TCP scaffolds were significantly upregulated compared with conventional bioactive glass (BG)-modified β-TCP (BG-β-TCP) and pure β-TCP scaffolds. Furthermore, MBG-β-TCP scaffolds significantly enhanced the formation of new bone in vivo as compared to BG-β-TCP and β-TCP scaffolds. The results suggest that application of the MBG nanolayer to modify 3D-printed bioceramic scaffolds offers a new strategy to construct hierarchically porous scaffolds with significantly improved physicochemical and biological properties, such as mechanical properties, osteogenesis, angiogenesis and protein expression for bone tissue

Treatment of osteomyelitis defects by a vancomycin-loaded gelatin/β-tricalcium phosphate composite scaffold

PubMed Central

Zhou, J.; Zhou, X. G.; Wang, J. W.; Zhou, H.; Dong, J.

2018-01-01

Objective In the present study, we aimed to assess whether gelatin/β-tricalcium phosphate (β-TCP) composite porous scaffolds could be used as a local controlled release system for vancomycin. We also investigated the efficiency of the scaffolds in eliminating infections and repairing osteomyelitis defects in rabbits. Methods The gelatin scaffolds containing differing amounts of of β-TCP (0%, 10%, 30% and 50%) were prepared for controlled release of vancomycin and were labelled G-TCP0, G-TCP1, G-TCP3 and G-TCP5, respectively. The Kirby-Bauer method was used to examine the release profile. Chronic osteomyelitis models of rabbits were established. After thorough debridement, the osteomyelitis defects were implanted with the scaffolds. Radiographs and histological examinations were carried out to investigate the efficiency of eliminating infections and repairing bone defects. Results The prepared gelatin/β-TCP scaffolds exhibited a homogeneously interconnected 3D porous structure. The G-TCP0 scaffold exhibited the longest duration of vancomycin release with a release duration of eight weeks. With the increase of β-TCP contents, the release duration of the β-TCP-containing composite scaffolds was decreased. The complete release of vancomycin from the G-TCP5 scaffold was achieved within three weeks. In the treatment of osteomyelitis defects in rabbits, the G-TCP3 scaffold showed the most efficacious performance in eliminating infections and repairing bone defects. Conclusions The composite scaffolds could achieve local therapeutic drug levels over an extended duration. The G-TCP3 scaffold possessed the optimal porosity, interconnection and controlled release performance. Therefore, this scaffold could potentially be used in the treatment of chronic osteomyelitis defects. Cite this article: J. Zhou, X. G. Zhou, J. W. Wang, H. Zhou, J. Dong. Treatment of osteomyelitis defects by a vancomycin-loaded gelatin/β-tricalcium phosphate composite scaffold. Bone Joint Res

Strength reliability and in vitro degradation of three-dimensional powder printed strontium-substituted magnesium phosphate scaffolds.

PubMed

Meininger, Susanne; Mandal, Sourav; Kumar, Alok; Groll, Jürgen; Basu, Bikramjit; Gbureck, Uwe

2016-02-01

Strontium ions (Sr(2+)) are known to prevent osteoporosis and also encourage bone formation. Such twin requirements have motivated researchers to develop Sr-substituted biomaterials for orthopaedic applications. The present study demonstrates a new concept of developing Sr-substituted Mg3(PO4)2 - based biodegradable scaffolds. In particular, this work reports the fabrication, mechanical properties with an emphasis on strength reliability as well as in vitro degradation of highly biodegradable strontium-incorporated magnesium phosphate cements. These implantable scaffolds were fabricated using three-dimensional powder printing, followed by high temperature sintering and/or chemical conversion, a technique adaptable to develop patient-specific implants. A moderate combination of strength properties of 36.7MPa (compression), 24.2MPa (bending) and 10.7MPa (tension) were measured. A reasonably modest Weibull modulus of up to 8.8 was recorded after uniaxial compression or diametral tensile tests on 3D printed scaffolds. A comparison among scaffolds with varying compositions or among sintered or chemically hardened scaffolds reveals that the strength reliability is not compromised in Sr-substituted scaffolds compared to baseline Mg3(PO4)2. The micro-computed tomography analysis reveals the presence of highly interconnected porous architecture in three-dimension with lognormal pore size distribution having median in the range of 17.74-26.29μm for the investigated scaffolds. The results of extensive in vitro ion release study revealed passive degradation with a reduced Mg(2+) release and slow but sustained release of Sr(2+) from strontium-substituted magnesium phosphate scaffolds. Taken together, the present study unequivocally illustrates that the newly designed Sr-substituted magnesium phosphate scaffolds with good strength reliability could be used for biomedical applications requiring consistent Sr(2+)- release, while the scaffold degrades in physiological medium. The

3D Microperiodic Hydrogel Scaffolds for Robust Neuronal Cultures

PubMed Central

Hanson Shepherd, Jennifer N.; Parker, Sara T.; Shepherd, Robert F.; Gillette, Martha U.; Lewis, Jennifer A.; Nuzzo, Ralph G.

2011-01-01

Three-dimensional (3D) microperiodic scaffolds of poly(2-hydroxyethyl methacrylate) (pHEMA) have been fabricated by direct-write assembly of a photopolymerizable hydrogel ink. The ink is initially composed of physically entangled pHEMA chains dissolved in a solution of HEMA monomer, comonomer, photoinitiator and water. Upon printing 3D scaffolds of varying architecture, the ink filaments are exposed to UV light, where they are transformed into an interpenetrating hydrogel network of chemically cross-linked and physically entangled pHEMA chains. These 3D microperiodic scaffolds are rendered growth compliant for primary rat hippocampal neurons by absorption of polylysine. Neuronal cells thrive on these scaffolds, forming differentiated, intricately branched networks. Confocal laser scanning microscopy reveals that both cell distribution and extent of neuronal process alignment depend upon scaffold architecture. This work provides an important step forward in the creation of suitable platforms for in vitro study of sensitive cell types. PMID:21709750

In vitro study of 3D PLGA/n-HAp/β-TCP composite scaffolds with etched oxygen plasma surface modification in bone tissue engineering

NASA Astrophysics Data System (ADS)

Roh, Hee-Sang; Jung, Sang-Chul; Kook, Min-Suk; Kim, Byung-Hoon

2016-12-01

Three-dimensional (3D) scaffolds have many advantageous properties for bone tissue engineering application, due to its controllable properties such as pore size, structural shape and interconnectivity. In this study, effects on oxygen plasma surface modification and adding of nano-hydroxyapatite (n-HAp) and β-tricalcium phosphate (β-TCP) on the 3D PLGA/n-HAp/β-TCP scaffolds for improving preosteoblast cell (MC3T3-E1) adhesion, proliferation and differentiation were investigated. The 3D PLGA/n-HAp/β-TCP scaffolds were fabricated by 3D Bio-Extruder equipment. The 3D scaffolds were prepared with 0°/90° architecture and pore size of approximately 300 μm. In addition 3D scaffolds surface were etched by oxygen plasma to enhance the hydrophilic property and surface roughness. After oxygen plasma treatment, the surface chemistry and morphology were investigated by Fourier transform infrared spectroscopy, scanning electron microscopy, and atomic force microscopy. And also hydrophilic property was measured by contact angle. The MC3T3-E1 cell proliferation and differentiation were investigated by MTT assay and ALP activity. In present work, the 3D PLGA/HAp/beta-TCP composite scaffold with suitable structure for the growth of osteoblast cells was successfully fabricated by 3D rapid prototyping technique. The surface hydrophilicity and roughness of 3D scaffold increased by oxygen plasma treatment had a positive effect on cell adhesion, proliferation, and differentiation. Furthermore, the differentiation of MC3T3-E1 cell was significantly enhanced by adding of n-HAp and β-TCP on 3D PLGA scaffold. As a result, combination of bioceramics and oxygen plasma treatment showed a synergistic effect on biocompatibility of 3D scaffolds. This result confirms that this technique was useful tool for improving the biocompatibility in bone tissue engineering application.

Production of new 3D scaffolds for bone tissue regeneration by rapid prototyping.

PubMed

Fradique, R; Correia, T R; Miguel, S P; de Sá, K D; Figueira, D R; Mendonça, A G; Correia, I J

2016-04-01

The incidence of bone disorders, whether due to trauma or pathology, has been trending upward with the aging of the worldwide population. The currently available treatments for bone injuries are rather limited, involving mainly bone grafts and implants. A particularly promising approach for bone regeneration uses rapid prototyping (RP) technologies to produce 3D scaffolds with highly controlled structure and orientation, based on computer-aided design models or medical data. Herein, tricalcium phosphate (TCP)/alginate scaffolds were produced using RP and subsequently their physicochemical, mechanical and biological properties were characterized. The results showed that 60/40 of TCP and alginate formulation was able to match the compression and present a similar Young modulus to that of trabecular bone while presenting an adequate biocompatibility. Moreover, the biomineralization ability, roughness and macro and microporosity of scaffolds allowed cell anchoring and proliferation at their surface, as well as cell migration to its interior, processes that are fundamental for osteointegration and bone regeneration.

Direct deposited porous scaffolds of calcium phosphate cement with alginate for drug delivery and bone tissue engineering.

PubMed

Lee, Gil-Su; Park, Jeong-Hui; Shin, Ueon Sang; Kim, Hae-Won

2011-08-01

This study reports the preparation of novel porous scaffolds of calcium phosphate cement (CPC) combined with alginate, and their potential usefulness as a three-dimensional (3-D) matrix for drug delivery and tissue engineering of bone. An α-tricalcium phosphate-based powder was mixed with sodium alginate solution and then directly injected into a fibrous structure in a Ca-containing bath. A rapid hardening reaction of the alginate with Ca(2+) helps to shape the composite into a fibrous form with diameters of hundreds of micrometers, and subsequent pressing in a mold allows the formation of 3-D porous scaffolds with different porosity levels. After transformation of the CPC into a calcium-deficient hydroxyapatite phase in simulated biological fluid the scaffold was shown to retain its mechanical stability. During the process biological proteins, such as bovine serum albumin and lysozyme, used as model proteins, were observed to be effectively loaded onto and released from the scaffolds for up to more than a month, proving the efficacy of the scaffolds as a drug delivering matrix. Mesenchymal stem cells (MSCs) were isolated from rat bone marrow and then cultured on the CPC-alginate porous scaffolds to investigate the ability to support proliferation of cells and their subsequent differentiation along the osteogenic lineage. It was shown that MSCs increasingly actively populated and also permeated into the porous network with time of culture. In particular, cells cultured within a scaffold with a relatively high porosity level showed favorable proliferation and osteogenic differentiation. An in vivo pilot study of the CPC-alginate porous scaffolds after implantation into the rat calvarium for 6 weeks revealed the formation of new bone tissue within the scaffold, closing the defect almost completely. Based on these results, the newly developed CPC-alginate porous scaffolds could be potentially useful as a 3-D matrix for drug delivery and tissue engineering of bone

Effect of Chemistry on Osteogenesis and Angiogenesis Towards Bone Tissue Engineering Using 3D Printed Scaffolds.

PubMed

Bose, Susmita; Tarafder, Solaiman; Bandyopadhyay, Amit

2017-01-01

The functionality or survival of tissue engineering constructs depends on the adequate vascularization through oxygen transport and metabolic waste removal at the core. This study reports the presence of magnesium and silicon in direct three dimensional printed (3DP) tricalcium phosphate (TCP) scaffolds promotes in vivo osteogenesis and angiogenesis when tested in rat distal femoral defect model. Scaffolds with three different interconnected macro pore sizes were fabricated using direct three dimensional printing. In vitro ion release in phosphate buffer for 30 days showed sustained Mg 2+  and Si 4+  release from these scaffolds. Histolomorphology and histomorphometric analysis from the histology tissue sections revealed a significantly higher bone formation, between 14 and 20% for 4-16 weeks, and blood vessel formation, between 3 and 6% for 4-12 weeks, due to the presence of magnesium and silicon in TCP scaffolds compared to bare TCP scaffolds. The presence of magnesium in these 3DP TCP scaffolds also caused delayed TRAP activity. These results show that magnesium and silicon incorporated 3DP TCP scaffolds with multiscale porosity have huge potential for bone tissue repair and regeneration.

The design of 3D scaffold for tissue engineering using automated scaffold design algorithm.

PubMed

Mahmoud, Shahenda; Eldeib, Ayman; Samy, Sherif

2015-06-01

Several progresses have been introduced in the field of bone regenerative medicine. A new term tissue engineering (TE) was created. In TE, a highly porous artificial extracellular matrix or scaffold is required to accommodate cells and guide their growth in three dimensions. The design of scaffolds with desirable internal and external structure represents a challenge for TE. In this paper, we introduce a new method known as automated scaffold design (ASD) for designing a 3D scaffold with a minimum mismatches for its geometrical parameters. The method makes use of k-means clustering algorithm to separate the different tissues and hence decodes the defected bone portions. The segmented portions of different slices are registered to construct the 3D volume for the data. It also uses an isosurface rendering technique for 3D visualization of the scaffold and bones. It provides the ability to visualize the transplanted as well as the normal bone portions. The proposed system proves good performance in both the segmentation results and visualizations aspects.

Photoinitiator-free 3D scaffolds fabricated by excimer laser photocuring.

PubMed

Farkas, Balázs; Dante, Silvia; Brandi, Fernando

2017-01-20

Photoinitiator-free fabrication of poly(ethylene glycol) diacrylate (PEGDA) scaffolds is achieved using a novel three-dimensional (3D) printing method called mask projected excimer laser stereolithography (MPExSL). The spatial resolution of photoinitiator-free curing is suitable for 3D layer-by-layer fabrication with a single layer thickness well controllable at tens to hundreds of microns using 248 nm wavelength for the irradiation. The photoinitiator-free scaffolds are superior compared to their counterparts fabricated by using photoinitiator molecules, showing a higher level of biocompatibility. A release of toxic chemicals from the photoinitiator containing scaffolds is proven by cell proliferation tests. In contrast, no toxic release is found from the photoinitiator-free scaffolds, resulting in the very same level of cell proliferation as the control sample. The demonstration of photoinitiator-free PEGDA scaffolds enables the fabrication of 3D scaffolds with the highest level of biocompatibility for both in vitro and in vivo applications.

3D printed porous ceramic scaffolds for bone tissue engineering: a review.

PubMed

Wen, Yu; Xun, Sun; Haoye, Meng; Baichuan, Sun; Peng, Chen; Xuejian, Liu; Kaihong, Zhang; Xuan, Yang; Jiang, Peng; Shibi, Lu

2017-08-22

This study summarizes the recent research status and development of three-dimensional (3D)-printed porous ceramic scaffolds in bone tissue engineering. Recent literature on 3D-printed porous ceramic scaffolds was reviewed. Compared with traditional processing and manufacturing technologies, 3D-printed porous ceramic scaffolds have obvious advantages, such as enhancement of the controllability of the structure or improvement of the production efficiency. More sophisticated scaffolds were fabricated by 3D printing technology. 3D printed bioceramics have broad application prospects in bone tissue engineering. Through understanding the advantages and limitations of different 3D-printing approaches, new classes of bone graft substitutes can be developed.

In Vivo Evaluation of 3D-Printed Polycaprolactone Scaffold Implantation Combined with β-TCP Powder for Alveolar Bone Augmentation in a Beagle Defect Model

PubMed Central

Park, Su A.; Lee, Hyo-Jung; Kim, Keun-Suh; Lee, Jung-Tae; Kim, Sung-Yeol; Chang, Na-Hee

2018-01-01

Insufficient bone volume is one of the major challenges encountered by dentists after dental implant placement. This study aimed to evaluate the efficacy of a customized three-dimensional polycaprolactone (3D PCL) scaffold implant fabricated with a 3D bio-printing system to facilitate rapid alveolar bone regeneration. Saddle-type bone defects were surgically created on the healed site after extracting premolars from the mandibles of four beagle dogs. The defects were radiologically examined using computed tomography for designing a customized 3D PCL scaffold block to fit the defect site. After fabricating 3D PCL scaffolds using rapid prototyping, the scaffolds were implanted into the alveolar bone defects along with β-tricalcium phosphate powder. In vivo analysis showed that the PCL blocks maintained the physical space and bone conductivity around the defects. In addition, no inflammatory infiltrates were observed around the scaffolds. However, new bone formation occurred adjacent to the scaffolds, rather than directly in contact with them. More new bone was observed around PCL blocks with 400/1200 lattices than around blocks with 400/400 lattices, but the difference was not significant. These results indicated the potential of 3D-printed porous PCL scaffolds to promote alveolar bone regeneration for defect healing in dentistry. PMID:29401707

A Novel Polymer-Synthesized Ceramic Composite Based System for Bone Repair: Osteoblast Growth on Scaffolds with Varied Calcium Phosphate Content

DTIC Science & Technology

2005-01-01

demongtrated the synthesis of degradable scaffolds from PLAGA /calcium phosphate composite microspheres in which an amorphous calcium phosphate is...EXPERIMENTAL DETAILS Scaffold Preparation Scaffolds were prepared as described in detail previously [3]. Briefly, PLAGA /calcium phosphate composite...culture polystyrene (TCPS) 2- pure PLAGA microspheres 64 3- composite microsphere matrices with a low polymer/ceramic ratio 4- composite microsphere

Electrospun nanofibrous 3D scaffold for bone tissue engineering.

PubMed

Eap, Sandy; Ferrand, Alice; Palomares, Carlos Mendoza; Hébraud, Anne; Stoltz, Jean-François; Mainard, Didier; Schlatter, Guy; Benkirane-Jessel, Nadia

2012-01-01

Tissue engineering aims at developing functional substitutes for damaged tissues by mimicking natural tissues. In particular, tissue engineering for bone regeneration enables healing of some bone diseases. Thus, several methods have been developed in order to produce implantable biomaterial structures that imitate the constitution of bone. Electrospinning is one of these methods. This technique produces nonwoven scaffolds made of nanofibers which size and organization match those of the extracellular matrix. Until now, seldom electrospun scaffolds were produced with thickness exceeding one millimeter. This article introduces a new kind of electrospun membrane called 3D scaffold of thickness easily exceeding one centimeter. The manufacturing involves a solution of poly(ε-caprolactone) in DMF/DCM system. The aim is to establish parameters for electrospinning in order to characterize these 3D scaffolds and, establish whether such scaffolds are potentially interesting for bone regeneration.

Partially nanofibrous architecture of 3D tissue engineering scaffolds.

PubMed

Wei, Guobao; Ma, Peter X

2009-11-01

An ideal tissue-engineering scaffold should provide suitable pores and appropriate pore surface to induce desired cellular activities and to guide 3D tissue regeneration. In the present work, we have developed macroporous polymer scaffolds with varying pore wall architectures from smooth (solid), microporous, partially nanofibrous, to entirely nanofibrous ones. All scaffolds are designed to have well-controlled interconnected macropores, resulting from leaching sugar sphere template. We examine the effects of material composition, solvent, and phase separation temperature on the pore surface architecture of 3D scaffolds. In particular, phase separation of PLLA/PDLLA or PLLA/PLGA blends leads to partially nanofibrous scaffolds, in which PLLA forms nanofibers and PDLLA or PLGA forms the smooth (solid) surfaces on macropore walls, respectively. Specific surface areas are measured for scaffolds with similar macroporosity but different macropore wall architectures. It is found that the pore wall architecture predominates the total surface area of the scaffolds. The surface area of a partially nanofibrous scaffold increases linearly with the PLLA content in the polymer blend. The amounts of adsorbed proteins from serum increase with the surface area of the scaffolds. These macroporous scaffolds with adjustable pore wall surface architectures may provide a platform for investigating the cellular responses to pore surface architecture, and provide us with a powerful tool to develop superior scaffolds for various tissue-engineering applications.

3D bioactive composite scaffolds for bone tissue engineering.

PubMed

Turnbull, Gareth; Clarke, Jon; Picard, Frédéric; Riches, Philip; Jia, Luanluan; Han, Fengxuan; Li, Bin; Shu, Wenmiao

2018-09-01

Bone is the second most commonly transplanted tissue worldwide, with over four million operations using bone grafts or bone substitute materials annually to treat bone defects. However, significant limitations affect current treatment options and clinical demand for bone grafts continues to rise due to conditions such as trauma, cancer, infection and arthritis. Developing bioactive three-dimensional (3D) scaffolds to support bone regeneration has therefore become a key area of focus within bone tissue engineering (BTE). A variety of materials and manufacturing methods including 3D printing have been used to create novel alternatives to traditional bone grafts. However, individual groups of materials including polymers, ceramics and hydrogels have been unable to fully replicate the properties of bone when used alone. Favourable material properties can be combined and bioactivity improved when groups of materials are used together in composite 3D scaffolds. This review will therefore consider the ideal properties of bioactive composite 3D scaffolds and examine recent use of polymers, hydrogels, metals, ceramics and bio-glasses in BTE. Scaffold fabrication methodology, mechanical performance, biocompatibility, bioactivity, and potential clinical translations will be discussed.

Rapid prototyping for tissue-engineered bone scaffold by 3D printing and biocompatibility study.

PubMed

He, Hui-Yu; Zhang, Jia-Yu; Mi, Xue; Hu, Yang; Gu, Xiao-Yu

2015-01-01

The prototyping of tissue-engineered bone scaffold (calcined goat spongy bone-biphasic ceramic composite/PVA gel) by 3D printing was performed, and the biocompatibility of the fabricated bone scaffold was studied. Pre-designed STL file was imported into the GXYZ303010-XYLE 3D printing system, and the tissue-engineered bone scaffold was fabricated by 3D printing using gel extrusion. Rabbit bone marrow stromal cells (BMSCs) were cultured in vitro and then inoculated to the sterilized bone scaffold obtained by 3D printing. The growth of rabbit BMSCs on the bone scaffold was observed under the scanning electron microscope (SEM). The effect of the tissue-engineered bone scaffold on the proliferation and differentiation of rabbit BMSCs using MTT assay. Universal testing machine was adopted to test the tensile strength of the bone scaffold. The leachate of the bone scaffold was prepared and injected into the New Zealand rabbits. Cytotoxicity test, acute toxicity test, pyrogenic test and intracutaneous stimulation test were performed to assess the biocompatibility of the bone scaffold. Bone scaffold manufactured by 3D printing had uniform pore size with the porosity of about 68.3%. The pores were well interconnected, and the bone scaffold showed excellent mechanical property. Rabbit BMSCs grew and proliferated on the surface of the bone scaffold after adherence. MTT assay indicated that the proliferation and differentiation of rabbit BMSCs on the bone scaffold did not differ significantly from that of the cells in the control. In vivo experiments proved that the bone scaffold fabricated by 3D printing had no acute toxicity, pyrogenic reaction or stimulation. Bone scaffold manufactured by 3D printing allows the rabbit BMSCs to adhere, grow and proliferate and exhibits excellent biomechanical property and high biocompatibility. 3D printing has a good application prospect in the prototyping of tissue-engineered bone scaffold.

Rapid prototyping for tissue-engineered bone scaffold by 3D printing and biocompatibility study

PubMed Central

He, Hui-Yu; Zhang, Jia-Yu; Mi, Xue; Hu, Yang; Gu, Xiao-Yu

2015-01-01

The prototyping of tissue-engineered bone scaffold (calcined goat spongy bone-biphasic ceramic composite/PVA gel) by 3D printing was performed, and the biocompatibility of the fabricated bone scaffold was studied. Pre-designed STL file was imported into the GXYZ303010-XYLE 3D printing system, and the tissue-engineered bone scaffold was fabricated by 3D printing using gel extrusion. Rabbit bone marrow stromal cells (BMSCs) were cultured in vitro and then inoculated to the sterilized bone scaffold obtained by 3D printing. The growth of rabbit BMSCs on the bone scaffold was observed under the scanning electron microscope (SEM). The effect of the tissue-engineered bone scaffold on the proliferation and differentiation of rabbit BMSCs using MTT assay. Universal testing machine was adopted to test the tensile strength of the bone scaffold. The leachate of the bone scaffold was prepared and injected into the New Zealand rabbits. Cytotoxicity test, acute toxicity test, pyrogenic test and intracutaneous stimulation test were performed to assess the biocompatibility of the bone scaffold. Bone scaffold manufactured by 3D printing had uniform pore size with the porosity of about 68.3%. The pores were well interconnected, and the bone scaffold showed excellent mechanical property. Rabbit BMSCs grew and proliferated on the surface of the bone scaffold after adherence. MTT assay indicated that the proliferation and differentiation of rabbit BMSCs on the bone scaffold did not differ significantly from that of the cells in the control. In vivo experiments proved that the bone scaffold fabricated by 3D printing had no acute toxicity, pyrogenic reaction or stimulation. Bone scaffold manufactured by 3D printing allows the rabbit BMSCs to adhere, grow and proliferate and exhibits excellent biomechanical property and high biocompatibility. 3D printing has a good application prospect in the prototyping of tissue-engineered bone scaffold. PMID:26380018

3D Printed Silicone-Hydrogel Scaffold with Enhanced Physicochemical Properties.

PubMed

Mohanty, Soumyaranjan; Alm, Martin; Hemmingsen, Mette; Dolatshahi-Pirouz, Alireza; Trifol, Jon; Thomsen, Peter; Dufva, Martin; Wolff, Anders; Emnéus, Jenny

2016-04-11

Scaffolds with multiple functionalities have attracted widespread attention in the field of tissue engineering due to their ability to control cell behavior through various cues, including mechanical, chemical, and electrical. Fabrication of such scaffolds from clinically approved materials is currently a huge challenge. The goal of this work was to fabricate a tissue engineering scaffold from clinically approved materials with the capability of delivering biomolecules and direct cell fate. We have used a simple 3D printing approach, that combines polymer casting with supercritical fluid technology to produce 3D interpenetrating polymer network (IPN) scaffold of silicone-poly(2-hydroxyethyl methacrylate)-co-poly(ethylene glycol) methyl ether acrylate (pHEMA-co-PEGMEA). The pHEMA-co-PEGMEA IPN materials were employed to support growth of human mesenchymal stem cells (hMSC), resulting in high cell viability and metabolic activity over a 3 weeks period. In addition, the IPN scaffolds support 3D tissue formation inside the porous scaffold with well spread cell morphology on the surface of the scaffold. As a proof of concept, sustained doxycycline (DOX) release from pHEMA-co-PEGMEA IPN was demonstrated and the biological activity of released drug from IPN was confirmed using a DOX regulated green fluorescent reporter (GFP) gene expression assay with HeLa cells. Given its unique mechanical and drug releasing characteristics, IPN scaffolds may be used for directing stem cell differentiation by releasing various chemicals from its hydrogel network.

Development of a novel alginate-polyvinyl alcohol-hydroxyapatite hydrogel for 3D bioprinting bone tissue engineered scaffolds.

PubMed

Bendtsen, Stephanie T; Quinnell, Sean P; Wei, Mei

2017-05-01

Three-dimensional printed biomaterials used as personalized tissue substitutes have the ability to promote and enhance regeneration in areas of defected tissue. The challenge with 3D printing for bone tissue engineering remains the selection of a material with optimal rheological properties for printing in addition to biocompatibility and capacity for uniform cell incorporation. Hydrogel biomaterials may provide sufficient printability to allow cell encapsulation and bioprinting of scaffolds with uniform cell distribution. In this study, a novel alginate-polyvinyl alcohol (PVA)-hydroxyapatite (HA) hydrogel formulation with optimal rheological properties for 3D bioprinting of mouse calvaria 3T3-E1 (MC3T3) cells into scaffolds of high shape fidelity has been developed. A systematic investigation was conducted to determine the effect of varying concentrations of alginate, phosphate, calcium, and the PVA-HA suspension in the formulation on the resulting viscosity and thus printability of the hydrogel. HA, the main mineral component in natural bone, was incorporated into the hydrogel formulation to create a favorable bone-forming environment due to its excellent osteoconductivity. Degradation studies in α-MEM cell culture media showed that the 3D printed alginate-PVA-HA scaffolds remained in-tact for 14 days. MC3T3 cells were well distributed and encapsulated throughout the optimal hydrogel formulation and expressed high viability through the completion of the 3D printing process. Thus, the development of this novel, osteoconductive, biodegradable, alginate-PVA-HA formulation and its ability to 3D bioprint tissue engineered scaffolds make it a promising candidate for treating personalized bone defects. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1457-1468, 2017. © 2017 Wiley Periodicals, Inc.

Modeling and Reconstruction of Micro-structured 3D Chitosan/Gelatin Porous Scaffolds Using Micro-CT

NASA Astrophysics Data System (ADS)

Gong, Haibo; Li, Dichen; He, Jiankang; Liu, Yaxiong; Lian, Qin; Zhao, Jinna

2008-09-01

Three dimensional (3D) channel networks are the key to promise the uniform distribution of nutrients inside 3D hepatic tissue engineering scaffolds and prompt elimination of metabolic products out of the scaffolds. 3D chitosan/gelatin porous scaffolds with predefined internal channels were fabricated and a combination of light microscope, laser confocal microscopy and micro-CT were employed to characterize the structure of porous scaffolds. In order to evaluate the flow field distribution inside the micro-structured 3D scaffolds, a computer reconstructing method based on Micro-CT was proposed. According to this evaluating method, a contrast between 3D porous scaffolds with and without predefined internal channels was also performed to assess scaffolds' fluid characters. Results showed that the internal channel of the 3D scaffolds formed the 3D fluid channel network; the uniformity of flow field distribution of the scaffolds fabricated in this paper was better than the simple porous scaffold without micro-fluid channels.

Superelastic, superabsorbent and 3D nanofiber-assembled scaffold for tissue engineering.

PubMed

Chen, Weiming; Ma, Jun; Zhu, Lei; Morsi, Yosry; -Ei-Hamshary, Hany; Al-Deyab, Salem S; Mo, Xiumei

2016-06-01

Fabrication of 3D scaffold to mimic the nanofibrous structure of the nature extracellular matrix (ECM) with appropriate mechanical properties and excellent biocompatibility, remain an important technical challenge in tissue engineering. The present study reports the strategy to fabricate a 3D nanofibrous scaffold with similar structure to collagen in ECM by combining electrospinning and freeze-drying technique. With the technique reported here, a nanofibrous structure scaffold with hydrophilic and superabsorbent properties can be readily prepared by Gelatin and Polylactic acid (PLA). In wet state the scaffold also shows a super-elastic property, which could bear a compressive strain as high as 80% and recovers its original shape afterwards. Moreover, after 6 days of culture, L-929 cells grow, proliferate and infiltrated into the scaffold. The results suggest that this 3D nanofibrous scaffold would be promising for varied field of tissue engineering application. Copyright © 2016 Elsevier B.V. All rights reserved.

Bone Regeneration in Critical Bone Defects Using Three-Dimensionally Printed β-Tricalcium Phosphate/Hydroxyapatite Scaffolds Is Enhanced by Coating Scaffolds with Either Dipyridamole or BMP-2

PubMed Central

Ishack, Stephanie; Mediero, Aranzazu; Wilder, Tuere; Ricci, John L.; Cronstein, Bruce N.

2017-01-01

Bone defects resulting from trauma or infection need timely and effective treatments to restore damaged bone. Using specialized three-dimensional (3-D) printing technology we have created custom 3-D scaffolds of hydroxyapatite (HA)/Beta-Tri-Calcium Phosphate (β-TCP) to promote bone repair. To further enhance bone regeneration we have coated the scaffolds with dipyridamole, an agent that increases local adenosine levels by blocking cellular uptake of adenosine. 15% HA:85% β-TCP scaffolds were designed using Robocad software, fabricated using a 3-D Robocasting system, and sintered at 1100°C for 4h. Scaffolds were coated with BMP-2 (200ng/ml), Dypiridamole 100µM or saline and implanted in C57B6 and adenosine A2A receptor knockout (A2AKO) mice with 3mm cranial critical bone defects for 2-8 weeks. Dipyridamole release from scaffold was assayed spectrophotometrically. MicroCT and histological analysis were performed. micro-computed tomography (microCT) showed significant bone formation and remodeling in HA/β-TCP- dipyridamole and HA/β-TCP -BMP-2 scaffolds when compared to scaffolds immersed in vehicle at 2, 4 and 8 weeks (n=5 per group; p≤ 0.05, p≤ 0.05 and p≤ 0.01, respectively). Histological analysis showed increased bone formation and a trend toward increased remodeling in HA/β-TCP- dipyridamole and HA/β-TCP-BMP-2 scaffolds. coating scaffolds with dipyridamole did not enhance bone regeneration in A2AKO mice. In conclusion, scaffolds printed with HA/β-TCP promote bone regeneration in critical bone defects and coating these scaffolds with agents that stimulate A2A receptors and growth factors can further enhance bone regeneration. These coated scaffolds may be very useful for treating critical bone defects due to trauma, infection or other causes. PMID:26513656

Polycaprolactone- and polycaprolactone/ceramic-based 3D-bioplotted porous scaffolds for bone regeneration: A comparative study.

PubMed

Gómez-Lizárraga, K K; Flores-Morales, C; Del Prado-Audelo, M L; Álvarez-Pérez, M A; Piña-Barba, M C; Escobedo, C

2017-10-01

One of the critical challenges that scaffolding faces in the organ and tissue regeneration field lies in mimicking the structure, and the chemical and biological properties of natural tissue. A high-level control over the architecture, mechanical properties and composition of the materials in contact with cells is essential to overcome such challenge. Therefore, definition of the method, materials and parameters for the production of scaffolds during the fabrication stage is critical. With the recent emergence of rapid prototyping (RP), it is now possible to create three-dimensional (3D) scaffolds with the essential characteristics for the proliferation and regeneration of tissues, such as porosity, mechanical strength, pore size and pore interconnectivity, and biocompatibility. In this study, we employed 3D bioplotting, a RP technology, to fabricate scaffolds made from (i) pure polycaprolactone (PCL) and (ii) a composite based on PCL and ceramic micro-powder. The ceramics used for the composite were bovine bone filling Nukbone® (NKB), and hydroxyapatite (HA) with 5%, 10% or 20% wt. The scaffolds were fabricated in a cellular lattice structure (i.e. meshing mode) using a 0/90° lay down pattern with a continuous contour filament in order to achieve interconnected porous reticular structures. We varied the temperature, as well as injection speed and pressure during the bioplotting process to achieve scaffolds with pore size ranging between 200 and 400μm and adequate mechanical stability. The resulting scaffolds had an average pore size of 323μm and an average porosity of 32%. Characterization through ATR-FTIR revealed the presence of the characteristic bands of hydroxyapatite in the PCL matrix, and presented an increase of the intensity of the phosphate and carbonyl bands as the ceramic content increased. The bioplotted 3D scaffolds have a Young's modulus (E) in the range between 0.121 and 0.171GPa, which is compatible with the modulus of natural bone. PCL

Robotic deposition and in vitro characterization of 3D gelatin-bioactive glass hybrid scaffolds for biomedical applications.

PubMed

Gao, Chunxia; Rahaman, Mohamed N; Gao, Qiang; Teramoto, Akira; Abe, Koji

2013-07-01

The development of inorganic-organic hybrid scaffolds with controllable degradation and bioactive properties is receiving considerable interest for bone and tissue regeneration. The objective of this study was to create hybrid scaffolds of gelatin and bioactive glass (BG) with a controlled, three-dimensional (3D) architecture by a combined sol-gel and robotic deposition (robocasting) method and evaluate their mechanical response, bioactivity, and response to cells in vitro. Inks for robotic deposition of the scaffolds were prepared by dissolving gelatin in a sol-gel precursor solution of the bioactive glass (70SiO2 -25CaO-5P2 O5 ; mol%) and aging the solution to form a gel with the requisite viscosity. After drying and crosslinking, the gelatin-BG scaffolds, with a grid-like architecture (filament diameter ∼350 µm; pore width ∼550 µm), showed an elasto-plastic response, with a compressive strength of 5.1 ± 0.6 MPa, in the range of values for human trabecular bone (2-12 MPa). When immersed in phosphate-buffered saline, the crosslinked scaffolds rapidly absorbed water (∼440% of its dry weight after 2 h) and showed an elastic response at deformations up to ∼60%. Immersion of the scaffolds in a simulated body fluid resulted in the formation of a hydroxyapatite-like surface layer within 5 days, indicating their bioactivity in vitro. The scaffolds supported the proliferation, alkaline phosphatase activity, and mineralization of osteogenic MC3T3-E1 cells in vitro, showing their biocompatibility. Altogether, the results indicate that these gelatin-BG hybrid scaffolds with a controlled, 3D architecture of inter-connected pores have potential for use as implants for bone regeneration. Copyright © 2012 Wiley Periodicals, Inc.

Extrusion-based, three-dimensional printing of calcium-phosphate scaffolds

NASA Astrophysics Data System (ADS)

Witek, Lukasz

Small or large bone defects, can occur due to a variety of reasons: congenital disorders, infections, tumors, or traumas which can lead to significant disabilities. There is an assortment of bone grafting procedures, each having their own respective advantages and disadvantages and exhibiting certain essential characteristics. Among the available grafts, autogenous (autograft), allograft, xenograft, and alloplasts, all exhibit a minimum of two-thirds of the essential characteristics and have been proven useful in fully or partially repairing skeletal defects. However, different host-to-grafting material responses have been reported and should be taken into consideration when determining treatment options. A large range of physical and chemical properties can be achieved with calcium phosphate based materials, which possess two of the ideal characteristics for grafting procedures: osteoconduction and osseointegration. Calcium phosphate based scaffolds composed of hydroxyapatite (HA), beta-tri-calcium phosphate (beta-TCP), or a combination of both (HA/beta-TCP) were investigated as materials for three-dimensional printing process to create layer-by-layer structures for use as bone regeneration scaffolds. Different calcium-phosphate phases will result in different degrees of in vivo dissolution and/or cell-mediated resorption. There has been a growing interest in BCP because it has been shown that this material improves the formation of new bone inside the implanted scaffold. The literature indicates that the faster dissolution rate of ?-TCP would be greatly responsible of this enhancement. However, in vitro tests indicate that fast dissolution can decrease the mechanical strength of BCP scaffolds. Furthermore, studies reported that HA has higher mechanical strength and lower degradation rate than beta-TCP. Therefore, the HA/beta-TCP ratio is a key parameter controlling the performance of the scaffold for bone repair applications, since it determines degradation rate

Hybrid braided 3-D scaffold for bioartificial liver assist devices.

PubMed

Hoque, M E; Mao, H Q; Ramakrishna, S

2007-01-01

Three-dimensional ex vivo hepatocyte culture is a tissue-engineering approach to improve the treatment of liver disease. The extracorporeal bioartificial liver (BAL) assists devices that are used in patients until they either recover or receive a liver transplant. The 3-D scaffold plays a key role in the design of bioreactor that is the most important component of the BAL. Presently available 3-D scaffolds used in BAL have shown good performance. However, existing scaffolds are considered to be less than ideal in terms of high-density cultures of hepatocytes maintaining long-term metabolic functions. This study aims to develop a 3-D hybrid scaffold for a BAL support system that would facilitate high-density hepatocyte anchorage with long-term metabolic functions. The scaffolds were fabricated by interlacing polyethylene terephthalate (PET) fibers onto the polysulfone hollow fibers utilizing a modern microbraiding technique. Scaffolds with various pore sizes and porosities were developed by varying braiding angle which was controlled by the gear ratio of the microbraiding machine. The morphological characteristics (pore size and porosity) of the scaffolds were found to be regulated by the gear ratio. Smaller braiding angle yields larger pore and higher porosity. On the other hand, a larger braiding angle causes smaller pore and lower porosity. In hepatocyte culture it was investigated how the morphological characteristics (pore size and porosity) of scaffolds influenced the cell anchorage and metabolic functions. Scaffolds with larger pores and higher porosity resulted in more cell anchorage and higher cellular functions, like albumin and urea secretion, compared to that of smaller pores and lower porosity.

3D Electrospun scaffolds promote a cytotrophic phenotype of cultured primary astrocytes.

PubMed

Lau, Chew L; Kovacevic, Michelle; Tingleff, Tine S; Forsythe, John S; Cate, Holly S; Merlo, Daniel; Cederfur, Cecilia; Maclean, Francesca L; Parish, Clare L; Horne, Malcolm K; Nisbet, David R; Beart, Philip M

2014-07-01

Astrocytes are a target for regenerative neurobiology because in brain injury their phenotype arbitrates brain integrity, neuronal death and subsequent repair and reconstruction. We explored the ability of 3D scaffolds to direct astrocytes into phenotypes with the potential to support neuronal survival. Poly-ε-caprolactone scaffolds were electrospun with random and aligned fibre orientations on which murine astrocytes were sub-cultured and analysed at 4 and 12 DIV. Astrocytes survived, proliferated and migrated into scaffolds adopting 3D morphologies, mimicking in vivo stellated phenotypes. Cells on random poly-ε-caprolactone scaffolds grew as circular colonies extending processes deep within sub-micron fibres, whereas astrocytes on aligned scaffolds exhibited rectangular colonies with processes following not only the direction of fibre alignment but also penetrating the scaffold. Cell viability was maintained over 12 DIV, and cytochemistry for F-/G-actin showed fewer stress fibres on bioscaffolds relative to 2D astrocytes. Reduced cytoskeletal stress was confirmed by the decreased expression of glial fibrillary acidic protein. PCR demonstrated up-regulation of genes (excitatory amino acid transporter 2, brain-derived neurotrophic factor and anti-oxidant) reflecting healthy biologies of mature astrocytes in our extended culture protocol. This study illustrates the therapeutic potential of bioengineering strategies using 3D electrospun scaffolds which direct astrocytes into phenotypes supporting brain repair. Astrocytes exist in phenotypes with pro-survival and destructive components, and their biology can be modulated by changing phenotype. Our findings demonstrate murine astrocytes adopt a healthy phenotype when cultured in 3D. Astrocytes proliferate and extend into poly-ε-caprolactone scaffolds displaying 3D stellated morphologies with reduced GFAP expression and actin stress fibres, plus a cytotrophic gene profile. Bioengineered 3D scaffolds have potential

Impregnation of β-tricalcium phosphate robocast scaffolds by in situ polymerization.

PubMed

Martínez-Vázquez, Francisco J; Perera, Fidel H; van der Meulen, Inge; Heise, Andreas; Pajares, Antonia; Miranda, Pedro

2013-11-01

Ring-opening polymerization of ε-caprolactone (ε-CL) and L-lactide (LLA) was performed to impregnate β-tricalcium phosphate (β-TCP) scaffolds fabricated by robocasting. Concentrated colloidal inks prepared from β-TCP commercial powders were used to fabricate porous structures consisting of a 3D mesh of interpenetrating rods. ε-CL and LLA were in situ polymerized within the ceramic structure by using a lipase and stannous octanoate, respectively, as catalysts. The results show that both the macropores inside the ceramic mesh and the micropores within the ceramic rods are full of polymer in either case. The mechanical properties of scaffolds impregnated by in situ polymerization (ISP) are significantly increased over those of the bare structures, exhibiting similar values than those obtained by other, more aggressive, impregnation methods such as melt-immersion (MI). ISP using enzymatic catalysts requires a reduced processing temperature which could facilitate the incorporation of growth factors and other drugs into the polymer composition, thus enhancing the bioactivity of the composite scaffold. The implications of these results for the optimization of the mechanical and biological performance of scaffolds for bone tissue engineering applications are discussed. Copyright © 2013 Wiley Periodicals, Inc.

Spiral-structured, nanofibrous, 3D scaffolds for bone tissue engineering.

PubMed

Wang, Junping; Valmikinathan, Chandra M; Liu, Wei; Laurencin, Cato T; Yu, Xiaojun

2010-05-01

Polymeric nanofiber matrices have already been widely used in tissue engineering. However, the fabrication of nanofibers into complex three-dimensional (3D) structures is restricted due to current manufacturing techniques. To overcome this limitation, we have incorporated nanofibers onto spiral-structured 3D scaffolds made of poly (epsilon-caprolactone) (PCL). The spiral structure with open geometries, large surface areas, and porosity will be helpful for improving nutrient transport and cell penetration into the scaffolds, which are otherwise limited in conventional tissue-engineered scaffolds for large bone defects repair. To investigate the effect of structure and fiber coating on the performance of the scaffolds, three groups of scaffolds including cylindrical PCL scaffolds, spiral PCL scaffolds (without fiber coating), and spiral-structured fibrous PCL scaffolds (with fiber coating) have been prepared. The morphology, porosity, and mechanical properties of the scaffolds have been characterized. Furthermore, human osteoblast cells are seeded on these scaffolds, and the cell attachment, proliferation, differentiation, and mineralized matrix deposition on the scaffolds are evaluated. The results indicated that the spiral scaffolds possess porosities within the range of human trabecular bone and an appropriate pore structure for cell growth, and significantly lower compressive modulus and strength than cylindrical scaffolds. When compared with the cylindrical scaffolds, the spiral-structured scaffolds demonstrated enhanced cell proliferation, differentiation, and mineralization and allowed better cellular growth and penetration. The incorporation of nanofibers onto spiral scaffolds further enhanced cell attachment, proliferation, and differentiation. These studies suggest that spiral-structured nanofibrous scaffolds may serve as promising alternatives for bone tissue engineering applications. Copyright 2009 Wiley Periodicals, Inc.

Improved resolution of 3D printed scaffolds by shrinking.

PubMed

Chia, Helena N; Wu, Benjamin M

2015-10-01

Three-dimensional printing (3DP) uses inkjet printheads to selectively deposit liquid binder to adjoin powder particles in a layer-by-layer fashion to create a computer-modeled 3D object. Two general approaches for 3DP have been described for biomedical applications (direct and indirect 3DP). The two approaches offer competing advantages, and both are limited by print resolution. This study describes a materials processing strategy to enhance 3DP resolution by controlled shrinking net-shape scaffolds. Briefly, porogen preforms are printed and infused with the desired monomer or polymer solution. After solidification or polymerization, the porogen is leached and the polymer is allowed to shrink by controlled drying. Heat treatment is performed to retain the dimensions against swelling forces. The main objective of this study is to determine the effects of polymer content and post-processing on dimension, microstructure, and thermomechanical properties of the scaffold. For polyethylene glycol diacrylate (PEG-DA), reducing polymer content corresponded with greater shrinkage with maximum shrinkage of ∼80 vol% at 20% vol% PEG-DA. The secondary heat treatment retains the microarchitecture and new dimensions of the scaffolds, even when the heat-treated scaffolds are immersed into water. To demonstrate shrinkage predictability, 3D components with interlocking positive and negative features were printed, processed, and fitted. This material processing strategy provides an alternative method to enhance the resolution of 3D scaffolds, for a wide range of polymers, without optimizing the binder-powder interaction physics to print each material combination. © 2014 Wiley Periodicals, Inc.

Soft chitosan microbeads scaffold for 3D functional neuronal networks.

PubMed

Tedesco, Maria Teresa; Di Lisa, Donatella; Massobrio, Paolo; Colistra, Nicolò; Pesce, Mattia; Catelani, Tiziano; Dellacasa, Elena; Raiteri, Roberto; Martinoia, Sergio; Pastorino, Laura

2018-02-01

The availability of 3D biomimetic in vitro neuronal networks of mammalian neurons represents a pivotal step for the development of brain-on-a-chip experimental models to study neuronal (dys)functions and particularly neuronal connectivity. The use of hydrogel-based scaffolds for 3D cell cultures has been extensively studied in the last years. However, limited work on biomimetic 3D neuronal cultures has been carried out to date. In this respect, here we investigated the use of a widely popular polysaccharide, chitosan (CHI), for the fabrication of a microbead based 3D scaffold to be coupled to primary neuronal cells. CHI microbeads were characterized by optical and atomic force microscopies. The cell/scaffold interaction was deeply characterized by transmission electron microscopy and by immunocytochemistry using confocal microscopy. Finally, a preliminary electrophysiological characterization by micro-electrode arrays was carried out. Copyright © 2017 Elsevier Ltd. All rights reserved.

3D conductive nanocomposite scaffold for bone tissue engineering.

PubMed

Shahini, Aref; Yazdimamaghani, Mostafa; Walker, Kenneth J; Eastman, Margaret A; Hatami-Marbini, Hamed; Smith, Brenda J; Ricci, John L; Madihally, Sundar V; Vashaee, Daryoosh; Tayebi, Lobat

2014-01-01

Bone healing can be significantly expedited by applying electrical stimuli in the injured region. Therefore, a three-dimensional (3D) ceramic conductive tissue engineering scaffold for large bone defects that can locally deliver the electrical stimuli is highly desired. In the present study, 3D conductive scaffolds were prepared by employing a biocompatible conductive polymer, ie, poly(3,4-ethylenedioxythiophene) poly(4-styrene sulfonate) (PEDOT:PSS), in the optimized nanocomposite of gelatin and bioactive glass. For in vitro analysis, adult human mesenchymal stem cells were seeded in the scaffolds. Material characterizations using hydrogen-1 nuclear magnetic resonance, in vitro degradation, as well as thermal and mechanical analysis showed that incorporation of PEDOT:PSS increased the physiochemical stability of the composite, resulting in improved mechanical properties and biodegradation resistance. The outcomes indicate that PEDOT:PSS and polypeptide chains have close interaction, most likely by forming salt bridges between arginine side chains and sulfonate groups. The morphology of the scaffolds and cultured human mesenchymal stem cells were observed and analyzed via scanning electron microscope, micro-computed tomography, and confocal fluorescent microscope. Increasing the concentration of the conductive polymer in the scaffold enhanced the cell viability, indicating the improved microstructure of the scaffolds or boosted electrical signaling among cells. These results show that these conductive scaffolds are not only structurally more favorable for bone tissue engineering, but also can be a step forward in combining the tissue engineering techniques with the method of enhancing the bone healing by electrical stimuli.

Effects of SiO2 and ZnO doping on mechanical and biological properties of 3D printed TCP scaffolds

PubMed Central

Fielding, Gary A.; Bandyopadhyay, Amit; Bose, Susmita

2011-01-01

Objectives To evaluate the effects of SiO2 (0.5 wt %) and ZnO (0.25 wt %) dopants on the mechanical and biological properties of tricalcium phosphate (TCP) scaffolds with three dimensionally (3D) interconnected pores. Methods Scaffolds were created with a commercial 3D printer. Post sintering phase analysis was determined by x-ray diffraction. Surface morphology of the scaffolds was examined by field emission electron microscopy. Mechanical strength was evaluated with a screw driven universal testing machine. MTT assay was used for cellular proliferation characteristics and cellular morphology was examined by field emission electron microscopy. Results Addition of dopants into TCP increased the average density of pure TCP from 90.8 ± 0.8% to 94.1 ± 1.6% and retarded the β to α phase transformation at high sintering temperatures, which resulted in up to 2.5 fold increase in compressive strength. In vitro cell-materials interaction studies, carried out using hFOB cells, confirmed that the addition of SiO2 and ZnO to the scaffolds facilitates faster cell proliferation when compared to pure TCP scaffolds. Significance Addition of SiO2 and ZnO dopants to the TCP scaffolds showed increased mechanical strength as well as increased cellular proliferation. PMID:22047943

Fabrication and Evaluation of Electrospun, 3D-Bioplotted, and Combination of Electrospun/3D-Bioplotted Scaffolds for Tissue Engineering Applications

PubMed Central

Mellor, Liliana F.; Huebner, Pedro; Cai, Shaobo; Taylor, Michael A.; Spang, Jeffrey

2017-01-01

Electrospun scaffolds provide a dense framework of nanofibers with pore sizes and fiber diameters that closely resemble the architecture of native extracellular matrix. However, it generates limited three-dimensional structures of relevant physiological thicknesses. 3D printing allows digitally controlled fabrication of three-dimensional single/multimaterial constructs with precisely ordered fiber and pore architecture in a single build. However, this approach generally lacks the ability to achieve submicron resolution features to mimic native tissue. The goal of this study was to fabricate and evaluate 3D printed, electrospun, and combination of 3D printed/electrospun scaffolds to mimic the native architecture of heterogeneous tissue. We assessed their ability to support viability and proliferation of human adipose derived stem cells (hASC). Cells had increased proliferation and high viability over 21 days on all scaffolds. We further tested implantation of stacked-electrospun scaffold versus combined electrospun/3D scaffold on a cadaveric pig knee model and found that stacked-electrospun scaffold easily delaminated during implantation while the combined scaffold was easier to implant. Our approach combining these two commonly used scaffold fabrication technologies allows for the creation of a scaffold with more close resemblance to heterogeneous tissue architecture, holding great potential for tissue engineering and regenerative medicine applications of osteochondral tissue and other heterogeneous tissues. PMID:28536700

3D Printing of Scaffolds for Tissue Regeneration Applications.

PubMed

Do, Anh-Vu; Khorsand, Behnoush; Geary, Sean M; Salem, Aliasger K

2015-08-26

The current need for organ and tissue replacement, repair, and regeneration for patients is continually growing such that supply is not meeting demand primarily due to a paucity of donors as well as biocompatibility issues leading to immune rejection of the transplant. In order to overcome these drawbacks, scientists have investigated the use of scaffolds as an alternative to transplantation. These scaffolds are designed to mimic the extracellular matrix (ECM) by providing structural support as well as promoting attachment, proliferation, and differentiation with the ultimate goal of yielding functional tissues or organs. Initial attempts at developing scaffolds were problematic and subsequently inspired an interest in 3D printing as a mode for generating scaffolds. Utilizing three-dimensional printing (3DP) technologies, ECM-like scaffolds can be produced with a high degree of complexity, where fine details can be included at a micrometer level. In this Review, the criteria for printing viable and functional scaffolds, scaffolding materials, and 3DP technologies used to print scaffolds for tissue engineering are discussed. Creating biofunctional scaffolds could potentially help to meet the demand by patients for tissues and organs without having to wait or rely on donors for transplantation. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

3D conductive nanocomposite scaffold for bone tissue engineering

PubMed Central

Shahini, Aref; Yazdimamaghani, Mostafa; Walker, Kenneth J; Eastman, Margaret A; Hatami-Marbini, Hamed; Smith, Brenda J; Ricci, John L; Madihally, Sundar V; Vashaee, Daryoosh; Tayebi, Lobat

2014-01-01

Bone healing can be significantly expedited by applying electrical stimuli in the injured region. Therefore, a three-dimensional (3D) ceramic conductive tissue engineering scaffold for large bone defects that can locally deliver the electrical stimuli is highly desired. In the present study, 3D conductive scaffolds were prepared by employing a biocompatible conductive polymer, ie, poly(3,4-ethylenedioxythiophene) poly(4-styrene sulfonate) (PEDOT:PSS), in the optimized nanocomposite of gelatin and bioactive glass. For in vitro analysis, adult human mesenchymal stem cells were seeded in the scaffolds. Material characterizations using hydrogen-1 nuclear magnetic resonance, in vitro degradation, as well as thermal and mechanical analysis showed that incorporation of PEDOT:PSS increased the physiochemical stability of the composite, resulting in improved mechanical properties and biodegradation resistance. The outcomes indicate that PEDOT:PSS and polypeptide chains have close interaction, most likely by forming salt bridges between arginine side chains and sulfonate groups. The morphology of the scaffolds and cultured human mesenchymal stem cells were observed and analyzed via scanning electron microscope, micro-computed tomography, and confocal fluorescent microscope. Increasing the concentration of the conductive polymer in the scaffold enhanced the cell viability, indicating the improved microstructure of the scaffolds or boosted electrical signaling among cells. These results show that these conductive scaffolds are not only structurally more favorable for bone tissue engineering, but also can be a step forward in combining the tissue engineering techniques with the method of enhancing the bone healing by electrical stimuli. PMID:24399874

Microsphere-Based Scaffolds Encapsulating Tricalcium Phosphate And Hydroxyapatite For Bone Regeneration

PubMed Central

Gupta, Vineet; Lyne, Dina V.; Barragan, Marilyn; Berkland, Cory J.; Detamore, Michael S.

2016-01-01

Bioceramic mixtures of tricalcium phosphate (TCP) and hydroxyapatite (HAp) are widely used for bone regeneration because of their excellent cytocompatibility, osteoconduction, and osteoinduction. Therefore, we hypothesized that incorporation of a mixture of TCP and HAp in microsphere-based scaffolds would enhance osteogenesis of rat bone marrow stromal cells (rBMSCs) compared to a positive control of scaffolds with encapsulated bone-morphogenic protein-2 (BMP-2). Poly(D,L-lactic-co-glycolic acid) (PLGA) microsphere-based scaffolds encapsulating TCP and HAp mixtures in two different ratios (7:3 and 1:1) were fabricated with the same net ceramic content (30 wt%) to evaluate how incorporation of these ceramic mixtures would affect the osteogenesis in rBMSCs. Encapsulation of TCP/HAp mixtures impacted microsphere morphologies and the compressive moduli of the scaffolds. Additionally, TCP/HAp mixtures enhanced the end-point secretion of extracellular matrix (ECM) components relevant to bone tissue compared to the “blank” (PLGA-only) microsphere-based scaffolds as evidenced by the biochemical, gene expression, histology, and immunohistochemical characterization. Moreover, the TCP/HAp mixture groups even surpassed the BMP-2 positive control group in some instances in terms of matrix synthesis and gene expression. Lastly, gene expression data suggested that the rBMSCs responded differently to different TCP/HAp ratios presented to them. Altogether, it can be concluded that TCP/HAp mixtures stimulated the differentiation of rBMSCs toward an osteoblastic phenotype, and therefore may be beneficial in gradient microsphere-based scaffolds for osteochondral regeneration. PMID:27272903

Bone regeneration in critical bone defects using three-dimensionally printed β-tricalcium phosphate/hydroxyapatite scaffolds is enhanced by coating scaffolds with either dipyridamole or BMP-2.

PubMed

Ishack, Stephanie; Mediero, Aranzazu; Wilder, Tuere; Ricci, John L; Cronstein, Bruce N

2017-02-01

Bone defects resulting from trauma or infection need timely and effective treatments to restore damaged bone. Using specialized three-dimensional (3D) printing technology we have created custom 3D scaffolds of hydroxyapatite (HA)/beta-tri-calcium phosphate (β-TCP) to promote bone repair. To further enhance bone regeneration we have coated the scaffolds with dipyridamole, an agent that increases local adenosine levels by blocking cellular uptake of adenosine. Nearly 15% HA:85% β-TCP scaffolds were designed using Robocad software, fabricated using a 3D Robocasting system, and sintered at 1100°C for 4 h. Scaffolds were coated with BMP-2 (200 ng mL -1 ), dypiridamole 100 µM or saline and implanted in C57B6 and adenosine A2A receptor knockout (A2AKO) mice with 3 mm cranial critical bone defects for 2-8 weeks. Dipyridamole release from scaffold was assayed spectrophotometrically. MicroCT and histological analysis were performed. Micro-computed tomography (microCT) showed significant bone formation and remodeling in HA/β-TCP-dipyridamole and HA/β-TCP-BMP-2 scaffolds when compared to scaffolds immersed in vehicle at 2, 4, and 8 weeks (n = 5 per group; p ≤ 0.05, p ≤ 0.05, and p ≤ 0.01, respectively). Histological analysis showed increased bone formation and a trend toward increased remodeling in HA/β-TCP- dipyridamole and HA/β-TCP-BMP-2 scaffolds. Coating scaffolds with dipyridamole did not enhance bone regeneration in A2AKO mice. In conclusion, scaffolds printed with HA/β-TCP promote bone regeneration in critical bone defects and coating these scaffolds with agents that stimulate A2A receptors and growth factors can further enhance bone regeneration. These coated scaffolds may be very useful for treating critical bone defects due to trauma, infection or other causes. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 366-375, 2017. © 2015 Wiley Periodicals, Inc.

Friction of sodium alginate hydrogel scaffold fabricated by 3-D printing.

PubMed

Yang, Qian; Li, Jian; Xu, Heng; Long, Shijun; Li, Xuefeng

2017-04-01

A rapid prototyping technology, formed by three-dimensional (3-D) printing and then crosslinked by spraying Ca 2+ solution, is developed to fabricate a sodium alginate (SA) hydrogel scaffold. The porosity, swelling ratio, and compression modulus of the scaffold are investigated. A friction mechanism is developed by studying the reproducible friction behavior. Our results show that the scaffold can have 3-D structure with a porosity of 52%. The degree of swelling of the SA hydrogel scaffold is 8.5, which is nearly the same as bulk SA hydrogel. SA hydrogel exhibits better compressive resilience than bulk hydrogel despite its lower compressive modulus compared to bulk hydrogel. The SA hydrogel scaffold exhibits a higher frictional force at low sliding velocity (10 -6 to 10 -3  m/s) compared to bulk SA hydrogel, and they are equal at high sliding velocity (10 -2 to 1 m/s). For a small pressure (0.3 kPa), the SA hydrogel scaffold shows good friction reproducibility. In contrast, bulk SA hydrogel shows poor reproducibility with respect to friction behavior. The differences in friction behaviors between the SA hydrogel scaffold and bulk SA hydrogel are related to the structure of the scaffold, which can keep a stable hydrated lubrication layer.

Oxygen Plasma Treatment on 3D-Printed Chitosan/Gelatin/Hydroxyapatite Scaffolds for Bone Tissue Engineering.

PubMed

Lee, Chang-Min; Yang, Seong-Won; Jung, Sang-Chul; Kim, Byung-Hoon

2017-04-01

The 3D hydroxyapatite/gelatin/chitosan composite scaffolds were fabricated by 3D printing technique. The scaffolds were treated by oxygen plasma to improve the bioactivity and its surface characterization and in vitro cell culture were investigated. The scaffolds exhibited the good porosity and interconnectivity of pores. After oxygen plasma etching, roughness and wettability on the scaffolds surface are increased. Plasma treated scaffolds showed higher proliferation than that of untreated scaffolds. Oxygen plasma treatment could be used as potential tool to enhance the biocompatibility on the 3D composite scaffolds.

3D chitosan-gelatin-chondroitin porous scaffold improves osteogenic differentiation of mesenchymal stem cells.

PubMed

Machado, C B; Ventura, J M G; Lemos, A F; Ferreira, J M F; Leite, M F; Goes, A M

2007-06-01

A porous 3D scaffold was developed to support and enhance the differentiation process of mesenchymal stem cells (MSC) into osteoblasts in vitro. The 3D scaffold was made with chitosan, gelatin and chondroitin and it was crosslinked by EDAC. The scaffold physicochemical properties were evaluated. SEM revealed the high porosity and interconnection of pores in the scaffold; rheological measurements show that the scaffold exhibits a characteristic behavior of strong gels. The elastic modulus found in compressive tests of the crosslinked scaffold was about 50 times higher than the non-crosslinked one. After 21 days, the 3D matrix submitted to hydrolytic degradation loses above 40% of its weight. MSC were collected from rat bone marrow and seeded in chitosan-gelatin-chondroitin 3D scaffolds and in 2D culture plates as well. MSC were differentiated into osteoblasts for 21 days. Cell proliferation and alkaline phosphatase activity were followed weekly during the osteogenic process. The osteogenic differentiation of MSC was improved in 3D culture as shown by MTT assay and alkaline phosphatase activity. On the 21st day, bone markers, osteopontin and osteocalcin, were detected by the PCR analysis. This study shows that the chitosan-gelatin-chondroitin 3D structure provides a good environment for the osteogenic process and enhances cellular proliferation.

3D printing process of oxidized nanocellulose and gelatin scaffold.

PubMed

Xu, Xiaodong; Zhou, Jiping; Jiang, Yani; Zhang, Qi; Shi, Hongcan; Liu, Dongfang

2018-08-01

For tissue engineering applications tissue scaffolds need to have a porous structure to meet the needs of cell proliferation/differentiation, vascularisation and sufficient mechanical strength for the specific tissue. Here we report the results of a study of the 3D printing process for composite materials based on oxidized nanocellulose and gelatin, that was optimised through measuring rheological properties of different batches of materials after different crosslinking times, simulation of the pneumatic extrusion process and 3D scaffolds fabrication with Solidworks Flow Simulation, observation of its porous structure by SEM, measurement of pressure-pull performance, and experiments aimed at finding out the vitro cytotoxicity and cell morphology. The materials printed are highly porous scaffolds with good mechanical properties.

Scaffold-free trachea regeneration by tissue engineering with bio-3D printing.

PubMed

Taniguchi, Daisuke; Matsumoto, Keitaro; Tsuchiya, Tomoshi; Machino, Ryusuke; Takeoka, Yosuke; Elgalad, Abdelmotagaly; Gunge, Kiyofumi; Takagi, Katsunori; Taura, Yasuaki; Hatachi, Go; Matsuo, Naoto; Yamasaki, Naoya; Nakayama, Koichi; Nagayasu, Takeshi

2018-05-01

Currently, most of the artificial airway organs still require scaffolds; however, such scaffolds exhibit several limitations. Alternatively, the use of an autologous artificial trachea without foreign materials and immunosuppressants may solve these issues and constitute a preferred tool. The rationale of this study was to develop a new scaffold-free approach for an artificial trachea using bio-3D printing technology. Here, we assessed the circumferential tracheal replacement using scaffold-free trachea-like grafts generated from isolated cells in an inbred animal model. Chondrocytes and mesenchymal stem cells were isolated from F344 rats. Rat lung microvessel endothelial cells were purchased. Our bio-3D printer generates spheroids consisting of several types of cells to create 3D structures. The bio-3D-printed artificial trachea from spheroids was matured in a bioreactor and transplanted into F344 rats as a tracheal graft under general anaesthesia. The mechanical strength of the artificial trachea was measured, and histological and immunohistochemical examinations were performed. Tracheal transplantation was performed in 9 rats, which were followed up postoperatively for 23 days. The average tensile strength of artificial tracheas before transplantation was 526.3 ± 125.7 mN. The bio-3D-printed scaffold-free artificial trachea had sufficient strength to transplant into the trachea with silicone stents that were used to prevent collapse of the artificial trachea and to support the graft until sufficient blood supply was obtained. Chondrogenesis and vasculogenesis were observed histologically. The scaffold-free isogenic artificial tracheas produced by a bio-3D printer could be utilized as tracheal grafts in rats.

3D Printing of Scaffolds for Tissue Regeneration Applications

PubMed Central

Do, Anh-Vu; Khorsand, Behnoush; Geary, Sean M.; Salem, Aliasger K.

2015-01-01

The current need for organ and tissue replacement, repair and regeneration for patients is continually growing such that supply is not meeting the high demand primarily due to a paucity of donors as well as biocompatibility issues that lead to immune rejection of the transplant. In an effort to overcome these drawbacks, scientists working in the field of tissue engineering and regenerative medicine have investigated the use of scaffolds as an alternative to transplantation. These scaffolds are designed to mimic the extracellular matrix (ECM) by providing structural support as well as promoting attachment, proliferation, and differentiation with the ultimate goal of yielding functional tissues or organs. Initial attempts at developing scaffolds were problematic and subsequently inspired a growing interest in 3D printing as a mode for generating scaffolds. Utilizing three-dimensional printing (3DP) technologies, ECM-like scaffolds can be produced with a high degree of complexity and precision, where fine details can be included at a micron level. In this review, we discuss the criteria for printing viable and functional scaffolds, scaffolding materials, and 3DP technologies used to print scaffolds for tissue engineering. A hybrid approach, employing both natural and synthetic materials, as well as multiple printing processes may be the key to yielding an ECM-like scaffold with high mechanical strength, porosity, interconnectivity, biocompatibility, biodegradability, and high processability. Creating such biofunctional scaffolds could potentially help to meet the demand by patients for tissues and organs without having to wait or rely on donors for transplantation. PMID:26097108

Clinical application of a 3D-printed scaffold in chronic wound treatment: a case series.

PubMed

Sun, Haining; Lv, Huayao; Qiu, Fanghui; Sun, Duolun; Gao, Yue; Chen, Ning; Zheng, YongKe; Deng, Kunxue; Yang, Yaya; Zhang, Haitao; Xu, Tao; Ren, Dongni

2018-05-02

This case series evaluates the safety and effectiveness of 3D-printed scaffold in chronic wounds. The scaffold is a composite of natural and synthetic materials, and can be prepared in the form of powder or membrane. We recruited patients with pressure ulcera (PU) and/or a diabetic foot ulcers (DFU). We used two methods: 3D-printed scaffolds alone, or 3D-printing powder mixed with platelet-rich fibrinogen (PRF). Clinicians and patients were asked to rate the scaffold's ease of application and comfort during use. A total of five patients were recruited; four with a PU and one with a DFU. For the patient treated with the 3D-printed scaffold membrane (n=1), their PU healed in 28 days, and for patients treated with the 3D-printed scaffold powder (n=2), their PUs healed in 54 days. For the patients treated with the 3D-printing powder mixed with PRF (n=2), the patient with a PU healed in 11 days, and the patient with the DFU healed in 14 days. All clinicians rated the 3D-printed scaffold as 'easy' or 'very easy' to use, and patients rated their comfort during wear and at dressing change as 'good' or 'very good'. This study demonstrated that 3D-printed scaffold was convenient to use, have the potential to improve wound healing rates, and provided a safe and effective way for treating chronic wounds.

Validation of an in vitro 3D bone culture model with perfused and mechanically stressed ceramic scaffold.

PubMed

Bouet, G; Cruel, M; Laurent, C; Vico, L; Malaval, L; Marchat, D

2015-05-15

An engineered three dimensional (3D) in vitro cell culture system was designed with the goal of inducing and controlling in vitro osteogenesis in a reproducible manner under conditions more similar to the in vivo bone microenvironment than traditional two-dimensional (2D) models. This bioreactor allows efficient mechanical loading and perfusion of an original cubic calcium phosphate bioceramic of highly controlled composition and structure. This bioceramic comprises an internal portion containing homogeneously interconnected macropores surrounded by a dense layer, which minimises fluid flow bypass around the scaffold. This dense and flat layer permits the application of a homogeneous loading on the bioceramic while also enhancing its mechanical strength. Numerical modelling of constraints shows that the system provides direct mechanical stimulation of cells within the scaffold. Experimental results establish that under perfusion at a steady flow of 2 µL/min, corresponding to 3 ≤ Medium velocity ≤ 23 µm/s, mouse calvarial cells grow and differentiate as osteoblasts in a reproducible manner, and lay down a mineralised matrix. Moreover, cells respond to mechanical loading by increasing C-fos expression, which demonstrates the effective mechanical stimulation of the culture within the scaffold. In summary, we provide a "proof-of-concept" for osteoblastic cell culture in a controlled 3D culture system under perfusion and mechanical loading. This model will be a tool to analyse bone cell functions in vivo, and will provide a bench testing system for the clinical assessment of bioactive bone-targeting molecules under load.

3D silicon doped hydroxyapatite scaffolds decorated with Elastin-like Recombinamers for bone regenerative medicine.

PubMed

Vila, Mercedes; García, Ana; Girotti, Alessandra; Alonso, Matilde; Rodríguez-Cabello, Jose Carlos; González-Vázquez, Arlyng; Planell, Josep A; Engel, Elisabeth; Buján, Julia; García-Honduvilla, Natalio; Vallet-Regí, María

2016-11-01

The current study reports on the manufacturing by rapid prototyping technique of three-dimensional (3D) scaffolds based on silicon substituted hydroxyapatite with Elastin-like Recombinamers (ELRs) functionalized surfaces. Silicon doped hydroxyapatite (Si-HA), with Ca 10 (PO 4 ) 5.7 (SiO 4 ) 0.3 (OH) 1.7 h 0.3 nominal formula, was surface functionalized with two different types of polymers designed by genetic engineering: ELR-RGD that contain cell attachment specific sequences and ELR-SN A 15/RGD with both hydroxyapatite and cells domains that interact with the inorganic phase and with the cells, respectively. These hybrid materials were subjected to in vitro assays in order to clarify if the ELRs coating improved the well-known biocompatible and bone regeneration properties of calcium phosphates materials. The in vitro tests showed that there was a total and homogeneous colonization of the 3D scaffolds by Bone marrow Mesenchymal Stromal Cells (BMSCs). In addition, the BMSCs were viable and able to proliferate and differentiate into osteoblasts. Bone tissue engineering is an area of increasing interest because its main applications are directly related to the rising life expectancy of the population, which promotes higher rates of several bone pathologies, so innovative strategies are needed for bone tissue regeneration therapies. Here we use the rapid prototyping technology to allow moulding ceramic 3D scaffolds and we use different bio-polymers for the functionalization of their surfaces in order to enhance the biological response. Combining the ceramic material (silicon doped hydroxyapatite, Si-HA) and the Elastin like Recombinamers (ELRs) polymers with the presence of the integrin-mediate adhesion domain alone or in combination with SNA15 peptide that possess high affinity for hydroxyapatite, provided an improved Bone marrow Mesenchymal Stromal Cells (BMSCs) differentiation into osteoblastic linkage. Copyright © 2016 Acta Materialia Inc. Published by Elsevier

Development of 3D printed fibrillar collagen scaffold for tissue engineering.

PubMed

Nocera, Aden Díaz; Comín, Romina; Salvatierra, Nancy Alicia; Cid, Mariana Paula

2018-02-27

Collagen is widely used in tissue engineering because it can be extracted in large quantities, and has excellent biocompatibility, good biodegradability, and weak antigenicity. In the present study, we isolated printable collagen from bovine Achilles tendon and examined the purity of the isolated collagen using sodium dodecyl sulfate polyacrylamide gel electrophoresis. The bands obtained corresponded to α 1 , α 2 and β chains with little contamination from other small proteins. Furthermore, rheological measurements of collagen dispersions (60 mg per ml of PBS) at pH 7 revealed values of viscosity of 35.62 ± 1.42 Pa s at shear rate of 10 s - 1 and a shear thinning behavior. Collagen gels and solutions can be used for building scaffolds by three-dimensional (3D) printing. After designing and fabricating a low-cost 3D printer we assayed the collagen printing and obtaining 3D printed scaffolds of collagen at pH 7. The porosity of the scaffold was 90.22% ± 0.88% and the swelling ratio was 1437% ± 146%. The microstructure of the scaffolds was studied using scanning electron microscopy, and a porous mesh of fibrillar collagen was observed. In addition, the 3D printed collagen scaffold was not cytotoxic with cell viability higher than 70% using Vero and NIH 3 T3 cells. In vitro evaluation using both cells lines demonstrated that the collagen scaffolds had the ability to support cell attachment and proliferation. Also a fibrillar collagen mesh was observed after two weeks of culture at 37 °C. Overall, these results are promising since they show the capability of the presented protocol to obtain printable fibrillar collagen at pH 7 and the potential of the printing technique for building low-cost biocompatible 3D plotted structures which maintained the fibrillar collagen structure after incubation in culture media without using additional strategies as crosslinking.

Interwoven Aligned Conductive Nanofiber Yarn/Hydrogel Composite Scaffolds for Engineered 3D Cardiac Anisotropy.

PubMed

Wu, Yaobin; Wang, Ling; Guo, Baolin; Ma, Peter X

2017-06-27

Mimicking the anisotropic cardiac structure and guiding 3D cellular orientation play a critical role in designing scaffolds for cardiac tissue regeneration. Significant advances have been achieved to control cellular alignment and elongation, but it remains an ongoing challenge for engineering 3D cardiac anisotropy using these approaches. Here, we present a 3D hybrid scaffold based on aligned conductive nanofiber yarns network (NFYs-NET, composition: polycaprolactone, silk fibroin, and carbon nanotubes) within a hydrogel shell for mimicking the native cardiac tissue structure, and further demonstrate their great potential for engineering 3D cardiac anisotropy for cardiac tissue engineering. The NFYs-NET structures are shown to control cellular orientation and enhance cardiomyocytes (CMs) maturation. 3D hybrid scaffolds were then fabricated by encapsulating NFYs-NET layers within hydrogel shell, and these 3D scaffolds performed the ability to promote aligned and elongated CMs maturation on each layer and individually control cellular orientation on different layers in a 3D environment. Furthermore, endothelialized myocardium was constructed by using this hybrid strategy via the coculture of CMs on NFYs-NET layer and endothelial cells within hydrogel shell. Therefore, these 3D hybrid scaffolds, containing NFYs-NET layer inducing cellular orientation, maturation, and anisotropy and hydrogel shell providing a suitable 3D environment for endothelialization, has great potential in engineering 3D cardiac anisotropy.

3D-Printed Bioactive Ca3SiO5 Bone Cement Scaffolds with Nano Surface Structure for Bone Regeneration.

PubMed

Yang, Chen; Wang, Xiaoya; Ma, Bing; Zhu, Haibo; Huan, Zhiguang; Ma, Nan; Wu, Chengtie; Chang, Jiang

2017-02-22

Silicate bioactive materials have been widely studied for bone regeneration because of their eminent physicochemical properties and outstanding osteogenic bioactivity, and different methods have been developed to prepare porous silicate bioactive ceramics scaffolds for bone-tissue engineering applications. Among all of these methods, the 3D-printing technique is obviously the most efficient way to control the porous structure. However, 3D-printed bioceramic porous scaffolds need high-temperature sintering, which will cause volume shrinkage and reduce the controllability of the pore structure accuracy. Unlike silicate bioceramic, bioactive silicate cements such as tricalcium silicate (Ca 3 SiO 5 and C 3 S) can be self-set in water to obtain high mechanical strength under mild conditions. Another advantage of using C 3 S to prepare 3D scaffolds is the possibility of simultaneous drug loading. Herein, we, for the first time, demonstrated successful preparation of uniform 3D-printed C 3 S bone cement scaffolds with controllable 3D structure at room temperature. The scaffolds were loaded with two model drugs and showed a loading location controllable drug-release profile. In addition, we developed a surface modification process to create controllable nanotopography on the surface of pore wall of the scaffolds, which showed activity to enhance rat bone-marrow stem cells (rBMSCs) attachment, spreading, and ALP activities. The in vivo experiments revealed that the 3D-printed C 3 S bone cement scaffolds with nanoneedle-structured surfaces significantly improved bone regeneration, as compared to pure C 3 S bone cement scaffolds, suggesting that 3D-printed C 3 S bone cement scaffolds with controllable nanotopography surface are bioactive implantable biomaterials for bone repair.

Multifunctional pH sensitive 3D scaffolds for treatment and prevention of bone infection.

PubMed

Cicuéndez, Mónica; Doadrio, Juan C; Hernández, Ana; Portolés, M Teresa; Izquierdo-Barba, Isabel; Vallet-Regí, María

2018-01-01

Multifunctional-therapeutic three-dimensional (3D) scaffolds have been prepared. These biomaterials are able to destroy the S. aureus bacterial biofilm and to allow bone regeneration at the same time. The present study is focused on the design of pH sensitive 3D hierarchical meso-macroporous 3D scaffolds based on MGHA nanocomposite formed by a mesostructured glassy network with embedded hydroxyapatite nanoparticles, whose mesopores have been loaded with levofloxacin (Levo) as antibacterial agent. These 3D platforms exhibit controlled and pH-dependent Levo release, sustained over time at physiological pH (7.4) and notably increased at infection pH (6.7 and 5.5), which is due to the different interaction rate between diverse Levo species and the silica matrix. These 3D systems are able to inhibit the S. aureus growth and to destroy the bacterial biofilm without cytotoxic effects on human osteoblasts and allowing an adequate colonization and differentiation of preosteoblastic cells on their surface. These findings suggest promising applications of these hierarchical MGHA nanocomposite 3D scaffolds for the treatment and prevention of bone infection. Multifunctional 3D nanocomposite scaffolds with the ability for loading and sustained delivery of an antimicrobial agent, to eliminate and prevent bone infection and at the same time to contribute to bone regeneration process without cytotoxic effects on the surrounding tissue has been proposed. These 3D scaffolds exhibit a sustained levofloxacin delivery at physiological pH (pH 7.4), which increasing notably when pH decreases to characteristic values of bone infection process (pH 6.7 and pH 5.5). In vitro competitive assays between preosteoblastic and bacteria onto the 3D scaffold surface demonstrated an adequate osteoblast colonization in entire scaffold surface together with the ability to eliminate bacteria contamination. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Bioengineered 3D Scaffolds in Cancer Research: Focus on Epithelial to Mesenchymal Transition and Drug Screening.

PubMed

Xu, Xiaoli; Tang, LiLing

2017-01-01

The living environment of cancer cells is complicated and information-rich. Thus, traditional 2D culture mold in vitro cannot mimic the microenvironment of cancer cells exactly. Currently, bioengineered 3D scaffolds have been developed which can better simulate the microenvironment of tumors and fill the gap between 2D culture and clinical application. In this review, we discuss the scaffold materials used for fabrication techniques, biological behaviors of cancer cells in 3D scaffolds and the scaffold-based drug screening. A major emphasis is placed on the description of scaffold-based epithelial to mesenchymal transition and drug screening in 3D culture. By overcoming the defects of traditional 2D culture, 3D scaffolds culture can provide a simpler, safer and more reliable approach for cancer research. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Fabrication and Handling of 3D Scaffolds Based on Polymers and Decellularized Tissues.

PubMed

Shpichka, Anastasia; Koroleva, Anastasia; Kuznetsova, Daria; Dmitriev, Ruslan I; Timashev, Peter

2017-01-01

Polymeric, ceramic and hybrid material-based three-dimensional (3D) scaffold or matrix structures are important for successful tissue engineering. While the number of approaches utilizing the use of cell-based scaffold and matrix structures is constantly growing, it is essential to provide a framework of their typical preparation and evaluation for tissue engineering. This chapter describes the fabrication of 3D scaffolds using two-photon polymerization, decellularization and cell encapsulation methods and easy-to-use protocols allowing assessing the cell morphology, cytotoxicity and viability in these scaffolds.

[Mechanical properties of polylactic acid/beta-tricalcium phosphate composite scaffold with double channels based on three-dimensional printing technique].

PubMed

Lian, Qin; Zhuang, Pei; Li, Changhai; Jin, Zhongmin; Li, Dichen

2014-03-01

To improve the poor mechanical strength of porous ceramic scaffold, an integrated method based on three-dimensional (3-D) printing technique is developed to incorporate the controlled double-channel porous structure into the polylactic acid/beta-tricalcium phosphate (PLA/beta-TCP) reinforced composite scaffolds (double-channel composite scaffold) to improve their tissue regeneration capability and the mechanical properties. The designed double-channel structure inside the ceramic scaffold consisted of both primary and secondary micropipes, which parallel but un-connected. The set of primary channels was used for cell ingrowth, while the set of secondary channels was used for the PLA perfusion. Integration technology of 3-D printing technique and gel-casting was firstly used to fabricate the double-channel ceramic scaffolds. PLA/beta-TCP composite scaffolds were obtained by the polymer gravity perfusion process to pour PLA solution into the double-channel ceramic scaffolds through the secondary channel set. Microscope, porosity, and mechanical experiments for the standard samples were used to evaluate the composite properties. The ceramic scaffold with only the primary channel (single-channel scaffold) was also prepared as a control. Morphology observation results showed that there was no PLA inside the primary channels of the double-channel composite scaffolds but a dense interface layer between PLA and beta-TCP obviously formed on the inner wall of the secondary channels by the PLA penetration during the perfusion process. Finite element simulation found that the compressive strength of the double-channel composite scaffold was less than that of the single-channel scaffold; however, mechanical tests found that the maximum compressive strength of the double-channel composite scaffold [(21.25 +/- 1.15) MPa] was higher than that of the single-channel scaffold[ (9.76 +/- 0.64) MPa]. The double-channel composite scaffolds fabricated by 3-D printing technique have

3D printing of PLGA scaffolds for tissue engineering.

PubMed

Mironov, Anton V; Grigoryev, Aleksey M; Krotova, Larisa I; Skaletsky, Nikolaj N; Popov, Vladimir K; Sevastianov, Viktor I

2017-01-01

We proposed a novel method of generation of bioresorbable polymeric scaffolds with specified architectonics for tissue engineering using extrusion three-dimensional (3D) printing with solutions of polylactoglycolide in tetraglycol with their subsequent solidifying in aqueous medium. On the basis of 3D computer models, we obtained the matrix structures with interconnected system of pores ranging in size from 0.5 to 500 µm. The results of in vitro studies using cultures of line NIH 3Т3 mouse fibroblasts, floating islet cultures of newborn rabbit pancreas, and mesenchymal stem cells of human adipose tissue demonstrated the absence of cytotoxicity and good adhesive properties of scaffolds in regard to the cell cultures chosen. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 104-109, 2017. © 2016 Wiley Periodicals, Inc.

Cryogenic 3D printing for producing hierarchical porous and rhBMP-2-loaded Ca-P/PLLA nanocomposite scaffolds for bone tissue engineering.

PubMed

Wang, Chong; Zhao, Qilong; Wang, Min

2017-06-07

The performance of bone tissue engineering scaffolds can be assessed through cell responses to scaffolds, including cell attachment, infiltration, morphogenesis, proliferation, differentiation, etc, which are determined or heavily influenced by the composition, structure, mechanical properties, and biological properties (e.g. osteoconductivity and osteoinductivity) of scaffolds. Although some promising 3D printing techniques such as fused deposition modeling and selective laser sintering could be employed to produce biodegradable bone tissue engineering scaffolds with customized shapes and tailored interconnected pores, effective methods for fabricating scaffolds with well-designed hierarchical porous structure (both interconnected macropores and surface micropores) and tunable osteoconductivity/osteoinductivity still need to be developed. In this investigation, a novel cryogenic 3D printing technique was investigated and developed for producing hierarchical porous and recombinant human bone morphogenetic protein-2 (rhBMP-2)-loaded calcium phosphate (Ca-P) nanoparticle/poly(L-lactic acid) nanocomposite scaffolds, in which the Ca-P nanoparticle-incorporated scaffold layer and rhBMP-2-encapsulated scaffold layer were deposited alternatingly using different types of emulsions as printing inks. The mechanical properties of the as-printed scaffolds were comparable to those of human cancellous bone. Sustained releases of Ca 2+ ions and rhBMP-2 were achieved and the biological activity of rhBMP-2 was well-preserved. Scaffolds with a desirable hierarchical porous structure and dual delivery of Ca 2+ ions and rhBMP-2 exhibited superior performance in directing the behaviors of human bone marrow-derived mesenchymal stem cells and caused improved cell viability, attachment, proliferation, and osteogenic differentiation, which has suggested their great potential for bone tissue engineering.

Design of 3D scaffolds for tissue engineering testing a tough polylactide-based graft copolymer.

PubMed

Dorati, R; Colonna, C; Tomasi, C; Genta, I; Bruni, G; Conti, B

2014-01-01

The aim of this research was to investigate a tough polymer to develop 3D scaffolds and 2D films for tissue engineering applications, in particular to repair urethral strictures or defects. The polymer tested was a graft copolymer of polylactic acid (PLA) synthesized with the rationale to improve the toughness of the related PLA homopolymer. The LMP-3055 graft copolymer (in bulk) demonstrated to have negligible cytotoxicity (bioavailability >85%, MTT test). Moreover, the LMP-3055 sterilized through gamma rays resulted to be cytocompatible and non-toxic, and it has a positive effect on cell biofunctionality, promoting the cell growth. 3D scaffolds and 2D film were prepared using different LMP-3055 polymer concentrations (7.5, 10, 12.5 and 15%, w/v), and the effect of polymer concentration on pore size, porosity and interconnectivity of the 3D scaffolds and 2D film was investigated. 3D scaffolds got better results for fulfilling structural and biofunctional requirements: porosity, pore size and interconnectivity, cell attachment and proliferation. 3D scaffolds obtained with 10 and 12.5% polymer solutions (3D-2 and 3D-3, respectively) were identified as the most suitable construct for the cell attachment and proliferation presenting pore size ranged between 100 and 400μm, high porosity (77-78%) and well interconnected pores. In vitro cell studies demonstrated that all the selected scaffolds were able to support the cell proliferation, the cell attachment and growth resulting to their dependency on the polymer concentration and structural features. The degradation test revealed that the degradation of polymer matrix (ΔMw) and water uptake of 3D scaffolds exceed those of 2D film and raw polymer (used as control reference), while the mass loss of samples (3D scaffold and 2D film) resulted to be controlled, they showed good stability and capacity to maintain the physical integrity during the incubation time. © 2013.

Immunological compatibility status of placenta-derived stem cells is mediated by scaffold 3D structure.

PubMed

Azizian, Sara; Khatami, Fatemeh; Modaresifar, Khashayar; Mosaffa, Nariman; Peirovi, Habibollah; Tayebi, Lobat; Bahrami, Soheyl; Redl, Heinz; Niknejad, Hassan

2018-02-23

Placenta-derived amniotic epithelial cells (AECs), a great cell source for tissue engineering and stem cell therapy, are immunologically inert in their native state; however, immunological changes in these cells after culture and differentiation have challenged their applications. The aim of this study was to investigate the effect of 2D and 3D scaffolds on human lymphocyte antigens (HLA) expression by AECs. The effect of different preparation parameters including pre-freezing time and temperature was evaluated on 3D chitosan-gelatine scaffolds properties. Evaluation of MHC class I, HLA-DR and HLA-G expression in AECs after 7 d culture on 2D bed and 3D scaffold of chitosan-gelatine showed that culture of AECs on the 2D substrate up-regulated MHC class I and HLA-DR protein markers on AECs surface and down-regulated HLA-G protein. In contrast, 3D scaffold did not increase protein expression of MHC class I and HLA-DR. Moreover, HLA-G protein expression remained unchanged in 3D culture. These results confirm that 3D scaffold can remain AECs in their native immunological state and modification of physical properties of the scaffold is a key regulator of immunological markers at the gene and protein expression levels; a strategy which circumvents rejection challenge of amniotic stem cells to be translated into the clinic.

Tissue-engineered bone with 3-dimensionally printed β-tricalcium phosphate and polycaprolactone scaffolds and early implantation: an in vivo pilot study in a porcine mandible model.

PubMed

Konopnicki, Sandra; Sharaf, Basel; Resnick, Cory; Patenaude, Adam; Pogal-Sussman, Tracy; Hwang, Kyung-Gyun; Abukawa, Harutsugi; Troulis, Maria J

2015-05-01

Deep bone penetration into implanted scaffolds remains a challenge in tissue engineering. The purpose of this study was to evaluate bone penetration depth within 3-dimensionally (3D) printed β-tricalcium phosphate (β-TCP) and polycaprolactone (PCL) scaffolds, seeded with porcine bone marrow progenitor cells (pBMPCs), and implanted early in vivo. Scaffolds were 3D printed with 50% β-TCP and 50% PCL. The pBMPCs were harvested, isolated, expanded, and differentiated into osteoblasts. Cells were seeded into the scaffolds and constructs were incubated in a rotational oxygen-permeable bioreactor system for 14 days. Six 2- × 2-cm defects were created in each mandible (N = 2 minipigs). In total, 6 constructs were placed within defects and 6 defects were used as controls (unseeded scaffolds, n = 3; empty defects, n = 3). Eight weeks after surgery, specimens were harvested and analyzed by hematoxylin and eosin (H&E), 4',6-diamidino-2-phenylindole (DAPI), and CD31 staining. Analysis included cell counts, bone penetration, and angiogenesis at the center of the specimens. All specimens (N = 12) showed bone formation similar to native bone at the periphery. Of 6 constructs, 4 exhibited bone formation in the center. Histomorphometric analysis of the H&E-stained sections showed an average of 22.1% of bone in the center of the constructs group compared with 1.87% in the unseeded scaffolds (P < .05). The 2 remaining constructs, which did not display areas of mature bone in the center, showed massive cell penetration depth by DAPI staining, with an average of 2,109 cells/0.57 mm(2) in the center compared with 1,114 cells/0.57 mm(2) in the controls (P < .05). CD31 expression was greater in the center of the constructs compared with the unseeded scaffolds (P < .05). 3D printed β-TCP and PCL scaffolds seeded with pBMPCs and implanted early into porcine mandibular defects display good bone penetration depth. Further study with a larger sample and larger bone defects should be

Scaffolds with a standardized macro-architecture fabricated from several calcium phosphate ceramics using an indirect rapid prototyping technique

PubMed Central

Wilson, C. E.; van Blitterswijk, C. A.; Verbout, A. J.; de Bruijn, J. D.

2010-01-01

Calcium phosphate ceramics, commonly applied as bone graft substitutes, are a natural choice of scaffolding material for bone tissue engineering. Evidence shows that the chemical composition, macroporosity and microporosity of these ceramics influences their behavior as bone graft substitutes and bone tissue engineering scaffolds but little has been done to optimize these parameters. One method of optimization is to place focus on a particular parameter by normalizing the influence, as much as possible, of confounding parameters. This is difficult to accomplish with traditional fabrication techniques. In this study we describe a design based rapid prototyping method of manufacturing scaffolds with virtually identical macroporous architectures from different calcium phosphate ceramic compositions. Beta-tricalcium phosphate, hydroxyapatite (at two sintering temperatures) and biphasic calcium phosphate scaffolds were manufactured. The macro- and micro-architectures of the scaffolds were characterized as well as the influence of the manufacturing method on the chemistries of the calcium phosphate compositions. The structural characteristics of the resulting scaffolds were remarkably similar. The manufacturing process had little influence on the composition of the materials except for the consistent but small addition of, or increase in, a beta-tricalcium phosphate phase. Among other applications, scaffolds produced by the method described provide a means of examining the influence of different calcium phosphate compositions while confidently excluding the influence of the macroporous structure of the scaffolds. PMID:21069558

Modelling glioma invasion using 3D bioprinting and scaffold-free 3D culture.

PubMed

van Pel, Derek M; Harada, Kaori; Song, Dandan; Naus, Christian C; Sin, Wun Chey

2018-06-16

Glioma is a highly aggressive form of brain cancer, with some subtypes having 5-year survival rates of less than 5%. Tumour cell invasion into the surrounding parenchyma seems to be the primary driver of these poor outcomes, as most gliomas recur within 2 cm of the original surgically-resected tumour. Many current approaches to the development of anticancer therapy attempt to target genetic weaknesses in a particular cancer, but may not take into account the microenvironment experienced by a tumour and the patient-specific genetic differences in susceptibility to treatment. Here we demonstrate the use of complementary approaches, 3D bioprinting and scaffold-free 3D tissue culture, to examine the invasion of glioma cells into neural-like tissue with 3D confocal microscopy. We found that, while both approaches were successful, the use of 3D tissue culture for organoid development offers the advantage of broad accessibility. As a proof-of-concept of our approach, we developed a system in which we could model the invasion of human glioma cells into mouse neural progenitor cell-derived spheroids. We show that we can follow invasion of human tumour cells using cell-tracking dyes and 3D laser scanning confocal microscopy, both in real time and in fixed samples. We validated these results using conventional cryosectioning. Our scaffold-free 3D approach has broad applicability, as we were easily able to examine invasion using different neural progenitor cell lines, thus mimicking differences that might be observed in patient brain tissue. These results, once applied to iPSC-derived cerebral organoids that incorporate the somatic genetic variability of patients, offer the promise of truly personalized treatments for brain cancer.

Nanostructured thick 3D nanofibrous scaffold can induce bone.

PubMed

Eap, Sandy; Morand, David; Clauss, François; Huck, Olivier; Stoltz, Jean-François; Lutz, Jean-Christophe; Gottenberg, Jacques-Eric; Benkirane-Jessel, Nadia; Keller, Laetitia; Fioretti, Florence

2015-01-01

Designing unique nanostructured biomimetic materials is a new challenge in modern regenerative medicine. In order to develop functional substitutes for damaged organs or tissues, several methods have been used to create implants able to regenerate robust and durable bone. Electrospinning produces nonwoven scaffolds based on polymer nanofibers mimicking the fibrillar organization of bone extracellular matrix. Here, we describe a biomimetic 3D thick nanofibrous scaffold obtained by electrospinning of the biodegradable, bioresorbable and FDA-approved polymer, poly(ε-caprolactone). Such scaffold presents a thickness reaching one centimeter. We report here the demonstration that the designed nanostructured implant is able to induce in vivo bone regeneration.

Coating electrospun poly(epsilon-caprolactone) fibers with gelatin and calcium phosphate and their use as biomimetic scaffolds for bone tissue engineering.

PubMed

Li, Xiaoran; Xie, Jingwei; Yuan, Xiaoyan; Xia, Younan

2008-12-16

Electrospinning was employed to fabricate fibrous scaffolds of poly(epsilon-caprolactone) in the form of nonwoven mats. The surfaces of the fibers were then coated with gelatin through layer-by-layer self-assembly, followed by functionalization with a uniform coating of bonelike calcium phosphate by mineralization in the 10 times concentrated simulated body fluid for 2 h. Transmission electron microscopy, water contact angle, and scanning electron microscopy measurements confirmed the presence of gelatin and calcium phosphate coating layers, and X-ray diffraction results suggested that the deposited mineral phase was a mixture of dicalcium phosphate dehydrate (a precursor to apatite) and apatite. It was also demonstrated that the incorporation of gelatin promoted nucleation and growth of calcium phosphate. The porous scaffolds could mimic the structure, composition, and biological function of bone extracellular matrix. It was found that the preosteoblastic MC3T3-E1 cells attached, spread, and proliferated well with a flat morphology on the mineralized scaffolds. The proliferation rate of the cells on the mineralized scaffolds was significantly higher (by 1.9-fold) than that on the pristine fibrous scaffolds after culture for 7 days. These results indicated that the hybrid system containing poly(epsilon-caprolactone), gelatin, and calcium phosphate could serve as a new class of biomimetic scaffolds for bone tissue engineering.

3D printing nano conductive multi-walled carbon nanotube scaffolds for nerve regeneration

NASA Astrophysics Data System (ADS)

Lee, Se-Jun; Zhu, Wei; Nowicki, Margaret; Lee, Grace; Nyoung Heo, Dong; Kim, Junghoon; Zuo, Yi Y.; Zhang, Lijie Grace

2018-02-01

Objective. Nanomaterials, such as carbon nanotubes (CNTs), have been introduced to modify the surface properties of scaffolds, thus enhancing the interaction between the neural cells and biomaterials. In addition to superior electrical conductivity, CNTs can provide nanoscale structures similar to those present in the natural neural environment. The primary objective of this study is to investigate the proliferative capability and differential potential of neural stem cells (NSCs) seeded on a CNT incorporated scaffold. Approach. Amine functionalized multi-walled carbon nanotubes (MWCNTs) were incorporated with a PEGDA polymer to provide enhanced electrical properties as well as nanofeatures on the surface of the scaffold. A stereolithography 3D printer was employed to fabricate a well-dispersed MWCNT-hydrogel composite neural scaffold with a tunable porous structure. 3D printing allows easy fabrication of complex 3D scaffolds with extremely intricate microarchitectures and controlled porosity. Main results. Our results showed that MWCNT-incorporated scaffolds promoted neural stem cell proliferation and early neuronal differentiation when compared to those scaffolds without the MWCNTs. Furthermore, biphasic pulse stimulation with 500 µA current promoted neuronal maturity quantified through protein expression analysis by quantitative polymerase chain reaction. Significance. Results of this study demonstrated that an electroconductive MWCNT scaffold, coupled with electrical stimulation, may have a synergistic effect on promoting neurite outgrowth for therapeutic application in nerve regeneration.

Functionalization of 3D scaffolds with protein-releasing biomaterials for intracellular delivery.

PubMed

Seras-Franzoso, Joaquin; Steurer, Christoph; Roldán, Mònica; Vendrell, Meritxell; Vidaurre-Agut, Carla; Tarruella, Anna; Saldaña, Laura; Vilaboa, Nuria; Parera, Marc; Elizondo, Elisa; Ratera, Imma; Ventosa, Nora; Veciana, Jaume; Campillo-Fernández, Alberto J; García-Fruitós, Elena; Vázquez, Esther; Villaverde, Antonio

2013-10-10

Appropriate combinations of mechanical and biological stimuli are required to promote proper colonization of substrate materials in regenerative medicine. In this context, 3D scaffolds formed by compatible and biodegradable materials are under continuous development in an attempt to mimic the extracellular environment of mammalian cells. We have here explored how novel 3D porous scaffolds constructed by polylactic acid, polycaprolactone or chitosan can be decorated with bacterial inclusion bodies, submicron protein particles formed by releasable functional proteins. A simple dipping-based decoration method tested here specifically favors the penetration of the functional particles deeper than 300μm from the materials' surface. The functionalized surfaces support the intracellular delivery of biologically active proteins to up to more than 80% of the colonizing cells, a process that is slightly influenced by the chemical nature of the scaffold. The combination of 3D soft scaffolds and protein-based sustained release systems (Bioscaffolds) offers promise in the fabrication of bio-inspired hybrid matrices for multifactorial control of cell proliferation in tissue engineering under complex architectonic setting-ups. © 2013.

3D printing of novel osteochondral scaffolds with graded microstructure

NASA Astrophysics Data System (ADS)

Nowicki, Margaret A.; Castro, Nathan J.; Plesniak, Michael W.; Zhang, Lijie Grace

2016-10-01

Osteochondral tissue has a complex graded structure where biological, physiological, and mechanical properties vary significantly over the full thickness spanning from the subchondral bone region beneath the joint surface to the hyaline cartilage region at the joint surface. This presents a significant challenge for tissue-engineered structures addressing osteochondral defects. Fused deposition modeling (FDM) 3D bioprinters present a unique solution to this problem. The objective of this study is to use FDM-based 3D bioprinting and nanocrystalline hydroxyapatite for improved bone marrow human mesenchymal stem cell (hMSC) adhesion, growth, and osteochondral differentiation. FDM printing parameters can be tuned through computer aided design and computer numerical control software to manipulate scaffold geometries in ways that are beneficial to mechanical performance without hindering cellular behavior. Additionally, the ability to fine-tune 3D printed scaffolds increases further through our investment casting procedure which facilitates the inclusion of nanoparticles with biochemical factors to further elicit desired hMSC differentiation. For this study, FDM was used to print investment-casting molds innovatively designed with varied pore distribution over the full thickness of the scaffold. The mechanical and biological impacts of the varied pore distributions were compared and evaluated to determine the benefits of this physical manipulation. The results indicate that both mechanical properties and cell performance improve in the graded pore structures when compared to homogeneously distributed porous and non-porous structures. Differentiation results indicated successful osteogenic and chondrogenic manipulation in engineered scaffolds.

Periodontal tissue engineering by nano beta-tricalcium phosphate scaffold and fibroblast growth factor-2 in one-wall infrabony defects of dogs.

PubMed

Ogawa, K; Miyaji, H; Kato, A; Kosen, Y; Momose, T; Yoshida, T; Nishida, E; Miyata, S; Murakami, S; Takita, H; Fugetsu, B; Sugaya, T; Kawanami, M

2016-12-01

Nanoparticle bioceramics are being investigated for biomedical applications. We fabricated a regenerative scaffold comprising type I collagen and beta-tricalcium phosphate (β-TCP) nanoparticles. Fibroblast growth factor-2 (FGF-2) is a bioeffective signaling molecule that stimulates cell proliferation and wound healing. This study examined the effects, on bioactivity, of a nano-β-TCP/collagen scaffold loaded with FGF-2, particularly on periodontal tissue wound healing. Beta-tricalcium phosphate was pulverized into nanosize particles (84 nm) and was then dispersed. A nano-β-TCP scaffold was prepared by coating the surface of a collagen scaffold with a nanosize β-TCP dispersion. Scaffolds were characterized using scanning electron microscopy, compressive testing, cell seeding and rat subcutaneous implant testing. Then, nano-β-TCP scaffold, nano-β-TCP scaffold loaded with FGF-2 and noncoated collagen scaffold were implanted into a dog one-wall infrabony defect model. Histological observations were made at 10 d and 4 wk postsurgery. Scanning electron microscopy images show that TCP nanoparticles were attached to collagen fibers. The nano-β-TCP scaffold showed higher compressive strength and cytocompatibility compared with the noncoated collagen scaffold. Rat subcutaneous implant tests showed that the DNA contents of infiltrating cells in the nano-β-TCP scaffold and the FGF-2-loaded scaffold were approximately 2.8-fold and 3.7-fold greater, respectively, than in the collagen scaffold. Histological samples from the periodontal defect model showed about five-fold greater periodontal tissue repair following implantation of the nano-β-TCP scaffold loaded with FGF-2 compared with the collagen scaffold. The β-TCP nanoparticle coating strongly improved the collagen scaffold bioactivity. Nano-β-TCP scaffolds containing FGF-2 are anticipated for use in periodontal tissue engineering. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Bioengineered silk scaffolds in 3D tissue modeling with focus on mammary tissues.

PubMed

Maghdouri-White, Yas; Bowlin, Gary L; Lemmon, Christopher A; Dréau, Didier

2016-02-01

In vitro generation of three-dimensional (3D) biological tissues and organ-like structures is a promising strategy to study and closely model complex aspects of the molecular, cellular, and physiological interactions of tissue. In particular, in vitro 3D tissue modeling holds promises to further our understanding of breast development. Indeed, biologically relevant 3D structures that combine mammary cells and engineered matrices have improved our knowledge of mammary tissue growth, organization, and differentiation. Several polymeric biomaterials have been used as scaffolds to engineer 3D mammary tissues. Among those, silk fibroin-based biomaterials have many biologically relevant properties and have been successfully used in multiple medical applications. Here, we review the recent advances in engineered scaffolds with an emphasis on breast-like tissue generation and the benefits of modified silk-based scaffolds. Copyright © 2015 Elsevier B.V. All rights reserved.

3D-Printing Composite Polycaprolactone-Decellularized Bone Matrix Scaffolds for Bone Tissue Engineering Applications.

PubMed

Rindone, Alexandra N; Nyberg, Ethan; Grayson, Warren L

2017-05-11

Millions of patients worldwide require bone grafts for treatment of large, critically sized bone defects from conditions such as trauma, cancer, and congenital defects. Tissue engineered (TE) bone grafts have the potential to provide a more effective treatment than current bone grafts since they would restore fully functional bone tissue in large defects. Most bone TE approaches involve a combination of stem cells with porous, biodegradable scaffolds that provide mechanical support and degrade gradually as bone tissue is regenerated by stem cells. 3D-printing is a key technique in bone TE that can be used to fabricate functionalized scaffolds with patient-specific geometry. Using 3D-printing, composite polycaprolactone (PCL) and decellularized bone matrix (DCB) scaffolds can be produced to have the desired mechanical properties, geometry, and osteoinductivity needed for a TE bone graft. This book chapter will describe the protocols for fabricating and characterizing 3D-printed PCL:DCB scaffolds. Moreover, procedures for culturing adipose-derived stem cells (ASCs) in these scaffolds in vitro will be described to demonstrate the osteoinductivity of the scaffolds.

Parameters optimization for the fabrication of phosphate glass/hydroxyapatite nanocomposite scaffold

NASA Astrophysics Data System (ADS)

Govindan, R.; Girija, E. K.

2015-06-01

Three-dimensional, highly porous, bioactive and biodegradable phosphate glass and nanohydroxyapatite (n-HA) composite scaffolds was fabricated by the polymer foam replication technique. Polyurethane foam (PU) and polyvinyl alcohol (PVA) were used as template and binder, respectively. Optimization of composition and sintering temperature is carried out for tissue engineering scaffold fabrication.

Alendronate-Eluting Biphasic Calcium Phosphate (BCP) Scaffolds Stimulate Osteogenic Differentiation

PubMed Central

Kim, Sung Eun; Lee, Deok-Won; Kang, Eun Young; Jeong, Won Jae; Lee, Boram; Jeong, Myeong Seon; Kim, Hak Jun; Park, Kyeongsoon; Song, Hae-Ryong

2015-01-01

Biphasic calcium phosphate (BCP) scaffolds have been widely used in orthopedic and dental fields as osteoconductive bone substitutes. However, BCP scaffolds are not satisfactory for the stimulation of osteogenic differentiation and maturation. To enhance osteogenic differentiation, we prepared alendronate- (ALN-) eluting BCP scaffolds. The coating of ALN on BCP scaffolds was confirmed by scanning electron microscopy (FE-SEM), energy-dispersive X-ray spectroscopy (EDS), and attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR). An in vitro release study showed that release of ALN from ALN-eluting BCP scaffolds was sustained for up to 28 days. In vitro results revealed that MG-63 cells grown on ALN-eluting BCP scaffolds exhibited increased ALP activity and calcium deposition and upregulated gene expression of Runx2, ALP, OCN, and OPN compared with the BCP scaffold alone. Therefore, this study suggests that ALN-eluting BCP scaffolds have the potential to effectively stimulate osteogenic differentiation. PMID:26221587

SiO2 and ZnO Dopants in 3D Printed TCP Scaffolds Enhances Osteogenesis and Angiogenesis in vivo

PubMed Central

Fielding, Gary; Bose, Susmita

2013-01-01

Calcium phosphate (CaP) scaffolds with three dimensionally (3D) interconnected pores play an important role in mechanical interlocking and biological fixation in bone implant applications. CaPs alone, however, are only osteoconductive (ability to guide bone growth). Much attention has been given to the incorporation of biologics and pharmacologics to add osteoinductive (ability to cause new bone growth) properties to CaP materials. Because biologics and pharmacologics are generally delicate compounds and also subject to increased regulatory scrutiny, there is a need to investigate alternative methods to introduce osteoinductivity to CaP materials. In this study silica (SiO2) and zinc oxide (ZnO) have been incorporated in to 3D printed β-tricalcium phosphate (TCP) scaffolds to investigate their potential to trigger osteoinduction in vivo. Silicon and zinc are trace elements that are common to bone and have also been shown to have many beneficial properties from increased bone regeneration to angiogenesis. Implants were placed in bicortical femur defects introduced to a murine model for up to 16 weeks. Addition of dopants into TCP increased the capacity for new early bone formation by modulating collagen I production and osteocalcin production. Neovascularization was found to be up to three times more than the pure TCP control group. The findings from this study indicate that the combination of SiO2 and ZnO dopants in TCP may be a viable alternative to introduce osteoinductive properties to CaPs. PMID:23871941

Calcium phosphate coated Keratin-PCL scaffolds for potential bone tissue regeneration.

PubMed

Zhao, Xinxin; Lui, Yuan Siang; Choo, Caleb Kai Chuen; Sow, Wan Ting; Huang, Charlotte Liwen; Ng, Kee Woei; Tan, Lay Poh; Loo, Joachim Say Chye

2015-04-01

The incorporation of hydroxyapatite (HA) nanoparticles within or on the surface of electrospun polymeric scaffolds is a popular approach for bone tissue engineering. However, the fabrication of osteoconductive composite scaffolds via benign processing conditions still remains a major challenge to date. In this work, a new method was developed to achieve a uniform coating of calcium phosphate (CaP) onto electrospun keratin-polycaprolactone composites (Keratin-PCL). Keratin within PCL was crosslinked to decrease its solubility, before coating of CaP. A homogeneous coating was achieved within a short time frame (~10min) by immersing the scaffolds into Ca(2+) and (PO4)(3-) solutions separately. Results showed that the incorporation of keratin into PCL scaffolds not only provided nucleation sites for Ca(2+) adsorption and subsequent homogeneous CaP surface deposition, but also facilitated cell-matrix interactions. An improvement in the mechanical strength of the resultant composite scaffold, as compared to other conventional coating methods, was also observed. This approach of developing a biocompatible bone tissue engineering scaffold would be adopted for further in vitro osteogenic differentiation studies in the future. Copyright © 2015 Elsevier B.V. All rights reserved.

Biomimetic Graphene-Based 3D Scaffold for Long-Term Cell Culture and Real-Time Electrochemical Monitoring.

PubMed

Hu, Xue-Bo; Liu, Yan-Ling; Wang, Wen-Jie; Zhang, Hai-Wei; Qin, Yu; Guo, Shan; Zhang, Xin-Wei; Fu, Lei; Huang, Wei-Hua

2018-01-16

Current achievements on electrochemical monitoring of cells are often gained on two-dimensional (2D) substrates, which fail in mimicking the cellular environments and accurately reproducing the cellular functions within a three-dimensional (3D) tissue. In this regard, 3D scaffold concurrently integrated with the function of cell culture and electrochemical sensing is conceivably a promising platform to monitor cells in real time under their in vivo-like 3D microenvironments. However, it is particularly challenging to construct such a multifunctional scaffold platform. Herein, we developed a 3-aminophenylboronic acid (APBA) functionalized graphene foam (GF) network, which combines the biomimetic property of APBA with the mechanical and electrochemical properties of GF. Hence, the GF network can serve as a 3D scaffold to culture cells for a long period with high viability and simultaneously as an electrode for highly sensitive electrochemical sensing. This allows monitoring of gaseous messengers H 2 S released from the cells cultured on the 3D scaffold in real time. This work represents considerable progress in fabricating 3D cell culture scaffold with electrochemical properties, thereby facilitating future studies of physiologically relevant processes.

3D Printed Polycaprolactone Carbon Nanotube Composite Scaffolds for Cardiac Tissue Engineering.

PubMed

Ho, Chee Meng Benjamin; Mishra, Abhinay; Lin, Pearlyn Teo Pei; Ng, Sum Huan; Yeong, Wai Yee; Kim, Young-Jin; Yoon, Yong-Jin

2017-04-01

Fabrication of tissue engineering scaffolds with the use of novel 3D printing has gained lot of attention, however systematic investigation of biomaterials for 3D printing have not been widely explored. In this report, well-defined structures of polycaprolactone (PCL) and PCL- carbon nanotube (PCL-CNT) composite scaffolds have been designed and fabricated using a 3D printer. Conditions for 3D printing has been optimized while the effects of varying CNT percentages with PCL matrix on the thermal, mechanical and biological properties of the printed scaffolds are studied. Raman spectroscopy is used to characterise the functionalized CNTs and its interactions with PCL matrix. Mechanical properties of the composites are characterised using nanoindentation. Maximum peak load, elastic modulus and hardness increases with increasing CNT content. Differential scanning calorimetry (DSC) studies reveal the thermal and crystalline behaviour of PCL and its CNT composites. Biodegradation studies are performed in Pseudomonas Lipase enzymatic media, showing its specificity and effect on degradation rate. Cell imaging and viability studies of H9c2 cells from rat origin on the scaffolds are performed using fluorescence imaging and MTT assay, respectively. PCL and its CNT composites are able to show cell proliferation and have the potential to be used in cardiac tissue engineering. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Influence of crosslinking on the mechanical behavior of 3D printed alginate scaffolds: Experimental and numerical approaches.

PubMed

Naghieh, Saman; Karamooz-Ravari, Mohammad Reza; Sarker, M D; Karki, Eva; Chen, Xiongbiao

2018-04-01

Tissue scaffolds fabricated by three-dimensional (3D) bioprinting are attracting considerable attention for tissue engineering applications. Because the mechanical properties of hydrogel scaffolds should match the damaged tissue, changing various parameters during 3D bioprinting has been studied to manipulate the mechanical behavior of the resulting scaffolds. Crosslinking scaffolds using a cation solution (such as CaCl 2 ) is also important for regulating the mechanical properties, but has not been well documented in the literature. Here, the effect of varied crosslinking agent volume and crosslinking time on the mechanical behavior of 3D bioplotted alginate scaffolds was evaluated using both experimental and numerical methods. Compression tests were used to measure the elastic modulus of each scaffold, then a finite element model was developed and a power model used to predict scaffold mechanical behavior. Results showed that crosslinking time and volume of crosslinker both play a decisive role in modulating the mechanical properties of 3D bioplotted scaffolds. Because mechanical properties of scaffolds can affect cell response, the findings of this study can be implemented to modulate the elastic modulus of scaffolds according to the intended application. Copyright © 2018 Elsevier Ltd. All rights reserved.

3D-printed scaffolds based on PLA/HA nanocomposites for trabecular bone reconstruction

NASA Astrophysics Data System (ADS)

Niaza, K. V.; Senatov, F. S.; Kaloshkin, S. D.; Maksimkin, A. V.; Chukov, D. I.

2016-08-01

In the present work porous PLA scaffolds filled with micro- and nano- HA were studied. Both composites with micro- and nano-HA were obtained by extrusion in the same conditions. Scaffolds were obtained by 3D-printing by fused filament fabrication method. Structure of porous scaffolds was pre-modeled by computer software. Compression and three - point flexural tests were used to study mechanical properties of the scaffolds.

Effect of 3D-scaffold formation on differentiation and survival in human neural progenitor cells.

PubMed

Ortinau, Stefanie; Schmich, Jürgen; Block, Stephan; Liedmann, Andrea; Jonas, Ludwig; Weiss, Dieter G; Helm, Christiane A; Rolfs, Arndt; Frech, Moritz J

2010-11-11

3D-scaffolds have been shown to direct cell growth and differentiation in many different cell types, with the formation and functionalisation of the 3D-microenviroment being important in determining the fate of the embedded cells. Here we used a hydrogel-based scaffold to investigate the influences of matrix concentration and functionalisation with laminin on the formation of the scaffolds, and the effect of these scaffolds on human neural progenitor cells cultured within them. In this study we used different concentrations of the hydrogel-based matrix PuraMatrix. In some experiments we functionalised the matrix with laminin I. The impact of concentration and treatment with laminin on the formation of the scaffold was examined with atomic force microscopy. Cells from a human fetal neural progenitor cell line were cultured in the different matrices, as well as in a 2D culture system, and were subsequently analysed with antibody stainings against neuronal markers. In parallel, the survival rate of the cells was determined by a live/dead assay. Atomic force microscopy measurements demonstrated that the matrices are formed by networks of isolated PuraMatrix fibres and aggregates of fibres. An increase of the hydrogel concentration led to a decrease in the mesh size of the scaffolds and functionalisation with laminin promoted aggregation of the fibres (bundle formation), which further reduces the density of isolated fibres. We showed that laminin-functionalisation is essential for human neural progenitor cells to build up 3D-growth patterns, and that proliferation of the cells is also affected by the concentration of matrix. In addition we found that 3D-cultures enhanced neuronal differentiation and the survival rate of the cells compared to 2D-cultures. Taken together, we have demonstrated a direct influence of the 3D-scaffold formation on the survival and neuronal differentiation of human neural progenitor cells. These findings emphasize the importance of optimizing 3

3D-Printed ABS and PLA Scaffolds for Cartilage and Nucleus Pulposus Tissue Regeneration.

PubMed

Rosenzweig, Derek H; Carelli, Eric; Steffen, Thomas; Jarzem, Peter; Haglund, Lisbet

2015-07-03

Painful degeneration of soft tissues accounts for high socioeconomic costs. Tissue engineering aims to provide biomimetics recapitulating native tissues. Biocompatible thermoplastics for 3D printing can generate high-resolution structures resembling tissue extracellular matrix. Large-pore 3D-printed acrylonitrile butadiene styrene (ABS) and polylactic acid (PLA) scaffolds were compared for cell ingrowth, viability, and tissue generation. Primary articular chondrocytes and nucleus pulposus (NP) cells were cultured on ABS and PLA scaffolds for three weeks. Both cell types proliferated well, showed high viability, and produced ample amounts of proteoglycan and collagen type II on both scaffolds. NP generated more matrix than chondrocytes; however, no difference was observed between scaffold types. Mechanical testing revealed sustained scaffold stability. This study demonstrates that chondrocytes and NP cells can proliferate on both ABS and PLA scaffolds printed with a simplistic, inexpensive desktop 3D printer. Moreover, NP cells produced more proteoglycan than chondrocytes, irrespective of thermoplastic type, indicating that cells maintain individual phenotype over the three-week culture period. Future scaffold designs covering larger pore sizes and better mimicking native tissue structure combined with more flexible or resorbable materials may provide implantable constructs with the proper structure, function, and cellularity necessary for potential cartilage and disc tissue repair in vivo.

Microstereolithography-Based Fabrication of Anatomically Shaped Beta-Tricalcium Phosphate Scaffolds for Bone Tissue Engineering

PubMed Central

Du, Dajiang; Asaoka, Teruo; Shinohara, Makoto; Kageyama, Tomonori; Ushida, Takashi; Furukawa, Katsuko Sakai

2015-01-01

Porous ceramic scaffolds with shapes matching the bone defects may result in more efficient grafting and healing than the ones with simple geometries. Using computer-assisted microstereolithography (MSTL), we have developed a novel gelcasting indirect MSTL technology and successfully fabricated two scaffolds according to CT images of rabbit femur. Negative resin molds with outer 3D dimensions conforming to the femur and an internal structure consisting of stacked meshes with uniform interconnecting struts, 0.5 mm in diameter, were fabricated by MSTL. The second mold type was designed for cortical bone formation. A ceramic slurry of beta-tricalcium phosphate (β-TCP) with room temperature vulcanization (RTV) silicone as binder was cast into the molds. After the RTV silicone was completely cured, the composite was sintered at 1500°C for 5 h. Both gross anatomical shape and the interpenetrating internal network were preserved after sintering. Even cortical structure could be introduced into the customized scaffolds, which resulted in enhanced strength. Biocompatibility was confirmed by vital staining of rabbit bone marrow mesenchymal stromal cells cultured on the customized scaffolds for 5 days. This fabrication method could be useful for constructing bone substitutes specifically designed according to local anatomical defects. PMID:26504839

Biomechanical properties of 3D-printed bone scaffolds are improved by treatment with CRFP.

PubMed

Helguero, Carlos G; Mustahsan, Vamiq M; Parmar, Sunjit; Pentyala, Sahana; Pfail, John L; Kao, Imin; Komatsu, David E; Pentyala, Srinivas

2017-12-22

One of the major challenges in orthopedics is to develop implants that overcome current postoperative problems such as osteointegration, proper load bearing, and stress shielding. Current implant techniques such as allografts or endoprostheses never reach full bone integration, and the risk of fracture due to stress shielding is a major concern. To overcome this, a novel technique of reverse engineering to create artificial scaffolds was designed and tested. The purpose of the study is to create a new generation of implants that are both biocompatible and biomimetic. 3D-printed scaffolds based on physiological trabecular bone patterning were printed. MC3T3 cells were cultured on these scaffolds in osteogenic media, with and without the addition of Calcitonin Receptor Fragment Peptide (CRFP) in order to assess bone formation on the surfaces of the scaffolds. Integrity of these cell-seeded bone-coated scaffolds was tested for their mechanical strength. The results show that cellular proliferation and bone matrix formation are both supported by our 3D-printed scaffolds. The mechanical strength of the scaffolds was enhanced by trabecular patterning in the order of 20% for compression strength and 60% for compressive modulus. Furthermore, cell-seeded trabecular scaffolds modulus increased fourfold when treated with CRFP. Upon mineralization, the cell-seeded trabecular implants treated with osteo-inductive agents and pretreated with CRFP showed a significant increase in the compressive modulus. This work will lead to creating 3D structures that can be used in the replacement of not only bone segments, but entire bones.

Enhancement of neurite outgrowth in neuron cancer stem cells by growth on 3-D collagen scaffolds

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Chih-Hao; Neurosurgery, Department of Surgery, Kaohsiung Veterans General Hospital, Taiwan, ROC; Department of Biomedical Engineering, I-Shou University, Taiwan, ROC

Highlights: Black-Right-Pointing-Pointer Neuron cancer stem cells (NCSCs) behave high multiply of growth on collagen scaffold. Black-Right-Pointing-Pointer Enhancement of NCSCs neurite outgrowth on porous collagen scaffold. Black-Right-Pointing-Pointer 3-D collagen culture of NCSCs shows an advance differentiation than 2-D culture. -- Abstract: Collagen is one component of the extracellular matrix that has been widely used for constructive remodeling to facilitate cell growth and differentiation. The 3-D distribution and growth of cells within the porous scaffold suggest a clinical significance for nerve tissue engineering. In the current study, we investigated proliferation and differentiation of neuron cancer stem cells (NCSCs) on a 3-D porousmore » collagen scaffold that mimics the natural extracellular matrix. We first generated green fluorescence protein (GFP) expressing NCSCs using a lentiviral system to instantly monitor the transitions of morphological changes during growth on the 3-D scaffold. We found that proliferation of GFP-NCSCs increased, and a single cell mass rapidly grew with unrestricted expansion between days 3 and 9 in culture. Moreover, immunostaining with neuronal nuclei (NeuN) revealed that NCSCs grown on the 3-D collagen scaffold significantly enhanced neurite outgrowth. Our findings confirmed that the 80 {mu}m porous collagen scaffold could enhance attachment, viability and differentiation of the cancer neural stem cells. This result could provide a new application for nerve tissue engineering and nerve regeneration.« less

3D X-Ray Nanotomography of Cells Grown on Electrospun Scaffolds.

PubMed

Bradley, Robert S; Robinson, Ian K; Yusuf, Mohammed

2017-02-01

Here, it is demonstrated that X-ray nanotomography with Zernike phase contrast can be used for 3D imaging of cells grown on electrospun polymer scaffolds. The scaffold fibers and cells are simultaneously imaged, enabling the influence of scaffold architecture on cell location and morphology to be studied. The high resolution enables subcellular details to be revealed. The X-ray imaging conditions were optimized to reduce scan times, making it feasible to scan multiple regions of interest in relatively large samples. An image processing procedure is presented which enables scaffold characteristics and cell location to be quantified. The procedure is demonstrated by comparing the ingrowth of cells after culture for 3 and 6 days. © 2016 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Design and Characterization of Calcium Phosphate Ceramic Scaffolds for Bone Tissue Engineering

PubMed Central

Kuhn, Liisa T.

2015-01-01

Objectives Our goal is to review design strategies for the fabrication of calcium phosphate ceramic scaffolds (CPS), in light of their transient role in bone tissue engineering and associated requirements for effective bone regeneration. Methods We examine the various design options available to meet mechanical and biological requirements of CPS and later focus on the importance of proper characterization of CPS in terms of architecture, mechanical properties and time-sensitive properties such as biodegradability. Finally, relationships between in vitro vs. in vivo testing are addressed, with an attempt to highlight reliable performance predictors. Results A combinatory design strategy should be used with CPS taking into consideration 3D architecture, adequate surface chemistry and topography, all of which are needed to promote bone formation. CPS represent the media of choice for delivery of osteogenic factors and anti-infectives. Non-osteoblast mediated mineral deposition can confound in vitro osteogenesis testing of CPS and therefore the expression of a variety of proteins or genes including collagen type I, bone sialoprotein and osteocalcin should be confirmed in addition to increased mineral content. Conclusions CPS are a superior scaffold material for bone regeneration because they actively promote osteogenesis. Biodegradability of CPS via calcium and phosphate release represents a unique asset. Structural control of CPS at the macro, micro and nanoscale and their combination with cells and polymeric materials is likely to lead to significant developments in bone tissue engineering. PMID:26423007

Design and characterization of calcium phosphate ceramic scaffolds for bone tissue engineering.

PubMed

Denry, Isabelle; Kuhn, Liisa T

2016-01-01

Our goal is to review design strategies for the fabrication of calcium phosphate ceramic scaffolds (CPS), in light of their transient role in bone tissue engineering and associated requirements for effective bone regeneration. We examine the various design options available to meet mechanical and biological requirements of CPS and later focus on the importance of proper characterization of CPS in terms of architecture, mechanical properties and time-sensitive properties such as biodegradability. Finally, relationships between in vitro versus in vivo testing are addressed, with an attempt to highlight reliable performance predictors. A combinatory design strategy should be used with CPS, taking into consideration 3D architecture, adequate surface chemistry and topography, all of which are needed to promote bone formation. CPS represent the media of choice for delivery of osteogenic factors and anti-infectives. Non-osteoblast mediated mineral deposition can confound in vitro osteogenesis testing of CPS and therefore the expression of a variety of proteins or genes including collagen type I, bone sialoprotein and osteocalcin should be confirmed in addition to increased mineral content. CPS are a superior scaffold material for bone regeneration because they actively promote osteogenesis. Biodegradability of CPS via calcium and phosphate release represents a unique asset. Structural control of CPS at the macro, micro and nanoscale and their combination with cells and polymeric materials is likely to lead to significant developments in bone tissue engineering. Copyright © 2015 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

An approach to architecture 3D scaffold with interconnective microchannel networks inducing angiogenesis for tissue engineering.

PubMed

Sun, Jiaoxia; Wang, Yuanliang; Qian, Zhiyong; Hu, Chenbo

2011-11-01

The angiogenesis of 3D scaffold is one of the major current limitations in clinical practice tissue engineering. The new strategy of construction 3D scaffold with microchannel circulation network may improve angiogenesis. In this study, 3D poly(D: ,L: -lactic acid) scaffolds with controllable microchannel structures were fabricated using sacrificial sugar structures. Melt drawing sugar-fiber network produced by a modified filament spiral winding method was used to form the microchannel with adjustable diameters and porosity. This fabrication process was rapid, inexpensive, and highly scalable. The porosity, microchannel diameter, interconnectivity and surface topographies of the scaffold were characterized by scanning electron microscopy. Mechanical properties were evaluated by compression tests. The mean porosity values of the scaffolds were in the 65-78% and the scaffold exhibited microchannel structure with diameter in the 100-200 μm range. The results showed that the scaffolds exhibited an adequate porosity, interconnective microchannel network, and mechanical properties. The cell culture studies with endothelial cells (ECs) demonstrated that the scaffold allowed cells to proliferate and penetrate into the volume of the entire scaffold. Overall, these findings suggest that the fabrication process offers significant advantages and flexibility in generating a variety of non-cytotoxic tissue engineering scaffolds with controllable distributions of porosity and physical properties that could provide the necessary physical cues for ECs and further improve angiogenesis for tissue engineering.

C 3-symmetric opioid scaffolds are pH-responsive DNA condensation agents.

PubMed

McStay, Natasha; Molphy, Zara; Coughlan, Alan; Cafolla, Attilio; McKee, Vickie; Gathergood, Nicholas; Kellett, Andrew

2017-01-25

Herein we report the synthesis of tripodal C 3 -symmetric opioid scaffolds as high-affinity condensation agents of duplex DNA. Condensation was achieved on both supercoiled and canonical B-DNA structures and identified by agarose electrophoresis, viscosity, turbidity and atomic force microscopy (AFM) measurements. Structurally, the requirement of a tris-opioid scaffold for condensation is demonstrated as both di- (C 2 -symmetric) and mono-substituted (C 1 -symmetric) mesitylene-linked opioid derivatives poorly coordinate dsDNA. Condensation, observed by toroidal and globule AFM aggregation, arises from surface-binding ionic interactions between protonated, cationic, tertiary amine groups on the opioid skeleton and the phosphate nucleic acid backbone. Indeed, by converting the 6-hydroxyl group of C 3 -morphine ( MC3: ) to methoxy substituents in C 3 -heterocodeine ( HC3: ) and C 3 -oripavine ( OC3: ) molecules, dsDNA compaction is retained thus negating the possibility of phosphate-hydroxyl surface-binding. Tripodal opioid condensation was identified as pH dependent and strongly influenced by ionic strength with further evidence of cationic amine-phosphate backbone coordination arising from thermal melting analysis and circular dichroism spectroscopy, with compaction also witnessed on synthetic dsDNA co-polymers poly[d(A-T) 2 ] and poly[d(G-C) 2 ]. On-chip microfluidic analysis of DNA condensed by C 3 -agents provided concentration-dependent protection (inhibition) to site-selective excision by type II restriction enzymes: BamHI, HindIII, SalI and EcoRI, but not to the endonuclease DNase I. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Raloxifene microsphere-embedded collagen/chitosan/β-tricalcium phosphate scaffold for effective bone tissue engineering.

PubMed

Zhang, Ming-Lei; Cheng, Ji; Xiao, Ye-Chen; Yin, Ruo-Feng; Feng, Xu

2017-02-25

Engineering novel scaffolds that can mimic the functional extracellular matrix (ECM) would be a great achievement in bone tissue engineering. This paper reports the fabrication of novel collagen/chitosan/β-tricalcium phosphate (CCTP) based tissue engineering scaffold. In order to improve the regeneration ability of scaffold, we have embedded raloxifene (RLX)-loaded PLGA microsphere in the CCTP scaffold. The average pore of scaffold was in the range of 150-200μm with ideal mechanical strength and swelling/degradation characteristics. The release rate of RLX from the microsphere (MS) embedded scaffold was gradual and controlled. Also a significantly enhanced cell proliferation was observed in RLX-MS exposed cell group suggesting that microsphere/scaffold could be an ideal biomaterial for bone tissue engineering. Specifically, RLX-MS showed a significantly higher Alizarin red staining indicating the higher mineralization capacity of this group. Furthermore, a high alkaline phosphatase (ALP) activity for RLX-MS exposed group after 15days incubation indicates the bone regeneration capacity of MC3T3-E1 cells. Overall, present study showed that RLX-loaded microsphere embedded scaffold has the promising potential for bone tissue engineering applications. Copyright © 2016. Published by Elsevier B.V.

Chitosan-based hydrogel tissue scaffolds made by 3D plotting promotes osteoblast proliferation and mineralization.

PubMed

Liu, I-Hsin; Chang, Shih-Hsin; Lin, Hsin-Yi

2015-05-13

A 3D plotting system was used to make chitosan-based tissue scaffolds with interconnected pores using pure chitosan (C) and chitosan cross-linked with pectin (CP) and genipin (CG). A freeze-dried chitosan scaffold (CF/D) was made to compare with C, to observe the effects of structural differences. The fiber size, pore size, porosity, compression strength, swelling ratio, drug release efficacy, and cumulative weight loss of the scaffolds were measured. Osteoblasts were cultured on the scaffolds and their proliferation, type I collagen production, alkaline phosphatase activity, calcium deposition, and morphology were observed. C had a lower swelling ratio, degradation, porosity and drug release efficacy and a higher compressional stiffness and cell proliferation compared to CF/D (p < 0.05). Of the 3D-plotted samples, cells on CP exhibited the highest degree of mineralization after 21 d (p < 0.05). CP also had the highest swelling ratio and fastest drug release, followed by C and CG (p < 0.05). Both CP and CG were stiffer and degraded more slowly in saline solution than C (p < 0.05). In summary, 3D-plotted scaffolds were stronger, less likely to degrade and better promoted osteoblast cell proliferation in vitro compared to the freeze-dried scaffolds. C, CP and CG were structurally similar, and the different crosslinking caused significant changes in their physical and biological performances.

Fabrication of a Highly Aligned Neural Scaffold via a Table Top Stereolithography 3D Printing and Electrospinning^.

PubMed

Lee, Se-Jun; Nowicki, Margaret; Harris, Brent; Zhang, Lijie Grace

2017-06-01

Three-dimensional (3D) bioprinting is a rapidly emerging technique in the field of tissue engineering to fabricate extremely intricate and complex biomimetic scaffolds in the range of micrometers. Such customized 3D printed constructs can be used for the regeneration of complex tissues such as cartilage, vessels, and nerves. However, the 3D printing techniques often offer limited control over the resolution and compromised mechanical properties due to short selection of printable inks. To address these limitations, we combined stereolithography and electrospinning techniques to fabricate a novel 3D biomimetic neural scaffold with a tunable porous structure and embedded aligned fibers. By employing two different types of biofabrication methods, we successfully utilized both synthetic and natural materials with varying chemical composition as bioink to enhance biocompatibilities and mechanical properties of the scaffold. The resulting microfibers composed of polycaprolactone (PCL) polymer and PCL mixed with gelatin were embedded in 3D printed hydrogel scaffold. Our results showed that 3D printed scaffolds with electrospun fibers significantly improve neural stem cell adhesion when compared to those without the fibers. Furthermore, 3D scaffolds embedded with aligned fibers showed an enhancement in cell proliferation relative to bare control scaffolds. More importantly, confocal microscopy images illustrated that the scaffold with PCL/gelatin fibers greatly increased the average neurite length and directed neurite extension of primary cortical neurons along the fiber. The results of this study demonstrate the potential to create unique 3D neural tissue constructs by combining 3D bioprinting and electrospinning techniques.

Morphology and characterization of 3D micro-porous structured chitosan scaffolds for tissue engineering.

PubMed

Hsieh, Wen-Chuan; Chang, Chih-Pong; Lin, Shang-Ming

2007-06-15

This research studies the morphology and characterization of three-dimensional (3D) micro-porous structures produced from biodegradable chitosan for use as scaffolds for cells culture. The chitosan 3D micro-porous structures were produced by a simple liquid hardening method, which includes the processes of foaming by mechanical stirring without any chemical foaming agent added, and hardening by NaOH cross linking. The pore size and porosity were controlled with mechanical stirring strength. This study includes the morphology of chitosan scaffolds, the characterization of mechanical properties, water absorption properties and in vitro enzymatic degradation of the 3D micro-porous structures. The results show that chitosan 3D micro-porous structures were successfully produced. Better formation samples were obtained when chitosan concentration is at 1-3%, and concentration of NaOH is at 5%. Faster stirring rate would produce samples of smaller pore diameter, but when rotation speed reaches 4000 rpm and higher the changes in pore size is minimal. Water absorption would reduce along with the decrease of chitosan scaffolds' pore diameter. From stress-strain analysis, chitosan scaffolds' mechanical properties are improved when it has smaller pore diameter. From in vitro enzymatic degradation results, it shows that the disintegration rate of chitosan scaffolds would increase along with the processing time increase, but approaching equilibrium when the disintegration rate reaches about 20%.

3D Scaffolds with Different Stiffness but the Same Microstructure for Bone Tissue Engineering.

PubMed

Chen, Guobao; Dong, Chanjuan; Yang, Li; Lv, Yonggang

2015-07-29

A growing body of evidence has shown that extracellular matrix (ECM) stiffness can modulate stem cell adhesion, proliferation, migration, differentiation, and signaling. Stem cells can feel and respond sensitively to the mechanical microenvironment of the ECM. However, most studies have focused on classical two-dimensional (2D) or quasi-three-dimensional environments, which cannot represent the real situation in vivo. Furthermore, most of the current methods used to generate different mechanical properties invariably change the fundamental structural properties of the scaffolds (such as morphology, porosity, pore size, and pore interconnectivity). In this study, we have developed novel three-dimensional (3D) scaffolds with different degrees of stiffness but the same 3D microstructure that was maintained by using decellularized cancellous bone. Mixtures of collagen and hydroxyapatite [HA: Ca10(PO4)6(OH)2] with different proportions were coated on decellularized cancellous bone to vary the stiffness (local stiffness, 13.00 ± 5.55 kPa, 13.87 ± 1.51 kPa, and 37.7 ± 19.6 kPa; bulk stiffness, 6.74 ± 1.16 kPa, 8.82 ± 2.12 kPa, and 23.61 ± 8.06 kPa). Microcomputed tomography (μ-CT) assay proved that there was no statistically significant difference in the architecture of the scaffolds before or after coating. Cell viability, osteogenic differentiation, cell recruitment, and angiogenesis were determined to characterize the scaffolds and evaluate their biological responses in vitro and in vivo. The in vitro results indicate that the scaffolds developed in this study could sustain adhesion and growth of rat mesenchymal stem cells (MSCs) and promote their osteogenic differentiation. The in vivo results further demonstrated that these scaffolds could help to recruit MSCs from subcutaneous tissue, induce them to differentiate into osteoblasts, and provide the 3D environment for angiogenesis. These findings showed that the method we developed can build scaffolds with

3D fabrication and characterization of phosphoric acid scaffold with a HA/β-TCP weight ratio of 60:40 for bone tissue engineering applications.

PubMed

Wang, Yanen; Wang, Kai; Li, Xinpei; Wei, Qinghua; Chai, Weihong; Wang, Shuzhi; Che, Yu; Lu, Tingli; Zhang, Bo

2017-01-01

A key requirement for three-dimensional printing (3-DP) at room temperature of medical implants depends on the availability of printable and biocompatible binder-powder systems. Different concentration polyvinyl alcohol (PVA) and phosphoric acid solutions were chosen as the binders to make the artificial stent biocompatible with sufficient compressive strength. In order to achieve an optimum balance between the bioceramic powder and binder solution, the biocompatibility and mechanical properties of these artificial stent samples were tested using two kinds of binder solutions. This study demonstrated the printable binder formulation at room temperature for the 3D artificial bone scaffolds. 0.6 wt% PVA solution was ejected easily via inkjet printing, with a supplementation of 0.25 wt% Tween 80 to reduce the surface tension of the polyvinyl alcohol solution. Compared with the polyvinyl alcohol scaffolds, the phosphoric acid scaffolds had better mechanical properties. Though both scaffolds supported the cell proliferation, the absorbance of the polyvinyl alcohol scaffolds was higher than that of the phosphoric acid scaffolds. The artificial stents with a hydroxyapatite/beta-tricalcium phosphate (HA/β-TCP) weight ratios of 60:40 depicted good biocompatibility for both scaffolds. Considering the scaffolds' mechanical and biocompatible properties, the phosphoric acid scaffolds with a HA/β-TCP weight ratio of 60:40 may be the best combination for bone tissue engineering applications.

Printability of calcium phosphate: calcium sulfate powders for the application of tissue engineered bone scaffolds using the 3D printing technique.

PubMed

Zhou, Zuoxin; Buchanan, Fraser; Mitchell, Christina; Dunne, Nicholas

2014-05-01

In this study, calcium phosphate (CaP) powders were blended with a three-dimensional printing (3DP) calcium sulfate (CaSO4)-based powder and the resulting composite powders were printed with a water-based binder using the 3DP technology. Application of a water-based binder ensured the manufacture of CaP:CaSO4 constructs on a reliable and repeatable basis, without long term damage of the printhead. Printability of CaP:CaSO4 powders was quantitatively assessed by investigating the key 3DP process parameters, i.e. in-process powder bed packing, drop penetration behavior and the quality of printed solid constructs. Effects of particle size, CaP:CaSO4 ratio and CaP powder type on the 3DP process were considered. The drop penetration technique was used to reliably identify powder formulations that could be potentially used for the application of tissue engineered bone scaffolds using the 3DP technique. Significant improvements (p<0.05) in the 3DP process parameters were found for CaP (30-110 μm):CaSO4 powders compared to CaP (<20 μm):CaSO4 powders. Higher compressive strength was obtained for the powders with the higher CaP:CaSO4 ratio. Hydroxyapatite (HA):CaSO4 powders showed better results than beta-tricalcium phosphate (β-TCP):CaSO4 powders. Solid and porous constructs were manufactured using the 3DP technique from the optimized CaP:CaSO4 powder formulations. High-quality printed constructs were manufactured, which exhibited appropriate green compressive strength and a high level of printing accuracy. Copyright © 2014 Elsevier B.V. All rights reserved.

Development and Assessment of a 3D-Printed Scaffold with rhBMP-2 for an Implant Surgical Guide Stent and Bone Graft Material: A Pilot Animal Study.

PubMed

Bae, Ji Cheol; Lee, Jin-Ju; Shim, Jin-Hyung; Park, Keun-Ho; Lee, Jeong-Seok; Bae, Eun-Bin; Choi, Jae-Won; Huh, Jung-Bo

2017-12-16

In this study, a new concept of a 3D-printed scaffold was introduced for the accurate placement of an implant and the application of a recombinant human bone morphogenetic protein-2 (rhBMP-2)-loaded bone graft. This preliminary study was conducted using two adult beagles to evaluate the 3D-printed polycaprolactone (PCL)/ β -tricalcium phosphate ( β -TCP)/bone decellularized extracellular matrix (bdECM) scaffold conjugated with rhBMP-2 for the simultaneous use as an implant surgical guide stent and bone graft material that promotes new bone growth. Teeth were extracted from the mandible of the beagle model and scanned by computed tomography (CT) to fabricate a customized scaffold that would fit the bone defect. After positioning the implant guide scaffold, the implant was placed and rhBMP-2 was injected into the scaffold of the experimental group. The two beagles were sacrificed after three months. The specimen block was obtained and scanned by micro-CT. Histological analysis showed that the control and experimental groups had similar new bone volume (NBV, %) but the experimental group with BMP exhibited a significantly higher bone-to-implant contact ratio (BIC, %). Within the limitations of this preliminary study, a 3D-printed scaffold conjugated with rhBMP-2 can be used simultaneously as an implant surgical guide and a bone graft in a large bone defect site. Further large-scale studies will be needed to confirm these results.

Formation of Neural Networks in 3D Scaffolds Fabricated by Means of Laser Microstereolithography.

PubMed

Vedunova, M V; Timashev, P S; Mishchenko, T A; Mitroshina, E V; Koroleva, A V; Chichkov, B N; Panchenko, V Ya; Bagratashvili, V N; Mukhina, I V

2016-08-01

We developed and tested new 3D scaffolds for neurotransplantation. Scaffolds of predetermined architectonic were prepared using microstereolithography technique. Scaffolds were highly biocompatible with the nervous tissue cells. In vitro studies showed that the material of fabricated scaffolds is not toxic for dissociated brain cells and promotes the formation of functional neural networks in the matrix. These results demonstrate the possibility of fabrication of tissue-engineering constructs for neurotransplantation based on created scaffolds.

3D-Printed Atsttrin-Incorporated Alginate/Hydroxyapatite Scaffold Promotes Bone Defect Regeneration with TNF/TNFR Signaling Involvement.

PubMed

Wang, Quan; Xia, Qingqing; Wu, Yan; Zhang, Xiaolei; Wen, Feiqiu; Chen, Xiaowen; Zhang, Shufang; Heng, Boon Chin; He, Yong; Ouyang, Hong-Wei

2015-08-05

High expression levels of pro-inflammatory tumor necrosis factor (TNF)-α within bone defects can decelerate and impair bone regeneration. However, there are few available bone scaffolds with anti-inflammatory function. The progranulin (PGRN)-derived engineered protein, Atsttrin, is known to exert antagonistic effects on the TNF-α function. Hence, this study investigates whether 3D-printed Atsttrin-incorporated alginate(Alg)/hydroxyapatite(nHAp) scaffolds can facilitate bone healing through affecting the TNF/TNFR signaling. A 3D bioprinting system is used to fabricate Atsttrin-Alg/nHAp composite scaffolds, and the Atsttrin release from this scaffold is characterized, followed by evaluation of its efficacy on bone regeneration both in vitro and in vivo. The 3D-printed Atsttrin-Alg/nHAp scaffold exhibits a precisely defined structure, can sustain Atsttrin release for at least 5 days, has negligible cytotoxicity, and supports cell adhesion. Atsttrin can also attenuate the suppressive effects of TNF-α on BMP-2-induced osteoblastic differentiation in vitro. The 3D-printed Atsttrin-Alg/nHAp scaffold significantly reduces the number of TNF-α positive cells within wound sites, 7 days after post-calvarial defect surgery. Additionally, histological staining and X-ray scanning results also show that the 3D-printed Atsttrin-Alg/nHAp scaffold enhances the regeneration of mice calvarial bone defects. These findings thus demonstrate that the precise structure and anti-inflammatory properties of 3D-printed Atsttrin-Alg/nHAp scaffolds may promote bone defect repair. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Engineering Human TMJ Discs with Protein-Releasing 3D-Printed Scaffolds.

PubMed

Legemate, K; Tarafder, S; Jun, Y; Lee, C H

2016-07-01

The temporomandibular joint (TMJ) disc is a heterogeneous fibrocartilaginous tissue positioned between the mandibular condyle and glenoid fossa of the temporal bone, with important roles in TMJ functions. Tissue engineering TMJ discs has emerged as an alternative approach to overcoming limitations of current treatments for TMJ disorders. However, the anisotropic collagen orientation and inhomogeneous fibrocartilaginous matrix distribution present challenges in the tissue engineering of functional TMJ discs. Here, we developed 3-dimensional (3D)-printed anatomically correct scaffolds with region-variant microstrand alignment, mimicking anisotropic collagen alignment in the TMJ disc and corresponding mechanical properties. Connective tissue growth factor (CTGF) and transforming growth factor beta 3 (TGFβ3) were then delivered in the scaffolds by spatially embedding CTGF- or TGFβ3-encapsulated microspheres (µS) to reconstruct the regionally variant fibrocartilaginous matrix in the native TMJ disc. When cultured with human mesenchymal stem/progenitor cells (MSCs) for 6 wk, 3D-printed scaffolds with CTGF/TGFβ3-µS resulted in a heterogeneous fibrocartilaginous matrix with overall distribution of collagen-rich fibrous structure in the anterior/posterior (AP) bands and fibrocartilaginous matrix in the intermediate zone, reminiscent of the native TMJ disc. High dose of CTGF/TGFβ3-µS (100 mg µS/g of scaffold) showed significantly more collagen II and aggrecan in the intermediate zone than a low dose (50 mg µS/g of scaffold). Similarly, a high dose of CTGF/TGFβ3-µS yielded significantly higher collagen I expression in the AP bands compared with the low-dose and empty µS. From stress relaxation tests, the ratio of relaxation modulus to instantaneous modulus was significantly smaller with CTGF/TGFβ3-µS than empty µS. Similarly, a significantly higher coefficient of viscosity was achieved with the high dose of CTGF/TGFβ3-µS compared with the low-dose and empty

Functionalization of PCL-3D Electrospun Nanofibrous Scaffolds for Improved BMP2-Induced Bone Formation.

PubMed

Miszuk, Jacob M; Xu, Tao; Yao, Qingqing; Fang, Fang; Childs, Josh D; Hong, Zhongkui; Tao, Jianning; Fong, Hao; Sun, Hongli

2018-03-01

Bone morphogenic protein 2 (BMP2) is a key growth factor for bone regeneration, possessing FDA approval for orthopedic applications. BMP2 is often required in supratherapeutic doses clinically, yielding adverse side effects and substantial treatment costs. Considering the crucial role of materials for BMPs delivery and cell osteogenic differentiation, we devote to engineering an innovative bone-matrix mimicking niche to improve low dose of BMP2-induced bone formation. Our previous work describes a novel technique, named thermally induced nanofiber self-agglomeration (TISA), for generating 3D electrospun nanofibrous (NF) polycaprolactone (PCL) scaffolds. TISA process could readily blend PCL with PLA, leading to increased osteogenic capabilities in vitro , however, these bio-inert synthetic polymers produced limited BMP2-induced bone formation in vivo. We therefore hypothesize that functionalization of NF 3D PCL scaffolds with bone-like hydroxyapatite (HA) and BMP2 signaling activator phenamil will provide a favorable osteogenic niche for bone formation at low doses of BMP2. Compared to PCL-3D scaffolds, PCL/HA-3D scaffolds demonstrated synergistically enhanced osteogenic differentiation capabilities of C2C12 cells with phenamil. Importantly, in vivo studies showed this synergism was able to generate significantly increased new bone in an ectopic mouse model, suggesting PCL/HA-3D scaffolds act as a favorable synthetic extracellular matrix for bone regeneration.

Highly defined 3D printed chitosan scaffolds featuring improved cell growth.

PubMed

Elviri, Lisa; Foresti, Ruben; Bergonzi, Carlo; Zimetti, Francesca; Marchi, Cinzia; Bianchera, Annalisa; Bernini, Franco; Silvestri, Marco; Bettini, Ruggero

2017-07-12

The augmented demand for medical devices devoted to tissue regeneration and possessing a controlled micro-architecture means there is a need for industrial scale-up in the production of hydrogels. A new 3D printing technique was applied to the automation of a freeze-gelation method for the preparation of chitosan scaffolds with controlled porosity. For this aim, a dedicated 3D printer was built in-house: a preliminary effort has been necessary to explore the printing parameter space to optimize the printing results in terms of geometry, tolerances and mechanical properties of the product. Analysed parameters included viscosity of the starting chitosan solution, which was measured with a Brookfield viscometer, and temperature of deposition, which was determined by filming the process with a cryocooled sensor thermal camera. Optimized parameters were applied to the production of scaffolds from solutions of chitosan alone or with the addition of raffinose as a viscosity modifier. Resulting hydrogels were characterized in terms of morphology and porosity. In vitro cell culture studies comparing 3D printed scaffolds with their homologous produced by solution casting evidenced an improvement in biocompatibility deriving from the production technique as well as from the solid state modification of chitosan stemming from the addition of the viscosity modifier.

A computational modeling approach for the characterization of mechanical properties of 3D alginate tissue scaffolds.

PubMed

Nair, K; Yan, K C; Sun, W

2008-01-01

Scaffold guided tissue engineering is an innovative approach wherein cells are seeded onto biocompatible and biodegradable materials to form 3-dimensional (3D) constructs that, when implanted in the body facilitate the regeneration of tissue. Tissue scaffolds act as artificial extracellular matrix providing the environment conducive for tissue growth. Characterization of scaffold properties is necessary to understand better the underlying processes involved in controlling cell behavior and formation of functional tissue. We report a computational modeling approach to characterize mechanical properties of 3D gellike biomaterial, specifically, 3D alginate scaffold encapsulated with cells. Alginate inherent nonlinearity and variations arising from minute changes in its concentration and viscosity make experimental evaluation of its mechanical properties a challenging and time consuming task. We developed an in silico model to determine the stress-strain relationship of alginate based scaffolds from experimental data. In particular, we compared the Ogden hyperelastic model to other hyperelastic material models and determined that this model was the most suitable to characterize the nonlinear behavior of alginate. We further propose a mathematical model that represents the alginate material constants in Ogden model as a function of concentrations and viscosity. This study demonstrates the model capability to predict mechanical properties of 3D alginate scaffolds.

Fabrication of a customized bone scaffold using a homemade medical 3D printer for comminuted fractures

NASA Astrophysics Data System (ADS)

Yoon, Do-Kun; Jung, Joo-Young; Shin, Han-Back; Kim, Moo-Sub; Choe, Bo-Young; Kim, Sunmi; Suh, Tae Suk; Lee, Keum Sil; Xing, Lei

2016-09-01

The purpose of this study was to show a 3D printed reconstruction model of a bone destroyed by a comminuted fracture. After a thoracic limb of a cow with a comminuted fracture was scanned by using computed tomography, a scaffold was designed by using a 3D modeling tool for its reconstruction and fabricated by using a homemade medical 3D printer. The homemade medical 3D printer was designed for medical use. In order to reconstruct the geometry of the destroyed bone, we use the geometry of a similar section (reference geometry) of normal bone in the 3D modeling process. The missing part between the destroyed ridge and the reference geometry was filled with an effective space by using a manual interpolation. Inexpensive materials and free software were used to construct the medical 3D printer system. The fabrication of the scaffold progressed according to the design of reconstructed bone by using this medical 3D printer. The material of the scaffold was biodegradable material, and could be transplanted into the human body. The fabricated scaffold was correctly inserted into the fractured bone in place of the destroyed portion, with good agreement. According to physical stress test results, the performance of printing resolution was 0.1 mm. The average geometrical error of the scaffold was below 0.3 mm. The reconstructed bone by using the fabricated scaffold was able to support the weight of the human body. No process used to obtain the result was complex or required many resources. The methods and results in this study show several possible clinical applications in fields such as orthopedics or oncology without a need to purchase high-price instruments for 3D printing.

Fabrication of chitosan/gallic acid 3D microporous scaffold for tissue engineering applications.

PubMed

Thangavel, Ponrasu; Ramachandran, Balaji; Muthuvijayan, Vignesh

2016-05-01

This study explores the potential of gallic acid incorporated chitosan (CS/GA) 3D scaffolds for tissue engineering applications. Scaffolds were prepared by freezing and lyophilization technique and characterized. FTIR spectra confirmed the presence of GA in chitosan (CS) gel. DSC and TGA analysis revealed that the structure of chitosan was not altered due to the incorporation of GA, but thermal stability was significantly increased compared to the CS scaffold. SEM micrographs showed smooth, homogeneous, and microporous architecture of the scaffolds with good interconnectivity. CS/GA scaffolds exhibited approximately 90% porosity on average, increased swelling (600-900%) and controlled biodegradation (15-40%) in PBS (pH 7.4 at 37°C) with 1 mg/mL of lysozyme. CS/GA scaffolds showed 2-4 fold decrease in CFUs (p < 0.05) for both gram positive and gram negative bacteria compared to the CS scaffold. Cytotoxicity of these scaffolds was evaluated using NIH 3T3 L1 fibroblast cells. CS/GA 0.25% scaffold showed similar viability with CS scaffold at 24 and 48 h. CS/GA scaffolds (0.5-1.0%) showed 60-75% viability at 24 h and 90% at 48 h. SEM images showed that an increased cell attachment was observed for CS/GA scaffolds compared to CS scaffolds. These findings authenticate that CS/GA scaffolds were cytocompatible and would be useful for tissue engineering applications. © 2015 Wiley Periodicals, Inc.

Expansion of Bone Marrow Mesenchymal Stromal Cells in Perfused 3D Ceramic Scaffolds Enhances In Vivo Bone Formation.

PubMed

Hoch, Allison I; Duhr, Ralph; Di Maggio, Nunzia; Mehrkens, Arne; Jakob, Marcel; Wendt, David

2017-12-01

Bone marrow-derived mesenchymal stromal cells (BMSC), when expanded directly within 3D ceramic scaffolds in perfusion bioreactors, more reproducibly form bone when implanted in vivo as compared to conventional expansion on 2D polystyrene dishes/flasks. Since the bioreactor-based expansion on 3D ceramic scaffolds encompasses multiple aspects that are inherently different from expansion on 2D polystyrene, we aimed to decouple the effects of specific parameters among these two model systems. We assessed the effects of the: 1) 3D scaffold vs. 2D surface; 2) ceramic vs. polystyrene materials; and 3) BMSC niche established within the ceramic pores during in vitro culture, on subsequent in vivo bone formation. While BMSC expanded on 3D polystyrene scaffolds in the bioreactor could maintain their in vivo osteogenic potential, results were similar as BMSC expanded in monolayer on 2D polystyrene, suggesting little influence of the scaffold 3D environment. Bone formation was most reproducible when BMSC are expanded on 3D ceramic, highlighting the influence of the ceramic substrate. The presence of a pre-formed niche within the scaffold pores had negligible effects on the in vivo bone formation. The results of this study allow a greater understanding of the parameters required for perfusion bioreactor-based manufacturing of osteogenic grafts for clinical applications. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Low intensity pulse ultrasound stimulate chondrocytes growth in a 3-D alginate scaffold through improved porosity and permeability.

PubMed

Guo, Gepu; Lu, Lu; Ji, Hongfei; Ma, Yong; Dong, Rui; Tu, Juan; Guo, Xiasheng; Qiu, Yuanyuan; Wu, Junru; Zhang, Dong

2015-04-01

A 3-D scaffold culture system has been used to promote in producing functional chondrocytes for repairing damaged cartilage. In the present study, the low intensity pulse ultrasound (LIPUS) (P(-)=0, 0.055, 0.085 and 0.11 MPa) was applied to improve the porosity and permeability of a 3-D alginate scaffold which was beneficial for the nutrition supply and metabolism during cell growth in 3-D alginate scaffold. The porosity and permeability of the scaffold was quantitatively analyzed based on scanning electron microscopy examination and fluorescence image observation. The results suggest that, for the scaffold exposed to LIPUS, its porosity and permeability could be significantly enhanced by the increasing LIPUS amplitude, which might be induced by the microstreaming shear stress generated by ultrasound-driven microbubble oscillations. Furthermore, the assessments of cell proliferation and collagen II expression confirmed that chondrocytes growth could be effectively promoted in 3-D alginate scaffolds treated by LIPUS, because of the improved scaffold porosity and permeability might benefit cell growth space and nutrition supply. It should also be noticed that appropriate LIPUS driving parameters should be adapted to achieve optimized chondrocytes culture effect in 3-D alginate scaffold. Copyright © 2014 Elsevier B.V. All rights reserved.

Rapid 3D printing of anatomically accurate and mechanically heterogeneous aortic valve hydrogel scaffolds

PubMed Central

Hockaday, L A; Kang, K H; Colangelo, N W; Cheung, P Y C; Duan, B; Malone, E; Wu, J; Girardi, L N; Bonassar, L J; Lipson, H; Chu, C C; Butcher, J T

2013-01-01

The aortic valve exhibits complex three-dimensional (3D) anatomy and heterogeneity essential for long-term efficient biomechanical function. These are, however, challenging to mimic in de novo engineered living tissue valve strategies. We present a novel simultaneous 3D-printing/photocrosslinking technique for rapidly engineering complex, heterogeneous aortic valve scaffolds. Native anatomic and axisymmetric aortic valve geometries (root wall and tri-leaflets) with 12 to 22 mm inner diameters (ID) were 3D printed with poly-ethylene glycol-diacrylate (PEG-DA) hydrogels (700 or 8000 MW) supplemented with alginate. 3D printing geometric accuracy was quantified and compared using Micro-CT. Porcine aortic valve interstitial cells (PAVIC) seeded scaffolds were cultured for up to 21 days. Results showed that blended PEG-DA scaffolds could achieve over 10-fold range in elastic modulus (5.3±0.9 to 74.6±1.5 kPa). 3D printing times for valve conduits with mechanically contrasting hydrogels were optimized to 14 to 45 minutes, increasing linearly with conduit diameter. Larger printed valves had greater shape fidelity (93.3±2.6, 85.1±2.0, and 73.3±5.2% for 22, 17, and 12 mm ID porcine valves; 89.1±4.0, 84.1±5.6, and 66.6±5.2% for simplified valves). PAVIC seeded scaffolds maintained near 100% viability over 21 days. These results demonstrate that 3D hydrogel printing with controlled photocrosslinking can rapidly fabricate anatomical heterogeneous valve conduits that support cell engraftment. PMID:22914604

Chitosan porous 3D scaffolds embedded with resolvin D1 to improve in vivo bone healing.

PubMed

Vasconcelos, Daniela P; Costa, Madalena; Neves, Nuno; Teixeira, José H; Vasconcelos, Daniel M; Santos, Susana G; Águas, Artur P; Barbosa, Mário A; Barbosa, Judite N

2018-06-01

The aim of this study was to investigate the effect chitosan (Ch) porous 3D scaffolds embedded with resolvin D1 (RvD1), an endogenous pro-resolving lipid mediator, on bone tissue healing. These scaffolds previous developed by us have demonstrated to have immunomodulatory properties namely in the modulation of the macrophage inflammatory phenotypic profile in an in vivo model of inflammation. Herein, results obtained in an in vivo rat femoral defect model demonstrated that two months after Ch + RvD1 scaffolds implantation, an increase in new bone formation, in bone trabecular thickness, and in collagen type I and Coll I/Coll III ratio were observed. These results suggest that Ch scaffolds embedded with RvD1 were able to lead to the formation of new bone with improvement of trabecular thickness. This study shows that the presence of RvD1 in the acute phase of the inflammatory response to the implanted biomaterial had a positive role in the subsequent bone tissue repair, thus demonstrating the importance of innovative approaches for the control of immune responses to biomedical implants in the design of advanced strategies for regenerative medicine. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1626-1633, 2018. © 2018 Wiley Periodicals, Inc.

Cold atmospheric plasma (CAP) surface nanomodified 3D printed polylactic acid (PLA) scaffolds for bone regeneration.

PubMed

Wang, Mian; Favi, Pelagie; Cheng, Xiaoqian; Golshan, Negar H; Ziemer, Katherine S; Keidar, Michael; Webster, Thomas J

2016-12-01

Three-dimensional (3D) printing is a new fabrication method for tissue engineering which can precisely control scaffold architecture at the micron-scale. However, scaffolds not only need 3D biocompatible structures that mimic the micron structure of natural tissues, they also require mimicking of the nano-scale extracellular matrix properties of the tissue they intend to replace. In order to achieve this, the objective of the present in vitro study was to use cold atmospheric plasma (CAP) as a quick and inexpensive way to modify the nano-scale roughness and chemical composition of a 3D printed scaffold surface. Water contact angles of a normal 3D printed poly-lactic-acid (PLA) scaffold dramatically dropped after CAP treatment from 70±2° to 24±2°. In addition, the nano-scale surface roughness (Rq) of the untreated 3D PLA scaffolds drastically increased (up to 250%) after 1, 3, and 5min of CAP treatment from 1.20nm to 10.50nm, 22.90nm, and 27.60nm, respectively. X-ray photoelectron spectroscopy (XPS) analysis showed that the ratio of oxygen to carbon significantly increased after CAP treatment, which indicated that the CAP treatment of PLA not only changed nano-scale roughness but also chemistry. Both changes in hydrophilicity and nano-scale roughness demonstrated a very efficient plasma treatment, which in turn significantly promoted both osteoblast (bone forming cells) and mesenchymal stem cell attachment and proliferation. These promising results suggest that CAP surface modification may have potential applications for enhancing 3D printed PLA bone tissue engineering materials (and all 3D printed materials) in a quick and an inexpensive manner and, thus, should be further studied. Three-dimensional (3D) printing is a new fabrication method for tissue engineering which can precisely control scaffold architecture at the micron-scale. Although their success is related to their ability to exactly mimic the structure of natural tissues and control mechanical

Chondrogenesis of infrapatellar fat pad derived adipose stem cells in 3D printed chitosan scaffold.

PubMed

Ye, Ken; Felimban, Raed; Traianedes, Kathy; Moulton, Simon E; Wallace, Gordon G; Chung, Johnson; Quigley, Anita; Choong, Peter F M; Myers, Damian E

2014-01-01

Infrapatellar fat pad adipose stem cells (IPFP-ASCs) have been shown to harbor chondrogenic potential. When combined with 3D polymeric structures, the stem cells provide a source of stem cells to engineer 3D tissues for cartilage repair. In this study, we have shown human IPFP-ASCs seeded onto 3D printed chitosan scaffolds can undergo chondrogenesis using TGFβ3 and BMP6. By week 4, a pearlescent, cartilage-like matrix had formed that penetrated the top layers of the chitosan scaffold forming a 'cap' on the scaffold. Chondrocytic morphology showed typical cells encased in extracellular matrix which stained positively with toluidine blue. Immunohistochemistry demonstrated positive staining for collagen type II and cartilage proteoglycans, as well as collagen type I. Real time PCR analysis showed up-regulation of collagen type II, aggrecan and SOX9 genes when IPFP-ASCs were stimulated by TGFβ3 and BMP6. Thus, IPFP-ASCs can successfully undergo chondrogenesis using TGFβ3 and BMP6 and the cartilage-like tissue that forms on the surface of 3D-printed chitosan scaffold may prove useful as an osteochondral graft.

Chitosan-alginate 3D scaffolds as a mimic of the glioma tumor microenvironment.

PubMed

Kievit, Forrest M; Florczyk, Stephen J; Leung, Matthew C; Veiseh, Omid; Park, James O; Disis, Mary L; Zhang, Miqin

2010-08-01

Despite recent advances in the understanding of its cell biology, glioma remains highly lethal. Development of effective therapies requires a cost-effective in vitro tumor model that more accurately resembles the in vivo tumor microenvironment as standard two-dimensional (2D) tissue culture conditions do so poorly. Here we report on the use of a three-dimensional (3D) chitosan-alginate (CA) scaffold to serve as an extracellular matrix that promotes the conversion of cultured cancer cells to a more malignant in vivo-like phenotype. Human U-87 MG and U-118 MG glioma cells and rat C6 glioma cells were chosen for the study. In vitro tumor cell proliferation and secretion of factors that promote tumor malignancy, including VEGF, MMP-2, fibronectin, and laminin, were assessed. The scaffolds pre-cultured with U-87 MG and C6 cells were then implanted into nude mice to evaluate tumor growth and blood vessel recruitment compared to the standard 2D cell culture and 3D Matrigel matrix xenograft controls. Our results indicate that while the behavior of C6 cells showed minimal differences due to their highly malignant and invasive nature, U-87 MG and U-118 MG cells exhibited notably higher malignancy when cultured in CA scaffolds. CA scaffolds provide a 3D microenvironment for glioma cells that is more representative of the in vivo tumor, thus can serve as a more effective platform for development and study of anticancer therapeutics. This unique CA scaffold platform may offer a valuable alternative strategy to the time-consuming and costly animal studies for a wide variety of experimental designs. Copyright 2010 Elsevier Ltd. All rights reserved.

Antimicrobial 3D Porous Scaffolds Prepared by Additive Manufacturing and Breath Figures.

PubMed

Vargas-Alfredo, Nelson; Dorronsoro, Ane; Cortajarena, Aitziber L; Rodríguez-Hernández, Juan

2017-10-25

We describe herein a novel strategy for the fabrication of efficient 3D printed antibacterial scaffolds. For this purpose, both the surface topography as well as the chemical composition of 3D scaffolds fabricated by additive manufacturing were modified. The scaffolds were fabricated by fused deposition modeling (FDM) using high-impact polystyrene (HIPS) filaments. The surface of the objects was then topographically modified providing materials with porous surfaces by means of the Breath Figures approach. The strategy involves the immersion of the scaffold in a polymer solution during a precise period of time. This approach permitted the modification of the pore size varying the immersion time as well as the solution concentration. Moreover, by using polymer blend solutions of polystyrene and polystyrene-b-poly(acrylic acid) (PS 23 -b-PAA 18 ) and a quaternized polystyrene-b-poly(dimethylaminoethyl methacrylate) (PS 42 -b-PDMAEMAQ 17 ), the scaffolds were simultaneously chemically modified. The surfaces were characterized by scanning electron microscopy and infrared spectroscopy. Finally, the biological response toward bacteria was explored. Porous surfaces prepared using quaternized PDMAEMA as well as those prepared using PAA confer antimicrobial activity to the films, i.e., were able to kill on contact Staphylococcus aureus employed as model bacteria.

Lead-oriented synthesis: Investigation of organolithium-mediated routes to 3-D scaffolds and 3-D shape analysis of a virtual lead-like library.

PubMed

Lüthy, Monique; Wheldon, Mary C; Haji-Cheteh, Chehasnah; Atobe, Masakazu; Bond, Paul S; O'Brien, Peter; Hubbard, Roderick E; Fairlamb, Ian J S

2015-06-01

Synthetic routes to six 3-D scaffolds containing piperazine, pyrrolidine and piperidine cores have been developed. The synthetic methodology focused on the use of N-Boc α-lithiation-trapping chemistry. Notably, suitably protected and/or functionalised medicinal chemistry building blocks were synthesised via concise, connective methodology. This represents a rare example of lead-oriented synthesis. A virtual library of 190 compounds was then enumerated from the six scaffolds. Of these, 92 compounds (48%) fit the lead-like criteria of: (i) -1⩽AlogP⩽3; (ii) 14⩽number of heavy atoms⩽26; (iii) total polar surface area⩾50Å(2). The 3-D shapes of the 190 compounds were analysed using a triangular plot of normalised principal moments of inertia (PMI). From this, 46 compounds were identified which had lead-like properties and possessed 3-D shapes in under-represented areas of pharmaceutical space. Thus, the PMI analysis of the 190 member virtual library showed that whilst scaffolds which may appear on paper to be 3-D in shape, only 24% of the compounds actually had 3-D structures in the more interesting areas of 3-D drug space. Copyright © 2015 Elsevier Ltd. All rights reserved.

A bioprosthetic ovary created using 3D printed microporous scaffolds restores ovarian function in sterilized mice.

PubMed

Laronda, Monica M; Rutz, Alexandra L; Xiao, Shuo; Whelan, Kelly A; Duncan, Francesca E; Roth, Eric W; Woodruff, Teresa K; Shah, Ramille N

2017-05-16

Emerging additive manufacturing techniques enable investigation of the effects of pore geometry on cell behavior and function. Here, we 3D print microporous hydrogel scaffolds to test how varying pore geometry, accomplished by manipulating the advancing angle between printed layers, affects the survival of ovarian follicles. 30° and 60° scaffolds provide corners that surround follicles on multiple sides while 90° scaffolds have an open porosity that limits follicle-scaffold interaction. As the amount of scaffold interaction increases, follicle spreading is limited and survival increases. Follicle-seeded scaffolds become highly vascularized and ovarian function is fully restored when implanted in surgically sterilized mice. Moreover, pups are born through natural mating and thrive through maternal lactation. These findings present an in vivo functional ovarian implant designed with 3D printing, and indicate that scaffold pore architecture is a critical variable in additively manufactured scaffold design for functional tissue engineering.

Extrusion-based 3D printing of poly(propylene fumarate) scaffolds with hydroxyapatite gradients

PubMed Central

Trachtenberg, Jordan E.; Placone, Jesse K.; Smith, Brandon T.; Fisher, John P.; Mikos, Antonios G.

2017-01-01

The primary focus of this work is to present the current challenges of printing scaffolds with concentration gradients of nanoparticles with an aim to improve the processing of these scaffolds. Furthermore, we address how print fidelity is related to material composition and emphasize the importance of considering this relationship when developing complex scaffolds for bone implants. The ability to create complex tissues is becoming increasingly relevant in the tissue engineering community. For bone tissue engineering applications, this work demonstrates the ability to use extrusion-based printing techniques to control the spatial deposition of hydroxyapatite (HA) nanoparticles in a 3D composite scaffold. In doing so, we combined the benefits of synthetic, degradable polymers, such as poly(propylene fumarate) (PPF), with osteoconductive HA nanoparticles that provide robust compressive mechanical properties. Furthermore, the final 3D printed scaffolds consisted of well-defined layers with interconnected pores, two critical features for a successful bone implant. To demonstrate a controlled gradient of HA, thermogravimetric analysis was carried out to quantify HA on a per-layer basis. Moreover, we non-destructively evaluated the tendency of HA particles to aggregate within PPF using micro-computed tomography (µCT). This work provides insight for proper fabrication and characterization of composite scaffolds containing particle gradients and has broad applicability for future efforts in fabricating complex scaffolds for tissue engineering applications. PMID:28125380

Extrusion-based 3D printing of poly(propylene fumarate) scaffolds with hydroxyapatite gradients.

PubMed

Trachtenberg, Jordan E; Placone, Jesse K; Smith, Brandon T; Fisher, John P; Mikos, Antonios G

2017-04-01

The primary focus of this work is to present the current challenges of printing scaffolds with concentration gradients of nanoparticles with an aim to improve the processing of these scaffolds. Furthermore, we address how print fidelity is related to material composition and emphasize the importance of considering this relationship when developing complex scaffolds for bone implants. The ability to create complex tissues is becoming increasingly relevant in the tissue engineering community. For bone tissue engineering applications, this work demonstrates the ability to use extrusion-based printing techniques to control the spatial deposition of hydroxyapatite (HA) nanoparticles in a 3D composite scaffold. In doing so, we combined the benefits of synthetic, degradable polymers, such as poly(propylene fumarate) (PPF), with osteoconductive HA nanoparticles that provide robust compressive mechanical properties. Furthermore, the final 3D printed scaffolds consisted of well-defined layers with interconnected pores, two critical features for a successful bone implant. To demonstrate a controlled gradient of HA, thermogravimetric analysis was carried out to quantify HA on a per-layer basis. Moreover, we non-destructively evaluated the tendency of HA particles to aggregate within PPF using micro-computed tomography (μCT). This work provides insight for proper fabrication and characterization of composite scaffolds containing particle gradients and has broad applicability for future efforts in fabricating complex scaffolds for tissue engineering applications.

Physicomechanical, In Vitro and In Vivo Performance of 3D Printed Doped Tricalcium Phosphate Scaffolds for Bone Tissue Engineering and Drug Delivery

NASA Astrophysics Data System (ADS)

Tarafder, Solaiman

Although tricalcium phosphate (TCP) is widely used in bone tissue engineering, the strength degradation kinetics is not well controlled. This study focuses on the underlying mechanism of strength degradation kinetics by incorporating trace elements in TCP. The objective of this research is to modify the mechanical properties of TCP to achieve the desired degradation rate for the specific need, and improve the in vivo bioactivity for early wound healing by incorporating trace elements such as strontium (Sr2+), magnesium (Mg2+) and silicon (Si4+) as dopants. The hypothesis of this research is that the presence of different trace elements in TCP will influence its phase stability, microstructure, mechanical strength, and both in vitro and in vivo bioactivity. Direct three dimensional printing (3DP) was used to fabricate designed interconnected macroporous pure and doped TCP scaffolds. Microwave sintering as opposed to conventional sintering was also used for better densification and higher mechanical strength. A maximum compressive strength of 10.95 +/- 1.28 MPa and 12.01 +/- 1.56 MPa were achieved for pure and Sr2+-Mg2+ doped TCP scaffolds with 500 microm designed pores (˜400 microm after sintering) sintered in microwave furnace, respectively. Substitution of Mg2+ and Sr2+ into calcium (Ca2+) sites of TCP crystal lattice contributed to phase stability and controlled gradual degradation. On the other hand, Si4+ substitution into phosphorous (P5+) sites destabilized the crystal structure and accelerated degradation of TCP. Interconnected macroporous beta-TCP scaffolds facilitated in vivo guided bone tissue regeneration through infiltration of cells and extracellular matrix into the designed pores. Presence of Sr2+, Mg2+ and Si4+ into beta-TCP induced increased in vivo early bone formation and better bone remodeling through increased extracellular matrix production such as, collagen and osteocalcin, when tested in rat and rabbit distal femur model. The presence of Si4

Enrichment of glioma stem cell-like cells on 3D porous scaffolds composed of different extracellular matrix.

PubMed

Wang, Xuanzhi; Dai, Xingliang; Zhang, Xinzhi; Li, Xinda; Xu, Tao; Lan, Qing

2018-04-15

Cancer stem cells (CSCs), being tumor-initiating with self-renewal capacity and heterogeneity, are most likely the cause of tumor resistance, reoccurrence and metastasis. To further investigate the role of CSCs in tumor biology, there is a need to develop an effective culture system to grow, maintain and enrich CSCs. Three-dimensional (3D) cell culture model has been widely used in tumor research and drug screening. Recently, researchers have begun to utilize 3D models to culture cancer cells for CSCs enrichment. In this study, glioma cell line was cultured with 3D porous chitosan (CS) scaffolds or chitosan-hyaluronic acid (CS-HA) scaffolds to explore the possibility of glioma stem cells (GSCs)-like cells enrichment, to study the morphology, gene expression, and in vivo tumorigenicity of 3D scaffolds cells, and to compare results to 2D controls. Results showed that glioma cells on both CS and CS-HA scaffolds could form tumor cell spheroids and increased the expression of GSCs biomarkers compared to conventional 2D monolayers. Furthermore, cells in CS-HA scaffolds had higher expression levels of epithelial-to-mesenchymal transition (EMT)-related gene. Specifically, the in vivo tumorigenicity capability of CS-HA scaffold cultured cells was greater than 2D cells or CS scaffold cultured cells. It is indicated that the chemical composition of scaffold plays an important role in the enrichment of CSCs. Our results suggest that CS-HA scaffolds have a better capability to enrich GSCs-like cells and can serve as a simple and effective way to cultivate and enrich CSCs in vitro to support the study of CSCs biology and development of novel anti-cancer therapies. Copyright © 2018 Elsevier Inc. All rights reserved.

Effect of the biodegradation rate controlled by pore structures in magnesium phosphate ceramic scaffolds on bone tissue regeneration in vivo.

PubMed

Kim, Ju-Ang; Lim, Jiwon; Naren, Raja; Yun, Hui-Suk; Park, Eui Kyun

2016-10-15

P) scaffold by combination of the 3D printing process, self-setting reaction and salt-leaching technique, and first studied the effect of pore structures of bioceramic scaffolds on bone tissue regeneration through both in vitro and in vivo studies (rabbit calvarial model). The MgP scaffolds with higher porosity promoted more rapid biodegradation and enhanced new bone formation and remodeling activities at the same time. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Highly porous 3D nanofiber scaffold using an electrospinning technique.

PubMed

Kim, Geunhyung; Kim, WanDoo

2007-04-01

A successful 3D tissue-engineering scaffold must have a highly porous structure and good mechanical stability. High porosity and optimally designed pore size provide structural space for cell accommodation and migration and enable the exchange of nutrients between the scaffold and environment. Poly(epsilon-carprolactone) fibers were electrospun using an auxiliary electrode and chemical blowing agent (BA), and characterized according to porosity, pore size, and their mechanical properties. We also investigated the effect of the BA on the electrospinning processability. The growth characteristic of human dermal fibroblasts cells cultured in the webs showed the good adhesion with the blown web relative to a normal electrospun mat. The blown nanofiber web had good tensile properties and high porosity compared to a typical electrospun nanofiber scaffold. (c) 2006 Wiley Periodicals, Inc.

3D printing scaffold coupled with low level light therapy for neural tissue regeneration.

PubMed

Zhu, Wei; George, Jonathan K; Sorger, Volker J; Grace Zhang, Lijie

2017-04-12

3D printing has shown promise for neural regeneration by providing customized nerve scaffolds to structurally support and bridge the defect gap as well as deliver cells or various bioactive substances. Low-level light therapy (LLLT) exhibits positive effects on rehabiliation of degenerative nerves and neural disorders. With this in mind, we postulate that 3D printed neural scaffold coupling with LLLT will generate a new strategy to repair neural degeneration. To achieve this goal, we applied red laser light to stimualte neural stem cells on 3D printed scaffolds and investigated the subsequent cell response with respect to cell proliferation and differentiation. Here we show that cell prolifeartion rate and intracellular reactive oxgen species synthesis were significantly increased after 15 s laser stimulation follwed by 1 d culture. Over culturing time of 14 d in vitro, the laser stimulation promoted neuronal differentiation of neural stem cells, while the glial differentiation was suppressed based on results of both immunocytochemistry studies and real-time quantitative reverse transcription polymerase chain reaction testing. These findings suggest that integration of 3D printing and LLLT might provide a powerful methodology for neural tissue engineering.

Water-based polyurethane 3D printed scaffolds with controlled release function for customized cartilage tissue engineering.

PubMed

Hung, Kun-Che; Tseng, Ching-Shiow; Dai, Lien-Guo; Hsu, Shan-hui

2016-03-01

Conventional 3D printing may not readily incorporate bioactive ingredients for controlled release because the process often involves the use of heat, organic solvent, or crosslinkers that reduce the bioactivity of the ingredients. Water-based 3D printing materials with controlled bioactivity for customized cartilage tissue engineering is developed in this study. The printing ink contains the water dispersion of synthetic biodegradable polyurethane (PU) elastic nanoparticles, hyaluronan, and bioactive ingredients TGFβ3 or a small molecule drug Y27632 to replace TGFβ3. Compliant scaffolds are printed from the ink at low temperature. These scaffolds promote the self-aggregation of mesenchymal stem cells (MSCs) and, with timely release of the bioactive ingredients, induce the chondrogenic differentiation of MSCs and produce matrix for cartilage repair. Moreover, the growth factor-free controlled release design may prevent cartilage hypertrophy. Rabbit knee implantation supports the potential of the novel 3D printing scaffolds in cartilage regeneration. We consider that the 3D printing composite scaffolds with controlled release bioactivity may have potential in customized tissue engineering. Copyright © 2016 Elsevier Ltd. All rights reserved.

Electrodeposition on nanofibrous polymer scaffolds: Rapid mineralization, tunable calcium phosphate composition and topography

PubMed Central

He, Chuanglong; Xiao, Guiyong; Jin, Xiaobing; Sun, Chenghui; Ma, Peter X.

2011-01-01

We developed a straightforward, fast, and versatile technique to fabricate mineralized nanofibrous polymer scaffolds for bone regeneration in this work. Nanofibrous poly(l-lactic acid) scaffolds were fabricated using both electrospinning and phase separation techniques. An electrodeposition process was designed to deposit calcium phosphate on the nanofibrous scaffolds. Such scaffolds contain a high quality mineral coating on the fiber surface with tunable surface topography and chemical composition by varying the processing parameters, which can mimic the composition and structure of natural bone extracellular matrix and provide a more biocompatible interface for bone regeneration. PMID:21673827

Enhancement of viability of muscle precursor cells on 3D scaffold in a perfusion bioreactor.

PubMed

Cimetta, E; Flaibani, M; Mella, M; Serena, E; Boldrin, L; De Coppi, P; Elvassore, N

2007-05-01

The aim of this study was to develop a methodology for the in vitro expansion of skeletal-muscle precursor cells (SMPC) in a three-dimensional (3D) environment in order to fabricate a cellularized artificial graft characterized by high density of viable cells and uniform cell distribution over the entire 3D domain. Cell seeding and culture within 3D porous scaffolds by conventional static techniques can lead to a uniform cell distribution only on the scaffold surface, whereas dynamic culture systems have the potential of allowing a uniform growth of SMPCs within the entire scaffold structure. In this work, we designed and developed a perfusion bioreactor able to ensure long-term culture conditions and uniform flow of medium through 3D collagen sponges. A mathematical model to assist the design of the experimental setup and of the operative conditions was developed. The effects of dynamic vs static culture in terms of cell viability and spatial distribution within 3D collagen scaffolds were evaluated at 1, 4 and 7 days and for different flow rates of 1, 2, 3.5 and 4.5 ml/min using C2C12 muscle cell line and SMPCs derived from satellite cells. C2C12 cells, after 7 days of culture in our bioreactor, perfused applying a 3.5 ml/min flow rate, showed a higher viability resulting in a three-fold increase when compared with the same parameter evaluated for cultures kept under static conditions. In addition, dynamic culture resulted in a more uniform 3D cell distribution. The 3.5 ml/min flow rate in the bioreactor was also applied to satellite cell-derived SMPCs cultured on 3D collagen scaffolds. The dynamic culture conditions improved cell viability leading to higher cell density and uniform distribution throughout the entire 3D collagen sponge for both C2C12 and satellite cells.

Chitosan-g-lactide copolymers for fabrication of 3D scaffolds for tissue engineering

NASA Astrophysics Data System (ADS)

Demina, T. S.; Zaytseva-Zotova, D. S.; Timashev, P. S.; Bagratashvili, V. N.; Bardakova, K. N.; Sevrin, Ch; Svidchenko, E. A.; Surin, N. M.; Markvicheva, E. A.; Grandfils, Ch; Akopova, T. A.

2015-07-01

Chitosan-g-oligo (L, D-lactide) copolymers were synthesized and assessed to fabricate a number of 3D scaffolds using a variety of technologies such as oil/water emulsion evaporation technique, freeze-drying and two-photon photopolymerization. Solid-state copolymerization method allowed us to graft up to 160 wt-% of oligolactide onto chitosan backbone via chitosan amino group acetylation with substitution degree reaching up to 0.41. Grafting of hydrophobic oligolactide side chains with polymerization degree up to 10 results in chitosan amphiphilic properties. The synthesized chitosan-g-lactide copolymers were used to design 3D scaffolds for tissue engineering such as spherical microparticles and macroporous hydrogels.

3D-printed PCL scaffolds for the cultivation of mesenchymal stem cells.

PubMed

Steffens, Daniela; Rezende, Rodrigo Alvarenga; Santi, Bruna; Pereira, Frederico David Alencar de Sena; Inforçatti Neto, Paulo; da Silva, Jorge Vicente Lopes; Pranke, Patricia

2016-04-06

Tissue engineering is a field which is currently under a great deal of investigation for the development and/or restoration of tissue and organs, through the combination of cell therapy with biomaterials. Rapid prototyping or additive manufacturing is a versatile technology which makes possible the fabrication of three dimensional (3D) structures from a wide range of materials with complex geometry and accuracy, such as scaffolds. The aim of this study has been to investigate the interaction between mesenchymal stem cells with poly (ε-caprolactone) (PCL) biomaterials used for obtaining scaffolds through additive manufacturing. Scanning electron microscopy, confocal microscopy and biological assays were performed to analyse the successful interaction between the cells and the biomaterials. As a result, the number of viable cells attached to the scaffolds was lower when compared to the control group; however, it was possible to observe cells in the scaffolds since day 1 of analysis, with regions of confluence after 21 days of seeding. To conclude, these biomaterials are interesting if used as medical artifacts, principally in tissue with prolonged regeneration time and which requires 3D supports with good mechanical properties.

Realization and testing of multi-material 3D printer for bone scaffold fabrication

NASA Astrophysics Data System (ADS)

Whulanza, Yudan; Hidayaturrahmi, Pretty; Kurniawati, Tri; AJ, Rahyussalim

2017-02-01

This research realized 3D constructs by integrating more than one material with multi fabrication system within a single session. A commercial rapid prototyping system, RepRap MendelTM, is modified so that it enables us to realize microenvironment composed of multi materials namely gelatin hydrogel and polylactic acid. Firstly, the session is preceded by realization of 3D scaffold using polylactic acid (PLA) with porosity and modulus elasticity as characterized. Later, the gelatin extrusion took place to seed the cellular in determined spatial arrangement. The results show that our apparatus able to realized scaffold that using PLA as matrix filled with gelatin that act as cell carrier in future application. The scaffolds have porous around 0.25 mm2 porosity with a modulus of elasticity around 160 MPa.

Design, construction and mechanical testing of digital 3D anatomical data-based PCL-HA bone tissue engineering scaffold.

PubMed

Yao, Qingqiang; Wei, Bo; Guo, Yang; Jin, Chengzhe; Du, Xiaotao; Yan, Chao; Yan, Junwei; Hu, Wenhao; Xu, Yan; Zhou, Zhi; Wang, Yijin; Wang, Liming

2015-01-01

The study aims to investigate the techniques of design and construction of CT 3D reconstructional data-based polycaprolactone (PCL)-hydroxyapatite (HA) scaffold. Femoral and lumbar spinal specimens of eight male New Zealand white rabbits were performed CT and laser scanning data-based 3D printing scaffold processing using PCL-HA powder. Each group was performed eight scaffolds. The CAD-based 3D printed porous cylindrical stents were 16 piece × 3 groups, including the orthogonal scaffold, the Pozi-hole scaffold and the triangular hole scaffold. The gross forms, fiber scaffold diameters and porosities of the scaffolds were measured, and the mechanical testing was performed towards eight pieces of the three kinds of cylindrical scaffolds, respectively. The loading force, deformation, maximum-affordable pressure and deformation value were recorded. The pore-connection rate of each scaffold was 100 % within each group, there was no significant difference in the gross parameters and micro-structural parameters of each scaffold when compared with the design values (P > 0.05). There was no significant difference in the loading force, deformation and deformation value under the maximum-affordable pressure of the three different cylinder scaffolds when the load was above 320 N. The combination of CT and CAD reverse technology could accomplish the design and manufacturing of complex bone tissue engineering scaffolds, with no significant difference in the impacts of the microstructures towards the physical properties of different porous scaffolds under large load.

Exosomes/tricalcium phosphate combination scaffolds can enhance bone regeneration by activating the PI3K/Akt signaling pathway.

PubMed

Zhang, Jieyuan; Liu, Xiaolin; Li, Haiyan; Chen, Chunyuan; Hu, Bin; Niu, Xin; Li, Qing; Zhao, Bizeng; Xie, Zongping; Wang, Yang

2016-09-20

Recently, accumulating evidence has shown that exosomes, the naturally secreted nanocarriers of cells, can exert therapeutic effects in various disease models in the absence of parent cells. However, application of exosomes in bone defect repair and regeneration has been rarely reported, and little is known regarding their underlying mechanisms. Exosomes derived from human-induced pluripotent stem cell-derived mesenchymal stem cells (hiPS-MSC-Exos) were combined with tricalcium phosphate (β-TCP) to repair critical-sized calvarial bone defects, and the efficacy was assessed by histological examination. We evaluated the in vitro effects of hiPSC-MSC-Exos on the proliferation, migration, and osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hBMSCs) by cell-counting, scratch assays, and qRT-PCR, respectively. Gene expression profiling and bioinformatics analyses were also used to identify the underlying mechanisms in the repair. We found that the exosome/β-TCP combination scaffolds could enhance osteogenesis as compared to pure β-TCP scaffolds. In vitro assays showed that the exosomes could release from β-TCP and could be internalized by hBMSCs. In addition, the internalization of exosomes into hBMSCs could profoundly enhance the proliferation, migration, and osteogenic differentiation of hBMSCs. Furthermore, gene expression profiling and bioinformatics analyses demonstrated that exosome/β-TCP combination scaffolds significantly altered the expression of a network of genes involved in the PI3K/Akt signaling pathway. Functional studies further confirmed that the PI3K/Akt signaling pathway was the critical mediator during the exosome-induced osteogenic responses of hBMSCs. We propose that the exosomes can enhance the osteoinductivity of β-TCP through activating the PI3K/Akt signaling pathway of hBMSCs, which means that the exosome/β-TCP combination scaffolds possess better osteogenesis activity than pure β-TCP scaffolds. These

Novel resorbable glass-ceramic scaffolds for hard tissue engineering: from the parent phosphate glass to its bone-like macroporous derivatives.

PubMed

Bretcanu, Oana; Baino, Francesco; Verné, Enrica; Vitale-Brovarone, Chiara

2014-05-01

One of the major challenges of hard tissue engineering research focuses on the development of scaffolds that can match the mechanical properties of the host bone and resorb at the same rate as the bone is repaired. The aim of this work was the synthesis and characterization of a resorbable phosphate glass, as well as its application for the fabrication of three dimensional (3-D) scaffolds for bone regeneration. The glass microstructure and behaviour upon heating were analysed by X-ray diffraction, differential scanning calorimetry and hot stage microscopy. The glass solubility was investigated according to relevant ISO standards using distilled water, simulated body fluid (SBF) and Tris-HCl as testing media. The glass underwent progressive dissolution over time in all three media but the formation of a hydroxyapatite-like layer was also observed on the samples soaked in SBF and Tris-HCl, which demonstrated the bioactivity of the material. The glass powder was used to fabricate 3-D macroporous bone-like glass-ceramic scaffolds by adopting polyethylene particles as pore formers: during thermal treatment, the polymer additive was removed and the sintering of glass particles was allowed. The obtained scaffolds exhibited high porosity (87 vol.%) and compressive strength around 1.5 MPa. After soaking for 4 months in SBF, the scaffolds mass loss was 76 wt.% and the pH of the solution did not exceed the 7.55 value, thereby remaining in a physiological range. The produced scaffolds, being resorbable, bioactive, architecturally similar to trabecular bone and exhibiting interesting mechanical properties, can be proposed as promising candidates for bone repair applications.

C 3-symmetric opioid scaffolds are pH-responsive DNA condensation agents

PubMed Central

McStay, Natasha; Molphy, Zara; Coughlan, Alan; Cafolla, Attilio; McKee, Vickie; Gathergood, Nicholas; Kellett, Andrew

2017-01-01

Herein we report the synthesis of tripodal C3-symmetric opioid scaffolds as high-affinity condensation agents of duplex DNA. Condensation was achieved on both supercoiled and canonical B-DNA structures and identified by agarose electrophoresis, viscosity, turbidity and atomic force microscopy (AFM) measurements. Structurally, the requirement of a tris-opioid scaffold for condensation is demonstrated as both di- (C2-symmetric) and mono-substituted (C1-symmetric) mesitylene-linked opioid derivatives poorly coordinate dsDNA. Condensation, observed by toroidal and globule AFM aggregation, arises from surface-binding ionic interactions between protonated, cationic, tertiary amine groups on the opioid skeleton and the phosphate nucleic acid backbone. Indeed, by converting the 6-hydroxyl group of C3-morphine (MC3) to methoxy substituents in C3-heterocodeine (HC3) and C3-oripavine (OC3) molecules, dsDNA compaction is retained thus negating the possibility of phosphate—hydroxyl surface-binding. Tripodal opioid condensation was identified as pH dependent and strongly influenced by ionic strength with further evidence of cationic amine-phosphate backbone coordination arising from thermal melting analysis and circular dichroism spectroscopy, with compaction also witnessed on synthetic dsDNA co-polymers poly[d(A-T)2] and poly[d(G-C)2]. On-chip microfluidic analysis of DNA condensed by C3-agents provided concentration-dependent protection (inhibition) to site-selective excision by type II restriction enzymes: BamHI, HindIII, SalI and EcoRI, but not to the endonuclease DNase I. PMID:27899572

3D printed scaffolds of calcium silicate-doped β-TCP synergize with co-cultured endothelial and stromal cells to promote vascularization and bone formation.

PubMed

Deng, Yuan; Jiang, Chuan; Li, Cuidi; Li, Tao; Peng, Mingzheng; Wang, Jinwu; Dai, Kerong

2017-07-17

Synthetic bone scaffolds have potential application in repairing large bone defects, however, inefficient vascularization after implantation remains the major issue of graft failure. Herein, porous β-tricalcium phosphate (β-TCP) scaffolds with calcium silicate (CS) were 3D printed, and pre-seeded with co-cultured human umbilical cord vein endothelial cells (HUVECs) and human bone marrow stromal cells (hBMSCs) to construct tissue engineering scaffolds with accelerated vascularization and better bone formation. Results showed that in vitro β-TCP scaffolds doped with 5% CS (5%CS/β-TCP) were biocompatible, and stimulated angiogenesis and osteogenesis. The results also showed that 5%CS/β-TCP scaffolds not only stimulated co-cultured cells angiogenesis on Matrigel, but also stimulated co-cultured cells to form microcapillary-like structures on scaffolds, and promoted migration of BMSCs by stimulating co-cultured cells to secrete PDGF-BB and CXCL12 into the surrounding environment. Moreover, 5%CS/β-TCP scaffolds enhanced vascularization and osteoinduction in comparison with β-TCP, and synergized with co-cultured cells to further increase early vessel formation, which was accompanied by earlier and better ectopic bone formation when implanted subcutaneously in nude mice. Thus, our findings suggest that porous 5%CS/β-TCP scaffolds seeded with co-cultured cells provide new strategy for accelerating tissue engineering scaffolds vascularization and osteogenesis, and show potential as treatment for large bone defects.

Micropore-induced capillarity enhances bone distribution in vivo in biphasic calcium phosphate scaffolds.

PubMed

Rustom, Laurence E; Boudou, Thomas; Lou, Siyu; Pignot-Paintrand, Isabelle; Nemke, Brett W; Lu, Yan; Markel, Mark D; Picart, Catherine; Wagoner Johnson, Amy J

2016-10-15

The increasing demand for bone repair solutions calls for the development of efficacious bone scaffolds. Biphasic calcium phosphate (BCP) scaffolds with both macropores and micropores (MP) have improved healing compared to those with macropores and no micropores (NMP), but the role of micropores is unclear. Here, we evaluate capillarity induced by micropores as a mechanism that can affect bone growth in vivo. Three groups of cylindrical scaffolds were implanted in pig mandibles for three weeks: MP were implanted either dry (MP-Dry), or after submersion in phosphate buffered saline, which fills pores with fluid and therefore suppresses micropore-induced capillarity (MP-Wet); NMP were implanted dry. The amount and distribution of bone in the scaffolds were quantified using micro-computed tomography. MP-Dry had a more homogeneous bone distribution than MP-Wet, although the average bone volume fraction, BVF‾, was not significantly different for these two groups (0.45±0.03 and 0.37±0.03, respectively). There was no significant difference in the radial bone distribution of NMP and MP-Wet, but the BVF‾, of NMP was significantly lower among the three groups (0.25±0.02). These results suggest that micropore-induced capillarity enhances bone regeneration by improving the homogeneity of bone distribution in BCP scaffolds. The explicit design and use of capillarity in bone scaffolds may lead to more effective treatments of large and complex bone defects. The increasing demand for bone repair calls for more efficacious bone scaffolds and calcium phosphate-based materials are considered suitable for this application. Macropores (>100μm) are necessary for bone ingrowth and vascularization. However, studies have shown that microporosity (<20μm) also enhances growth, but there is no consensus on the controlling mechanisms. In previous in vitro work, we suggested that micropore-induced capillarity had the potential to enhance bone growth in vivo. This work illustrates the

Impact of Particle Size of Ceramic Granule Blends on Mechanical Strength and Porosity of 3D Printed Scaffolds

PubMed Central

Spath, Sebastian; Drescher, Philipp; Seitz, Hermann

2015-01-01

3D printing is a promising method for the fabrication of scaffolds in the field of bone tissue engineering. To date, the mechanical strength of 3D printed ceramic scaffolds is not sufficient for a variety of applications in the reconstructive surgery. Mechanical strength is directly in relation with the porosity of the 3D printed scaffolds. The porosity is directly influenced by particle size and particle-size distribution of the raw material. To investigate this impact, a hydroxyapatite granule blend with a wide particle size distribution was fractioned by sieving. The specific fractions and bimodal mixtures of the sieved granule blend were used to 3D print specimens. It has been shown that an optimized arrangement of fractions with large and small particles can provide 3D printed specimens with good mechanical strength due to a higher packing density. An increase of mechanical strength can possibly expand the application area of 3D printed hydroxyapatite scaffolds. PMID:28793467

Efficacy of rhBMP-2 Loaded PCL/β-TCP/bdECM Scaffold Fabricated by 3D Printing Technology on Bone Regeneration.

PubMed

Bae, Eun-Bin; Park, Keun-Ho; Shim, Jin-Hyung; Chung, Ho-Yun; Choi, Jae-Won; Lee, Jin-Ju; Kim, Chang-Hwan; Jeon, Ho-Jun; Kang, Seong-Soo; Huh, Jung-Bo

2018-01-01

This study was undertaken to evaluate the effect of 3D printed polycaprolactone (PCL)/ β -tricalcium phosphate ( β -TCP) scaffold containing bone demineralized and decellularized extracellular matrix (bdECM) and human recombinant bone morphogenetic protein-2 (rhBMP-2) on bone regeneration. Scaffolds were divided into PCL/ β -TCP, PCL/ β -TCP/bdECM, and PCL/ β -TCP/bdECM/BMP groups. In vitro release kinetics of rhBMP-2 were determined with respect to cell proliferation and osteogenic differentiation. These three reconstructive materials were implanted into 8 mm diameter calvarial bone defect in male Sprague-Dawley rats. Animals were sacrificed four weeks after implantation for micro-CT, histologic, and histomorphometric analyses. The findings obtained were used to calculate new bone volumes (mm 3 ) and new bone areas (%). Excellent cell bioactivity was observed in the PCL/ β -TCP/bdECM and PCL/ β -TCP/bdECM/BMP groups, and new bone volume and area were significantly higher in the PCL/ β -TCP/bdECM/BMP group than in the other groups ( p < .05). Within the limitations of this study, bdECM printed PCL/ β -TCP scaffolds can reproduce microenvironment for cells and promote adhering and proliferating the cells onto scaffolds. Furthermore, in the rat calvarial defect model, the scaffold which printed rhBMP-2 loaded bdECM stably carries rhBMP-2 and enhances bone regeneration confirming the possibility of bdECM as rhBMP-2 carrier.

In vivo bone formation by human marrow stromal cells in biodegradable scaffolds that release dexamethasone and ascorbate-2-phosphate.

PubMed

Kim, Hyongbum; Suh, Hwal; Jo, Sangmee Ahn; Kim, Hyun Woo; Lee, Jung Min; Kim, Eun Hae; Reinwald, Yvonne; Park, Sang-Hyug; Min, Byoung-Hyun; Jo, Inho

2005-07-15

An unsolved problem with stem cell-based engineering of bone tissue is how to provide a microenvironment that promotes the osteogenic differentiation of multipotent stem cells. Previously, we fabricated porous poly(D,L-lactide-co-glycolide) (PLGA) scaffolds that released biologically active dexamethasone (Dex) and ascorbate-2-phosphate (AsP), and that acted as osteogenic scaffolds. To determine whether these osteogenic scaffolds can be used for bone formation in vivo, we seeded multipotent human marrow stromal cells (hMSCs) onto the scaffolds and implanted them subcutaneously into athymic mice. Higher alkaline phosphatase expression was observed in hMSCs in the osteogenic scaffolds compared with that of hMSCs in control scaffolds. Furthermore, there was more calcium deposition and stronger von Kossa staining in the osteogenic scaffolds, which suggested that there was enhanced mineralized bone formation. We failed to detect cartilage in the osteogenic scaffolds (negative Safranin O staining), which implied that there was intramembranous ossification. This is the first study to demonstrate the successful formation of mineralized bone tissue in vivo by hMSCs in PLGA scaffolds that release Dex and AsP.

Effect of pore architecture on oxygen diffusion in 3D scaffolds for tissue engineering.

PubMed

Ahn, Geunseon; Park, Jeong Hun; Kang, Taeyun; Lee, Jin Woo; Kang, Hyun-Wook; Cho, Dong-Woo

2010-10-01

The aim of this study was to maximize oxygen diffusion within a three-dimensional scaffold in order to improve cell viability and proliferation. To evaluate the effect of pore architecture on oxygen diffusion, we designed a regular channel shape with uniform diameter, referred to as cylinder shaped, and a new channel shape with a channel diameter gradient, referred to as cone shaped. A numerical analysis predicted higher oxygen concentration in the cone-shaped channels than in the cylinder-shaped channels, throughout the scaffold. To confirm these numerical results, we examined cell proliferation and viability in 2D constructs and 3D scaffolds. Cell culture experiments revealed that cell proliferation and viability were superior in the constructs and scaffolds with cone-shaped channels.

Preparation of 3D fibroin/chitosan blend porous scaffold for tissue engineering via a simplified method.

PubMed

Ruan, Yuhui; Lin, Hong; Yao, Jinrong; Chen, Zhengrong; Shao, Zhengzhong

2011-03-10

In this work, we developed a simple and flexible method to manufacture a 3D porous scaffold based on the blend of regenerated silk fibroin (RSF) and chitosan (CS). No crosslinker or other toxic reagents were used in this method. The pores of resulted 3D scaffolds were connected with each other, and their sizes could be easily controlled by the concentration of the mixed solution. Compared with pure RSF scaffolds, the water absorptivities of these RSF/CS blend scaffolds with significantly enhanced mechanical properties were greatly increased. The results of MTT and RT-PCR tests indicated that the chondrocytes grew very well in these blend RSF/CS porous scaffolds. This suggested that the RSF/CS blend scaffold prepared by this new method could be a promising candidate for applications in tissue engineering. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

3D printed alendronate-releasing poly(caprolactone) porous scaffolds enhance osteogenic differentiation and bone formation in rat tibial defects.

PubMed

Kim, Sung Eun; Yun, Young-Pil; Shim, Kyu-Sik; Kim, Hak-Jun; Park, Kyeongsoon; Song, Hae-Ryong

2016-09-29

The aim of this study was to evaluate the in vitro osteogenic effects and in vivo new bone formation of three-dimensional (3D) printed alendronate (Aln)-releasing poly(caprolactone) (PCL) (Aln/PCL) scaffolds in rat tibial defect models. 3D printed Aln/PCL scaffolds were fabricated via layer-by-layer deposition. The fabricated Aln/PCL scaffolds had high porosity and an interconnected pore structure and showed sustained Aln release. In vitro studies showed that MG-63 cells seeded on the Aln/PCL scaffolds displayed increased alkaline phosphatase (ALP) activity and calcium content in a dose-dependent manner when compared with cell cultures in PCL scaffolds. In addition, in vivo animal studies and histologic evaluation showed that Aln/PCL scaffolds implanted in a rat tibial defect model markedly increased new bone formation and mineralized bone tissues in a dose-dependent manner compared to PCL-only scaffolds. Our results show that 3D printed Aln/PCL scaffolds are promising templates for bone tissue engineering applications.

3D printing facilitated scaffold-free tissue unit fabrication.

PubMed

Tan, Yu; Richards, Dylan J; Trusk, Thomas C; Visconti, Richard P; Yost, Michael J; Kindy, Mark S; Drake, Christopher J; Argraves, William Scott; Markwald, Roger R; Mei, Ying

2014-06-01

Tissue spheroids hold great potential in tissue engineering as building blocks to assemble into functional tissues. To date, agarose molds have been extensively used to facilitate fusion process of tissue spheroids. As a molding material, agarose typically requires low temperature plates for gelation and/or heated dispenser units. Here, we proposed and developed an alginate-based, direct 3D mold-printing technology: 3D printing microdroplets of alginate solution into biocompatible, bio-inert alginate hydrogel molds for the fabrication of scaffold-free tissue engineering constructs. Specifically, we developed a 3D printing technology to deposit microdroplets of alginate solution on calcium containing substrates in a layer-by-layer fashion to prepare ring-shaped 3D hydrogel molds. Tissue spheroids composed of 50% endothelial cells and 50% smooth muscle cells were robotically placed into the 3D printed alginate molds using a 3D printer, and were found to rapidly fuse into toroid-shaped tissue units. Histological and immunofluorescence analysis indicated that the cells secreted collagen type I playing a critical role in promoting cell-cell adhesion, tissue formation and maturation.

3D printed hyperelastic "bone" scaffolds and regional gene therapy: A novel approach to bone healing.

PubMed

Alluri, Ram; Jakus, Adam; Bougioukli, Sofia; Pannell, William; Sugiyama, Osamu; Tang, Amy; Shah, Ramille; Lieberman, Jay R

2018-04-01

The purpose of this study was to evaluate the viability of human adipose-derived stem cells (ADSCs) transduced with a lentiviral (LV) vector to overexpress bone morphogenetic protein-2 (BMP-2) loaded onto a novel 3D printed scaffold. Human ADSCs were transduced with a LV vector carrying the cDNA for BMP-2. The transduced cells were loaded onto a 3D printed Hyperelastic "Bone" (HB) scaffold. In vitro BMP-2 production was assessed using enzyme-linked immunosorbent assay analysis. The ability of ADSCs loaded on the HB scaffold to induce in vivo bone formation in a hind limb muscle pouch model was assessed in the following groups: ADSCs transduced with LV-BMP-2, LV-green fluorescent protein, ADSCs alone, and empty HB scaffolds. Bone formation was assessed using radiographs, histology and histomorphometry. Transduced ADSCs BMP-2 production on the HB scaffold at 24 hours was similar on 3D printed HB scaffolds versus control wells with transduced cells alone, and continued to increase after 1 and 2 weeks of culture. Bone formation was noted in LV-BMP-2 animals on plain radiographs at 2 and 4 weeks after implantation; no bone formation was noted in the other groups. Histology demonstrated that the LV-BMP-2 group was the only group that formed woven bone and the mean bone area/tissue area was significantly greater when compared with the other groups. 3D printed HB scaffolds are effective carriers for transduced ADSCs to promote bone repair. The combination of gene therapy and tissue engineered scaffolds is a promising multidisciplinary approach to bone repair with significant clinical potential. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1104-1110, 2018. © 2018 Wiley Periodicals, Inc.

Engineering anatomically shaped vascularized bone grafts with hASCs and 3D-printed PCL scaffolds.

PubMed

Temple, Joshua P; Hutton, Daphne L; Hung, Ben P; Huri, Pinar Yilgor; Cook, Colin A; Kondragunta, Renu; Jia, Xiaofeng; Grayson, Warren L

2014-12-01

The treatment of large craniomaxillofacial bone defects is clinically challenging due to the limited availability of transplantable autologous bone grafts and the complex geometry of the bones. The ability to regenerate new bone tissues that faithfully replicate the anatomy would revolutionize treatment options. Advances in the field of bone tissue engineering over the past few decades offer promising new treatment alternatives using biocompatible scaffold materials and autologous cells. This approach combined with recent advances in three-dimensional (3D) printing technologies may soon allow the generation of large, bioartificial bone grafts with custom, patient-specific architecture. In this study, we use a custom-built 3D printer to develop anatomically shaped polycaprolactone (PCL) scaffolds with varying internal porosities. These scaffolds are assessed for their ability to support induction of human adipose-derived stem cells (hASCs) to form vasculature and bone, two essential components of functional bone tissue. The development of functional tissues is assessed in vitro and in vivo. Finally, we demonstrate the ability to print large mandibular and maxillary bone scaffolds that replicate fine details extracted from patient's computed tomography scans. The findings of this study illustrate the capabilities and potential of 3D printed scaffolds to be used for engineering autologous, anatomically shaped, vascularized bone grafts. © 2014 Wiley Periodicals, Inc.

Degradability, biocompatibility, and osteogenesis of biocomposite scaffolds containing nano magnesium phosphate and wheat protein both in vitro and in vivo for bone regeneration.

PubMed

Xia, Yan; Zhou, Panyu; Wang, Fei; Qiu, Chao; Wang, Panfeng; Zhang, Yuntong; Zhao, Liming; Xu, Shuogui

2016-01-01

In this study, bioactive scaffold of nano magnesium phosphate (nMP)/wheat protein (WP) composite (MWC) was fabricated. The results revealed that the MWC scaffolds had interconnected not only macropores (sized 400-600 μm) but also micropores (sized 10-20 μm) on the walls of macropores. The MWC scaffolds containing 40 w% nMP had an appropriate degradability in phosphate-buffered saline and produced a weak alkaline microenvironment. In cell culture experiments, the results revealed that the MWC scaffolds significantly promoted the MC3T3-E1 cell proliferation, differentiation, and growth into the scaffolds. The results of synchrotron radiation microcomputed tomography and analysis of the histological sections of the in vivo implantation revealed that the MWC scaffolds evidently improved the new bone formation and bone defects repair as compared with WP scaffolds. Moreover, it was found that newly formed bone tissue continued to increase with the gradual reduction of materials residual in the MWC scaffolds. Furthermore, the immunohistochemical analysis further offered the evidence of the stimulatory effects of MWC scaffolds on osteogenic-related cell differentiation and new bone regeneration. The results indicated that MWC scaffolds with good biocompability and degradability could promote osteogenesis in vivo, which would have potential for bone tissue repair.

Cancer cell migration within 3D layer-by-layer microfabricated photocrosslinked PEG scaffolds with tunable stiffness.

PubMed

Soman, Pranav; Kelber, Jonathan A; Lee, Jin Woo; Wright, Tracy N; Vecchio, Kenneth S; Klemke, Richard L; Chen, Shaochen

2012-10-01

Our current understanding of 3-dimensional (3D) cell migration is primarily based on results from fibrous scaffolds with randomly organized internal architecture. Manipulations that change the stiffness of these 3D scaffolds often alter other matrix parameters that can modulate cell motility independently or synergistically, making observations less predictive of how cells behave when migrating in 3D. In order to decouple microstructural influences and stiffness effects, we have designed and fabricated 3D polyethylene glycol (PEG) scaffolds that permit orthogonal tuning of both elastic moduli and microstructure. Scaffolds with log-pile architectures were used to compare the 3D migration properties of normal breast epithelial cells (HMLE) and Twist-transformed cells (HMLET). Our results indicate that the nature of cell migration is significantly impacted by the ability of cells to migrate in the third dimension. 2D ECM-coated PEG substrates revealed no statistically significant difference in cell migration between HMLE and HMLET cells among substrates of different stiffness. However, when cells were allowed to move along the third dimension, substantial differences were observed for cell displacement, velocity and path straightness parameters. Furthermore, these differences were sensitive to both substrate stiffness and the presence of the Twist oncogene. Importantly, these 3D modes of migration provide insight into the potential for oncogene-transformed cells to migrate within and colonize tissues of varying stiffness. Copyright © 2012 Elsevier Ltd. All rights reserved.

Polymer-Ceramic Composite Scaffolds: The Effect of Hydroxyapatite and β-tri-Calcium Phosphate

PubMed Central

Caetano, Guilherme; Vyas, Cian; Diver, Carl; Bártolo, Paulo

2018-01-01

The design of bioactive scaffolds with improved mechanical and biological properties is an important topic of research. This paper investigates the use of polymer-ceramic composite scaffolds for bone tissue engineering. Different ceramic materials (hydroxyapatite (HA) and β-tri-calcium phosphate (TCP)) were mixed with poly-ε-caprolactone (PCL). Scaffolds with different material compositions were produced using an extrusion-based additive manufacturing system. The produced scaffolds were physically and chemically assessed, considering mechanical, wettability, scanning electron microscopy and thermal gravimetric tests. Cell viability, attachment and proliferation tests were performed using human adipose derived stem cells (hADSCs). Results show that scaffolds containing HA present better biological properties and TCP scaffolds present improved mechanical properties. It was also possible to observe that the addition of ceramic particles had no effect on the wettability of the scaffolds. PMID:29342890

Application of quality by design for 3D printed bone prostheses and scaffolds

PubMed Central

Martinez-Marquez, Daniel; Mirnajafizadeh, Ali; Carty, Christopher P.

2018-01-01

3D printing is an emergent manufacturing technology recently being applied in the medical field for the development of custom bone prostheses and scaffolds. However, successful industry transformation to this new design and manufacturing approach requires technology integration, concurrent multi-disciplinary collaboration, and a robust quality management framework. This latter change enabler is the focus of this study. While a number of comprehensive quality frameworks have been developed in recent decades to ensure that the manufacturing of medical devices produces reliable products, they are centred on the traditional context of standardised manufacturing techniques. The advent of 3D printing technologies and the prospects for mass customisation provides significant market opportunities, but also presents a serious challenge to regulatory bodies tasked with managing and assuring product quality and safety. Before 3D printing bone prostheses and scaffolds can gain traction, industry stakeholders, such as regulators, clients, medical practitioners, insurers, lawyers, and manufacturers, would all require a high degree of confidence that customised manufacturing can achieve the same quality outcomes as standardised manufacturing. A Quality by Design (QbD) approach to custom 3D printed prostheses can help to ensure that products are designed and manufactured correctly from the beginning without errors. This paper reports on the adaptation of the QbD approach for the development process of 3D printed custom bone prosthesis and scaffolds. This was achieved through the identification of the Critical Quality Attributes of such products, and an extensive review of different design and fabrication methods for 3D printed bone prostheses. Research outcomes include the development of a comprehensive design and fabrication process flow diagram, and categorised risks associated with the design and fabrication processes of such products. An extensive systematic literature review

Application of quality by design for 3D printed bone prostheses and scaffolds.

PubMed

Martinez-Marquez, Daniel; Mirnajafizadeh, Ali; Carty, Christopher P; Stewart, Rodney A

2018-01-01

3D printing is an emergent manufacturing technology recently being applied in the medical field for the development of custom bone prostheses and scaffolds. However, successful industry transformation to this new design and manufacturing approach requires technology integration, concurrent multi-disciplinary collaboration, and a robust quality management framework. This latter change enabler is the focus of this study. While a number of comprehensive quality frameworks have been developed in recent decades to ensure that the manufacturing of medical devices produces reliable products, they are centred on the traditional context of standardised manufacturing techniques. The advent of 3D printing technologies and the prospects for mass customisation provides significant market opportunities, but also presents a serious challenge to regulatory bodies tasked with managing and assuring product quality and safety. Before 3D printing bone prostheses and scaffolds can gain traction, industry stakeholders, such as regulators, clients, medical practitioners, insurers, lawyers, and manufacturers, would all require a high degree of confidence that customised manufacturing can achieve the same quality outcomes as standardised manufacturing. A Quality by Design (QbD) approach to custom 3D printed prostheses can help to ensure that products are designed and manufactured correctly from the beginning without errors. This paper reports on the adaptation of the QbD approach for the development process of 3D printed custom bone prosthesis and scaffolds. This was achieved through the identification of the Critical Quality Attributes of such products, and an extensive review of different design and fabrication methods for 3D printed bone prostheses. Research outcomes include the development of a comprehensive design and fabrication process flow diagram, and categorised risks associated with the design and fabrication processes of such products. An extensive systematic literature review

Electrospun conductive nanofibrous scaffolds for engineering cardiac tissue and 3D bioactuators.

PubMed

Wang, Ling; Wu, Yaobin; Hu, Tianli; Guo, Baolin; Ma, Peter X

2017-09-01

Mimicking the nanofibrous structure similar to extracellular matrix and conductivity for electrical propagation of native myocardium would be highly beneficial for cardiac tissue engineering and cardiomyocytes-based bioactuators. Herein, we developed conductive nanofibrous sheets with electrical conductivity and nanofibrous structure composed of poly(l-lactic acid) (PLA) blending with polyaniline (PANI) for cardiac tissue engineering and cardiomyocytes-based 3D bioactuators. Incorporating of varying contents of PANI from 0wt% to 3wt% into the PLA polymer, the electrospun nanofibrous sheets showed enhanced conductivity while maintaining the same fiber diameter. These PLA/PANI conductive nanofibrous sheets exhibited good cell viability and promoting effect on differentiation of H9c2 cardiomyoblasts in terms of maturation index and fusion index. Moreover, PLA/PANI nanofibrous sheets enhanced the cell-cell interaction, maturation and spontaneous beating of primary cardiomyocytes. Furthermore, the cardiomyocytes-laden PLA/PANI conductive nanofibrous sheets can form 3D bioactuators with tubular and folding shapes, and spontaneously beat with much higher frequency and displacement than that on cardiomyocytes-laden PLA nanofibrous sheets. Therefore, these PLA/PANI conductive nanofibrous sheets with conductivity and extracellular matrix like nanostructure demonstrated promising potential in cardiac tissue engineering and cardiomyocytes-based 3D bioactuators. Cardiomyocytes-based bioactuators have been paid more attention due to their spontaneous motion by integrating cardiomyocytes into polymer structures, but developing suitable scaffolds for bioactuators remains challenging. Electrospun nanofibrous scaffolds have been widely used in cardiac tissue engineering because they can mimic the extracellular matrix of myocardium. Developing conductive nanofibrous scaffolds by electrospinning would be beneficial for cardiomyocytes-based bioactuators, but such scaffolds have been

3D porous chitosan scaffolds suit survival and neural differentiation of dental pulp stem cells.

PubMed

Feng, Xingmei; Lu, Xiaohui; Huang, Dan; Xing, Jing; Feng, Guijuan; Jin, Guohua; Yi, Xin; Li, Liren; Lu, Yuanzhou; Nie, Dekang; Chen, Xiang; Zhang, Lei; Gu, Zhifeng; Zhang, Xinhua

2014-08-01

A key aspect of cell replacement therapy in brain injury treatment is construction of a suitable biomaterial scaffold that can effectively carry and transport the therapeutic cells to the target area. In the present study, we created small 3D porous chitosan scaffolds through freeze-drying, and showed that these can support and enhance the differentiation of dental pulp stem cells (DPSCs) to nerve cells in vitro. The DPSCs were collected from the dental pulp of adult human third molars. At a swelling rate of ~84.33 ± 10.92 %, the scaffold displayed high porosity and interconnectivity of pores, as revealed by SEM. Cell counting kit-8 assay established the biocompatibility of the chitosan scaffold, supporting the growth and survival of DPSCs. The successful neural differentiation of DPSCs was assayed by RT-PCR, western blotting, and immunofluorescence. We found that the scaffold-attached DPSCs showed high expression of Nestin that decreased sharply following induction of differentiation. Exposure to the differentiation media also increased the expression of neural molecular markers Microtubule-associated protein 2, glial fibrillary acidic protein, and 2',3'-cyclic nucleotide phosphodiesterase. This study demonstrates that the granular 3D chitosan scaffolds are non-cytotoxic, biocompatible, and provide a conducive and favorable micro-environment for attachment, survival, and neural differentiation of DPSCs. These scaffolds have enormous potential to facilitate future advances in treatment of brain injury.

Vascular Differentiation from Pluripotent Stem Cells in 3-D Auxetic Scaffolds.

PubMed

Song, Liqing; Ahmed, Mohammad Faisal; Li, Yan; Zeng, Changchun; Li, Yan

2018-05-10

Auxetic scaffolds, i.e. scaffolds that can display negative Poisson's ratio, have unique physical properties and can expand transversally when axially strained or contract under compression. Auxetic materials have been used for bioprostheses and artery stents due to the enhanced compressive strength and shear stiffness. In vascular tissue engineering, auxetic scaffolds allow the widening of blood vessels when blood flows through (creating compressive stress) to prevent the blockage. However, the influence of auxetic materials on the cellular fate decision in local environment is unclear. In this study, auxetic polyurethane foams were used to support vascular differentiation from pluripotent stem cells (PSCs). The expression of alkaline phosphatase, Oct-4 and Nanog was lower after four days of differentiation for the cells grown in auxetic scaffolds. Higher expression of vascular markers CD31 and VE-cadherin was observed for the cells from auxetic scaffolds compared to those from the scaffolds before auxetic conversion. Little influence on the expression of cardiac marker α-actinin was observed. The vascular cells secreted extracellular matrix proteins vitronectin and laminin and expressed membrane-bound matrix metalloproteinase 9. The examination of Yes-associated protein expression indicated more cytoplasmic retention in the cells from auxetic scaffolds compared to those from regular scaffolds, suggesting that the auxetic scaffolds may affect cellular contraction. This study demonstrates a novel 3-D culture based on auxetic scaffolds for vascular differentiation and provides a platform to study the influence of biophysical microenvironments on differentiation of PSCs. The outcome of this study has implications for regenerative medicine and drug discovery. This article is protected by copyright. All rights reserved.

Conversion of borate-based glass scaffold to hydroxyapatite in a dilute phosphate solution.

PubMed

Liu, Xin; Pan, Haobo; Fu, Hailuo; Fu, Qiang; Rahaman, Mohamed N; Huang, Wenhai

2010-02-01

Porous scaffolds of a borate-based glass (composition in mol%: 6Na2O, 8K2O, 8MgO, 22CaO, 36B2O3, 18SiO2, 2P2O5), with interconnected porosity of approximately 70% and pores of size 200-500 microm, were prepared by a polymer foam replication technique. The degradation of the scaffolds and conversion to a hydroxyapatite-type material in a 0.02 M K2HPO4 solution (starting pH = 7.0) at 37 degrees C were studied by measuring the weight loss of the scaffolds, as well as the pH and the boron concentration of the solution. X-ray diffraction, scanning electronic microscopy and energy dispersive x-ray analysis showed that a hydroxyapatite-type material was formed on the glass surface within 7 days of immersion in the phosphate solution. Cellular response to the scaffolds was assessed using murine MLO-A5 cells, an osteogenic cell line. Scanning electron microscopy showed that the scaffolds supported cell attachment and proliferation during the 6 day incubation. The results indicate that this borate-based glass could provide a promising degradable scaffold material for bone tissue engineering applications.

Micropore-induced Capillarity Enhances Bone Distribution in vivo in Biphasic Calcium Phosphate Scaffolds

PubMed Central

Rustom, Laurence E.; Boudou, Thomas; Lou, Siyu; Pignot-Paintrand, Isabelle; Nemke, Brett W.; Lu, Yan; Markel, Mark D.; Picart, Catherine; Wagoner Johnson, Amy J.

2016-01-01

The increasing demand for bone repair solutions calls for the development of efficacious bone scaffolds. Biphasic calcium phosphate (BCP) scaffolds with both macropores and micropores (MP) have improved healing compared to those with macropores and no micropores (NMP), but the role of micropores is unclear. Here, we evaluate capillarity induced by micropores as a mechanism that can affect bone growth in vivo. Three groups of cylindrical scaffolds were implanted in pig mandibles for three weeks: MP were implanted either dry (MP-Dry), or after submersion in phosphate buffered saline, which fills pores with fluid and therefore suppresses micropore-induced capillarity (MP-Wet); NMP were implanted dry. The amount and distribution of bone in the scaffolds were quantified using micro-computed tomography. MP-Dry had a more homogeneous bone distribution than MP-Wet, although the average bone volume fraction, BVF¯, was not significantly different for these two groups (0.45±0.03 and 0.37±0.03, respectively). There was no significant difference in the radial bone distribution of NMP and MP-Wet, but the BVF¯ of NMP was significantly lower among the three groups (0.25±0.02). These results suggest that micropore-induced capillarity enhances bone regeneration by improving the homogeneity of bone distribution in BCP scaffolds. The explicit design and use of capillarity in bone scaffolds may lead to more effective treatments of large and complex bone defects. PMID:27544807

3D Nanostructured materials: TiO2 nanoparticles incorporated gellan gum scaffold for photocatalyst and biomedical Applications

NASA Astrophysics Data System (ADS)

Hasmizam Razali, Mohd; Arifah Ismail, Nur; Zulkafli, Mohd Farhan Azly Mohd; Anuar Mat Amin, Khairul

2018-03-01

A unique three-dimensional (3D) nanostructured gellan gum (GG) is fabricated by incorporating TiO2 nanoparticles (GG + TiO2NPs) scaffold by freeze-drying. The fabricated GG + TiO2NPs were characterized using Fourier transform infrared (FTIR), x-ray diffraction (XRD), and scanning electron microscopy (SEM) to study their physiochemical properties. FTIR was used to investigate the intermolecular interactions in the scaffolds. The crystal structure was determined by bulk analysis using XRD and SEM for microstructure observation of scaffold surfaces. The performance of synthesized GG + TiO2NPs scaffold 3D nanostructured materials was evaluated as a photocatalyst for methyl orange (MO) degradation and for biomedical applications. The results showed that the scaffold possessed good photocatalytic activity for removal of methyl orange with 88.24% degradation after 3 h of UV irradiation. The scaffold also induces the cell growth, thus offering a good candidate for biomedical applications.

3D-printed gelatin scaffolds of differing pore geometry modulate hepatocyte function and gene expression.

PubMed

Lewis, Phillip L; Green, Richard M; Shah, Ramille N

2018-03-15

Three dimensional (3D) printing is highly amenable to the fabrication of tissue-engineered organs of a repetitive microstructure such as the liver. The creation of uniform and geometrically repetitive tissue scaffolds can also allow for the control over cellular aggregation and nutrient diffusion. However, the effect of differing geometries, while controlling for pore size, has yet to be investigated in the context of hepatocyte function. In this study, we show the ability to precisely control pore geometry of 3D-printed gelatin scaffolds. An undifferentiated hepatocyte cell line (HUH7) demonstrated high viability and proliferation when seeded on 3D-printed scaffolds of two different geometries. However, hepatocyte specific functions (albumin secretion, CYP activity, and bile transport) increases in more interconnected 3D-printed gelatin cultures compared to a less interconnected geometry and to 2D controls. Additionally, we also illustrate the disparity between gene expression and protein function in simple 2D culture modes, and that recreation of a physiologically mimetic 3D environment is necessary to induce both expression and function of cultured hepatocytes. Three dimensional (3D) printing provides tissue engineers the ability spatially pattern cells and materials in precise geometries, however the biological effects of scaffold geometry on soft tissues such as the liver have not been rigorously investigated. In this manuscript, we describe a method to 3D print gelatin into well-defined repetitive geometries that show clear differences in biological effects on seeded hepatocytes. We show that a relatively simple and widely used biomaterial, such as gelatin, can significantly modulate biological processes when fabricated into specific 3D geometries. Furthermore, this study expands upon past research into hepatocyte aggregation by demonstrating how it can be manipulated to enhance protein function, and how function and expression may not precisely correlate in

A multi-scale controlled tissue engineering scaffold prepared by 3D printing and NFES technology

NASA Astrophysics Data System (ADS)

Yan, Feifei; Liu, Yuanyuan; Chen, Haiping; Zhang, Fuhua; Zheng, Lulu; Hu, Qingxi

2014-03-01

The current focus in the field of life science is the use of tissue engineering scaffolds to repair human organs, which has shown great potential in clinical applications. Extracellular matrix morphology and the performance and internal structure of natural organs are required to meet certain requirements. Therefore, integrating multiple processes can effectively overcome the limitations of the individual processes and can take into account the needs of scaffolds for the material, structure, mechanical properties and many other aspects. This study combined the biological 3D printing technology and the near-field electro-spinning (NFES) process to prepare a multi-scale controlled tissue engineering scaffold. While using 3D printing technology to directly prepare the macro-scaffold, the compositing NFES process to build tissue micro-morphology ultimately formed a tissue engineering scaffold which has the specific extracellular matrix structure. This scaffold not only takes into account the material, structure, performance and many other requirements, but also focuses on resolving the controllability problems in macro- and micro-forming which further aim to induce cell directed differentiation, reproduction and, ultimately, the formation of target tissue organs. It has in-depth immeasurable significance to build ideal scaffolds and further promote the application of tissue engineering.

3D printed porous polycaprolactone/oyster shell powder (PCL/OSP) scaffolds for bone tissue engineering

NASA Astrophysics Data System (ADS)

Luo, Wenfeng; Zhang, Shuangying; Lan, Yuewei; Huang, Chen; Wang, Chao; Lai, Xuexu; Chen, Hanwei; Ao, Ningjian

2018-04-01

In this work, oyster shell powder (OSP) was used as the bio-filler and combined with polycaprolactone (PCL) through melt blending methodology. The PCL and PCL/OSP scaffolds were prepared using additive manufacturing process. All the 3D printed scaffolds hold a highly porosity and interconnected pore structures. OSP particles are dispersed in the polymer matrix, which helped to improve the degree of crystallinity and mineralization ability of the scaffolds. There was no significant cytotoxicity of the prepared scaffolds towards MG-63 cells, and all the scaffolds showed a well ALP activity. Therefore, PCL/OSP scaffolds had a high potential to be employed in the bone tissue engineering.

Prevascularization of 3D printed bone scaffolds by bioactive hydrogels and cell co-culture.

PubMed

Kuss, Mitchell A; Wu, Shaohua; Wang, Ying; Untrauer, Jason B; Li, Wenlong; Lim, Jung Yul; Duan, Bin

2017-09-13

Vascularization is a fundamental prerequisite for large bone construct development and remains one of the main challenges of bone tissue engineering. Our current study presents the combination of 3D printing technique with a hydrogel-based prevascularization strategy to generate prevascularized bone constructs. Human adipose derived mesenchymal stem cells (ADMSC) and human umbilical vein endothelial cells (HUVEC) were encapsulated within our bioactive hydrogels, and the effects of culture conditions on in vitro vascularization were determined. We further generated composite constructs by forming 3D printed polycaprolactone/hydroxyapatite scaffolds coated with cell-laden hydrogels and determined how the co-culture affected vascularization and osteogenesis. It was demonstrated that 3D co-cultured ADMSC-HUVEC generated capillary-like networks within the porous 3D printed scaffold. The co-culture systems promoted in vitro vascularization, but had no significant effects on osteogenesis. The prevascularized constructs were subcutaneously implanted into nude mice to evaluate the in vivo vascularization capacity and the functionality of engineered vessels. The hydrogel systems facilitated microvessel and lumen formation and promoted anastomosis of vascular networks of human origin with host murine vasculature. These findings demonstrate the potential of prevascularized 3D printed scaffolds with anatomical shape for the healing of larger bone defects. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2017. © 2017 Wiley Periodicals, Inc.

Fabrication and durable antibacterial properties of 3D porous wet electrospun RCSC/PCL nanofibrous scaffold with silver nanoparticles

NASA Astrophysics Data System (ADS)

Zhang, Mei; Lin, Han; Wang, Yilong; Yang, Guang; Zhao, He; Sun, Dahui

2017-08-01

Electrospunnanofibers are used as three-dimensional (3D) scaffold materials that can alter cell attachment and cell proliferation, change the antibacterial properties of materials, and can be used as wound dressings. But the fabrication of porous 3D scaffold structure and the antibacterial properties enhancing are challenges remained to improve. With the states here, a Ranachensinensis skin collagen (RCSC)/poly(ɛ-caprolactone) (PCL)AgNP-loaded3D nanofiber scaffold is fabricated as a wound dressing material by using an improved wet electrospinning method (blending). The nanoscale of the AgNPs is proved. The 3D porous morphologies of the materials with different AgNP loadings, are determined with field emission scanning electron microscopy (FESEM) and the presence and uniformity distribution of AgNPs is confirmed by Energy dispersive X-ray (EDX) spectroscopy. The silver-ion release rates, antibacterial properties, and cytotoxicities of dressing materials with different AgNP contents are evaluated using ICP-AES, the zone inhibition method, and MTT testing. These results showed that the improved wet electrospun is an effective way to fabricate AgNP loaded 3D scaffold materials with porous structure and nearly 90% porosity and the presence of AgNPs in dressing materials strengthen the antibacterial properties. The RCSC/PCL 3D scaffold materials containing 2.0%AgNP would be promising for dressing materials application nearly without cytotoxicities.

The promotion of angiogenesis induced by three-dimensional porous beta-tricalcium phosphate scaffold with different interconnection sizes via activation of PI3K/Akt pathways

NASA Astrophysics Data System (ADS)

Xiao, Xin; Wang, Wei; Liu, Dong; Zhang, Haoqiang; Gao, Peng; Geng, Lei; Yuan, Yulin; Lu, Jianxi; Wang, Zhen

2015-03-01

The porous architectural characteristics of biomaterials play an important role in scaffold revascularization. However, no consensus exists regarding optimal interconnection sizes for vascularization and its scaffold bioperformance with different interconnection sizes. Therefore, a series of disk-type beta-tricalcium phosphates with the same pore sizes and variable interconnections were produced to evaluate how the interconnection size influenced biomaterial vascularization in vitro and in vivo. We incubated human umbilical vein endothelial cells on scaffolds with interconnections of various sizes. Results showed that scaffolds with a 150 μm interconnection size ameliorated endothelial cell function evidenced by promoting cell adhesion and migration, increasing cell proliferation and enhancing expression of platelet-endothelial cell adhesion molecules and vascular endothelial growth factor. In vivo study was performed on rabbit implanted with scaffolds into the bone defect on femoral condyles. Implantation with scaffolds with 150 μm interconnection size significantly improved neovascularization as shown by micro-CT as compared to scaffolds with 100 and 120 μm interconnection sizes. Moreover, the aforementioned positive effects were abolished by blocking PI3K/Akt/eNOS pathway with LY-294002. Our study explicitly demonstrates that the scaffold with 150 μm interconnection size improves neovascularization via the PI3K/Akt pathway and provides a target for biomaterial inner structure modification to attain improved clinical performance in implant vascularization.

Functional 3-D cardiac co-culture model using bioactive chitosan nanofiber scaffolds.

PubMed

Hussain, Ali; Collins, George; Yip, Derek; Cho, Cheul H

2013-02-01

The in vitro generation of a three-dimensional (3-D) myocardial tissue-like construct employing cells, biomaterials, and biomolecules is a promising strategy in cardiac tissue regeneration, drug testing, and tissue engineering applications. Despite significant progress in this field, current cardiac tissue models are not yet able to stably maintain functional characteristics of cardiomyocytes for long-term culture and therapeutic purposes. The objective of this study was to fabricate bioactive 3-D chitosan nanofiber scaffolds using an electrospinning technique and exploring its potential for long-term cardiac function in the 3-D co-culture model. Chitosan is a natural polysaccharide biomaterial that is biocompatible, biodegradable, non-toxic, and cost effective. Electrospun chitosan was utilized to provide structural scaffolding characterized by scale and architectural resemblance to the extracellular matrix (ECM) in vivo. The chitosan fibers were coated with fibronectin via adsorption in order to enhance cellular adhesion to the fibers and migration into the interfibrous milieu. Ventricular cardiomyocytes were harvested from neonatal rats and studied in various culture conditions (i.e., mono- and co-cultures) for their viability and function. Cellular morphology and functionality were examined using immunofluorescent staining for alpha-sarcomeric actin (SM-actin) and gap junction protein, Connexin-43 (Cx43). Scanning electron microscopy (SEM) and light microscopy were used to investigate cellular morphology, spatial organization, and contractions. Calcium indicator was used to monitor calcium ion flux of beating cardiomyocytes. The results demonstrate that the chitosan nanofibers retained their cylindrical morphology in long-term cell cultures and exhibited good cellular attachment and spreading in the presence of adhesion molecule, fibronectin. Cardiomyocyte mono-cultures resulted in loss of cardiomyocyte polarity and islands of non-coherent contractions. However

The Effect of 3D Hydrogel Scaffold Modulus on Osteoblast Differentiation and Mineralization Revealed by Combinatorial Screening

PubMed Central

Chatterjee, Kaushik; Lin-Gibson, Sheng; Wallace, William E.; Parekh, Sapun H.; Lee, Young J.; Cicerone, Marcus T.; Young, Marian F.; Simon, Carl G.

2011-01-01

Cells are known to sense and respond to the physical properties of their environment and those of tissue scaffolds. Optimizing these cell-material interactions is critical in tissue engineering. In this work, a simple and inexpensive combinatorial platform was developed to rapidly screen three-dimensional (3D) tissue scaffolds and was applied to screen the effect of scaffold properties for tissue engineering of bone. Differentiation of osteoblasts was examined in poly(ethylene glycol) hydrogel gradients spanning a 30-fold range in compressive modulus (≈ 10 kPa to ≈ 300 kPa). Results demonstrate that material properties (gel stiffness) of scaffolds can be leveraged to induce cell differentiation in 3D culture as an alternative to biochemical cues such as soluble supplements, immobilized biomolecules and vectors, which are often expensive, labile and potentially carcinogenic. Gel moduli of ≈ 225 kPa and higher enhanced osteogenesis. Furthermore, it is proposed that material-induced cell differentiation can be modulated to engineer seamless tissue interfaces between mineralized bone tissue and softer tissues such as ligaments and tendons. This work presents a combinatorial method to screen biological response to 3D hydrogel scaffolds that more closely mimics the 3D environment experienced by cells in vivo. PMID:20378163

3D imaging of cells in scaffolds: direct labelling for micro CT.

PubMed

Shepherd, David V; Shepherd, Jennifer H; Best, Serena M; Cameron, Ruth E

2018-06-12

The development of in-vitro techniques to characterise the behaviour of cells in biomedical scaffolds is a rapidly developing field. However, until now it has not been possible to visualise, directly in 3D, the extent of cell migration using a desktop X-ray microCT. This paper describes a new technique based on cell labelling with a radio opacifier (barium sulphate), which permits cell tracking without the need for destructive sample preparation. The ability to track cells is highlighted via a comparison of cell migration through demonstrator lyophilised collagen scaffolds with contrasting pore size and interconnectivity. The results demonstrate the ease with which the technique can be used to characterise the effects of scaffold architecture on cell infiltration.

Silk scaffolds connected with different naturally occurring biomaterials for prostate cancer cell cultivation in 3D.

PubMed

Bäcker, Anne; Erhardt, Olga; Wietbrock, Lukas; Schel, Natalia; Göppert, Bettina; Dirschka, Marian; Abaffy, Paul; Sollich, Thomas; Cecilia, Angelica; Gruhl, Friederike J

2017-02-01

In the present work, different biopolymer blend scaffolds based on the silk protein fibroin from Bombyx mori (BM) were prepared via freeze-drying method. The chemical, structural, and mechanical properties of the three dimensional (3D) porous silk fibroin (SF) composite scaffolds of gelatin, collagen, and chitosan as well as SF from Antheraea pernyi (AP) and the recombinant spider silk protein spidroin (SSP1) have been systematically investigated, followed by cell culture experiments with epithelial prostate cancer cells (LNCaP) up to 14 days. Compared to the pure SF scaffold of BM, the blend scaffolds differ in porous morphology, elasticity, swelling behavior, and biochemical composition. The new composite scaffold with SSP1 showed an increased swelling degree and soft tissue like elastic properties. Whereas, in vitro cultivation of LNCaP cells demonstrated an increased growth behavior and spheroid formation within chitosan blended scaffolds based on its remarkable porosity, which supports nutrient supply matrix. Results of this study suggest that silk fibroin matrices are sufficient and certain SF composite scaffolds even improve 3D cell cultivation for prostate cancer research compared to matrices based on pure biomaterials or synthetic polymers. © 2016 Wiley Periodicals, Inc.

A novel 3D bone-mimetic scaffold composed of collagen/MTA/MWCNT modulates cell migration and osteogenesis.

PubMed

Valverde, Thalita M; Castro, Elisandra G; Cardoso, Maíssa H S; Martins-Júnior, Paulo A; Souza, Lívia M O; Silva, Patrícia P; Ladeira, Luiz O; Kitten, Gregory T

2016-10-01

This study characterized a three-dimensional (3D) biocomposite scaffolds produced using type I collagen, mineral trioxide aggregate (MTA) and multi-walled carbon nanotubes (MWCNT) to be used in bone tissue regeneration. The scaffolds were analyzed via scanning (SEM) and transmission (TEM) electron microscopy, as well as the viability and migration of osteoblasts and mineralization of the scaffolds. SEM and TEM analyses showed that MTA and MWCNT were distributed as both large agglomerates entrapped within the collagen network and as smaller accumulations or individual molecules dispersed throughout the scaffold. Ultrastructural analysis revealed that osteoblastic MC3T3-E1 cells grown in the biocomposite endocytosed MWCNT, which were localized in the cytoplasm and in vesicles. Analysis of cells grown in the 3D scaffolds demonstrated that >95% of the cells remained viable in all tested combinations and concentrations of the biocomposite. MC3T3-E1 osteoblasts migrated into scaffolds formed with concentrations of type I collagen between 1.75 and 3.0mg/mL. Cells displayed increased migration into scaffolds formed with collagen and a range of low to high concentrations of MTA. In contrast, the presence of MWCNT in the biocomposite had a slight negative effect on migration. Collagen gels containing specific concentrations of MTA, or MWCNT, or combinations of MTA/MWCNT, caused an increase in mineralization of scaffolds. Scaffolds composed of defined concentrations of type I collagen, MTA and MWCNT are biocompatible, promote migration and mineralization of osteoblasts, and hence may be useful as bone tissue mimetics. Copyright © 2016 Elsevier Inc. All rights reserved.

Porogen-based solid freeform fabrication of polycaprolactone-calcium phosphate scaffolds for tissue engineering.

PubMed

Mondrinos, Mark J; Dembzynski, Robert; Lu, Lin; Byrapogu, Venkata K C; Wootton, David M; Lelkes, Peter I; Zhou, Jack

2006-09-01

Drop on demand printing (DDP) is a solid freeform fabrication (SFF) technique capable of generating microscale physical features required for tissue engineering scaffolds. Here, we report results toward the development of a reproducible manufacturing process for tissue engineering scaffolds based on injectable porogens fabricated by DDP. Thermoplastic porogens were designed using Pro/Engineer and fabricated with a commercially available DDP machine. Scaffolds composed of either pure polycaprolactone (PCL) or homogeneous composites of PCL and calcium phosphate (CaP, 10% or 20% w/w) were subsequently fabricated by injection molding of molten polymer-ceramic composites, followed by porogen dissolution with ethanol. Scaffold pore sizes, as small as 200 microm, were attainable using the indirect (porogen-based) method. Scaffold structure and porosity were analyzed by scanning electron microscopy (SEM) and microcomputed tomography, respectively. We characterized the compressive strength of 90:10 and 80:20 PCL-CaP composite materials (19.5+/-1.4 and 24.8+/-1.3 Mpa, respectively) according to ASTM standards, as well as pure PCL scaffolds (2.77+/-0.26 MPa) fabricated using our process. Human embryonic palatal mesenchymal (HEPM) cells attached and proliferated on all scaffolds, as evidenced by fluorescent nuclear staining with Hoechst 33258 and the Alamar Blue assay, with increased proliferation observed on 80:20 PCL-CaP scaffolds. SEM revealed multilayer assembly of HEPM cells on 80:20 PCL-CaP composite, but not pure PCL, scaffolds. In summary, we have developed an SFF-based injection molding process for the fabrication of PCL and PCL-CaP scaffolds that display in vitro cytocompatibility and suitable mechanical properties for hard tissue repair.

Tailorable Surface Morphology of 3D Scaffolds by Combining Additive Manufacturing with Thermally Induced Phase Separation.

PubMed

Di Luca, Andrea; de Wijn, Joost R; van Blitterswijk, Clemens A; Camarero-Espinosa, Sandra; Moroni, Lorenzo

2017-08-01

The functionalization of biomaterials substrates used for cell culture is gearing towards an increasing control over cell activity. Although a number of biomaterials have been successfully modified by different strategies to display tailored physical and chemical surface properties, it is still challenging to step from 2D substrates to 3D scaffolds with instructive surface properties for cell culture and tissue regeneration. In this study, additive manufacturing and thermally induced phase separation are combined to create 3D scaffolds with tunable surface morphology from polymer gels. Surface features vary depending on the gel concentration, the exchanging temperature, and the nonsolvent used. When preosteoblasts (MC-3T3 cells) are cultured on these scaffolds, a significant increase in alkaline phosphatase activity is measured for submicron surface topography, suggesting a potential role on early cell differentiation. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

3D Printing of Aniline Tetramer-Grafted-Polyethylenimine and Pluronic F127 Composites for Electroactive Scaffolds.

PubMed

Dong, Shi-Lei; Han, Lu; Du, Cai-Xia; Wang, Xiao-Yu; Li, Lu-Hai; Wei, Yen

2017-02-01

Electroactive hydrogel scaffolds are fabricated by the 3D-printing technique using composites of 30% Pluronic F127 and aniline tetramer-grafted-polyethylenimine (AT-PEI) copolymers with various contents from 2.5% to 10%. The synthesized AT-PEI copolymers can self-assemble into nanoparticles with the diameter of ≈50 nm and display excellent electroactivity due to AT conjugation. The copolymers are then homogeneously distributed into 30% Pluronic F127 solution by virtue of the thermosensitivity of F127, denoted as F/AT-PEI composites. Macroscopic photographs of latticed scaffolds elucidate their excellent printability of F/AT-PEI hydrogels for the 3D-printing technique. The conductivities of the printed F/AT-PEI scaffolds are all higher than 2.0 × 10 -3 S cm -1 , which are significantly improved compared with that of F127 scaffold with only 0.94 × 10 -3 S cm -1 . Thus, the F/AT-PEI scaffolds can be considered as candidates for application in electrical stimulation of tissue regeneration such as repair of muscle and cardiac nerve tissue. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evaluating 3D-printed biomaterials as scaffolds for vascularized bone tissue engineering.

PubMed

Wang, Martha O; Vorwald, Charlotte E; Dreher, Maureen L; Mott, Eric J; Cheng, Ming-Huei; Cinar, Ali; Mehdizadeh, Hamidreza; Somo, Sami; Dean, David; Brey, Eric M; Fisher, John P

2015-01-07

There is an unmet need for a consistent set of tools for the evaluation of 3D-printed constructs. A toolbox developed to design, characterize, and evaluate 3D-printed poly(propylene fumarate) scaffolds is proposed for vascularized engineered tissues. This toolbox combines modular design and non-destructive fabricated design evaluation, evaluates biocompatibility and mechanical properties, and models angiogenesis. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evaluation of 3D-Printed Polycaprolactone Scaffolds Coated with Freeze-Dried Platelet-Rich Plasma for Bone Regeneration.

PubMed

Li, Junda; Chen, Meilin; Wei, Xiaoying; Hao, Yishan; Wang, Jinming

2017-07-19

Three-dimensional printing is one of the most promising techniques for the manufacturing of scaffolds for bone tissue engineering. However, a pure scaffold is limited by its biological properties. Platelet-rich plasma (PRP) has been shown to have the potential to improve the osteogenic effect. In this study, we improved the biological properties of scaffolds by coating 3D-printed polycaprolactone (PCL) scaffolds with freeze-dried and traditionally prepared PRP, and we evaluated these scaffolds through in vitro and in vivo experiments. In vitro, we evaluated the interaction between dental pulp stem cells (DPSCs) and the scaffolds by measuring cell proliferation, alkaline phosphatase (ALP) activity, and osteogenic differentiation. The results showed that freeze-dried PRP significantly enhanced ALP activity and the mRNA expression levels of osteogenic genes (ALP, RUNX2 (runt-related gene-2), OCN (osteocalcin), OPN (osteopontin)) of DPSCs ( p < 0.05). In vivo, 5 mm calvarial defects were created, and the PRP-PCL scaffolds were implanted. The data showed that compared with traditional PRP-PCL scaffolds or bare PCL scaffolds, the freeze-dried PRP-PCL scaffolds induced significantly greater bone formation ( p < 0.05). All these data suggest that coating 3D-printed PCL scaffolds with freeze-dried PRP can promote greater osteogenic differentiation of DPSCs and induce more bone formation, which may have great potential in future clinical applications.

Stem Cells and Calcium Phosphate Cement Scaffolds for Bone Regeneration

PubMed Central

Wang, P.; Zhao, L.; Chen, W.; Liu, X.; Weir, M.D.; Xu, H.H.K.

2014-01-01

Calcium phosphate cements (CPCs) have excellent biocompatibility and osteoconductivity for dental, craniofacial, and orthopedic applications. This article reviews recent developments in stem cell delivery via CPC for bone regeneration. This includes: (1) biofunctionalization of the CPC scaffold, (2) co-culturing of osteoblasts/endothelial cells and prevascularization of CPC, (3) seeding of CPC with different stem cell species, (4) human umbilical cord mesenchymal stem cell (hUCMSC) and bone marrow MSC (hBMSC) seeding on CPC for bone regeneration, and (5) human embryonic stem cell (hESC) and induced pluripotent stem cell (hiPSC) seeding with CPC for bone regeneration. Cells exhibited good attachment/proliferation in CPC scaffolds. Stem-cell-CPC constructs generated more new bone and blood vessels in vivo than did the CPC control without cells. hUCMSCs, hESC-MSCs, and hiPSC-MSCs in CPC generated new bone and blood vessels similar to those of hBMSCs; hence, they were viable cell sources for bone engineering. CPC with hESC-MSCs and hiPSC-MSCs generated new bone two- to three-fold that of the CPC control. Therefore, this article demonstrates that: (1) CPC scaffolds are suitable for delivering cells; (2) hUCMSCs, hESCs, and hiPSCs are promising alternatives to hBMSCs, which require invasive procedures to harvest with limited cell quantity; and (3) stem-cell-CPC constructs are highly promising for bone regeneration in dental, craniofacial, and orthopedic applications. PMID:24799422

3D printing of porous hydroxyapatite scaffolds intended for use in bone tissue engineering applications.

PubMed

Cox, Sophie C; Thornby, John A; Gibbons, Gregory J; Williams, Mark A; Mallick, Kajal K

2015-02-01

A systematic characterisation of bone tissue scaffolds fabricated via 3D printing from hydroxyapatite (HA) and poly(vinyl)alcohol (PVOH) composite powders is presented. Flowability of HA:PVOH precursor materials was observed to affect mechanical stability, microstructure and porosity of 3D printed scaffolds. Anisotropic behaviour of constructs and part failure at the boundaries of interlayer bonds was highlighted by compressive strength testing. A trade-off between the ability to facilitate removal of PVOH thermal degradation products during sintering and the compressive strength of green parts was revealed. The ultimate compressive strength of 55% porous green scaffolds printed along the Y-axis and dried in a vacuum oven for 6h was 0.88 ± 0.02 MPa. Critically, the pores of 3D printed constructs could be user designed, ensuring bulk interconnectivity, and the imperfect packing of powder particles created an inherent surface roughness and non-designed porosity within the scaffold. These features are considered promising since they are known to facilitate osteoconduction and osteointegration in-vivo. Characterisation techniques utilised in this study include two funnel flow tests, scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), compressive strength testing and computed tomography (CT). Copyright © 2014 Elsevier B.V. All rights reserved.

The ability of a collagen/calcium phosphate scaffold to act as its own vector for gene delivery and to promote bone formation via transfection with VEGF(165).

PubMed

Keeney, Michael; van den Beucken, Jeroen J J P; van der Kraan, Peter M; Jansen, John A; Pandit, Abhay

2010-04-01

Collagen/calcium phosphate scaffolds have been used for bone reconstruction due to their inherent similarities to the bone extracellular matrix. Calcium phosphate alone has also been used as a non-viral vector for gene delivery. The aim of this study was to determine the capability of a collagen/calcium phosphate scaffold to deliver naked plasmid DNA and mediate transfection in vivo. The second goal of the study was to deliver a plasmid encoding vascular endothelial growth factor(165) (pVEGF(165)) to promote angiogenesis, and hence bone formation, in a mouse intra-femoral model. The delivery of naked plasmid DNA resulted in a 7.6-fold increase in mRNA levels of beta-Galactosidase compared to the delivery of plasmid DNA complexed with a partially degraded PAMAM dendrimer (dPAMAM) in a subcutaneous murine model. When implanted in a muirne intra-femoral model, the delivery of pVEGF(165) resulted in a 2-fold increase in bone volume at the defect site relative to control scaffolds without pVEGF(165). It was concluded that a collagen/calcium phosphate scaffold can mediate transfection without the use of additional transfection vectors and can promote bone formation in a mouse model via the delivery of pVEGF(165). 2009 Elsevier Ltd. All rights reserved.

Structural Basis for Substrate Specificity in Phosphate Binding (beta/alpha)8-Barrels: D-Allulose 6-Phosphate 3-Epimerase from Escherichia coli K-12

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chan,K.; Fedorov, A.; Almo, S.

2008-01-01

Enzymes that share the ({beta}/{alpha})8-barrel fold catalyze a diverse range of reactions. Many utilize phosphorylated substrates and share a conserved C-terminal ({beta}/a)2-quarter barrel subdomain that provides a binding motif for the dianionic phosphate group. We recently reported functional and structural studies of d-ribulose 5-phosphate 3-epimerase (RPE) from Streptococcus pyogenes that catalyzes the equilibration of the pentulose 5-phosphates d-ribulose 5-phosphate and d-xylulose 5-phosphate in the pentose phosphate pathway [J. Akana, A. A. Fedorov, E. Fedorov, W. R. P. Novack, P. C. Babbitt, S. C. Almo, and J. A. Gerlt (2006) Biochemistry 45, 2493-2503]. We now report functional and structural studies ofmore » d-allulose 6-phosphate 3-epimerase (ALSE) from Escherichia coli K-12 that catalyzes the equilibration of the hexulose 6-phosphates d-allulose 6-phosphate and d-fructose 6-phosphate in a catabolic pathway for d-allose. ALSE and RPE prefer their physiological substrates but are promiscuous for each other's substrate. The active sites (RPE complexed with d-xylitol 5-phosphate and ALSE complexed with d-glucitol 6-phosphate) are superimposable (as expected from their 39% sequence identity), with the exception of the phosphate binding motif. The loop following the eighth {beta}-strand in ALSE is one residue longer than the homologous loop in RPE, so the binding site for the hexulose 6-phosphate substrate/product in ALSE is elongated relative to that for the pentulose 5-phosphate substrate/product in RPE. We constructed three single-residue deletion mutants of the loop in ALSE, ?T196, ?S197 and ?G198, to investigate the structural bases for the differing substrate specificities; for each, the promiscuity is altered so that d-ribulose 5-phosphate is the preferred substrate. The changes in kcat/Km are dominated by changes in kcat, suggesting that substrate discrimination results from differential transition state stabilization. In both ALSE and RPE, the

3D culturing and differentiation of SH-SY5Y neuroblastoma cells on bacterial nanocellulose scaffolds.

PubMed

Innala, Marcus; Riebe, Ilse; Kuzmenko, Volodymyr; Sundberg, Johan; Gatenholm, Paul; Hanse, Eric; Johannesson, Sara

2014-10-01

A new in vitro model, mimicking the complexity of nerve tissue, was developed based on a bacterial nanocellulose (BNC) scaffold that supports 3D culturing of neuronal cells. BNC is extracellularly excreted by Gluconacetobacter xylinus (G. xylinus) in the shape of long non-aggregated nanofibrils. The cellulose network created by G. xylinus has good mechanical properties, 99% water content, and the ability to be shaped into 3D structures by culturing in different molds. Surface modification with trimethyl ammonium beta-hydroxypropyl (TMAHP) to induce a positive surface charge, followed by collagen I coating, has been used to improve cell adhesion, growth, and differentiation on the scaffold. In the present study, we used SH-SY5Y neuroblastoma cells as a neuronal model. These cells attached and proliferated well on the BNC scaffold, as demonstrated by scanning electron microscopy (SEM) and the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS) assay. Following neuronal differentiation, we demonstrated functional action potentials (APs) by electrophysiological recordings, indicating the presence of mature neurons on the scaffolds. In conclusion, we have demonstrated for the first time that neurons can attach, proliferate, and differentiate on BNC. This 3D model based on BNC scaffolds could possibly be used for developing in vitro disease models, when combined with human induced pluripotent stem (iPS) cells (derived from diseased patients) for detailed investigations of neurodegenerative disease mechanisms and in the search for new therapeutics.

High porous titanium scaffolds showed higher compatibility than lower porous beta-tricalcium phosphate scaffolds for regulating human osteoblast and osteoclast differentiation.

PubMed

Hirota, Makoto; Hayakawa, Tohru; Shima, Takaki; Ametani, Akihiro; Tohnai, Iwai

2015-04-01

We compared osteoblast and osteoclast differentiation when using beta-tricalcium phosphate (βTCP) and titanium scaffolds by investigating human mesenchymal stem cells (hMSCs) and osteoclast progenitor cell activities. hMSCs were cultured for 7, 14, and 21days on titanium scaffolds with 60%, 73%, and 87% porosity and on βTCP scaffolds with 60% and 75% porosity. Human osteoclast progenitor cells were cultured with osteoblast for 14 and 21days on 87% titanium and 75% βTCP scaffolds. Viable cell numbers with 60% and 73% titanium were higher than with 87% titanium and βTCP scaffolds (P<0.05). An 87% titanium scaffold resulted in the highest osteocalcin production with calcification on day 14 (P<0.01) in titanium scaffolds. All titanium scaffolds resulted in higher osteocalcin production on days 7 and 14 compared to βTCP scaffolds (P<0.01). Osteoblasts cultured on 87% titanium scaffolds suppressed osteoclast differentiation on day 7 but enhanced osteoclast differentiation on day 14 compared to 75% βTCP scaffolds (P<0.01). These findings concluded that high porosity titanium scaffolds could enhance progression of hMSC/osteoblast differentiation and regulated osteoclast differentiation cooperating with osteoblast differentiation for calcification as compared with lower porous βTCP. Copyright © 2015 Elsevier B.V. All rights reserved.

Anti-biofouling 3D porous systems: the blend effect of oxazoline-based oligomers on chitosan scaffolds.

PubMed

Correia, Vanessa G; Coelho, Margarida; Barroso, Telma; Raje, Vivek P; Bonifácio, Vasco D B; Casimiro, Teresa; Pinho, Mariana G; Aguiar-Ricardo, Ana

2013-01-01

The production, characterization and anti-biofouling activity of 3D porous scaffolds combining different blends of chitosan and oxazoline-based antimicrobial oligomers is reported. The incorporation of ammonium quaternized oligo(2-oxazoline)s into the composition of the scaffold enhances the stability of the chitosan scaffold under physiological conditions as well as its ability to repel protein adsorption. The blended scaffolds showed mean pore sizes in the range of 18-32 μm, a good pore interconnectivity and high porosity, as well as a large surface area, ultimate key features for anti-biofouling applications. Bovine serum albumin (BSA) adhesion profiles showed that the composition of the scaffolds plays a critical role in the chitosan-oligooxazoline system. Oligobisoxazoline-enriched scaffolds (20% w/w, CB8020) decreased protein adsorption (BSA) by up to 70%. Moreover, 1 mg of CB8020 was able to kill 99.9% of Escherichia coli cells upon contact, demonstrating its potential as promising material for production of tailored non-fouling 3D structures to be used in the construction of novel devices with applications in the biomedical field and water treatment processes.

[Biological evaluation of three-dimensional printed co-poly lactic acid/glycolic acid/tri-calcium phosphate scaffold for bone reconstruction].

PubMed

Li, S Y; Zhou, M; Lai, Y X; Geng, Y M; Cao, S S; Chen, X M

2016-11-09

Objective: To biologically evaluate the three-dimensional(3D) printed co-poly lactic acid/glycolic acid/tri-calcium phosphate(PLGA/TCP) scaffold which could be used for repairing oral and maxillofacial bone defects, and to provide experimental evidence for its further research and clinical application. Methods: PLGA/TCP scaffolds were fabricated using low temperature rapid prototyping technique. Micro-CT and scanning electron microscope(SEM) were used to characterize the surface morphology. MC3T3-E1 cells were seeded onto the scaffold and stained with the rhodamine phalloidin and calcein acetomethoxy. After that, confocal laser scanning microscope was exploited to observe the features and viability of the cells. Moreover, the cells were co-cultured with the extract of PLGA/TCP and complete medium, respectively. The proliferation capability of the cells was assessed by the cell counting kit-8 (CCK-8) on the 1st, 2nd, and 3rd day. The PLGA/TCP scaffolds incorporated with recombinant human bone morphogenetic protein-2(rhBMP-2) of 0, 30, 60 μg(i.e. blank control group, low-dose group and high-dose group) were implanted into the latissimus dorsi muscle of the rats, and 6 weeks later, the samples were harvested to estimate the volume and pattern of new bone. Results: The 3D printed PLGA/TCP scaffold possessed a regular and well-defined porous stereo-structure with porosity of (73±3)%. Micro-CT and SEM showed that pore size were (379±32) and (453±29) μm respectively, and distance between layers were (452± 24) and (415±25) μm, and cylinder diameter were (342±24) and (350±28) μm. It also exhibited excellent cell adhesion and growth ability on the exterior and inner surface through rhodamine phalloidin and calcein acetomethoxy staining. The CCK-8 test demonstrated that the absorbance value of extract group on the 1st and 2nd day(0.51±0.08 and 0.63±0.09) were significantly higher than those in the blank control group(0.39± 0.05 and 0.53±0.05)( P< 0.05), while

Digital micromirror device (DMD)-based 3D printing of poly(propylene fumarate) scaffolds.

PubMed

Mott, Eric J; Busso, Mallory; Luo, Xinyi; Dolder, Courtney; Wang, Martha O; Fisher, John P; Dean, David

2016-04-01

Our recent investigations into the 3D printing of poly(propylene fumarate) (PPF), a linear polyester, using a DMD-based system brought us to a resin that used titanium dioxide (TiO2) as an ultraviolet (UV) filter for controlling cure depth. However, this material hindered the 3D printing process due to undesirable lateral or "dark" curing (i.e., in areas not exposed to light from the DMD chip). Well known from its use in sunscreen, another UV filter, oxybenzone, has previously been used in conjunction with TiO2. In this study we hypothesize that combining these two UV filters will result in a synergistic effect that controls cure depth and avoids dark cure. A resin mixture (i.e., polymer, initiator, UV filters) was identified that worked well. The resin was then further characterized through mechanical testing, cure testing, and cytotoxicity testing to investigate its use as a material for bone tissue engineering scaffolds. Results show that the final resin eliminated dark cure as shown through image analysis. Mechanically the new scaffolds proved to be far weaker than those printed from previous resins, with compressive strengths of 7.8 ± 0.5 MPa vs. 36.5 ± 1.6 MPa, respectively. The new scaffolds showed a 90% reduction in elastic modulus and a 74% increase in max strain. These properties may be useful in tissue engineering applications where resorption is required. Initial cytotoxicity evaluation was negative. As hypothesized, the use of TiO2 and oxybenzone showed synergistic effects in the 3D printing of PPF tissue engineering scaffolds. Copyright © 2015 Elsevier B.V. All rights reserved.

Histological evaluation of osteogenesis of 3D-printed poly-lactic-co-glycolic acid (PLGA) scaffolds in a rabbit model.

PubMed

Ge, Zigang; Tian, Xianfeng; Heng, Boon Chin; Fan, Victor; Yeo, Jin Fei; Cao, Tong

2009-04-01

Utilizing a suitable combination of lactide and glycolide in a copolymer would optimize the degradation rate of a scaffold upon implantation in situ. Moreover, 3D printing technology enables customizing the shape of the scaffold to biometric data from CT and MRI scans. A previous in vitro study has shown that novel 3D-printed poly-lactic-co-glycolic acid (PLGA) scaffolds had good biocompatibility and mechanical properties comparable with human cancellous bone, while they could support proliferation and osteogenic differentiation of osteoblasts. Based on the previous study, this study evaluated PLGA scaffolds for bone regeneration within a rabbit model. The scaffolds were implanted at two sites on the same animal, within the periosteum and within bi-cortical bone defects on the iliac crest. Subsequently, the efficacy of bone regeneration within the implanted scaffolds was evaluated at 4, 12 and 24 weeks post-surgery through histological analysis. In both the intra-periosteum and iliac bone defect models, the implanted scaffolds facilitated new bone tissue formation and maturation over the time course of 24 weeks, even though there was initially observed to be little tissue ingrowth within the scaffolds at 4 weeks post-surgery. Hence, the 3D-printed porous PLGA scaffolds investigated in this study displayed good biocompatibility and are osteoconductive in both the intra-periosteum and iliac bone defect models.

3D fiber-deposited scaffolds for tissue engineering: influence of pores geometry and architecture on dynamic mechanical properties.

PubMed

Moroni, L; de Wijn, J R; van Blitterswijk, C A

2006-03-01

One of the main issues in tissue engineering is the fabrication of scaffolds that closely mimic the biomechanical properties of the tissues to be regenerated. Conventional fabrication techniques are not sufficiently suitable to control scaffold structure to modulate mechanical properties. Within novel scaffold fabrication processes 3D fiber deposition (3DF) showed great potential for tissue engineering applications because of the precision in making reproducible 3D scaffolds, characterized by 100% interconnected pores with different shapes and sizes. Evidently, these features also affect mechanical properties. Therefore, in this study we considered the influence of different structures on dynamic mechanical properties of 3DF scaffolds. Pores were varied in size and shape, by changing fibre diameter, spacing and orientation, and layer thickness. With increasing porosity, dynamic mechanical analysis (DMA) revealed a decrease in elastic properties such as dynamic stiffness and equilibrium modulus, and an increase of the viscous parameters like damping factor and creep unrecovered strain. Furthermore, the Poisson's ratio was measured, and the shear modulus computed from it. Scaffolds showed an adaptable degree of compressibility between sponges and incompressible materials. As comparison, bovine cartilage was tested and its properties fell in the fabricated scaffolds range. This investigation showed that viscoelastic properties of 3DF scaffolds could be modulated to accomplish mechanical requirements for tailored tissue engineered applications.

Improved Human Bone Marrow Mesenchymal Stem Cell Osteogenesis in 3D Bioprinted Tissue Scaffolds with Low Intensity Pulsed Ultrasound Stimulation

PubMed Central

Zhou, Xuan; Castro, Nathan J.; Zhu, Wei; Cui, Haitao; Aliabouzar, Mitra; Sarkar, Kausik; Zhang, Lijie Grace

2016-01-01

3D printing and ultrasound techniques are showing great promise in the evolution of human musculoskeletal tissue repair and regeneration medicine. The uniqueness of the present study was to combine low intensity pulsed ultrasound (LIPUS) and advanced 3D printing techniques to synergistically improve growth and osteogenic differentiation of human mesenchymal stem cells (MSC). Specifically, polyethylene glycol diacrylate bioinks containing cell adhesive Arginine-Glycine-Aspartic acid-Serene (RGDS) peptide and/or nanocrystalline hydroxyapatite (nHA) were used to fabricate 3D scaffolds with different geometric patterns via novel table-top stereolithography 3D printer. The resultant scaffolds provide a highly porous and interconnected 3D environment to support cell proliferation. Scaffolds with small square pores were determined to be the optimal geometric pattern for MSC attachment and growth. The optimal LIPUS working parameters were determined to be 1.5 MHz, 20% duty cycle with 150 mW/cm2 intensity. Results demonstrated that RGDS peptide and nHA containing 3D printed scaffolds under LIPUS treatment can greatly promote MSC proliferation, alkaline phosphatase activity, calcium deposition and total protein content. These results illustrate the effectiveness of the combination of LIPUS and biomimetic 3D printing scaffolds as a valuable combinatorial tool for improved MSC function, thus make them promising for future clinical and various regenerative medicine application. PMID:27597635

Integrating biologically inspired nanomaterials and table-top stereolithography for 3D printed biomimetic osteochondral scaffolds

NASA Astrophysics Data System (ADS)

Castro, Nathan J.; O'Brien, Joseph; Zhang, Lijie Grace

2015-08-01

The osteochondral interface of an arthritic joint is notoriously difficult to regenerate due to its extremely poor regenerative capacity and complex stratified architecture. Native osteochondral tissue extracellular matrix is composed of numerous nanoscale organic and inorganic constituents. Although various tissue engineering strategies exist in addressing osteochondral defects, limitations persist with regards to tissue scaffolding which exhibit biomimetic cues at the nano to micro scale. In an effort to address this, the current work focused on 3D printing biomimetic nanocomposite scaffolds for improved osteochondral tissue regeneration. For this purpose, two biologically-inspired nanomaterials have been synthesized consisting of (1) osteoconductive nanocrystalline hydroxyapatite (nHA) (primary inorganic component of bone) and (2) core-shell poly(lactic-co-glycolic) acid (PLGA) nanospheres encapsulated with chondrogenic transforming growth-factor β1 (TGF-β1) for sustained delivery. Then, a novel table-top stereolithography 3D printer and the nano-ink (i.e., nHA + nanosphere + hydrogel) were employed to fabricate a porous and highly interconnected osteochondral scaffold with hierarchical nano-to-micro structure and spatiotemporal bioactive factor gradients. Our results showed that human bone marrow-derived mesenchymal stem cell adhesion, proliferation, and osteochondral differentiation were greatly improved in the biomimetic graded 3D printed osteochondral construct in vitro. The current work served to illustrate the efficacy of the nano-ink and current 3D printing technology for efficient fabrication of a novel nanocomposite hydrogel scaffold. In addition, tissue-specific growth factors illustrated a synergistic effect leading to increased cell adhesion and directed stem cell differentiation.

Evaluation of synovium-derived mesenchymal stem cells and 3D printed nanocomposite scaffolds for tissue engineering

NASA Astrophysics Data System (ADS)

Pan, Jian-Feng; Li, Shuo; Guo, Chang-An; Xu, Du-Liang; Zhang, Feng; Yan, Zuo-Qin; Mo, Xiu-Mei

2015-08-01

Stem cells and scaffolds play a very important role in tissue engineering. Here, we isolated synovium-derived mesenchymal stem cells (SMSCs) from synovial membrane tissue and characterized stem-cell properties. Gelatin nanoparticles (NP) were prepared using a two-step desolvation method and then pre-mixed into different host matrix (silk fibroin (SF), gelatin (Gel), or SF-Gel mixture) to generate various 3D printed nanocomposite scaffolds (NP/SF, NP/SF-Gel, NP/Gel-1, and NP/Gel-2). The microstructure was examined by scanning electron microscopy. Biocompatibility assessment was performed through CCK-8 assay by coculturing with SMSCs at 1, 3, 7 and 14 days. According to the results, SMSCs are similar to other MSCs in their surface epitope expression, which are negative for CD45 and positive for CD44, CD90, and CD105. After incubation in lineage-specific medium, SMSCs could differentiate into chondrocytes, osteocytes and adipocytes. 3D printed nanocomposite scaffolds exhibited a good biocompatibility in the process of coculturing with SMSCs and had no negative effect on cell behavior. The study provides a strategy to obtain SMSCs and fabricate 3D printed nanocomposite scaffolds, the combination of which could be used for practical applications in tissue engineering.

Investigation of different cross-linking approaches on 3D gelatin scaffolds for tissue engineering application: A comparative analysis.

PubMed

Shankar, K Gopal; Gostynska, Natalia; Montesi, Monica; Panseri, Silvia; Sprio, Simone; Kon, Elizaveta; Marcacci, Maurilio; Tampieri, Anna; Sandri, Monica

2017-02-01

The present study aims to investigate the physical-chemical and biological features exhibited by porous scaffolds for regeneration of cartilaginous tissues obtained through stabilization of 3D gelatin hydrogels by physical (DHT), chemical (BDDGE) and natural (Genipin) cross-linking approaches. The study aimed at comparatively assessing the porous microstructure and the long-term resistance of the scaffolds upon degradation in wet physiological conditions (37°C, pH=7.4). The degree of cross-linking increases as function of incorporation of cross-linkers which was maximum up to 73% for BDDGE. The infrared spectroscopy and thermal analysis confirmed the gelatin structure was preserved during the cross-linking treatments. Mechanical properties of the scaffolds were analysed by static and dynamic compression test, which showed different viscoelastic behaviour upon various cross-linking strategies. The biological performance of the scaffolds investigated using human chondrocytes showed good cell adhesion, viability and proliferation, as well as extensive 3D scaffold colonization. Besides, the analysis of gene expression related to the formation of new chondral tissue reported increasing ability with time in the formation of new extra-cellular matrix. In conclusion, out of three different cross-linking methods, the gelatin scaffolds subjected to dehydrothermal treatment (DHT) represented to be the most favourable 3D scaffold for cartilage regeneration. Copyright © 2016 Elsevier B.V. All rights reserved.

Platelet lysate 3D scaffold supports mesenchymal stem cell chondrogenesis: an improved approach in cartilage tissue engineering.

PubMed

Moroz, Andrei; Bittencourt, Renata Aparecida Camargo; Almeida, Renan Padron; Felisbino, Sérgio Luis; Deffune, Elenice

2013-01-01

Articular lesions are still a major challenge in orthopedics because of cartilage's poor healing properties. A major improvement in therapeutics was the development of autologous chondrocytes implantation (ACI), a biotechnology-derived technique that delivers healthy autologous chondrocytes after in vitro expansion. To obtain cartilage-like tissue, 3D scaffolds are essential to maintain chondrocyte differentiated status. Currently, bioactive 3D scaffolds are promising as they can deliver growth factors, cytokines, and hormones to the cells, giving them a boost to attach, proliferate, induce protein synthesis, and differentiate. Using mesenchymal stem cells (MSCs) differentiated into chondrocytes, one can avoid cartilage harvesting. Thus, we investigated the potential use of a platelet-lysate-based 3D bioactive scaffold to support chondrogenic differentiation and maintenance of MSCs. The MSCs from adult rabbit bone marrow (n = 5) were cultivated and characterized using three antibodies by flow cytometry. MSCs (1 × 10(5)) were than encapsulated inside 60 µl of a rabbit platelet-lysate clot scaffold and maintained in Dulbecco's Modified Eagle Medium Nutrient Mixture F-12 supplemented with chondrogenic inductors. After 21 days, the MSCs-seeded scaffolds were processed for histological analysis and stained with toluidine blue. This scaffold was able to maintain round-shaped cells, typical chondrocyte metachromatic extracellular matrix deposition, and isogenous group formation. Cells accumulated inside lacunae and cytoplasm lipid droplets were other observed typical chondrocyte features. In conclusion, the usage of a platelet-lysate bioactive scaffold, associated with a suitable chondrogenic culture medium, supports MSCs chondrogenesis. As such, it offers an alternative tool for cartilage engineering research and ACI.

Osteoimmunomodulatory properties of magnesium scaffolds coated with β-tricalcium phosphate.

PubMed

Chen, Zetao; Mao, Xueli; Tan, Lili; Friis, Thor; Wu, Chengtie; Crawford, Ross; Xiao, Yin

2014-10-01

The osteoimmunomodulatory property of bone biomaterials is a vital property determining the in vivo fate of the implants. Endowing bone biomaterials with favorable osteoimmunomodulatory properties is of great importance in triggering desired immune response and thus supports the bone healing process. Magnesium (Mg) has been recognized as a revolutionary metal for applications in orthopedics due to it being biodegradable, biocompatible, and having osteoconductive properties. However, Mg's high rate of degradation leads to an excessive inflammatory response and this has restricted its application in bone tissue engineering. In this study, β-tricalcium phosphate (β-TCP) was used to coat Mg scaffolds in an effort to modulate the detrimental osteoimmunomodulatory properties of Mg scaffolds, due to the reported favorable osteoimmunomodulatory properties of β-TCP. It was noted that macrophages switched to the M2 extreme phenotype in response to the Mg-β-TCP scaffolds, which could be due to the inhibition of the toll like receptor (TLR) signaling pathway. VEGF and BMP2 were significantly upregulated in the macrophages exposed to Mg-β-TCP scaffolds, indicating pro-osteogenic properties of macrophages in β-TCP modified Mg scaffolds. This was further demonstrated by the macrophage-mediated osteogenic differentiation of bone marrow stromal cells (BMSCs). When BMSCs were stimulated by conditioned medium from macrophages cultured on Mg-β-TCP scaffolds, osteogenic differentiation of BMSCs was significantly enhanced; whereas osteoclastogenesis was inhibited, as indicated by the downregualtion of MCSF, TRAP and inhibition of the RANKL/RANK system. These findings suggest that β-TCP coating of Mg scaffolds can modulate the scaffold's osteoimmunomodulatory properties, shift the immune microenvironment towards one that favors osteogenesis over osteoclastogenesis. Endowing bone biomaterials with favorable osteoimmunomodulatory properties can be a highly valuable strategy for

Two-photon polymerization technique for microfabrication of CAD-designed 3D scaffolds from commercially available photosensitive materials.

PubMed

Ovsianikov, Aleksandr; Schlie, Sabrina; Ngezahayo, Anaclet; Haverich, Axel; Chichkov, Boris N

2007-01-01

We report on recent advances in the fabrication of three-dimensional (3D) scaffolds for tissue engineering and regenerative medicine constructs using a two-photon polymerization technique (2PP). 2PP is a novel CAD/CAM technology allowing the fabrication of any computer-designed 3D structure from a photosensitive polymeric material. The flexibility of this technology and the ability to precisely define 3D construct geometry allows issues associated with vascularization and patient-specific tissue fabrication to be directly addressed. The fabrication of reproducible scaffold structures by 2PP is important for systematic studies of cellular processes and better understanding of in vitro tissue formation. In this study, 2PP was applied for the generation of 3D scaffold-like structures, using the photosensitive organic-inorganic hybrid polymer ORMOCER (ORganically MOdified CERamics) and epoxy-based SU8 materials. By comparing the proliferation rates of cells grown on flat material surfaces and under control conditions, it was demonstrated that ORMOCER and SU8 are not cytotoxic. Additional tests show that the DNA strand breaking of GFSHR-17 granulosa cells was not affected by the presence of ORMOCER. Furthermore, gap junction conductance measurements revealed that ORMOCER did not alter the formation of cell-cell junctions, critical for functional tissue growth. The possibilities of seeding 3D structures with cells were analysed. These studies demonstrate the great potential of 2PP technique for the manufacturing of scaffolds with controlled topology and properties.

Poly(lactide-co-glycolide acid)/biphasic calcium phosphate composite coating on a porous scaffold to deliver simvastatin for bone tissue engineering.

PubMed

Sadiasa, Alexander; Kim, Min Sung; Lee, Byong Taek

2013-09-01

In this study, simvastatin (SIM) drug incorporated poly(D,L-lactic-co-glycolide) (PLGA)/biphasic calcium phosphate (BCP) composite material (SPB) was coated on the BCP/ZrO2 (SPB-BCP/ZrO2) scaffold to enhance the mechanical and bioactive properties of the BCP/ZrO2 scaffold for bone engineering applications. The composite coating was prepared by combining different ratios of PLGA and BCP (1:2, 1:1, 2:1). After completion of the coating process, the compressive strength of the scaffolds was shown to increase with an increase in PLGA concentration from 8.5 ± 0.52 MPa for the SPB1-BCP/ZrO2 (1:2) to 11 ± 0.65 MPa for SPB3-BCP/ZrO2 (2:1) scaffolds when PLGA concentration was increased. Furthermore, the increase of PLGA in the coating composition corresponds to a decrease in porosity, degradation rate and weight loss of the scaffolds after 4 weeks. SIM release study demonstrated sustained release of the drug for the three kinds of scaffolds with improved biocompatibility. The increase of PLGA concentration also resulted in a lower release rate of SIM. Thus, the lower release rate of SIM brought upon by the increase of PLGA concentration further enhanced the performance of the scaffold in vitro making it a promising approach in the field of bone tissue regeneration.

Bio-mimetic hollow scaffolds for long bone replacement

NASA Astrophysics Data System (ADS)

Müller, Bert; Deyhle, Hans; Fierz, Fabienne C.; Irsen, Stephan H.; Yoon, Jin Y.; Mushkolaj, Shpend; Boss, Oliver; Vorndran, Elke; Gburek, Uwe; Degistirici, Özer; Thie, Michael; Leukers, Barbara; Beckmann, Felix; Witte, Frank

2009-08-01

The tissue engineering focuses on synthesis or regeneration of tissues and organs. The hierarchical structure of nearly all porous scaffolds on the macro, micro- and nanometer scales resembles that of engineering foams dedicated for technical applications, but differ from the complex architecture of long bone. A major obstacle of scaffold architecture in tissue regeneration is the limited cell infiltration as the result of the engineering approaches. The biological cells seeded on the three-dimensional constructs are finally only located on the scaffold's periphery. This paper reports on the successful realization of calcium phosphate scaffolds with an anatomical architecture similar to long bones. Two base materials, namely nano-porous spray-dried hydroxyapatite hollow spheres and tri-calcium phosphate powder, were used to manufacture cylindrically shaped, 3D-printed scaffolds with micro-passages and one central macro-canal following the general architecture of long bones. The macro-canal is built for the surgical placement of nerves or larger blood vessels. The micro-passages allow for cell migration and capillary formation through the entire scaffold. Finally, the nanoporosity is essential for the molecule transport crucial for signaling, any cell nutrition and waste removal.

3D bioprinting of BMSC-laden methacrylamide gelatin scaffolds with CBD-BMP2-collagen microfibers.

PubMed

Du, Mingchun; Chen, Bing; Meng, Qingyuan; Liu, Sumei; Zheng, Xiongfei; Zhang, Cheng; Wang, Heran; Li, Hongyi; Wang, Nuo; Dai, Jianwu

2015-12-18

Three-dimensional (3D) bioprinting combines biomaterials, cells and functional components into complex living tissues. Herein, we assembled function-control modules into cell-laden scaffolds using 3D bioprinting. A customized 3D printer was able to tune the microstructure of printed bone mesenchymal stem cell (BMSC)-laden methacrylamide gelatin scaffolds at the micrometer scale. For example, the pore size was adjusted to 282 ± 32 μm and 363 ± 60 μm. To match the requirements of the printing nozzle, collagen microfibers with a length of 22 ± 13 μm were prepared with a high-speed crusher. Collagen microfibers bound bone morphogenetic protein 2 (BMP2) with a collagen binding domain (CBD) as differentiation-control module, from which BMP2 was able to be controllably released. The differentiation behaviors of BMSCs in the printed scaffolds were compared in three microenvironments: samples without CBD-BMP2-collagen microfibers in the growth medium, samples without microfibers in the osteogenic medium and samples with microfibers in the growth medium. The results indicated that BMSCs showed high cell viability (>90%) during printing; CBD-BMP2-collagen microfibers induced BMSC differentiation into osteocytes within 14 days more efficiently than the osteogenic medium. Our studies suggest that these function-control modules are attractive biomaterials and have potential applications in 3D bioprinting.

3D fiber deposited polymeric scaffolds for external auditory canal wall.

PubMed

Mota, Carlos; Milazzo, Mario; Panetta, Daniele; Trombi, Luisa; Gramigna, Vera; Salvadori, Piero A; Giannotti, Stefano; Bruschini, Luca; Stefanini, Cesare; Moroni, Lorenzo; Berrettini, Stefano; Danti, Serena

2018-05-07

The external auditory canal (EAC) is an osseocartilaginous structure extending from the auricle to the eardrum, which can be affected by congenital, inflammatory, and neoplastic diseases, thus reconstructive materials are needed. Current biomaterial-based approaches for the surgical reconstruction of EAC posterior wall still suffer from resorption (biological) and extrusion (synthetic). In this study, 3D fiber deposited scaffolds based on poly(ethylene oxide terephthalate)/poly(butylene terephthalate) were designed and fabricated to replace the EAC wall. Fiber diameter and scaffold porosity were optimized, leading to 200 ± 33 µm and 55% ± 5%, respectively. The mechanical properties were evaluated, resulting in a Young's modulus of 25.1 ± 7.0 MPa. Finally, the EAC scaffolds were tested in vitro with osteo-differentiated human mesenchymal stromal cells (hMSCs) with different seeding methods to produce homogeneously colonized replacements of interest for otologic surgery. This study demonstrated the fabrication feasibility of EAC wall scaffolds aimed to match several important requirements for biomaterial application to the ear under the Tissue Engineering paradigm, including shape, porosity, surface area, mechanical properties and favorable in vitro interaction with osteoinduced hMSCs. This study demonstrated the fabrication feasibility of outer ear canal wall scaffolds via additive manufacturing. Aimed to match several important requirements for biomaterial application to ear replacements under the Tissue Engineering paradigm, including shape, porosity and pore size, surface area, mechanical properties and favorable in vitro interaction with osteo-differentiated mesenchymal stromal cells.

Poly(dopamine) coating of 3D printed poly(lactic acid) scaffolds for bone tissue engineering.

PubMed

Kao, Chia-Tze; Lin, Chi-Chang; Chen, Yi-Wen; Yeh, Chia-Hung; Fang, Hsin-Yuan; Shie, Ming-You

2015-11-01

3D printing is a versatile technique to generate large quantities of a wide variety of shapes and sizes of polymer. The aim of this study is to develop functionalized 3D printed poly(lactic acid) (PLA) scaffolds and use a mussel-inspired surface coating to regulate cell adhesion, proliferation and differentiation of human adipose-derived stem cells (hADSCs). We prepared PLA 3D scaffolds coated with polydopamine (PDA). The chemical composition and surface properties of PDA/PLA were characterized by XPS. PDA/PLA modulated hADSCs' responses in several ways. Firstly, adhesion and proliferation, and cell cycle of hADSCs cultured on PDA/PLA were significantly enhanced relative to those on PLA. In addition, the collagen I secreted from cells was increased and promoted cell attachment and cell cycle progression were depended on the PDA content. In osteogenesis assay, the ALP activity and osteocalcin of hADSCs cultured on PDA/PLA were significantly higher than seen in those cultured on pure PLA scaffolds. Moreover, hADSCs cultured on PDA/PLA showed up-regulation of the ang-1 and vWF proteins associated with angiogenic differentiation. Our results demonstrate that the bio-inspired coating synthetic PLA polymer can be used as a simple technique to render the surfaces of synthetic scaffolds active, thus enabling them to direct the specific responses of hADSCs. Copyright © 2015 Elsevier B.V. All rights reserved.

Fabrication and characterization of toughness-enhanced scaffolds comprising β-TCP/POC using the freeform fabrication system with micro-droplet jetting.

PubMed

Gao, Li; Li, Cuidi; Chen, Fangping; Liu, Changsheng

2015-06-24

A novel elastomeric material, poly(1,8-octanediol-co-citrate) (POC), has demonstrated tremendous versatility because of its advantageous toughness, tunable degradation properties, and efficient drug release capability. In this study, POC was used to improve the mechanical performance of β-tricalcium phosphate (β-Ca3(PO4)2, β-TCP). (3D) β-TCP/POC composite scaffolds were fabricated by a 3D printing technique based on the freeform fabrication system with micro-droplet jetting (FFS-MDJ). The physiochemical properties, compressive modulus, drug release behavior, and cell response of β-TCP/POC composite scaffolds were systematically investigated. The results showed that β-TCP/POC scaffolds had uniform macropores of 300-400 μm, porosity of approximately 45%, biodegradability in phosphate-buffered saline, and high compressive modulus of 50-75 MPa. With the incorporation of POC into β-TCP, the toughness of the composite scaffolds was improved significantly. Moreover, β-TCP/POC scaffolds exhibited sustained drug (ibuprofen (IBU)) release capability. Additionally, β-TCP/POC scaffolds facilitated C2C12 cell attachment and proliferation. It was indicated that the 3D-printed porous β-TCP/POC scaffolds with high compressive modulus and good drug delivery performance might be a promising candidate for bone defect repair.

Characterization of mechanical and biological properties of 3-D scaffolds reinforced with zinc oxide for bone tissue engineering.

PubMed

Feng, Pei; Wei, Pingpin; Shuai, Cijun; Peng, Shuping

2014-01-01

A scaffold for bone tissue engineering should have highly interconnected porous structure, appropriate mechanical and biological properties. In this work, we fabricated well-interconnected porous β-tricalcium phosphate (β-TCP) scaffolds via selective laser sintering (SLS). We found that the mechanical and biological properties of the scaffolds were improved by doping of zinc oxide (ZnO). Our data showed that the fracture toughness increased from 1.09 to 1.40 MPam(1/2), and the compressive strength increased from 3.01 to 17.89 MPa when the content of ZnO increased from 0 to 2.5 wt%. It is hypothesized that the increase of ZnO would lead to a reduction in grain size and an increase in density of the strut. However, the fracture toughness and compressive strength decreased with further increasing of ZnO content, which may be due to the sharp increase in grain size. The biocompatibility of the scaffolds was investigated by analyzing the adhesion and the morphology of human osteoblast-like MG-63 cells cultured on the surfaces of the scaffolds. The scaffolds exhibited better and better ability to support cell attachment and proliferation when the content of ZnO increased from 0 to 2.5 wt%. Moreover, a bone like apatite layer formed on the surfaces of the scaffolds after incubation in simulated body fluid (SBF), indicating an ability of osteoinduction and osteoconduction. In summary, interconnected porous β-TCP scaffolds doped with ZnO were successfully fabricated and revealed good mechanical and biological properties, which may be used for bone repair and replacement potentially.

In vitro degradation of a 3D porous Pennisetum purpureum/PLA biocomposite scaffold.

PubMed

Revati, R; Majid, M S Abdul; Ridzuan, M J M; Basaruddin, K S; Rahman Y, M N; Cheng, E M; Gibson, A G

2017-10-01

The in vitro degradation and mechanical properties of a 3D porous Pennisetum purpureum (PP)/polylactic acid (PLA)-based scaffold were investigated. In this study, composite scaffolds with PP to PLA ratios of 0%, 10%, 20%, and 30% were immersed in a PBS solution at 37°C for 40 days. Compression tests were conducted to evaluate the compressive strength and modulus of the scaffolds, according to ASTM F451-95. The compression strength of the scaffolds was found to increase from 1.94 to 9.32MPa, while the compressive modulus increased from 1.73 to 5.25MPa as the fillers' content increased from 0wt% to 30wt%. Moreover, field emission scanning electron microscopy (FESEM) and X-ray diffraction were employed to observe and analyse the microstructure and fibre-matrix interface. Interestingly, the degradation rate was reduced for the PLA/PP 20 scaffold, though insignificantly, this could be attributed to the improved mechanical properties and stronger fibre-matrix interface. Microstructure changes after degradation were observed using FESEM. The FESEM results indicated that a strong fibre-matrix interface was formed in the PLA/PP 20 scaffold, which reflected the addition of P. purpureum into PLA decreasing the degradation rate compared to in pure PLA scaffolds. The results suggest that the P. purpureum/PLA scaffold degradation rate can be altered and controlled to meet requirements imposed by a given tissue engineering application. Copyright © 2017 Elsevier Ltd. All rights reserved.

Development of various composition multicomponent chitosan/fish collagen/glycerin 3D porous scaffolds: Effect on morphology, mechanical strength, biostability and cytocompatibility.

PubMed

Ullah, Saleem; Zainol, Ismail; Chowdhury, Shiplu Roy; Fauzi, M B

2018-05-01

The various composition multicomponent chitosan/fish collagen/glycerin 3D porous scaffolds were developed and investigated the effect of various composition chitosan/fish collagen/glycerin on scaffolds morphology, mechanical strength, biostability and cytocompatibility. The scaffolds were fabricated via freeze-drying technique. The effects of various compositions consisting in 3D scaffolds were investigated via FT-IR analysis, porosity, swelling and mechanical tests, and effect on the morphology of scaffolds investigated microscopically. The biostability and cytocompatibility tests were used to explore the ability of scaffolds to use for tissue engineering application. The average pore sizes of scaffolds were in range of 100.73±27.62-116.01±52.06, porosity 71.72±3.46-91.17±2.42%, tensile modulus in dry environment 1.47±0.08-0.17±0.03MPa, tensile modulus in wet environment 0.32±0.03-0.14±0.04MPa and biodegradation rate (at day 30) 60.38±0.70-83.48±0.28%. In vitro culture of human fibroblasts and keratinocytes showed that the various composition multicomponent 3D scaffolds were good cytocompatibility however, the scaffolds contained high amount of fish collagen excellently facilitated cell proliferation and adhesion. It was found that the high amount fish collagen and glycerin scaffolds have high porosity, enough mechanical strength and biostability, and excellent cytocompatibility. Copyright © 2018 Elsevier B.V. All rights reserved.

Strain-controlled fatigue behaviors of porous PLA-based scaffolds by 3D-printing technology.

PubMed

Gong, Baoming; Cui, Shaohua; Zhao, Yun; Sun, Yongtao; Ding, Qian

2017-12-01

In the study, the low-cycle fatigue behaviors of 3D-printed poly lactic acid (PLA) scaffolds with 60% porosity and two kinds of geometrical pores were investigated under strain-controlled loading. The obtained Δε a -N f curves were fitted by Coffin-Manson relation. The mechanical stability of the porous structure under cyclic loading was studied. Both kinds of specimens undergo the strain softening after the initial cyclic hardening. The scaffold with circular pore exhibits stable resistance to the fatigue damage which is desirable for bone repairing. Regarding to the accumulation of inelastic deformation, the triangular-scaffold is more sensitive to the cyclic load. The superior fatigue behaviors of the scaffold with circular pore is attributed to homogeneous distribution of the applied mechanical stress and diminishing stress concentration by the introduction of circular pore.

Flow cytometric cell cycle analysis of muscle precursor cells cultured within 3D scaffolds in a perfusion bioreactor.

PubMed

Flaibani, Marina; Luni, Camilla; Sbalchiero, Elisa; Elvassore, Nicola

2009-01-01

It has been widely demonstrated that perfusion bioreactors improve in vitro three-dimensional (3D) cultures in terms of high cell density and uniformity of cell distribution; however, the studies reported in literature were primarily based on qualitative analysis (histology, immunofluorescent staining) or on quantitative data averaged on the whole population (DNA assay, PCR). Studies on the behavior, in terms of cell cycle, of a cell population growing in 3D scaffolds in static or dynamic conditions are still absent. In this work, a perfusion bioreactor suitable to culture C(2)C(12) muscle precursor cells within 3D porous collagen scaffolds was designed and developed and a method based on flowcytometric analyses for analyzing the cell cycle in the cell population was established. Cells were extracted by enzymatic digestion of the collagen scaffolds after 4, 7, and 10 days of culture, and flow cytometric live/dead and cell cycle analyses were performed with Propidium Iodide. A live/dead assay was used for validating the method for cell extraction and staining. Moreover, to investigate spatial heterogeneity of the cell population under perfusion conditions, two stacked scaffolds in the 3D domain, of which only the upstream layer was seeded, were analyzed separately. All results were compared with those obtained from static 3D cultures. The live/dead assay revealed the presence of less than 20% of dead cells, which did not affect the cell cycle analysis. Cell cycle analyses highlighted the increment of cell fractions in proliferating phases (S/G(2)/M) owing to medium perfusion in long-term cultures. After 7-10 days, the percentage of proliferating cells was 8-12% for dynamic cultures and 3-5% for the static controls. A higher fraction of proliferating cells was detected in the downstream scaffold. From a general perspective, this method provided data with a small standard deviation and detected the differences between static and dynamic cultures and between upper and

Pore architecture and cell viability on freeze dried 3D recombinant human collagen-peptide (RHC)-chitosan scaffolds.

PubMed

Zhang, Jing; Zhou, Aimei; Deng, Aipeng; Yang, Yang; Gao, Lihu; Zhong, Zhaocai; Yang, Shulin

2015-04-01

Pore architecture of 3D scaffolds used in tissue engineering plays a critical role in the maintenance of cell survival, proliferation and further promotion of tissue regeneration. We investigated the pore size and structure, porosity, swelling as well as cell viability of a series of recombinant human collagen-peptide-chitosan (RHCC) scaffolds fabricated by lyophilization. In this paper, freezing regime containing a final temperature of freezing (Tf) and cooling rates was applied to obtain scaffolds with pore size ranging from 100μm to 120μm. Other protocols of RHC/chitosan suspension concentration and ratio modification were studied to produce more homogenous and appropriate structural scaffolds. The mean pore size decreased along with the decline of Tf at a slow cooling rate of 0.7°C/min; a more rapid cooling rate under 5°C/min resulted to a smaller pore size and more homogenous microstructure. High concentration could reduce pore size and lead to thick well of scaffold, while improved the ratio of RHC, lamellar and fiber structure coexisted with cellular pores. Human umbilical vein endothelial cells (HUVECs) were seeded on these manufactured scaffolds, the cell viability represented a negative correlation to the pore size. This study provides an alternative method to fabricate 3D RHC-chitosan scaffolds with appropriate pores for potential tissue engineering. Copyright © 2014 Elsevier B.V. All rights reserved.

3-D loaded scaffolds obtained by supercritical CO2 assisted process

NASA Astrophysics Data System (ADS)

Cardea, S.; Reverchon, E.

2014-08-01

In this work, a supercritical CO2 (SC-CO2) drying process for the formation of 3-D PVDF-HFP loaded scaffolds was tested. Experiments at pressures ranging between 150 and 250 bar and at temperatures ranging between 35 and 55°C were performed. The PVDF-HFP- acetone-ethanol solution at 15% w/w polymer was selected as the base case. The drug (amoxicillin) concentration was varied from 20 to 30% w/w with respect to PVDF-HFP. SC- CO2 drying process was confirmed to be a valid alternative to generate loaded structures; indeed, scaffolds characterized by nanometric networks (with mean pore diameter of about 300 nm) with a homogeneous drug distribution were obtained. Drug controlled release experiments were also performed and a quasi-zero order release kinetic was observed.

3D-Printed Scaffolds and Biomaterials: Review of Alveolar Bone Augmentation and Periodontal Regeneration Applications

PubMed Central

Asa'ad, Farah; Giannì, Aldo Bruno; Giannobile, William V.; Rasperini, Giulio

2016-01-01

To ensure a successful dental implant therapy, the presence of adequate vertical and horizontal alveolar bone is fundamental. However, an insufficient amount of alveolar ridge in both dimensions is often encountered in dental practice due to the consequences of oral diseases and tooth loss. Although postextraction socket preservation has been adopted to lessen the need for such invasive approaches, it utilizes bone grafting materials, which have limitations that could negatively affect the quality of bone formation. To overcome the drawbacks of routinely employed grafting materials, bone graft substitutes such as 3D scaffolds have been recently investigated in the dental field. In this review, we highlight different biomaterials suitable for 3D scaffold fabrication, with a focus on “3D-printed” ones as bone graft substitutes that might be convenient for various applications related to implant therapy. We also briefly discuss their possible adoption for periodontal regeneration. PMID:27366149

Cell-secreted extracellular matrix formation and differentiation of adipose-derived stem cells in 3D alginate scaffolds with tunable properties.

PubMed

Guneta, Vipra; Loh, Qiu Li; Choong, Cleo

2016-05-01

Three dimensional (3D) alginate scaffolds with tunable mechanical and structural properties are explored for investigating the effect of the scaffold properties on stem cell behavior and extracellular matrix (ECM) formation. Varying concentrations of crosslinker (20 - 60%) are used to tune the stiffness, porosity, and the pore sizes of the scaffolds post-fabrication. Enhanced cell proliferation and adipogenesis occur in scaffolds with 3.52 ± 0.59 kPa stiffness, 87.54 ± 18.33% porosity and 68.33 ± 0.88 μm pore size. On the other hand, cells in scaffolds with stiffness greater than 11.61 ± 1.74 kPa, porosity less than 71.98 ± 6.25%, and pore size less than 64.15 ± 4.34 μm preferentially undergo osteogenesis. When cultured in differentiation media, adipose-derived stem cells (ASCs) undergoing terminal adipogenesis in 20% firming buffer (FB) scaffolds and osteogenesis in 40% and 60% FB scaffolds show the highest secretion of collagen as compared to other groups of scaffolds. Overall, this study demonstrates the three-way relationship between 3D scaffolds, ECM composition, and stem cell differentiation. © 2016 Wiley Periodicals, Inc.

Mechanical, thermal and morphological characterisation of 3D porous Pennisetum purpureum/PLA biocomposites scaffold.

PubMed

Revati, R; Abdul Majid, M S; Ridzuan, M J M; Normahira, M; Mohd Nasir, N F; Rahman Y, M N; Gibson, A G

2017-06-01

The mechanical, thermal, and morphological properties of a 3D porous Pennisetum purpureum (PP)/polylactic acid (PLA) based scaffold were investigated. In this study, a scaffold containing P. purpureum and PLA was produced using the solvent casting and particulate leaching method. P. purpureum fibre, also locally known as Napier grass, is composed of 46% cellulose, 34% hemicellulose, and 20% lignin. PLA composites with various P. purpureum contents (10%, 20%, and 30%) were prepared and subsequently characterised. The morphologies, structures and thermal behaviours of the prepared composite scaffolds were characterised using field-emission scanning electron microscopy (FESEM), Fourier transform infrared spectroscopy (FTIR), and thermogravimetric analysis (TGA). The morphology was studied using FESEM; the scaffold possessed 70-200μm-sized pores with a high level of interconnectivity. The moisture content and mechanical properties of the developed porous scaffolds were further characterised. The P. purpureum/PLA scaffold had a greater porosity factor (99%) and compression modulus (5.25MPa) than those of the pure PLA scaffold (1.73MPa). From the results, it can be concluded that the properties of the highly porous P. purpureum/PLA scaffold developed in this study can be controlled and optimised. This can be used to facilitate the construction of implantable tissue-engineered cartilage. Copyright © 2017 Elsevier B.V. All rights reserved.

Hierarchically porous structure, mechanical strength and cell biological behaviors of calcium phosphate composite scaffolds prepared by combination of extrusion and porogen burnout technique and enhanced by gelatin.

PubMed

Feng, Shenglei; He, Fupo; Ye, Jiandong

2018-01-01

In this study, hierarchically porous calcium phosphate scaffolds (HTCP) with unidirectional pores, transversely interconnected pores, and micropores were fabricated by the combination of extrusion and porogen burnout technique. Gelatin was incorporated into the HTCP scaffolds by vacuum-impregnation of gelatin solution and subsequent freeze-drying. The phase composition, microstructure, physical and cytobiological properties were analyzed. The results showed that the HTCP scaffolds were composed of β-tricalcium phosphate with minor hydroxyapatite. The HTCP scaffolds had unidirectional pores (~400μm), transversely interconnected pores (~130μm) and micropores (~1μm). The incorporation of gelatin significantly increased the compressive strength, toughness, and cell seeding of the HTCP scaffolds. The composite scaffolds showed excellent cytocompatibility. The hierarchically porous calcium phosphate composite scaffolds may have potential application prospects in bone tissue engineering. Copyright © 2017 Elsevier B.V. All rights reserved.

Characterization of a recombinant type II 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase from Helicobacter pylori.

PubMed

Webby, Celia J; Patchett, Mark L; Parker, Emily J

2005-08-15

DAH7P (3-Deoxy-D-arabino-heptulosonate 7-phosphate) synthase catalyses the condensation reaction between phosphoenolpyruvate (PEP) and D-erythrose 4-phosphate (E4P) as the first committed step in the biosynthesis of aromatic compounds in plants and micro-organisms. Previous work has identified two families of DAH7P synthases based on sequence similarity and molecular mass, with the majority of the mechanistic and structural studies being carried out on the type I paralogues from Escherichia coli. Whereas a number of organisms possess genes encoding both type I and type II DAH7P synthases, the pathogen Helicobacter pylori has only a single, type II, enzyme. Recombinant DAH7P synthase from H. pylori was partially solubilized by co-expression with chaperonins GroEL/GroES in E. coli, and purified to homogeneity. The enzyme reaction follows an ordered sequential mechanism with the following kinetic parameters: K(m) (PEP), 3 microM; K(m) (E4P), 6 microM; and kcat, 3.3 s(-1). The enzyme reaction involves interaction of the si face of PEP with the re face of E4P. H. pylori DAH7P synthase is not inhibited by phenylalanine, tyrosine, tryptophan or chorismate. EDTA inactivates the enzyme, and activity is restored by a range of bivalent metal ions, including (in order of decreasing effectiveness) Co2+, Mn2+, Ca2+, Mg2+, Cu2+ and Zn2+. Analysis of type II DAH7P synthase sequences reveals several highly conserved motifs, and comparison with the type I enzymes suggests that catalysis by these two enzyme types occurs on a similar active-site scaffold and that the two DAH7P synthase families may indeed be distantly related.

Constructing a novel hierarchical 3D flower-like nano/micro titanium phosphate with efficient hydrogen evolution from water splitting

NASA Astrophysics Data System (ADS)

Guo, Si-yao; Han, Song

2014-12-01

A novel nano/micro hierarchical structured titanium phosphate with unique 3D flower-like morphology has been prepared by a simple hydrothermal method without adding any surfactants. The shape of the titanium phosphate could be controlled by simply adjusting the concentration of phosphoric acid. The 3D flower-like titanium phosphate with diameter of 2-3 μm is characterized by the assembly of numerous porous and connected lamella structures. Interestingly, this novel hierarchical mesoporous 3D flower-like titanium exhibits enhanced hydrogen evolution from water splitting under xenon lamp irradiation in the presence of methanol as the sacrificial reagent, which is also the first example of 3D flower-like titanium phosphate with high photocatalytic activity for water splitting. Since the use of titanium phosphate as a photocatalyst has been mostly neglected up to now, this low-cost, simple procedure and large-scale yield of 3D nano/micro structure titanium phosphate could be expected to be applicable in the synthesis of controlled, reproducible and robust photocatalytic systems.

An engineering perspective on 3D printed personalized scaffolds for tracheal suspension technique

PubMed Central

An, Jia

2016-01-01

3D printing is a large family of many distinct technologies covering a wide range of topics. From an engineering point of view, there should be considerations for selection of design, material, and process when using 3D printing for surgical technique innovation such as personalized scaffolds. Moreover, cost should also be considered if there are equally effective alternatives to the innovation. Furthermore, engineering considerations and options should be clearly communicated and readily available to surgeons for advancement in future. PMID:28149624

An engineering perspective on 3D printed personalized scaffolds for tracheal suspension technique.

PubMed

An, Jia; Chua, Chee Kai

2016-12-01

3D printing is a large family of many distinct technologies covering a wide range of topics. From an engineering point of view, there should be considerations for selection of design, material, and process when using 3D printing for surgical technique innovation such as personalized scaffolds. Moreover, cost should also be considered if there are equally effective alternatives to the innovation. Furthermore, engineering considerations and options should be clearly communicated and readily available to surgeons for advancement in future.

Preparation and Evaluation of Gelatin-Chitosan-Nanobioglass 3D Porous Scaffold for Bone Tissue Engineering

PubMed Central

Maji, Kanchan; Dasgupta, Sudip; Pramanik, Krishna; Bissoyi, Akalabya

2016-01-01

The aim of the present study was to prepare and characterize bioglass-natural biopolymer based composite scaffold and evaluate its bone regeneration ability. Bioactive glass nanoparticles (58S) in the size range of 20–30 nm were synthesized using sol-gel method. Porous scaffolds with varying bioglass composition from 10 to 30 wt% in chitosan, gelatin matrix were fabricated using the method of freeze drying of its slurry at 40 wt% solids loading. Samples were cross-linked with glutaraldehyde to obtain interconnected porous 3D microstructure with improved mechanical strength. The prepared scaffolds exhibited >80% porosity with a mean pore size range between 100 and 300 microns. Scaffold containing 30 wt% bioglass (GCB 30) showed a maximum compressive strength of 2.2 ± 0.1 MPa. Swelling and degradation studies showed that the scaffold had excellent properties of hydrophilicity and biodegradability. GCB 30 scaffold was shown to be noncytotoxic and supported mesenchymal stem cell attachment, proliferation, and differentiation as indicated by MTT assay and RUNX-2 expression. Higher cellular activity was observed in GCB 30 scaffold as compared to GCB 0 scaffold suggesting the fact that 58S bioglass nanoparticles addition into the scaffold promoted better cell adhesion, proliferation, and differentiation. Thus, the study showed that the developed composite scaffolds are potential candidates for regenerating damaged bone tissue. PMID:26884764

Dietary vitamin D intake is not associated with 25-hydroxyvitamin D3 or parathyroid hormone in elderly subjects, whereas the calcium-to-phosphate ratio affects parathyroid hormone.

PubMed

Jungert, Alexandra; Neuhäuser-Berthold, Monika

2013-08-01

This cross-sectional study investigates whether serum 25-hydroxyvitamin D3 [25(OH)D3] and intact parathyroid hormone (iPTH) are affected by vitamin D, calcium, or phosphate intake in 140 independently living elderly subjects from Germany (99 women and 41 men; age, 66-96 years). We hypothesized that habitual dietary intakes of vitamin D, calcium, and phosphate are not associated with 25(OH)D3 or iPTH and that body mass index confounds these associations. Serum 25(OH)D3 and iPTH were measured by an electrochemiluminescence immunoassay. Dietary intake was determined using a 3-day estimated dietary record. The median dietary intake levels of vitamin D, calcium, and phosphate were 3 μg/d, 999 mg/d, and 1250 mg/d, respectively. Multiple regression analyses confirmed that dietary vitamin D and calcium did not affect 25(OH)D3 or iPTH; however, supplemental intakes of vitamin D and calcium were associated with 25(OH)D3 after adjustment for age, sex, body composition, sun exposure, physical activity, and smoking. In addition, phosphate intake and the calcium-to-phosphate ratio were associated with iPTH after multiple adjustments. In a subgroup analysis, calcium and vitamin D supplements, as well as phosphate intake, were associated with 25(OH)D3 and/or iPTH in normal-weight subjects only. Our results indicate that habitual dietary vitamin D and calcium intakes have no independent effects on 25(OH)D3 or iPTH in elderly subjects without vitamin D deficiency, whereas phosphate intake and the calcium-to-phosphate ratio affect iPTH. However, vitamin D and calcium supplements may increase 25(OH)D3 and decrease iPTH, even during the summer, but the impact of supplements may depend on body mass index. Copyright © 2013. Published by Elsevier Inc.

Effect of in vitro enzymatic degradation on 3D printed poly(ε-caprolactone) scaffolds: morphological, chemical and mechanical properties.

PubMed

Ferreira, Joana; Gloria, Antonio; Cometa, Stefania; Coelho, Jorge F J; Domingos, Marco

2017-07-27

In recent years, the tissue engineering (TE) field has significantly benefited from advanced techniques such as additive manufacturing (AM), for the design of customized 3D scaffolds with the aim of guided tissue repair. Among the wide range of materials available to biomanufacture 3D scaffolds, poly(ε-caprolactone) (PCL) clearly arises as the synthetic polymer with the greatest potential, due to its unique properties - namely, biocompatibility, biodegradability, thermal and chemical stability and processability. This study aimed for the first time to investigate the effect of pore geometry on the in vitro enzymatic chain cleavage mechanism of PCL scaffolds manufactured by the AM extrusion process. Methods: Morphological properties of 3D printed PCL scaffolds before and after degradation were evaluated using Scanning Electron Microscopy (SEM) and micro-computed tomography (μ-CT). Differential Scanning Calorimetry (DSC) was employed to determine possible variations in the crystallinity of the scaffolds during the degradation period. The molecular weight was assessed using Size Exclusion Chromatography (SEC) while the mechanical properties were investigated under static compression conditions. Morphological results suggested a uniform reduction of filament diameter, while increasing the scaffolds' porosity. DSC analysis revealed and increment in the crystallinity degree while the molecular weight, evaluated through SEC, remained almost constant during the incubation period (25 days). Mechanical analysis highlighted a decrease in the compressive modulus and maximum stress over time, probably related to the significant weight loss of the scaffolds. All of these results suggest that PCL scaffolds undergo enzymatic degradation through a surface erosion mechanism, which leads to significant variations in mechanical, physical and chemical properties, but which has little influence on pore geometry.

3D printing for the design and fabrication of polymer-based gradient scaffolds.

PubMed

Bracaglia, Laura G; Smith, Brandon T; Watson, Emma; Arumugasaamy, Navein; Mikos, Antonios G; Fisher, John P

2017-07-01

To accurately mimic the native tissue environment, tissue engineered scaffolds often need to have a highly controlled and varied display of three-dimensional (3D) architecture and geometrical cues. Additive manufacturing in tissue engineering has made possible the development of complex scaffolds that mimic the native tissue architectures. As such, architectural details that were previously unattainable or irreproducible can now be incorporated in an ordered and organized approach, further advancing the structural and chemical cues delivered to cells interacting with the scaffold. This control over the environment has given engineers the ability to unlock cellular machinery that is highly dependent upon the intricate heterogeneous environment of native tissue. Recent research into the incorporation of physical and chemical gradients within scaffolds indicates that integrating these features improves the function of a tissue engineered construct. This review covers recent advances on techniques to incorporate gradients into polymer scaffolds through additive manufacturing and evaluate the success of these techniques. As covered here, to best replicate different tissue types, one must be cognizant of the vastly different types of manufacturing techniques available to create these gradient scaffolds. We review the various types of additive manufacturing techniques that can be leveraged to fabricate scaffolds with heterogeneous properties and discuss methods to successfully characterize them. Additive manufacturing techniques have given tissue engineers the ability to precisely recapitulate the native architecture present within tissue. In addition, these techniques can be leveraged to create scaffolds with both physical and chemical gradients. This work offers insight into several techniques that can be used to generate graded scaffolds, depending on the desired gradient. Furthermore, it outlines methods to determine if the designed gradient was achieved. This review

Phage nanofibers induce vascularized osteogenesis in 3D printed bone scaffolds.

PubMed

Wang, Jianglin; Yang, Mingying; Zhu, Ye; Wang, Lin; Tomsia, Antoni P; Mao, Chuanbin

2014-08-06

A virus-activated matrix is developed to overcome the challenge of forming vascularized bone tissue. It is generated by filling a 3D printed bioceramic scaffold with phage nanofibers displaying high-density RGD peptide. After it is seeded with mesenchymal stem cells (MSCs) and implanted into a bone defect, the phage nanofibers induce osteogenesis and angiogenesis by activating endothelialization and osteogenic differentiation of MSCs. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

In vitro cytocompatibility evaluation of chitosan/graphene oxide 3D scaffold composites designed for bone tissue engineering.

PubMed

Dinescu, Sorina; Ionita, Mariana; Pandele, Andreea Madalina; Galateanu, Bianca; Iovu, Horia; Ardelean, Aurel; Costache, Marieta; Hermenean, Anca

2014-01-01

Extensively studied nowadays, graphene oxide (GO) has a benefic effect on cell proliferation and differentiation, thus holding promise for bone tissue engineering (BTE) approaches. The aim of this study was not only to design a chitosan 3D scaffold improved with GO for optimal BTE, but also to analyze its physicochemical properties and to evaluate its cytocompatibility and ability to support cell metabolic activity and proliferation. Overall results show that the addition of GO in the scaffold's composition improved mechanical properties and pore formation and enhanced the bioactivity of the scaffold material for tissue engineering. The new developed CHT/GO 3 wt% scaffold could be a potential candidate for further in vitro and in vivo osteogenesis studies and BTE approaches.

Comparative Chondrogenesis of Human Cell Sources in 3D Scaffolds

PubMed Central

Tıg̑lı, R. Seda; Ghosh, Sourabh; Laha, Michael M.; Shevde, Nirupama K.; Daheron, Laurence; Gimble, Jeffrey; Gümüşdereliog̑lu, Menemşe; Kaplan, David L.

2009-01-01

Cartilage tissue can be engineered by starting from a diversity of cell sources, including stem-cell based and primary cell-based platforms. Selecting an appropriate cell source for the process of cartilage tissue engineering or repair is critical and challenging due to the variety of cell options available. In this study, cellular responses of isolated human chondrocytes, human embryonic stem cells and mesenchymal stem cells (MSCs) derived from three sources, human embryonic stem cells, bone marrow and adipose tissue, were assessed for chondrogenic potential in 3D culture. All cell sources were characterized by FACS analysis to compare expression of some surface markers. The cells were differentiated in two different biomaterial matrices, silk and chitosan scaffolds, in the presence and absence of bone morphogenetic protein 6 (BMP-6) along with the standard chondrogenic differentiating factors. Embryonic stem cells derived MSCs showed unique characteristics with preserved chondrogenic phenotype in both scaffolds with regard to chondrogenesis, as determined by real time RT-PCR, histological and microscopic analyses. After 4 weeks of cultivation, embryonic stem cells derived MSCs were promising for chondrogenesis, particularly in the silk scaffolds with BMP-6. The results suggest that cell source differences are important to consider with regard to chondrogenic outcomes and with the variables addressed here, the human embryonic stem cells derived MSCs were the preferred cell source. PMID:19382119

Biomimetic fabrication of a three-level hierarchical calcium phosphate/collagen/hydroxyapatite scaffold for bone tissue engineering.

PubMed

Zhou, Changchun; Ye, Xingjiang; Fan, Yujiang; Ma, Liang; Tan, Yanfei; Qing, Fangzu; Zhang, Xingdong

2014-09-01

A three-level hierarchical calcium phosphate/collagen/hydroxyapatite (CaP/Col/HAp) scaffold for bone tissue engineering was developed using biomimetic synthesis. Porous CaP ceramics were first prepared as substrate materials to mimic the porous bone structure. A second-level Col network was then composited into porous CaP ceramics by vacuum infusion. Finally, a third-level HAp layer was achieved by biomimetic mineralization. The three-level hierarchical biomimetic scaffold was characterized using scanning electron microscopy, energy-dispersive x-ray spectra, x-ray diffraction and Fourier transform infrared spectroscopy, and the mechanical properties of the scaffold were evaluated using dynamic mechanical analysis. The results show that this scaffold exhibits a similar structure and composition to natural bone tissues. Furthermore, this three-level hierarchical biomimetic scaffold showed enhanced mechanical strength compared with pure porous CaP scaffolds. The biocompatibility and osteoinductivity of the biomimetic scaffolds were evaluated using in vitro and in vivo tests. Cell culture results indicated the good biocompatibility of this biomimetic scaffold. Faster and increased bone formation was observed in these scaffolds following a six-month implantation in the dorsal muscles of rabbits, indicating that this biomimetic scaffold exhibits better osteoinductivity than common CaP scaffolds.

Enhanced osteogenesis of β-tricalcium phosphate reinforced silk fibroin scaffold for bone tissue biofabrication.

PubMed

Lee, Dae Hoon; Tripathy, Nirmalya; Shin, Jae Hun; Song, Jeong Eun; Cha, Jae Geun; Min, Kyung Dan; Park, Chan Hum; Khang, Gilson

2017-02-01

Scaffolds, used for tissue regeneration are important to preserve their function and morphology during tissue healing. Especially, scaffolds for bone tissue engineering should have high mechanical properties to endure load of bone. Silk fibroin (SF) from Bombyx mori silk cocoon has potency as a type of biomaterials in the tissue engineering. β-tricalcium phosphate (β-TCP) as a type of bioceramics is also critical as biomaterials for bone regeneration because of its biocompatibility, osteoconductivity, and mechanical strength. The aim of this study was to fabricate three-dimensional SF/β-TCP scaffolds and access its availability for bone grafts through in vitro and in vivo test. The scaffolds were fabricated in each different ratios of SF and β-TCP (100:0, 75:25, 50:50, 25:75). The characterizations of scaffolds were conducted by FT-IR, compressive strength, porosity, and SEM. The in vitro and in vivo tests were carried out by MTT, ALP, RT-PCR, SEM, μ-CT, and histological staining. We found that the SF/β-TCP scaffolds have high mechanical strength and appropriate porosity for bone tissue engineering. The study showed that SF/β-TCP (75:25) scaffold exhibited the highest osteogenesis compared with other scaffolds. The results suggested that SF/β-TCP (75:25) scaffold can be applied as one of potential bone grafts for bone tissue engineering. Copyright © 2016. Published by Elsevier B.V.

New developments in 3D liquid crystal elastomers scaffolds for tissue engineering: from physical template to responsive substrate

NASA Astrophysics Data System (ADS)

Prévôt, Marianne E.; Bergquist, Leah E.; Sharma, Anshul; Mori, Taizo; Gao, Yungxiang; Bera, Tanmay; Zhu, Chenhui; Leslie, Michelle T.; Cukelj, Richard; Korley, LaShanda T. J.; Freeman, Ernest J.; McDonough, Jennifer A.; Clements, Robert J.; Hegmann, Elda

2017-08-01

We report here on cell growth and proliferation within a 3D architecture created using smectic liquid crystal elastomers (LCEs) leading to a responsive scaffold for tissue engineering. The investigated LCE scaffolds exhibit biocompatibility, controlled degradability, with mechanical properties and morphologies that can match development of the extracellular matrix. Moreover, the synthetic pathway and scaffold design offer a versatility of processing, allowing modifications of the surface such as adjusting the hydrophilic/hydrophobic balance and the mobility of the LC moieties to enhance the biomaterial performance. First, we succeeded in generating LCEs whose mechanical properties mimic muscle tissue. In films, our LCEs showed cell adhesion, proliferation, and alignment. We also achieved creating 3D LCE structures using either metallic template or microsphere scaffolds. Finally, we recorded a four times higher cell proliferation capability in comparison to conventional porous films and, most importantly, anisotropic cell growth that highlights the tremendous effect of liquid crystal moieties within LCEs on the cell environment.

Construction of a 3D rGO-collagen hybrid scaffold for enhancement of the neural differentiation of mesenchymal stem cells

NASA Astrophysics Data System (ADS)

Guo, Weibo; Wang, Shu; Yu, Xin; Qiu, Jichuan; Li, Jianhua; Tang, Wei; Li, Zhou; Mou, Xiaoning; Liu, Hong; Wang, Zhonglin

2016-01-01

The cell-material interface is one of the most important considerations in designing a high-performance tissue engineering scaffold because the surface of the scaffold can determine the fate of stem cells. A conductive surface is required for a scaffold to direct stem cells toward neural differentiation. However, most conductive polymers are toxic and not amenable to biological degradation, which restricts the design of neural tissue engineering scaffolds. In this study, we used a bioactive three-dimensional (3D) porcine acellular dermal matrix (PADM), which is mainly composed of type I collagen, as a basic material and successfully assembled a layer of reduced graphene oxide (rGO) nanosheets on the surface of the PADM channels to obtain a porous 3D, biodegradable, conductive and biocompatible PADM-rGO hybrid neural tissue engineering scaffold. Compared with the PADM scaffold, assembling the rGO into the scaffold did not induce a significant change in the microstructure but endowed the PADM-rGO hybrid scaffold with good conductivity. A comparison of the neural differentiation of rat bone-marrow-derived mesenchymal stem cells (MSCs) was performed by culturing the MSCs on PADM and PADM-rGO scaffolds in neuronal culture medium, followed by the determination of gene expression and immunofluorescence staining. The results of both the gene expression and protein level assessments suggest that the rGO-assembled PADM scaffold may promote the differentiation of MSCs into neuronal cells with higher protein and gene expression levels after 7 days under neural differentiation conditions. This study demonstrated that the PADM-rGO hybrid scaffold is a promising scaffold for neural tissue engineering; this scaffold can not only support the growth of MSCs at a high proliferation rate but also enhance the differentiation of MSCs into neural cells.The cell-material interface is one of the most important considerations in designing a high-performance tissue engineering scaffold

Anti-infective efficacy, cytocompatibility and biocompatibility of a 3D-printed osteoconductive composite scaffold functionalized with quaternized chitosan.

PubMed

Yang, Ying; Yang, Shengbing; Wang, Yugang; Yu, Zhifeng; Ao, Haiyong; Zhang, Hongbo; Qin, Ling; Guillaume, Olivier; Eglin, David; Richards, R Geoff; Tang, Tingting

2016-12-01

Contaminated or infected bone defects remain serious challenges in clinical trauma and orthopaedics, and a bone substitute with both osteoconductivity and antibacterial properties represents an improvement for treatment strategy. In this study, quaternized chitosan (hydroxypropyltrimethyl ammonium chloride chitosan, HACC) was grafted to 3D-printed scaffolds composed of polylactide-co-glycolide (PLGA) and hydroxyapatite (HA), in order to design bone engineering scaffolds endowed with antibacterial and osteoconductive properties. We found that both the PLGA/HA/HACC and PLGA/HACC composite scaffolds decreased bacterial adhesion and biofilm formation under in vitro and in vivo conditions. Additionally, ATP leakage assay indicated that immobilizing HACC on the scaffolds could effectively disrupt microbial membranes. Using human bone marrow-derived mesenchymal stem cells (hBMSCs), we demonstrated that HA incorporated scaffolds, including PLGA/HA and PLGA/HA/HACC, favoured cell attachment, proliferation, spreading and osteogenic differentiation compared to HA-free PLGA or PLGA/HACC scaffolds. Finally, an in vivo biocompatibility assay conducted on rats, showed that HA incorporated scaffolds (including PLGA/HA and PLGA/HA/HACC scaffolds) exhibited good neovascularization and tissue integration. Taken together, our findings support the approach for developing porous PLGA/HA/HACC composite scaffold with potential clinical application in the treatment of infected bone. Although plenty of conductive scaffold biomaterials have been exploited to improve bone regeneration under infection, potential tissue toxicity under high concentration and antibiotic-resistance are their main deficiencies. This study indicated that HACC-grafted PLGA/HA composite scaffold prepared using an innovative 3D-printing technique and covalent grafting strategy showed significantly enhanced antibacterial activities, especially against the antibiotic-resistant strains, together with good osteogenic

Mesoporous silicate nanoparticles/3D nanofibrous scaffold-mediated dual-drug delivery for bone tissue engineering.

PubMed

Yao, Qingqing; Liu, Yangxi; Selvaratnam, Balaranjan; Koodali, Ranjit T; Sun, Hongli

2018-04-09

Controlled delivery systems play a critical role in the success of bone morphogenetic proteins (i.e., BMP2 and BMP7) for challenged bone repair. Instead of single-drug release that is currently and commonly prevalent, dual-drug delivery strategies are highly desired to achieve effective bone regeneration because natural bone repair process is driven by multiple factors. Particularly, angiogenesis is essential for osteogenesis and requires more than just one factor (e.g., Vascular Endothelial Growth Factor, VEGF). Therefore, we developed a novel mesoporous silicate nanoparticles (MSNs) incorporated-3D nanofibrous gelatin (GF) scaffold for dual-delivery of BMP2 and deferoxamine (DFO). DFO is a hypoxia-mimetic drug that can activate hypoxia-inducible factor-1 alpha (HIF-1α), and trigger subsequent angiogenesis. Sustained BMP2 release system was achieved through encapsulation into large-pored MSNs, while the relative short-term release of DFO was engineered through covalent conjugation with chitosan to reduce its cytotoxicity and elongate its half-life. Both MSNs and DFO were incorporated onto a porous 3D GF scaffold to serve as a biomimetic osteogenic microenvironment. Our data indicated that DFO and BMP2 were released from a scaffold at different release rates (10 vs 28 days) yet maintained their angiogenic and osteogenic ability, respectively. Importantly, our data indicated that the released DFO significantly improved BMP2-induced osteogenic differentiation where the dose/duration was important for its effects in both mouse and human stem cell models. Thus, we developed a novel and tunable MSNs/GF 3D scaffold-mediated dual-drug delivery system and studied the potential application of the both FDA-approved DFO and BMP2 for bone tissue engineering. Copyright © 2018 Elsevier B.V. All rights reserved.

Production of Composite Scaffold Containing Silk Fibroin, Chitosan, and Gelatin for 3D Cell Culture and Bone Tissue Regeneration.

PubMed

Li, Jianqing; Wang, Qiuke; Gu, Yebo; Zhu, Yu; Chen, Liang; Chen, Yunfeng

2017-11-08

BACKGROUND Bone tissue engineering, a powerful tool to treat bone defects, is highly dependent on use of scaffolds. Both silk fibroin (SF) and chitosan (Cs) are biocompatible and actively studied for reconstruction of tissue engineering. Gelatin (Gel) is also widely applied in the biomedical field due to its low antigenicity and physicochemical stability. MATERIAL AND METHODS In this study, 4 different types of scaffolds were constructed - SF, SF/Cs, SF/Gel, and SF/Cs/Gel - and we compared their physical and chemical properties as well as biological characterization of these scaffolds to determine the most suitable scaffold for use in bone regeneration. First, these scaffolds were produced via chemical cross-linking method and freeze-drying technique. Next, the characterization of internal structure was studied using scanning electron microscopy and the porosity was evaluated by liquid displacement method. Then, we compared physicochemical properties such as water absorption rate and degradation property. Finally, MC3T3-E1 cells were inoculated on the scaffolds to study the biocompatibility and osteogenesis of the three-dimensional (3D) scaffolds in vitro. RESULTS The composite scaffold formed by all 3 components was the best for use in bone regeneration. CONCLUSIONS We conclude that the best scaffold among the 4 studied for MC3T3-E1 cells is our SF/Cs/Gel scaffold, suggesting a new choice for bone regeneration that can be used to treat bone defects or fractures in clinical practice.

4D printing smart biomedical scaffolds with novel soybean oil epoxidized acrylate

PubMed Central

Miao, Shida; Zhu, Wei; Castro, Nathan J.; Nowicki, Margaret; Zhou, Xuan; Cui, Haitao; Fisher, John P.; Zhang, Lijie Grace

2016-01-01

Photocurable, biocompatible liquid resins are highly desired for 3D stereolithography based bioprinting. Here we solidified a novel renewable soybean oil epoxidized acrylate, using a 3D laser printing technique, into smart and highly biocompatible scaffolds capable of supporting growth of multipotent human bone marrow mesenchymal stem cells (hMSCs). Porous scaffolds were readily fabricated by simply adjusting the printer infill density; superficial structures of the polymerized soybean oil epoxidized acrylate were significantly affected by laser frequency and printing speed. Shape memory tests confirmed that the scaffold fixed a temporary shape at −18 °C and fully recovered its original shape at human body temperature (37 °C), which indicated the great potential for 4D printing applications. Cytotoxicity analysis proved that the printed scaffolds had significant higher hMSC adhesion and proliferation than traditional polyethylene glycol diacrylate (PEGDA), and had no statistical difference from poly lactic acid (PLA) and polycaprolactone (PCL). This research is believed to significantly advance the development of biomedical scaffolds with renewable plant oils and advanced 3D fabrication techniques. PMID:27251982

Preparation, characterization and biological test of 3D-scaffolds based on chitosan, fibroin and hydroxyapatite for bone tissue engineering.

PubMed

Lima, Paulo Autran Leite; Resende, Cristiane Xavier; Soares, Glória Dulce de Almeida; Anselme, Karine; Almeida, Luís Eduardo

2013-08-01

This work describes the preparation and characterization of porous 3D-scaffolds based on chitosan (CHI), chitosan/silk fibroin (CHI/SF) and chitosan/silk fibroin/hydroxyapatite (CHI/SF/HA) by freeze drying. The biomaterials were characterized by X-ray diffraction, attenuated total reflection Fourier transform infrared spectroscopy, thermogravimetric analysis, differential scanning calorimetry, scanning electron microscopy and energy dispersive spectroscopy. In addition, studies of porosity, pore size, contact angle and biological response of SaOs-2osteoblastic cells were performed. The CHI scaffolds have a porosity of 94.2±0.9%, which is statistically higher than the one presented by CHI/SF/HA scaffolds, 89.7±2.6%. Although all scaffolds were able to promote adhesion, growth and maintenance of osteogenic differentiation of SaOs-2 cells, the new 3D-scaffold based on CHI/SF/HA showed a significantly higher cell growth at 7 days and 21 days and the level of alkaline phosphatase at 14 and 21 days was statistically superior compared to other tested materials. Copyright © 2013 Elsevier B.V. All rights reserved.

Marine Spongin: Naturally Prefabricated 3D Scaffold-Based Biomaterial

PubMed Central

Jesionowski, Teofil; Norman, Małgorzata; Żółtowska-Aksamitowska, Sonia; Petrenko, Iaroslav; Ehrlich, Hermann

2018-01-01

The biosynthesis, chemistry, structural features and functionality of spongin as a halogenated scleroprotein of keratosan demosponges are still paradigms. This review has the principal goal of providing thorough and comprehensive coverage of spongin as a naturally prefabricated 3D biomaterial with multifaceted applications. The history of spongin’s discovery and use in the form of commercial sponges, including their marine farming strategies, have been analyzed and are discussed here. Physicochemical and material properties of spongin-based scaffolds are also presented. The review also focuses on prospects and trends in applications of spongin for technology, materials science and biomedicine. Special attention is paid to applications in tissue engineering, adsorption of dyes and extreme biomimetics. PMID:29522478

Isolation of a Mutant of Salmonella typhimurium Dependent on D-Arabinose-5-phosphate for Growth and Synthesis of 3-Deoxy-D-mannoctulosonate (Ketodeoxyoctonate)

PubMed Central

Rick, P. D.; Osborn, M. J.

1972-01-01

A new type of auxotrophic mutant of Salmonella typhimurium has been isolated that is defective in the synthesis of the 3-deoxy-D-mannooctulosonate (ketodeoxyoctonate) region of the lipopolysaccharide and requires D-arabinose-5-phosphate for growth. Genetic and biochemical evidence indicated that the mutant defect was due to an altered ketodeoxyoctonate-8-phosphate synthetase with an apparent Km for D-arabinose-5-phosphate 35-fold higher than that of the parental enzyme. As a result of this enzymatic lesion, the mutant strain was dependent on exogenous D-arabinose-5-phosphate both for growth and for synthesis of a complete lipopolysaccharide. PMID:4566459

Doped Tricalcium Phosphate Scaffolds by Thermal Decomposition of Naphthalene: Mechanical Properties and In vivo Osteogenesis in a Rabbit Femur Model

PubMed Central

Ke, Dongxu; Dernell, William; Bandyopadhyay, Amit; Bose, Susmita

2015-01-01

Tricalcium phosphate (TCP) is a bioceramic that is widely used in orthopedic and dental applications. TCP structures show excellent biocompatibility as well as biodegradability. In this study, porous β-TCP scaffolds were prepared by thermal decomposition of naphthalene. Scaffolds with 57.64 ± 3.54 % density and a maximum pore size around 100 μm were fabricated via removing 30% naphthalene at 1150°C. The compressive strength for these scaffolds was 32.85 ± 1.41 MPa. Furthermore, by mixing 1 wt % SrO and 0.5 wt % SiO2, pore interconnectivity improved, but the compressive strength decreased to 22.40 ± 2.70 MPa. However, after addition of polycaprolactone (PCL) coating layers, the compressive strength of doped scaffolds increased to 29.57 ± 3.77 MPa. Porous scaffolds were implanted in rabbit femur defects to evaluate their biological property. The addition of dopants triggered osteoinduction by enhancing osteoid formation, osteocalcin expression and bone regeneration, especially at the interface of the scaffold and host bone. This study showed processing flexibility to make interconnected porous scaffolds with different pore size and volume fraction porosity with high compressive mechanical strength and better bioactivity. Results show that SrO/SiO2 doped porous TCP scaffolds have excellent potential to be used in bone tissue engineering applications. PMID:25504889

A microfluidic chip containing multiple 3D nanofibrous scaffolds for culturing human pluripotent stem cells

NASA Astrophysics Data System (ADS)

Wertheim, Lior; Shapira, Assaf; Amir, Roey J.; Dvir, Tal

2018-04-01

In microfluidics-based lab-on-a-chip systems, which are used for investigating the effect of drugs and growth factors on cells, the latter are usually cultured within the device’s channels in two-dimensional, and not in their optimal three-dimensional (3D) microenvironment. Herein, we address this shortfall by designing a microfluidic system, comprised of two layers. The upper layer of the system consists of multiple channels generating a gradient of soluble factors. The lower layer is comprised of multiple wells, each deposited with 3D, nanofibrous scaffold. We first used a mathematical model to characterize the fluid flow within the system. We then show that induced pluripotent stem cells can be seeded within the 3D scaffolds and be exposed to a well-mixed gradient of soluble factors. We believe that utilizing such system may enable in the future to identify new differentiation factors, investigate drug toxicity, and eventually allow to perform analyses on patient-specific tissues, in order to fit the appropriate combination and concentration of drugs.

From 2D to 3D: The morphology, proliferation and differentiation of MC3T3-E1 on silk fibroin/chitosan matrices.

PubMed

Li, Da-Wei; He, Feng-Li; He, Jin; Deng, Xudong; Liu, Ya-Li; Liu, Yang-Yang; Ye, Ya-Jing; Yin, Da-Chuan

2017-12-15

It has been widely accepted that cell culture in two-dimensional (2D) conditions may not be able to represent growth in three-dimensional (3D) conditions. Systematic comparisons between 2D and 3D cell cultures are needed to appropriately use the existing 2D results. In this work, we conducted a comparative study between 2D and 3D cell cultures of MC3T3-E1 using the same type of material (a mixture of silk fibroin (SF) and chitosan (CS)). Our results showed 3D SF/CS scaffold exhibited different effects on cell culture compared with the 2D cases. 1) The cells grown in 3D scaffold showed multiple morphologies. 2) The proliferation of cells in 3D scaffold was long-term and sustainable. 3) Cell differentiation occurred throughout the entire 3D scaffold. The results showed that cell culture in 3D SF/CS scaffold exhibited different features than 2D cases and 3D SF/CS scaffold could be a promising material for 3D cell culture. Copyright © 2017 Elsevier Ltd. All rights reserved.

Hybrid chitosan-ß-glycerol phosphate-gelatin nano-/micro fibrous scaffolds with suitable mechanical and biological properties for tissue engineering.

PubMed

Lotfi, Marzieh; Bagherzadeh, Roohollah; Naderi-Meshkin, Hojjat; Mahdipour, Elahe; Mafinezhad, Asghar; Sadeghnia, Hamid Reza; Esmaily, Habibollah; Maleki, Masoud; Hasssanzadeh, Halimeh; Ghayaour-Mobarhan, Majid; Bidkhori, Hamid Reza; Bahrami, Ahmad Reza

2016-03-01

Scaffold-based tissue engineering is considered as a promising approach in the regenerative medicine. Graft instability of collagen, by causing poor mechanical properties and rapid degradation, and their hard handling remains major challenges to be addressed. In this research, a composite structured nano-/microfibrous scaffold, made from a mixture of chitosan-ß-glycerol phosphate-gelatin (chitosan-GP-gelatin) using a standard electrospinning set-up was developed. Gelatin-acid acetic and chitosan ß-glycerol phosphate-HCL solutions were prepared at ratios of 30/70, 50/50, 70/30 (w/w) and their mechanical and biological properties were engineered. Furthermore, the pore structure of the fabricated nanofibrous scaffolds was investigated and predicted using a theoretical model. Higher gelatin concentrations in the polymer blend resulted in significant increase in mean pore size and its distribution. Interaction between the scaffold and the contained cells was also monitored and compared in the test and control groups. Scaffolds with higher chitosan concentrations showed higher rate of cell attachment with better proliferation property, compared with gelatin-only scaffolds. The fabricated scaffolds, unlike many other natural polymers, also exhibit non-toxic and biodegradable properties in the grafted tissues. In conclusion, the data clearly showed that the fabricated biomaterial is a biologically compatible scaffold with potential to serve as a proper platform for retaining the cultured cells for further application in cell-based tissue engineering, especially in wound healing practices. These results suggested the potential of using mesoporous composite chitosan-GP-gelatin fibrous scaffolds for engineering three-dimensional tissues with different inherent cell characteristics. © 2015 Wiley Periodicals, Inc.

Biofunctional Ionic-Doped Calcium Phosphates: Silk Fibroin Composites for Bone Tissue Engineering Scaffolding.

PubMed

Pina, S; Canadas, R F; Jiménez, G; Perán, M; Marchal, J A; Reis, R L; Oliveira, J M

2017-01-01

The treatment and regeneration of bone defects caused by traumatism or diseases have not been completely addressed by current therapies. Lately, advanced tools and technologies have been successfully developed for bone tissue regeneration. Functional scaffolding materials such as biopolymers and bioresorbable fillers have gained particular attention, owing to their ability to promote cell adhesion, proliferation, and extracellular matrix production, which promote new bone growth. Here, we present novel biofunctional scaffolds for bone regeneration composed of silk fibroin (SF) and β-tricalcium phosphate (β-TCP) and incorporating Sr, Zn, and Mn, which were successfully developed using salt-leaching followed by a freeze-drying technique. The scaffolds presented a suitable pore size, porosity, and high interconnectivity, adequate for promoting cell attachment and proliferation. The degradation behavior and compressive mechanical strengths showed that SF/ionic-doped TCP scaffolds exhibit improved characteristics for bone tissue engineering when compared with SF scaffolds alone. The in vitro bioactivity assays using a simulated body fluid showed the growth of an apatite layer. Furthermore, in vitro assays using human adipose-derived stem cells presented different effects on cell proliferation/differentiation when varying the doping agents in the biofunctional scaffolds. The incorporation of Zn into the scaffolds led to improved proliferation, while the Sr- and Mn-doped scaffolds presented higher osteogenic potential as demonstrated by DNA quantification and alkaline phosphatase activity. The combination of Sr with Zn led to an influence on cell proliferation and osteogenesis when compared with single ions. Our results indicate that biofunctional ionic-doped composite scaffolds are good candidates for further in vivo studies on bone tissue regeneration. © 2017 S. Karger AG, Basel.

Design and properties of 3D scaffolds for bone tissue engineering.

PubMed

Gómez, S; Vlad, M D; López, J; Fernández, E

2016-09-15

In this study, the Voronoi tessellation method has been used to design novel bone like three dimension (3D) porous scaffolds. The Voronoi method has been processed with computer design software to obtain 3D virtual isotropic porous interconnected models, exactly matching the main histomorphometric indices of trabecular bone (trabecular thickness, trabecular separation, trabecular number, bone volume to total volume ratio, bone surface to bone volume ratio, etc.). These bone like models have been further computed for mechanical (elastic modulus) and fluid mass transport (permeability) properties. The results show that the final properties of the scaffolds can be controlled during their microstructure and histomorphometric initial design stage. It is also shown that final properties can be tuned during the design stage to exactly match those of trabecular natural bone. Moreover, identical total porosity models can be designed with quite different specific bone surface area and thus, this specific microstructural feature can be used to favour cell adhesion, migration and, ultimately, new bone apposition (i.e. osteoconduction). Once the virtual models are fully characterized and optimized, these can be easily 3D printed by additive manufacturing and/or stereolitography technologies. The significance of this article goes far beyond the specific objectives on which it is focussed. In fact, it shows, in a guided way, the entire novel process that can be followed to design graded porous implants, whatever its external shape and geometry, but internally tuned to the exact histomorphometric indices needed to match natural human tissues microstructures and, consequently, their mechanical and fluid properties, among others. The significance is even more relevant nowadays thanks to the available new computing and design software that is easily linked to the 3D printing new technologies. It is this transversality, at the frontier of different disciplines, the main characteristic

Bone regeneration in 3D printing bioactive ceramic scaffolds with improved tissue/material interface pore architecture in thin-wall bone defect.

PubMed

Shao, Huifeng; Ke, Xiurong; Liu, An; Sun, Miao; He, Yong; Yang, Xianyan; Fu, Jianzhong; Liu, Yanming; Zhang, Lei; Yang, Guojing; Xu, Sanzhong; Gou, Zhongru

2017-04-12

Three-dimensional (3D) printing bioactive ceramics have demonstrated alternative approaches to bone tissue repair, but an optimized materials system for improving the recruitment of host osteogenic cells into the bone defect and enhancing targeted repair of the thin-wall craniomaxillofacial defects remains elusive. Herein we systematically evaluated the role of side-wall pore architecture in the direct-ink-writing bioceramic scaffolds on mechanical properties and osteogenic capacity in rabbit calvarial defects. The pure calcium silicate (CSi) and dilute Mg-doped CSi (CSi-Mg6) scaffolds with different layer thickness and macropore sizes were prepared by varying the layer deposition mode from single-layer printing (SLP) to double-layer printing (DLP) and then by undergoing one-, or two-step sintering. It was found that the dilute Mg doping and/or two-step sintering schedule was especially beneficial for improving the compressive strength (∼25-104 MPa) and flexural strength (∼6-18 MPa) of the Ca-silicate scaffolds. The histological analysis for the calvarial bone specimens in vivo revealed that the SLP scaffolds had a high osteoconduction at the early stage (4 weeks) but the DLP scaffolds displayed a higher osteogenic capacity for a long time stage (8-12 weeks). Although the DLP CSi scaffolds displayed somewhat higher osteogenic capacity at 8 and 12 weeks, the DLP CSi-Mg6 scaffolds with excellent fracture resistance also showed appreciable new bone tissue ingrowth. These findings demonstrate that the side-wall pore architecture in 3D printed bioceramic scaffolds is required to optimize for bone repair in calvarial bone defects, and especially the Mg doping wollastontie is promising for 3D printing thin-wall porous scaffolds for craniomaxillofacial bone defect treatment.

Polymer-Enriched 3D Graphene Foams for Biomedical Applications.

PubMed

Wang, Jun Kit; Xiong, Gordon Minru; Zhu, Minmin; Özyilmaz, Barbaros; Castro Neto, Antonio Helio; Tan, Nguan Soon; Choong, Cleo

2015-04-22

Graphene foams (GFs) are versatile nanoplatforms for biomedical applications because of their excellent physical, chemical, and mechanical properties. However, the brittleness and inflexibility of pristine GF (pGF) are some of the important factors restricting their widespread application. Here, a chemical-vapor-deposition-assisted method was used to synthesize 3D GFs, which were subsequently spin-coated with polymer to produce polymer-enriched 3D GFs with high conductivity and flexibility. Compared to pGF, both poly(vinylidene fluoride)-enriched GF (PVDF/GF) and polycaprolactone-enriched GF (PCL/GF) scaffolds showed improved flexibility and handleability. Despite the presence of the polymers, the polymer-enriched 3D GF scaffolds retained high levels of electrical conductivity because of the presence of microcracks that allowed for the flow of electrons through the material. In addition, polymer enrichment of GF led to an enhancement in the formation of calcium phosphate (Ca-P) compounds when the scaffolds were exposed to simulated body fluid. Between the two polymers tested, PCL enrichment of GF resulted in a higher in vitro mineralization nucleation rate because the oxygen-containing functional group of PCL had a higher affinity for Ca-P deposition and formation compared to the polar carbon-fluorine (C-F) bond in PVDF. Taken together, our current findings are a stepping stone toward future applications of polymer-enriched 3D GFs in the treatment of bone defects as well as other biomedical applications.

Microwave-induced biomimetic approach for hydroxyapatite coatings of chitosan scaffolds.

PubMed

Kaynak Bayrak, Gökçe; Demirtaş, T Tolga; Gümüşderelioğlu, Menemşe

2017-02-10

Simulated body fluid (SBF) can form calcium phosphates on osteoinductive materials, so it is widely used for coating of bone scaffolds to mimic natural extracellular matrix (ECM). However, difficulties of bulk coating in 3D scaffolds and the necessity of long process times are the common problems for coating with SBF. In the present study, a microwave-assisted process was developed for rapid and internal coating of chitosan scaffolds. The scaffolds were fabricated as superporous hydrogel (SPH) by combining microwave irradiation and gas foaming methods. Then, they were immersed into 10x  SBF-like solution and homogenous bone-like hydroxyapatite (HA) coating was achieved by microwave treatment at 600W without the need of any nucleating agent. Cell culture studies with MC3T3-E1 preosteoblasts showed that microwave-assisted biomimetic HA coating process could be evaluated as an efficient and rapid method to obtain composite scaffolds for bone tissue engineering. Copyright © 2016 Elsevier Ltd. All rights reserved.

Fabrication of scalable tissue engineering scaffolds with dual-pore microarchitecture by combining 3D printing and particle leaching.

PubMed

Mohanty, Soumyaranjan; Sanger, Kuldeep; Heiskanen, Arto; Trifol, Jon; Szabo, Peter; Dufva, Marin; Emnéus, Jenny; Wolff, Anders

2016-04-01

Limitations in controlling scaffold architecture using traditional fabrication techniques are a problem when constructing engineered tissues/organs. Recently, integration of two pore architectures to generate dual-pore scaffolds with tailored physical properties has attracted wide attention in tissue engineering community. Such scaffolds features primary structured pores which can efficiently enhance nutrient/oxygen supply to the surrounding, in combination with secondary random pores, which give high surface area for cell adhesion and proliferation. Here, we present a new technique to fabricate dual-pore scaffolds for various tissue engineering applications where 3D printing of poly(vinyl alcohol) (PVA) mould is combined with salt leaching process. In this technique the sacrificial PVA mould, determining the structured pore architecture, was filled with salt crystals to define the random pore regions of the scaffold. After crosslinking the casted polymer the combined PVA-salt mould was dissolved in water. The technique has advantages over previously reported ones, such as automated assembly of the sacrificial mould, and precise control over pore architecture/dimensions by 3D printing parameters. In this study, polydimethylsiloxane and biodegradable poly(ϵ-caprolactone) were used for fabrication. However, we show that this technique is also suitable for other biocompatible/biodegradable polymers. Various physical and mechanical properties of the dual-pore scaffolds were compared with control scaffolds with either only structured or only random pores, fabricated using previously reported methods. The fabricated dual-pore scaffolds supported high cell density, due to the random pores, in combination with uniform cell distribution throughout the scaffold, and higher cell proliferation and viability due to efficient nutrient/oxygen transport through the structured pores. In conclusion, the described fabrication technique is rapid, inexpensive, scalable, and compatible

3D bioprinting of scaffolds with living Schwann cells for potential nerve tissue engineering applications.

PubMed

Ning, Liqun; Sun, Haoying; Lelong, Tiphanie; Guilloteau, Romain; Zhu, Ning; Schreyer, David J; Chen, Daniel Xiongbiao

2018-06-18

Three-dimensional (3D) bioprinting of biomaterials shows great potential for producing cell-encapsulated scaffolds to repair nerves after injury or disease. For this, preparation of biomaterials and bioprinting itself are critical to create scaffolds with both biological and mechanical properties appropriate for nerve regeneration, yet remain unachievable. This paper presents our study on bioprinting Schwann cell-encapsulated scaffolds using composite hydrogels of alginate, fibrin, hyaluronic acid, and/or RGD peptide, for nerve tissue engineering applications. For the preparation of composite hydrogels, suitable hydrogel combinations were identified and prepared by adjusting the concentration of fibrin based on the morphological spreading of Schwann cells. In bioprinting, the effects of various printing process parameters (including the air pressure for dispensing, dispensing head movement speed, and crosslinking conditions) on printed structures were investigated and, by regulating these parameters, mechanically-stable scaffolds with fully interconnected pores were printed. The performance of Schwann cells within the printed scaffolds were examined in terms of viability, proliferation, orientation, and ability to produce laminin. Our results show that the printed scaffolds can promote the alignment of Schwann cells inside scaffolds and thus provide haptotactic cues to direct the extension of dorsal root ganglion neurites along the printed strands, demonstrating their great potential for applications in the field of nerve tissue engineering. © 2018 IOP Publishing Ltd.

3D structural patterns in scalable, elastomeric scaffolds guide engineered tissue architecture.

PubMed

Kolewe, Martin E; Park, Hyoungshin; Gray, Caprice; Ye, Xiaofeng; Langer, Robert; Freed, Lisa E

2013-08-27

Microfabricated elastomeric scaffolds with 3D structural patterns are created by semiautomated layer-by-layer assembly of planar polymer sheets with through-pores. The mesoscale interconnected pore architectures governed by the relative alignment of layers are shown to direct cell and muscle-like fiber orientation in both skeletal and cardiac muscle, enabling scale up of tissue constructs towards clinically relevant dimensions. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Hybrid Macro-Porous Titanium Ornamented by Degradable 3D Gel/nHA Micro-Scaffolds for Bone Tissue Regeneration

PubMed Central

Yin, Bo; Ma, Pei; Chen, Jun; Wang, Hai; Wu, Gui; Li, Bo; Li, Qiang; Huang, Zhifeng; Qiu, Guixing; Wu, Zhihong

2016-01-01

Porous titanium is a kind of promising material for bone substitution, while its bio-inert property results in demand of modifications to improve the osteointegration capacity. In this study, gelatin (Gel) and nano-hydroxyapatite (nHA) were used to construct 3D micro-scaffolds in the pores of porous titanium in the ratios of Gel:nHA = 1:0, Gel:nHA = 1:1, and Gel:nHA = 1:3, respectively. Cell attachment and proliferation, and gene and protein expression levels of osteogenic markers were evaluated in MC3T3-E1 cells, followed by bone regeneration assessment in a rabbit radius defect model. All hybrid scaffolds with different composition ratio were found to have significant promotional effects in cell adhesion, proliferation and differentiation, in which the group with Gel:nHA = 1:1 showed the best performance in vitro, as well as the most bone regeneration volume in vivo. This 3D micro-scaffolds modification may be an innovative method for porous titanium ornamentation and shows potential application values in clinic. PMID:27092492

Hybrid Macro-Porous Titanium Ornamented by Degradable 3D Gel/nHA Micro-Scaffolds for Bone Tissue Regeneration.

PubMed

Yin, Bo; Ma, Pei; Chen, Jun; Wang, Hai; Wu, Gui; Li, Bo; Li, Qiang; Huang, Zhifeng; Qiu, Guixing; Wu, Zhihong

2016-04-15

Porous titanium is a kind of promising material for bone substitution, while its bio-inert property results in demand of modifications to improve the osteointegration capacity. In this study, gelatin (Gel) and nano-hydroxyapatite (nHA) were used to construct 3D micro-scaffolds in the pores of porous titanium in the ratios of Gel:nHA = 1:0, Gel:nHA = 1:1, and Gel:nHA = 1:3, respectively. Cell attachment and proliferation, and gene and protein expression levels of osteogenic markers were evaluated in MC3T3-E1 cells, followed by bone regeneration assessment in a rabbit radius defect model. All hybrid scaffolds with different composition ratio were found to have significant promotional effects in cell adhesion, proliferation and differentiation, in which the group with Gel:nHA = 1:1 showed the best performance in vitro, as well as the most bone regeneration volume in vivo. This 3D micro-scaffolds modification may be an innovative method for porous titanium ornamentation and shows potential application values in clinic.

Alternating air-medium exposure in rotating bioreactors optimizes cell metabolism in 3D novel tubular scaffold polyurethane foams.

PubMed

Tresoldi, Claudia; Stefani, Ilaria; Ferracci, Gaia; Bertoldi, Serena; Pellegata, Alessandro F; Farè, Silvia; Mantero, Sara

2017-04-26

In vitro dynamic culture conditions play a pivotal role in developing engineered tissue grafts, where the supply of oxygen and nutrients, and waste removal must be permitted within construct thickness. For tubular scaffolds, mass transfer is enhanced by introducing a convective flow through rotating bioreactors with positive effects on cell proliferation, scaffold colonization and extracellular matrix deposition. We characterized a novel polyurethane-based tubular scaffold and investigated the impact of 3 different culture configurations over cell behavior: dynamic (i) single-phase (medium) rotation and (ii) double-phase exposure (medium-air) rotation; static (iii) single-phase static culture as control. A new mixture of polyol was tested to create polyurethane foams (PUFs) as 3D scaffold for tissue engineering. The structure obtained was morphologically and mechanically analyzed tested. Murine fibroblasts were externally seeded on the novel porous PUF scaffold, and cultured under different dynamic conditions. Viability assay, DNA quantification, SEM and histological analyses were performed at different time points. The PUF scaffold presented interesting mechanical properties and morphology adequate to promote cell adhesion, highlighting its potential for tissue engineering purposes. Results showed that constructs under dynamic conditions contain enhanced viability and cell number, exponentially increased for double-phase rotation; under this last configuration, cells uniformly covered both the external surface and the lumen. The developed 3D structure combined with the alternated exposure to air and medium provided the optimal in vitro biochemical conditioning with adequate nutrient supply for cells. The results highlight a valuable combination of material and dynamic culture for tissue engineering applications.

Composite Scaffolds Containing Silk Fibroin, Gelatin, and Hydroxyapatite for Bone Tissue Regeneration and 3D Cell Culturing

PubMed Central

Moisenovich, M. M.; Arkhipova, A. Yu.; Orlova, A. A.; Drutskaya, M. S; Volkova, S. V.; Zacharov, S. E.; Agapov, I. I.; Kirpichnikov, M. P.

2014-01-01

Three-dimensional (3D) silk fibroin scaffolds were modified with one of the major bone tissue derivatives (nano-hydroxyapatite) and/or a collagen derivative (gelatin). Adhesion and proliferation of mouse embryonic fibroblasts (MEF) within the scaffold were increased after modification with either nano-hydroxyapatite or gelatin. However, a significant increase in MEF adhesion and proliferation was observed when both additives were introduced into the scaffold. Such modified composite scaffolds provide a new and better platform to study wound healing, bone and other tissue regeneration, as well as artificial organ bioengineering. This system can further be applied to establish experimental models to study cell-substrate interactions, cell migration and other complex processes, which may be difficult to address using the conventional two-dimensional culture systems. PMID:24772332

Chitosan functionalized poly-ε-caprolactone electrospun fibers and 3D printed scaffolds as antibacterial materials for tissue engineering applications.

PubMed

Tardajos, Myriam G; Cama, Giuseppe; Dash, Mamoni; Misseeuw, Lara; Gheysens, Tom; Gorzelanny, Christian; Coenye, Tom; Dubruel, Peter

2018-07-01

Tissue engineering (TE) approaches often employ polymer-based scaffolds to provide support with a view to the improved regeneration of damaged tissues. The aim of this research was to develop a surface modification method for introducing chitosan as an antibacterial agent in both electrospun membranes and 3D printed poly-ε-caprolactone (PCL) scaffolds. The scaffolds were functionalized by grafting methacrylic acid N-hydroxysuccinimide ester (NHSMA) onto the surface after Ar-plasma/air activation. Subsequently, the newly-introduced NHS groups were used to couple with chitosan of various molecular weights (Mw). High Mw chitosan exhibited a better coverage of the surface as indicated by the higher N% detected by X-ray photoelectron spectroscopy (XPS) and the observations with either scanning electron microscopy (SEM)(for fibers) or Coomassie blue staining (for 3D-printed scaffolds). A lactate dehydrogenase assay (LDH) using L929 fibroblasts demonstrated the cell-adhesion and cell-viability capacity of the modified samples. The antibacterial properties against S. aureus ATCC 6538 and S. epidermidis ET13 revealed a slower bacterial growth rate on the surface of the chitosan modified scaffolds, regardless the chitosan Mw. Copyright © 2018 Elsevier Ltd. All rights reserved.

Multi-scale osteointegration and neovascularization of biphasic calcium phosphate bone scaffolds

NASA Astrophysics Data System (ADS)

Lan, Sheeny K.

Bone grafts are utilized clinically to guide tissue regeneration. Autologous bone and allogeneic bone are the current clinical standards. However, there are significant limitations to their use. To address the need for alternatives to autograft and allograft, researchers have worked to develop synthetic grafts, also referred to as scaffolds. Despite extensive efforts in this area, a gap persists between basic research and clinical application. In particular, solutions for repairing critical size and/or load-bearing defects are lacking. The aim of this thesis work was to address two critical barriers preventing design of successful tissue engineering constructs for bone regeneration within critical size and/or load-bearing defects. Those barriers are insufficient osteointegration and slow neovascularization. In this work, the effects of scaffold microporosity, recombinant human bone morphogenetic protein-2 delivery and endothelial colony forming cell vasculogenesis were evaluated in the context of bone formation in vivo. This was accomplished to better understand the role of these factors in bone regeneration, which may translate to improvements in tissue engineering construct design. Biphasic calcium phosphate (BCP) scaffolds with controlled macro- and microporosity were implanted in porcine mandibular defects. Evaluation of the BCP scaffolds after in vivo implantation showed, for the first time, osteocytes embedded in bone within scaffold micropores (< 10 microm) as well as the most extensive bone growth into micropores to date with bone penetration throughout rods 394 microm in diameter. The result is the first truly osteointegrated bone scaffolds with integration occurring at both the macro and micro length scales, leaving no "dead space" or discontinuities of bone in the defect site. The scaffold forms a living composite upon integration with regenerating bone and this has significant implications with regard to improved scaffold mechanical properties. The

Design control for clinical translation of 3D printed modular scaffolds.

PubMed

Hollister, Scott J; Flanagan, Colleen L; Zopf, David A; Morrison, Robert J; Nasser, Hassan; Patel, Janki J; Ebramzadeh, Edward; Sangiorgio, Sophia N; Wheeler, Matthew B; Green, Glenn E

2015-03-01

founded on 3D printing for developing tissue engineering therapies and (2) illustrate the design control process for modular implementation of two scaffold based tissue engineering therapies: airway reconstruction and bone tissue engineering based spine fusion.

Design Control for Clinical Translation of 3D Printed Modular Scaffolds

PubMed Central

Hollister, Scott J.; Flanagan, Colleen L.; Zopf, David A.; Morrison, Robert J.; Nasser, Hassan; Patel, Janki J.; Ebramzadeh, Edward; Sangiorgio, Sophia N.; Wheeler, Matthew B.; Green, Glenn E.

2015-01-01

founded on 3D printing for developing tissue engineering therapies and (2) illustrate the design control process for modular implementation of two scaffold based tissue engineering therapies: airway reconstruction and bone tissue engineering based spine fusion. PMID:25666115

Designing of PLA scaffolds for bone tissue replacement fabricated by ordinary commercial 3D printer.

PubMed

Gregor, Aleš; Filová, Eva; Novák, Martin; Kronek, Jakub; Chlup, Hynek; Buzgo, Matěj; Blahnová, Veronika; Lukášová, Věra; Bartoš, Martin; Nečas, Alois; Hošek, Jan

2017-01-01

The primary objective of Tissue engineering is a regeneration or replacement of tissues or organs damaged by disease, injury, or congenital anomalies. At present, Tissue engineering repairs damaged tissues and organs with artificial supporting structures called scaffolds. These are used for attachment and subsequent growth of appropriate cells. During the cell growth gradual biodegradation of the scaffold occurs and the final product is a new tissue with the desired shape and properties. In recent years, research workplaces are focused on developing scaffold by bio-fabrication techniques to achieve fast, precise and cheap automatic manufacturing of these structures. Most promising techniques seem to be Rapid prototyping due to its high level of precision and controlling. However, this technique is still to solve various issues before it is easily used for scaffold fabrication. In this article we tested printing of clinically applicable scaffolds with use of commercially available devices and materials. Research presented in this article is in general focused on "scaffolding" on a field of bone tissue replacement. Commercially available 3D printer and Polylactic acid were used to create originally designed and possibly suitable scaffold structures for bone tissue engineering. We tested printing of scaffolds with different geometrical structures. Based on the osteosarcoma cells proliferation experiment and mechanical testing of designed scaffold samples, it will be stated that it is likely not necessary to keep the recommended porosity of the scaffold for bone tissue replacement at about 90%, and it will also be clarified why this fact eliminates mechanical properties issue. Moreover, it is demonstrated that the size of an individual pore could be double the size of the recommended range between 0.2-0.35 mm without affecting the cell proliferation. Rapid prototyping technique based on Fused deposition modelling was used for the fabrication of designed scaffold

Physicochemical and osteoplastic characteristics of 3D printed bone grafts based on synthetic calcium phosphates and natural polymers

NASA Astrophysics Data System (ADS)

Nezhurina, E. K.; Karalkin, P. A.; Komlev, V. S.; Sviridova, I. K.; Kirsanova, V. A.; Akhmedova, S. A.; Shanskiy, Ya D.; Fedotov, A. Yu; Barinov, S. M.; Sergeeva, N. S.

2018-04-01

A creation of personalized implants for regeneration of bone tissue seems to be a very promising biomedical technological approach. We have studied the physicochemical characteristics, cyto- and biocompatibility of three-dimensional constructs based on sodium alginate and gelatin in combination with 2 types of calcium phosphate (tricalcium phosphate or octacalcium phosphate) obtained by inkjet 3D printing. In our experiments, we have studied the physical and chemical properties of the constructs – their porosity, chemical composition, microarchitecture of the surface and mechanical elasticity. The cytocompatibility of 3D constructs and matrix-for-cell properties were investigated in vitro on a model of human osteosarcoma MG-63 cell line by means of MTT assay. The biocompatibility of 3D constructs was studied on the model of subcutaneous implantation in mice up to 12 weeks. All types of 3D constructs were cytocompatible in vitro, demonstrated good matrix-for-cells properties, and had supported cell proliferation for 2 weeks. In results of subcutaneous in vivo test all constructs demonstrated biocompatibility with slow bioresorption of organic and inorganic components. Osteogenesis proceeded more actively in rat tibia model defects (marginal excision), substituted by 3D printed 3-component implants based on alginate, gelatin and octacalcium phosphate.

Fabrication of scalable and structured tissue engineering scaffolds using water dissolvable sacrificial 3D printed moulds.

PubMed

Mohanty, Soumyaranjan; Larsen, Layla Bashir; Trifol, Jon; Szabo, Peter; Burri, Harsha Vardhan Reddy; Canali, Chiara; Dufva, Marin; Emnéus, Jenny; Wolff, Anders

2015-10-01

One of the major challenges in producing large scale engineered tissue is the lack of ability to create large highly perfused scaffolds in which cells can grow at a high cell density and viability. Here, we explore 3D printed polyvinyl alcohol (PVA) as a sacrificial mould in a polymer casting process. The PVA mould network defines the channels and is dissolved after curing the polymer casted around it. The printing parameters determined the PVA filament density in the sacrificial structure and this density resulted in different stiffness of the corresponding elastomer replica. It was possible to achieve 80% porosity corresponding to about 150 cm(2)/cm(3) surface to volume ratio. The process is easily scalable as demonstrated by fabricating a 75 cm(3) scaffold with about 16,000 interconnected channels (about 1m(2) surface area) and with a channel to channel distance of only 78 μm. To our knowledge this is the largest scaffold ever to be produced with such small feature sizes and with so many structured channels. The fabricated scaffolds were applied for in-vitro culturing of hepatocytes over a 12-day culture period. Smaller scaffolds (6×4 mm) were tested for cell culturing and could support homogeneous cell growth throughout the scaffold. Presumably, the diffusion of oxygen and nutrient throughout the channel network is rapid enough to support cell growth. In conclusion, the described process is scalable, compatible with cell culture, rapid, and inexpensive. Copyright © 2015. Published by Elsevier B.V.

Induced migration of endothelial cells into 3D scaffolds by chemoattractants secreted by pro-inflammatory macrophages in situ.

PubMed

Li, Xuguang; Dai, Yuankun; Shen, Tao; Gao, Changyou

2017-06-01

Cell migration in scaffolds plays a crucial role in tissue regeneration, which can better mimic cell behaviors in vivo . In this study, a novel model has been proposed on controlling 3D cell migration in porous collagen-chitosan scaffolds with various pore structures under the stimulation of inflammatory cells to mimic the angiogenesis process. Endothelial cells (ECs) cultured atop the scaffolds in the Transwell molds which were placed into a well of a 24-well culture plate were promoted to migrate into the scaffolds by chemoattractants such as vascular endothelial growth factor (VEGF) and tumor necrosis factor-alpha (TNF-α) secreted by the pro-inflammatory macrophages incubated in the well culture plate. The phenotype of macrophages was mediated by 50 ng/ml interferon-gamma (IFN-γ) and different concentrations of lipopolysaccharide (LPS, 150-300 ng/ml). The cell migration depth had a positive correlation with LPS concentration, and thereby the TNF-α concentration. The ECs migrated easier to a deeper zone of the scaffolds prepared at - 10ºC (187 μm in pore diameter) than that at - 20ºC (108 μm in pore diameter) as well. The method provides a useful strategy to study the 3D cell migration, and is helpful to reveal the vascularization process during wound healing in the long run.

Induced migration of endothelial cells into 3D scaffolds by chemoattractants secreted by pro-inflammatory macrophages in situ

PubMed Central

Li, Xuguang; Dai, Yuankun; Shen, Tao

2017-01-01

Abstract Cell migration in scaffolds plays a crucial role in tissue regeneration, which can better mimic cell behaviors in vivo. In this study, a novel model has been proposed on controlling 3D cell migration in porous collagen-chitosan scaffolds with various pore structures under the stimulation of inflammatory cells to mimic the angiogenesis process. Endothelial cells (ECs) cultured atop the scaffolds in the Transwell molds which were placed into a well of a 24-well culture plate were promoted to migrate into the scaffolds by chemoattractants such as vascular endothelial growth factor (VEGF) and tumor necrosis factor-alpha (TNF-α) secreted by the pro-inflammatory macrophages incubated in the well culture plate. The phenotype of macrophages was mediated by 50 ng/ml interferon-gamma (IFN-γ) and different concentrations of lipopolysaccharide (LPS, 150–300 ng/ml). The cell migration depth had a positive correlation with LPS concentration, and thereby the TNF-α concentration. The ECs migrated easier to a deeper zone of the scaffolds prepared at − 10ºC (187 μm in pore diameter) than that at − 20ºC (108 μm in pore diameter) as well. The method provides a useful strategy to study the 3D cell migration, and is helpful to reveal the vascularization process during wound healing in the long run. PMID:28596912

3D polylactide-based scaffolds for studying human hepatocarcinoma processes in vitro

NASA Astrophysics Data System (ADS)

Scaffaro, Roberto; Lo Re, Giada; Rigogliuso, Salvatrice; Ghersi, Giulio

2012-08-01

We evaluated the combination of leaching techniques and melt blending of polymers and particles for the preparation of highly interconnected three-dimensional polymeric porous scaffolds for in vitro studies of human hepatocarcinoma processes. More specifically, sodium chloride and poly(ethylene glycol) (PEG) were used as water-soluble porogens to form porous and solvent-free poly(L,D-lactide) (PLA)-based scaffolds. Several characterization techniques, including porosimetry, image analysis and thermogravimetry, were combined to improve the reliability of measurements and mapping of the size, distribution and microarchitecture of pores. We also investigated the effect of processing, in PLA-based blends, on the simultaneous bulk/surface modifications and pore architectures in the scaffolds, and assessed the effects on human hepatocarcinoma viability and cell adhesion. The influence of PEG molecular weight on the scaffold morphology and cell viability and adhesion were also investigated. Morphological studies indicated that it was possible to obtain scaffolds with well-interconnected pores of assorted sizes. The analysis confirmed that SK-Hep1 cells adhered well to the polymeric support and emitted surface protrusions necessary to grow and differentiate three-dimensional systems. PEGs with higher molecular weight showed the best results in terms of cell adhesion and viability.

The Use of Silk as a Scaffold for Mature, Sustainable Unilocular Adipose 3D Tissue Engineered Systems.

PubMed

Abbott, Rosalyn D; Wang, Rebecca Y; Reagan, Michaela R; Chen, Ying; Borowsky, Francis E; Zieba, Adam; Marra, Kacey G; Rubin, J Peter; Ghobrial, Irene M; Kaplan, David L

2016-07-01

There is a critical need for monitoring physiologically relevant, sustainable, human adipose tissues in vitro to gain new insights into metabolic diseases. To support long-term culture, a 3D silk scaffold assisted culture system is developed that maintains mature unilocular adipocytes ex vivo in coculture with preadipocytes, endothelial cells, and smooth muscle cells obtained from small volumes of liquefied adipose samples. Without the silk scaffold, adipose tissue explants cannot be sustained in long-term culture (3 months) due to their fragility. Adjustments to media components are used to tune lipid metabolism and proliferation, in addition to responsiveness to an inflammatory stimulus. Interestingly, patient specific responses to TNFα stimulation are observed, providing a proof-of-concept translational technique for patient specific disease modeling in the future. In summary, this novel 3D scaffold assisted approach is required for establishing physiologically relevant, sustainable, human adipose tissue systems from small volumes of lipoaspirate, making this methodology of great value to studies of metabolism, adipokine-driven diseases, and other diseases where the roles of adipocytes are only now becoming uncovered. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Sulfophenyl-Functionalized Reduced Graphene Oxide Networks on Electrospun 3D Scaffold for Ultrasensitive NO₂ Gas Sensor.

PubMed

Zou, Bin; Guo, Yunlong; Shen, Nannan; Xiao, Anshan; Li, Mingjun; Zhu, Liang; Wan, Pengbo; Sun, Xiaoming

2017-12-19

Ultrasensitive room temperature real-time NO₂ sensors are highly desirable due to potential threats on environmental security and personal respiratory. Traditional NO₂ gas sensors with highly operated temperatures (200-600 °C) and limited reversibility are mainly constructed from semiconducting oxide-deposited ceramic tubes or inter-finger probes. Herein, we report the functionalized graphene network film sensors assembled on an electrospun three-dimensional (3D) nanonetwork skeleton for ultrasensitive NO₂ sensing. The functional 3D scaffold was prepared by electrospinning interconnected polyacrylonitrile (PAN) nanofibers onto a nylon window screen to provide a 3D nanonetwork skeleton. Then, the sulfophenyl-functionalized reduced graphene oxide (SFRGO) was assembled on the electrospun 3D nanonetwork skeleton to form SFRGO network films. The assembled functionalized graphene network film sensors exhibit excellent NO₂ sensing performance (10 ppb to 20 ppm) at room temperature, reliable reversibility, good selectivity, and better sensing cycle stability. These improvements can be ascribed to the functionalization of graphene with electron-withdrawing sulfophenyl groups, the high surface-to-volume ratio, and the effective sensing channels from SFRGO wrapping onto the interconnected 3D scaffold. The SFRGO network-sensing film has the advantages of simple preparation, low cost, good processability, and ultrasensitive NO₂ sensing, all advantages that can be utilized for potential integration into smart windows and wearable electronic devices for real-time household gas sensors.

Growth of human breast tissues from patient cells in 3D hydrogel scaffolds.

PubMed

Sokol, Ethan S; Miller, Daniel H; Breggia, Anne; Spencer, Kevin C; Arendt, Lisa M; Gupta, Piyush B

2016-03-01

Three-dimensional (3D) cultures have proven invaluable for expanding human tissues for basic research and clinical applications. In both contexts, 3D cultures are most useful when they (1) support the outgrowth of tissues from primary human cells that have not been immortalized through extensive culture or viral infection and (2) include defined, physiologically relevant components. Here we describe a 3D culture system with both of these properties that stimulates the outgrowth of morphologically complex and hormone-responsive mammary tissues from primary human breast epithelial cells. Primary human breast epithelial cells isolated from patient reduction mammoplasty tissues were seeded into 3D hydrogels. The hydrogel scaffolds were composed of extracellular proteins and carbohydrates present in human breast tissue and were cultured in serum-free medium containing only defined components. The physical properties of these hydrogels were determined using atomic force microscopy. Tissue growth was monitored over time using bright-field and fluorescence microscopy, and maturation was assessed using morphological metrics and by immunostaining for markers of stem cells and differentiated cell types. The hydrogel tissues were also studied by fabricating physical models from confocal images using a 3D printer. When seeded into these 3D hydrogels, primary human breast epithelial cells rapidly self-organized in the absence of stromal cells and within 2 weeks expanded to form mature mammary tissues. The mature tissues contained luminal, basal, and stem cells in the correct topological orientation and also exhibited the complex ductal and lobular morphologies observed in the human breast. The expanded tissues became hollow when treated with estrogen and progesterone, and with the further addition of prolactin produced lipid droplets, indicating that they were responding to hormones. Ductal branching was initiated by clusters of cells expressing putative mammary stem cell

Partial reactions of d-glucose 6-phosphate–1 l-myoinositol 1-phosphate cyclase

PubMed Central

Barnett, J. E. G.; Rasheed, A.; Corina, D. L.

1973-01-01

After removal of tightly bound NAD+ by using charcoal, a preparation of d-glucose 6-phosphate–1 l-myoinositol 1-phosphate cyclase catalysed the reduction of 5-keto-d-glucitol 6-phosphate and 5-keto-d-glucose 6-phosphate by [4-3H]NADH to give [5-3H]-glucitol 6-phosphate and [5-3H]glucose 6-phosphate respectively. The position of the tritium atom in the latter was shown by degradation. Both enzyme-catalysed reductions were strongly inhibited by 2-deoxy-d-glucose 6-phosphate, a powerful competitive inhibitor of inositol cyclase. The charcoal-treated enzyme preparation also converted 5-keto-d-glucose 6-phosphate into [3H]myoinositol 1-phosphate in the presence of [4-3H]NADH, but less effectively. These partial reactions of inositol cyclase are interpreted as providing strong evidence for the formation of 5-keto-d-glucose 6-phosphate as an enzyme-bound intermediate in the conversion of d-glucose 6-phosphate into 1 l-myoinositol 1-phosphate. The enzyme was partially inactivated by NaBH4 in the presence of NAD+. Glucose 6-phosphate did not increase the inactivation, and there was no inactivation in the absence of NAD+. There was no evidence for Schiff base formation during the cyclization. d-Glucitol 6-phosphate (l-sorbitol 1-phosphate) was a good inhibitor of the overall reaction. It did not inactivate the enzyme. The apparent molecular weight of inositol cyclase as determined by Sephadex chromatography was 2.15×105. PMID:4352864

Modeling of flow-induced shear stress applied on 3D cellular scaffolds: Implications for vascular tissue engineering.

PubMed

Lesman, Ayelet; Blinder, Yaron; Levenberg, Shulamit

2010-02-15

Novel tissue-culture bioreactors employ flow-induced shear stress as a means of mechanical stimulation of cells. We developed a computational fluid dynamics model of the complex three-dimensional (3D) microstructure of a porous scaffold incubated in a direct perfusion bioreactor. Our model was designed to predict high shear-stress values within the physiological range of those naturally sensed by vascular cells (1-10 dyne/cm(2)), and will thereby provide suitable conditions for vascular tissue-engineering experiments. The model also accounts for cellular growth, which was designed as an added cell layer grown on all scaffold walls. Five model variants were designed, with geometric differences corresponding to cell-layer thicknesses of 0, 50, 75, 100, and 125 microm. Four inlet velocities (0.5, 1, 1.5, and 2 cm/s) were applied to each model. Wall shear-stress distribution and overall pressure drop calculations were then used to characterize the relation between flow rate, shear stress, cell-layer thickness, and pressure drop. The simulations showed that cellular growth within 3D scaffolds exposes cells to elevated shear stress, with considerably increasing average values in correlation to cell growth and inflow velocity. Our results provide in-depth analysis of the microdynamic environment of cells cultured within 3D environments, and thus provide advanced control over tissue development in vitro. 2009 Wiley Periodicals, Inc.

Development of a Bioreactor to Culture Tissue Engineered Ureters Based on the Application of Tubular OPTIMAIX 3D Scaffolds.

PubMed

Seifarth, Volker; Gossmann, Matthias; Janke, Heinz Peter; Grosse, Joachim O; Becker, Christoph; Heschel, Ingo; Artmann, Gerhard M; Temiz Artmann, Aysegül

2015-01-01

Regenerative medicine, tissue engineering and biomedical research give hope to many patients who need bio-implants. Tissue engineering applications have already been developed based on bioreactors. Physiological ureter implants, however, do not still function sufficiently, as they represent tubular hollow structures with very specific cellular structures and alignments consisting of several cell types. The aim of this study was to a develop a new bioreactor system based on seamless, collagenous, tubular OPTIMAIX 3D prototype sponge as scaffold material for ex-vivo culturing of a tissue engineered ureter replacement for future urological applications. Particular emphasis was given to a great extent to mimic the physiological environment similar to the in vivo situation of a ureter. NIH-3T3 fibroblasts, C2C12, Urotsa and primary genitourinary tract cells were applied as co-cultures on the scaffold and the penetration of cells into the collagenous material was followed. By the end of this study, the bioreactor was functioning, physiological parameter as temperature and pH and the newly developed BIOREACTOR system is applicable to tubular scaffold materials with different lengths and diameters. The automatized incubation system worked reliably. The tubular OPTIMAIX 3D sponge was a suitable scaffold material for tissue engineering purposes and co-cultivation procedures. © 2015 S. Karger AG, Basel.

Differential osteogenic activity of osteoprogenitor cells on HA and TCP/HA scaffold of tissue engineered bone.

PubMed

Ng, Angela M H; Tan, K K; Phang, M Y; Aziyati, O; Tan, G H; Isa, M R; Aminuddin, B S; Naseem, M; Fauziah, O; Ruszymah, B H I

2008-05-01

Biomaterial, an essential component of tissue engineering, serves as a scaffold for cell attachment, proliferation, and differentiation; provides the three dimensional (3D) structure and, in some applications, the mechanical strength required for the engineered tissue. Both synthetic and naturally occurring calcium phosphate based biomaterial have been used as bone fillers or bone extenders in orthopedic and reconstructive surgeries. This study aims to evaluate two popular calcium phosphate based biomaterial i.e., hydroxyapatite (HA) and tricalcium phosphate/hydroxyapatite (TCP/HA) granules as scaffold materials in bone tissue engineering. In our strategy for constructing tissue engineered bone, human osteoprogenitor cells derived from periosteum were incorporated with human plasma-derived fibrin and seeded onto HA or TCP/HA forming 3D tissue constructs and further maintained in osteogenic medium for 4 weeks to induce osteogenic differentiation. Constructs were subsequently implanted intramuscularly in nude mice for 8 weeks after which mice were euthanized and constructs harvested for evaluation. The differential cell response to the biomaterial (HA or TCP/HA) adopted as scaffold was illustrated by the histology of undecalcified constructs and evaluation using SEM and TEM. Both HA and TCP/HA constructs showed evidence of cell proliferation, calcium deposition, and collagen bundle formation albeit lesser in the former. Our findings demonstrated that TCP/HA is superior between the two in early bone formation and hence is the scaffold material of choice in bone tissue engineering. Copyright 2007 Wiley Periodicals, Inc.

Highlighting the Importance of Surface Grafting in Combination with a Layer-by-Layer Approach for Fabricating Advanced 3D Poly(l-lactide) Microsphere Scaffolds

PubMed Central

2016-01-01

A combined surface treatment (i.e., surface grafting and a layer-by-layer (LbL) approach) is presented to create advanced biomaterials, i.e., 3D poly(l-lactide) (PLLA) microsphere scaffolds, at room temperature. The grafted surface plays a crucial role in assembling polyelectrolyte multilayers (PEMs) onto the surface of the microspheres, thus improving the physicochemical properties of the 3D microsphere scaffolds. The grafted surface of the PLLA microspheres demonstrates much better PEM adsorption, improved surface coverage at low pH, and smoother surfaces at high pH compared with those of nongrafted surfaces of PLLA microspheres during the assembly of PEMs. They induce more swelling than nongrafted surfaces after the assembly of the PEMs and exhibit blue emission after functionalization of the microsphere surface with a fluorescent dye molecule. The 3D scaffolds functionalized with and without nanosheets not only exhibit good mechanical performance similar to the compressive modulus of cancellous bone but also exhibit the porosity required for cancellous bone regeneration. The magnetic nanoparticle-functionalized 3D scaffolds result in an electrical conductivity in the high range of semiconducting materials (i.e., 1–250 S cm–1). Thus, these 3D microsphere scaffolds fabricated by surface grafting and the LbL approach are promising candidates for bone tissue engineering. PMID:29503506

Zein Increases the Cytoaffinity and Biodegradability of Scaffolds 3D-Printed with Zein and Poly(ε-caprolactone) Composite Ink.

PubMed

Jing, Linzhi; Wang, Xiang; Liu, Hang; Lu, Yuyun; Bian, Jinsong; Sun, Jie; Huang, Dejian

2018-05-25

Electrohydrodynamic printing (EHDP) has attracted extensive interests as a powerful technology to fabricate micro- to nano-scale fibrous scaffolds in a custom-tailored manner for biomedical applications. A few synthetic biopolymer inks are applicable to this EHDP technology, but the fabricated scaffolds suffered from low mechanical strength, biocompatibility, and biodegradability. In this study, a series of poly(ε-caprolactone) (PCL)/zein composite inks were developed and their printability was examined on a solution-based EHDP system for scaffold fabrication. Multilayer grid scaffolds were manufactured by PCL, PCL/zein-10, and PCL/zein-20 inks, respectively and characterized. The mechanical strength of scaffolds printed by PCL/zein composite inks was remarkably enhanced in terms of Young's modulus and yield stress. The enzyme-accelerated in vitro degradation study demonstrated that zein-containing scaffolds exhibited dose-responsive improvement on the degradation rate as evidenced by surface morphological change of fibers. Moreover, the biocompatibility of PCL/zein scaffolds, tested on mice embryonic fibroblast (NIH/3T3) and human nonsmall lung cancer cell (H1299), manifested better cell affinity. Our findings suggest that scaffolds fabricated by the solution-based EHDP with PCL/zein composite inks can significantly improve Young's modulus, yield stress, biocompatibility, and biodegradability and have potential applications in drug delivery systems, 3D cell culture modeling, or tissue engineering.

Visualizing Angiogenesis by Multiphoton Microscopy In Vivo in Genetically Modified 3D-PLGA/nHAp Scaffold for Calvarial Critical Bone Defect Repair.

PubMed

Li, Jian; Jahr, Holger; Zheng, Wei; Ren, Pei-Gen

2017-09-07

The reconstruction of critically sized bone defects remains a serious clinical problem because of poor angiogenesis within tissue-engineered scaffolds during repair, which gives rise to a lack of sufficient blood supply and causes necrosis of the new tissues. Rapid vascularization is a vital prerequisite for new tissue survival and integration with existing host tissue. The de novo generation of vasculature in scaffolds is one of the most important steps in making bone regeneration more efficient, allowing repairing tissue to grow into a scaffold. To tackle this problem, the genetic modification of a biomaterial scaffold is used to accelerate angiogenesis and osteogenesis. However, visualizing and tracking in vivo blood vessel formation in real-time and in three-dimensional (3D) scaffolds or new bone tissue is still an obstacle for bone tissue engineering. Multiphoton microscopy (MPM) is a novel bio-imaging modality that can acquire volumetric data from biological structures in a high-resolution and minimally-invasive manner. The objective of this study was to visualize angiogenesis with multiphoton microscopy in vivo in a genetically modified 3D-PLGA/nHAp scaffold for calvarial critical bone defect repair. PLGA/nHAp scaffolds were functionalized for the sustained delivery of a growth factor pdgf-b gene carrying lentiviral vectors (LV-pdgfb) in order to facilitate angiogenesis and to enhance bone regeneration. In a scaffold-implanted calvarial critical bone defect mouse model, the blood vessel areas (BVAs) in PHp scaffolds were significantly higher than in PH scaffolds. Additionally, the expression of pdgf-b and angiogenesis-related genes, vWF and VEGFR2, increased correspondingly. MicroCT analysis indicated that the new bone formation in the PHp group dramatically improved compared to the other groups. To our knowledge, this is the first time multiphoton microscopy was used in bone tissue-engineering to investigate angiogenesis in a 3D bio-degradable scaffold in

Co-delivery of cisplatin and doxorubicin from calcium phosphate beads/matrix scaffolds for osteosarcoma therapy.

PubMed

Hess, Ulrike; Shahabi, Shakiba; Treccani, Laura; Streckbein, Philipp; Heiss, Christian; Rezwan, Kurosch

2017-08-01

Bone substitute materials with a controlled drug release ability can fill cavities caused by the resection of bone tumours and thereby combat any leftover bone cancer cells. The combined release of different cytostatics seems to enhance their toxicity. In this study, calcium phosphate beads and matrix scaffolds are combined for a long-term co-delivery of cis-diamminedichloroplatinum (cisplatin, CDDP) and doxorubicin hydrochloride (DOX) as clinical relevant model drugs. Tricalcium phosphate/alginate beads as additional drug carrier are produced by droplet extrusion with ionotropic gelation and incorporated in scaffold matrix by freeze gelation without sintering. CDDP shows a short burst release while DOX has a continuous release measurable over the entire study period of 40days. Drug release from matrix is decreased by ~30% compared to release from beads. Nevertheless, all formulations follow the Korsmeyer-Peppas release kinetic model and show Fickian diffusion. Cytotoxic activity was conducted on MG-63 osteosarcoma cells after 1, 4, and 7days with WST-1 cell viability assay. Co-loaded composites enhance activity towards MG-63 cells up to ~75% toxicity while reducing the released drug quantity. The results suggest that co-loaded beads/matrix scaffolds are highly promising for osteosarcoma therapy due to synergistic effects over a long period of more than a month. Copyright © 2017 Elsevier B.V. All rights reserved.

Proliferation and enrichment of CD133(+) glioblastoma cancer stem cells on 3D chitosan-alginate scaffolds.

PubMed

Kievit, Forrest M; Florczyk, Stephen J; Leung, Matthew C; Wang, Kui; Wu, Jennifer D; Silber, John R; Ellenbogen, Richard G; Lee, Jerry S H; Zhang, Miqin

2014-11-01

Emerging evidence implicates cancer stem cells (CSCs) as primary determinants of the clinical behavior of human cancers, representing an ideal target for next-generation anti-cancer therapies. However CSCs are difficult to propagate in vitro, severely limiting the study of CSC biology and drug development. Here we report that growing cells from glioblastoma (GBM) cell lines on three dimensional (3D) porous chitosan-alginate (CA) scaffolds dramatically promotes the proliferation and enrichment of cells possessing the hallmarks of CSCs. CA scaffold-grown cells were found more tumorigenic in nude mouse xenografts than cells grown from monolayers. Growing in CA scaffolds rapidly promoted expression of genes involved in the epithelial-to-mesenchymal transition that has been implicated in the genesis of CSCs. Our results indicate that CA scaffolds have utility as a simple and inexpensive means to cultivate CSCs in vitro in support of studies to understand CSC biology and develop more effective anti-cancer therapies. Copyright © 2014 Elsevier Ltd. All rights reserved.

Physicochemical properties of 3D collagen-CS scaffolds for potential use in neural tissue engineering.

PubMed

Pietrucha, Krystyna

2015-09-01

Collagen-based composite scaffolds have considerable potential due to their well-known ability to regenerate skin, bone and cartilage. However, the precise composition and structure of scaffolds that optimize their interaction with neural cells remains incompletely understood and yet to be explored. In the present study, a new family of bi-component 3D scaffolds consisting of collagen (Col) and chondroitin sulphate (CS) were synthesized using a two-stage process: multiple freeze-drying followed by carbodiimide modification. Col-CS matrices had an average pore diameter of 31 μm and a relatively high surface area to pore volume ratio. Importantly, the FTIR data indicated that the ratio between the intensity of amide III and 1452 cm(-1) for Col-CS scaffold was 0.87, which indicates that the Col triple helix was preserved during the formation of the bond between Col and CS. All experiments also clearly showed that the Col-CS matrices have a lower enzyme sensitivity and higher thermal resistance than Col alone. These differences are likely due to the relatively large amount of CS in the collagen sponges, which hinders access for attack at specific active sites of the Col triple helix. Improved binary composite scaffolds were designed for neural tissue engineering applications. Copyright © 2015 Elsevier B.V. All rights reserved.

Effect of oxygen plasma etching on pore size-controlled 3D polycaprolactone scaffolds for enhancing the early new bone formation in rabbit calvaria.