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Sample records for 3d hydrogel scaffold

  1. Fabricating gradient hydrogel scaffolds for 3D cell culture.

    PubMed

    Chatterjee, Kaushik; Young, Marian F; Simon, Carl G

    2011-05-01

    Optimizing cell-material interactions is critical for maximizing regeneration in tissue engineering. Combinatorial and high-throughput (CHT) methods can be used to systematically screen tissue scaffolds to identify optimal biomaterial properties. Previous CHT platforms in tissue engineering have involved a two-dimensional (2D) cell culture format where cells were cultured on material surfaces. However, these platforms are inadequate to predict cellular response in a three-dimensional (3D) tissue scaffold. We have developed a simple CHT platform to screen cell-material interactions in 3D culture format that can be applied to screen hydrogel scaffolds. Herein we provide detailed instructions on a method to prepare gradients in elastic modulus of photopolymerizable hydrogels.

  2. 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.

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

    PubMed

    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

    2012-09-01

    The aortic valve exhibits complex three-dimensional (3D) anatomy and heterogeneity essential for the 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 12-22 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 tenfold 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 min, 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.

  4. 3D Hydrogel Scaffolds for Articular Chondrocyte Culture and Cartilage Generation

    PubMed Central

    Yang, Fan; Bhutani, Nidhi

    2015-01-01

    Human articular cartilage is highly susceptible to damage and has limited self-repair and regeneration potential. Cell-based strategies to engineer cartilage tissue offer a promising solution to repair articular cartilage. To select the optimal cell source for tissue repair, it is important to develop an appropriate culture platform to systematically examine the biological and biomechanical differences in the tissue-engineered cartilage by different cell sources. Here we applied a three-dimensional (3D) biomimetic hydrogel culture platform to systematically examine cartilage regeneration potential of juvenile, adult, and osteoarthritic (OA) chondrocytes. The 3D biomimetic hydrogel consisted of synthetic component poly(ethylene glycol) and bioactive component chondroitin sulfate, which provides a physiologically relevant microenvironment for in vitro culture of chondrocytes. In addition, the scaffold may be potentially used for cell delivery for cartilage repair in vivo. Cartilage tissue engineered in the scaffold can be evaluated using quantitative gene expression, immunofluorescence staining, biochemical assays, and mechanical testing. Utilizing these outcomes, we were able to characterize the differential regenerative potential of chondrocytes of varying age, both at the gene expression level and in the biochemical and biomechanical properties of the engineered cartilage tissue. The 3D culture model could be applied to investigate the molecular and functional differences among chondrocytes and progenitor cells from different stages of normal or aberrant development. PMID:26484414

  5. 3D differentiation of neural stem cells in macroporous photopolymerizable hydrogel scaffolds.

    PubMed

    Li, Hang; Wijekoon, Asanka; Leipzig, Nic D

    2012-01-01

    Neural stem/progenitor cells (NSPCs) are the stem cell of the adult central nervous system (CNS). These cells are able to differentiate into the major cell types found in the CNS (neurons, oligodendrocytes, astrocytes), thus NSPCs are the mechanism by which the adult CNS could potentially regenerate after injury or disorder. Microenviromental factors are critical for guiding NSPC differentiation and are thus important for neural tissue engineering. In this study, D-mannitol crystals were mixed with photocrosslinkable methacrylamide chitosan (MAC) as a porogen to enhance pore size during hydrogel formation. D-mannitol was admixed to MAC at 5, 10 and 20 wt% D-mannitol per total initial hydrogel weight. D-mannitol crystals were observed to dissolve and leave the scaffold within 1 hr. Quantification of resulting average pore sizes showed that D-mannitol addition resulted in larger average pore size (5 wt%, 4060±160 µm(2), 10 wt%, 6330±1160 µm(2), 20 wt%, 7600±1550 µm(2)) compared with controls (0 wt%, 3150±220 µm(2)). Oxygen diffusion studies demonstrated that larger average pore area resulted in enhanced oxygen diffusion through scaffolds. Finally, the differentiation responses of NSPCs to phenotypic differentiation conditions were studied for neurons, astrocytes and oligodendrocytes in hydrogels of varied porosity over 14 d. Quantification of total cell numbers at day 7 and 14, showed that cell numbers decreased with increased porosity and over the length of the culture. At day 14 immunohistochemistry quantification for primary cell types demonstrated significant differentiation to the desired cells types, and that total percentages of each cell type was greatest when scaffolds were more porous. These results suggest that larger pore sizes in MAC hydrogels effectively promote NSPC 3D differentiation.

  6. 3D Differentiation of Neural Stem Cells in Macroporous Photopolymerizable Hydrogel Scaffolds

    PubMed Central

    Li, Hang; Wijekoon, Asanka; Leipzig, Nic D.

    2012-01-01

    Neural stem/progenitor cells (NSPCs) are the stem cell of the adult central nervous system (CNS). These cells are able to differentiate into the major cell types found in the CNS (neurons, oligodendrocytes, astrocytes), thus NSPCs are the mechanism by which the adult CNS could potentially regenerate after injury or disorder. Microenviromental factors are critical for guiding NSPC differentiation and are thus important for neural tissue engineering. In this study, D-mannitol crystals were mixed with photocrosslinkable methacrylamide chitosan (MAC) as a porogen to enhance pore size during hydrogel formation. D-mannitol was admixed to MAC at 5, 10 and 20 wt% D-mannitol per total initial hydrogel weight. D-mannitol crystals were observed to dissolve and leave the scaffold within 1 hr. Quantification of resulting average pore sizes showed that D-mannitol addition resulted in larger average pore size (5 wt%, 4060±160 µm2, 10 wt%, 6330±1160 µm2, 20 wt%, 7600±1550 µm2) compared with controls (0 wt%, 3150±220 µm2). Oxygen diffusion studies demonstrated that larger average pore area resulted in enhanced oxygen diffusion through scaffolds. Finally, the differentiation responses of NSPCs to phenotypic differentiation conditions were studied for neurons, astrocytes and oligodendrocytes in hydrogels of varied porosity over 14 d. Quantification of total cell numbers at day 7 and 14, showed that cell numbers decreased with increased porosity and over the length of the culture. At day 14 immunohistochemistry quantification for primary cell types demonstrated significant differentiation to the desired cells types, and that total percentages of each cell type was greatest when scaffolds were more porous. These results suggest that larger pore sizes in MAC hydrogels effectively promote NSPC 3D differentiation. PMID:23144988

  7. Carboxy-Methyl-Cellulose (CMC) hydrogel-filled 3-D scaffold: Preliminary study through a 3-D antiproliferative activity of Centella asiatica extract

    NASA Astrophysics Data System (ADS)

    Aizad, Syazwan; Yahaya, Badrul Hisham; Zubairi, Saiful Irwan

    2015-09-01

    This study focuses on the effects of using the water extract from Centella asiatica on the mortality of human lung cancer cells (A549) with the use of novel 3-D scaffolds infused with CMC hydrogel. A biodegradable polymer, poly (hydroxybutyrate-co-hydroxyvalerate) (PHBV) was used in this study as 3-D scaffolds, with some modifications made by introducing the gel structure on its pore, which provides a great biomimetic microenvironment for cells to grow apart from increasing the interaction between the cells and cell-bioactive extracts. The CMC showed a good hydrophilic characteristic with mean contact angle of 24.30 ± 22.03°. To ensure the CMC gel had good attachments with the scaffolds, a surface treatment was made before the CMC gel was infused into the scaffolds. The results showed that these modified scaffolds contained 42.41 ± 0.14% w/w of CMC gel, which indicated that the gel had already filled up the entire pore of 3-D scaffolds. Besides, the infused hydrogel scaffolds took only 24 hours to be saturated when absorbing the water. The viability of cancer cells by MTS assay after being treated with Centella asiatica showed that the scaffolds infused with CMC hydrogel had the cell viability of 46.89 ± 1.20% followed by porous 3-D model with 57.30 ± 1.60% of cell viability, and the 2-D model with 67.10 ± 1.10% of cell viability. The inhibitory activity in cell viability between 2-D and 3-D models did not differ significantly (p>0.05) due to the limitation of time in incubating the extract with the cell in the 3-D model microenvironment. In conclusion, with the application of 3-D scaffolds infused with CMC hydrogel, the extracts of Centella asiatica has been proven to have the ability to kill cancer cells and have a great potential to become one of the alternative methods in treating cancer patients.

  8. 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.

  9. 3D hydrogel scaffold doped with 2D graphene materials for biosensors and bioelectronics.

    PubMed

    Song, Hyun Seok; Kwon, Oh Seok; Kim, Jae-Hong; Conde, João; Artzi, Natalie

    2017-03-15

    Hydrogels consisting of three-dimensional (3D) polymeric networks have found a wide range of applications in biotechnology due to their large water capacity, high biocompatibility, and facile functional versatility. The hydrogels with stimulus-responsive swelling properties have been particularly instrumental to realizing signal transduction in biosensors and bioelectronics. Graphenes are two-dimensional (2D) nanomaterials with unprecedented physical, optical, and electronic properties and have also found many applications in biosensors and bioelectronics. These two classes of materials present complementary strengths and limitations which, when effectively coupled, can result in significant synergism in their electrical, mechanical, and biocompatible properties. This report reviews recent advances made with hydrogel and graphene materials for the development of high-performance bioelectronics devices. The report focuses on the interesting intersection of these materials wherein 2D graphenes are hybridized with 3D hydrogels to develop the next generation biosensors and bioelectronics.

  10. The integration of 3-D cell printing and mesoscopic fluorescence molecular tomography of vascular constructs within thick hydrogel scaffolds.

    PubMed

    Zhao, Lingling; Lee, Vivian K; Yoo, Seung-Schik; Dai, Guohao; Intes, Xavier

    2012-07-01

    Developing methods that provide adequate vascular perfusion is an important step toward engineering large functional tissues. Meanwhile, an imaging modality to assess the three-dimensional (3-D) structures and functions of the vascular channels is lacking for thick matrices (>2 ≈ 3 mm). Herein, we report on an original approach to construct and image 3-D dynamically perfused vascular structures in thick hydrogel scaffolds. In this work, we integrated a robotic 3-D cell printing technology with a mesoscopic fluorescence molecular tomography imaging system, and demonstrated the capability of the platform to construct perfused collagen scaffolds with endothelial lining and to image both the fluid flow and fluorescent-labeled living endothelial cells at high-frame rates, with high sensitivity and accuracy. These results establish the potential of integrating both 3-D cell printing and fluorescence mesoscopic imaging for functional and molecular studies in complex tissue-engineered tissues.

  11. Design and Fabrication of Complex Scaffolds for Bone Defect Healing: Combined 3D Plotting of a Calcium Phosphate Cement and a Growth Factor-Loaded Hydrogel.

    PubMed

    Ahlfeld, Tilman; Akkineni, Ashwini Rahul; Förster, Yvonne; Köhler, Tino; Knaack, Sven; Gelinsky, Michael; Lode, Anja

    2017-01-01

    Additive manufacturing enables the fabrication of scaffolds with defined architecture. Versatile printing technologies such as extrusion-based 3D plotting allow in addition the incorporation of biological components increasing the capability to restore functional tissues. We have recently described the fabrication of calcium phosphate cement (CPC) scaffolds by 3D plotting of an oil-based CPC paste under mild conditions. In the present study, we have developed a strategy for growth factor loading based on multichannel plotting: a biphasic scaffold design was realised combining CPC with VEGF-laden, highly concentrated hydrogel strands. As hydrogel component, alginate and an alginate-gellan gum blend were evaluated; the blend exhibited a more favourable VEGF release profile and was chosen for biphasic scaffold fabrication. After plotting, two-step post-processing was performed for both, hydrogel crosslinking and CPC setting, which was shown to be compatible with both materials. Finally, a scaffold was designed and fabricated which can be applied for testing in a rat critical size femur defect. Optimization of CPC plotting enabled the fabrication of highly resolved structures with strand diameters of only 200 µm. Micro-computed tomography revealed a precise strand arrangement and an interconnected pore space within the biphasic scaffold even in swollen state of the hydrogel strands.

  12. 3D polymer scaffold arrays.

    PubMed

    Simon, Carl G; Yang, Yanyin; Dorsey, Shauna M; Ramalingam, Murugan; Chatterjee, Kaushik

    2011-01-01

    We have developed a combinatorial platform for fabricating tissue scaffold arrays that can be used for screening cell-material interactions. Traditional research involves preparing samples one at a time for characterization and testing. Combinatorial and high-throughput (CHT) methods lower the cost of research by reducing the amount of time and material required for experiments by combining many samples into miniaturized specimens. In order to help accelerate biomaterials research, many new CHT methods have been developed for screening cell-material interactions where materials are presented to cells as a 2D film or surface. However, biomaterials are frequently used to fabricate 3D scaffolds, cells exist in vivo in a 3D environment and cells cultured in a 3D environment in vitro typically behave more physiologically than those cultured on a 2D surface. Thus, we have developed a platform for fabricating tissue scaffold libraries where biomaterials can be presented to cells in a 3D format.

  13. 3D Cell Culture in Alginate Hydrogels

    PubMed Central

    Andersen, Therese; Auk-Emblem, Pia; Dornish, Michael

    2015-01-01

    This review compiles information regarding the use of alginate, and in particular alginate hydrogels, in culturing cells in 3D. Knowledge of alginate chemical structure and functionality are shown to be important parameters in design of alginate-based matrices for cell culture. Gel elasticity as well as hydrogel stability can be impacted by the type of alginate used, its concentration, the choice of gelation technique (ionic or covalent), and divalent cation chosen as the gel inducing ion. The use of peptide-coupled alginate can control cell–matrix interactions. Gelation of alginate with concomitant immobilization of cells can take various forms. Droplets or beads have been utilized since the 1980s for immobilizing cells. Newer matrices such as macroporous scaffolds are now entering the 3D cell culture product market. Finally, delayed gelling, injectable, alginate systems show utility in the translation of in vitro cell culture to in vivo tissue engineering applications. Alginate has a history and a future in 3D cell culture. Historically, cells were encapsulated in alginate droplets cross-linked with calcium for the development of artificial organs. Now, several commercial products based on alginate are being used as 3D cell culture systems that also demonstrate the possibility of replacing or regenerating tissue. PMID:27600217

  14. Research on the printability of hydrogels in 3D bioprinting

    PubMed Central

    He, Yong; Yang, FeiFei; Zhao, HaiMing; Gao, Qing; Xia, Bing; Fu, JianZhong

    2016-01-01

    As the biocompatible materials, hydrogels have been widely used in three- dimensional (3D) bioprinting/organ printing to load cell for tissue engineering. It is important to precisely control hydrogels deposition during printing the mimic organ structures. However, the printability of hydrogels about printing parameters is seldom addressed. In this paper, we systemically investigated the printability of hydrogels from printing lines (one dimensional, 1D structures) to printing lattices/films (two dimensional, 2D structures) and printing 3D structures with a special attention to the accurate printing. After a series of experiments, we discovered the relationships between the important factors such as air pressure, feedrate, or even printing distance and the printing quality of the expected structures. Dumbbell shape was observed in the lattice structures printing due to the hydrogel diffuses at the intersection. Collapses and fusion of adjacent layer would result in the error accumulation at Z direction which was an important fact that could cause printing failure. Finally, we successfully demonstrated a 3D printing hydrogel scaffold through harmonize with all the parameters. The cell viability after printing was compared with the casting and the results showed that our bioprinting method almost had no extra damage to the cells. PMID:27436509

  15. Research on the printability of hydrogels in 3D bioprinting

    NASA Astrophysics Data System (ADS)

    He, Yong; Yang, Feifei; Zhao, Haiming; Gao, Qing; Xia, Bing; Fu, Jianzhong

    2016-07-01

    As the biocompatible materials, hydrogels have been widely used in three- dimensional (3D) bioprinting/organ printing to load cell for tissue engineering. It is important to precisely control hydrogels deposition during printing the mimic organ structures. However, the printability of hydrogels about printing parameters is seldom addressed. In this paper, we systemically investigated the printability of hydrogels from printing lines (one dimensional, 1D structures) to printing lattices/films (two dimensional, 2D structures) and printing 3D structures with a special attention to the accurate printing. After a series of experiments, we discovered the relationships between the important factors such as air pressure, feedrate, or even printing distance and the printing quality of the expected structures. Dumbbell shape was observed in the lattice structures printing due to the hydrogel diffuses at the intersection. Collapses and fusion of adjacent layer would result in the error accumulation at Z direction which was an important fact that could cause printing failure. Finally, we successfully demonstrated a 3D printing hydrogel scaffold through harmonize with all the parameters. The cell viability after printing was compared with the casting and the results showed that our bioprinting method almost had no extra damage to the cells.

  16. Biofunctionalized Hydrogel Microscaffolds Promote 3D Hepatic Sheet Morphology.

    PubMed

    Kim, Myung Hee; Kumar, Supriya K; Shirahama, Hitomi; Seo, Jeongeun; Lee, Jae-Ho; Zhdanov, Vladimir P; Cho, Nam-Joon

    2016-03-01

    Development of artificial tissues providing the proper geometrical, mechanical, and environmental cues for cells is highly coveted in the field of tissue engineering. Recently, microfabrication strategies in combination with other chemistries have been utilized to capture the architectural complexity of intricate organs, such as the liver, in in vitro platforms. Here it is shown that a biofunctionalized poly (ethylene glycol) (PEG) hydrogel scaffold, fabricated using a sphere-template, facilitates hepatic sheet formation that follows the microscale patterns of the scaffold surface. The design takes advantage of the excellent diffusion properties of porous, uniform 3D hydrogel platforms, and the enhanced-cell-extracellular matrix interaction with the display of conjugated collagen type I, which in turn elicits favorable Huh-7.5 response. Collectively, the experimental findings and corresponding simulations demonstrate the importance of biofunctionalized porous scaffolds and indicate that the microscaffold shows promise in liver tissue engineering applications and provides distinct advantages over current cell sheet and hepatocyte spheroid technologies.

  17. Peptide hydrogelation and cell encapsulation for 3D culture of MCF-7 breast cancer cells.

    PubMed

    Huang, Hongzhou; Ding, Ying; Sun, Xiuzhi S; Nguyen, Thu A

    2013-01-01

    Three-dimensional (3D) cell culture plays an invaluable role in tumor biology by providing in vivo like microenviroment and responses to therapeutic agents. Among many established 3D scaffolds, hydrogels demonstrate a distinct property as matrics for 3D cell culture. Most of the existing pre-gel solutions are limited under physiological conditions such as undesirable pH or temperature. Here, we report a peptide hydrogel that shows superior physiological properties as an in vitro matrix for 3D cell culture. The 3D matrix can be accomplished by mixing a self-assembling peptide directly with a cell culture medium without any pH or temperature adjustment. Results of dynamic rheological studies showed that this hydrogel can be delivered multiple times via pipetting without permanently destroying the hydrogel architecture, indicating the deformability and remodeling ability of the hydrogel. Human epithelial cancer cells, MCF-7, are encapsulated homogeneously in the hydrogel matrix during hydrogelation. Compared with two-dimensional (2D) monolayer culture, cells residing in the hydrogel matrix grow as tumor-like clusters in 3D formation. Relevant parameters related to cell morphology, survival, proliferation, and apoptosis were analyzed using MCF-7 cells in 3D hydrogels. Interestingly, treatment of cisplatin, an anti-cancer drug, can cause a significant decrease of cell viability of MCF-7 clusters in hydrogels. The responses to cisplatin were dose- and time-dependent, indicating the potential usage of hydrogels for drug testing. Results of confocal microscopy and Western blotting showed that cells isolated from hydrogels are suitable for downstream proteomic analysis. The results provided evidence that this peptide hydrogel is a promising 3D cell culture material for drug testing.

  18. 3D Printing: 3D Printing of Highly Stretchable and Tough Hydrogels into Complex, Cellularized Structures.

    PubMed

    Hong, Sungmin; Sycks, Dalton; Chan, Hon Fai; Lin, Shaoting; Lopez, Gabriel P; Guilak, Farshid; Leong, Kam W; Zhao, Xuanhe

    2015-07-15

    X. Zhao and co-workers develop on page 4035 a new biocompatible hydrogel system that is extremely tough and stretchable and can be 3D printed into complex structures, such as the multilayer mesh shown. Cells encapsulated in the tough and printable hydrogel maintain high viability. 3D-printed structures of the tough hydrogel can sustain high mechanical loads and deformations.

  19. 3D high-resolution two-photon crosslinked hydrogel structures for biological studies.

    PubMed

    Brigo, Laura; Urciuolo, Anna; Giulitti, Stefano; Giustina, Gioia Della; Tromayer, Maximilian; Liska, Robert; Elvassore, Nicola; Brusatin, Giovanna

    2017-03-25

    Hydrogels are widely used as matrices for cell growth due to the their tuneable chemical and physical properties, which mimic the extracellular matrix of natural tissue. The microfabrication of hydrogels into arbitrarily complex 3D structures is becoming essential for numerous biological applications, and in particular for investigating the correlation between cell shape and cell function in a 3D environment. Micrometric and sub-micrometric resolution hydrogel scaffolds are required to deeply investigate molecular mechanisms behind cell-matrix interaction and downstream cellular processes. We report the design and development of high resolution 3D gelatin hydrogel woodpile structures by two-photon crosslinking. Hydrated structures of lateral linewidth down to 0.5 µm, lateral and axial resolution down to a few µm are demonstrated. According to the processing parameters, different degrees of polymerization are obtained, resulting in hydrated scaffolds of variable swelling and deformation. The 3D hydrogels are biocompatible and promote cell adhesion and migration. Interestingly, according to the polymerization degree, 3D hydrogel woodpile structures show variable extent of cell adhesion and invasion. Human BJ cell lines show capability of deforming 3D micrometric resolved hydrogel structures.

  20. 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.

  1. 3D Printing Facilitated Scaffold-free Tissue Unit Fabrication

    PubMed Central

    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

    2014-01-01

    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 micro-droplets 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 micro-droplets 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. PMID:24717646

  2. Capillary Origami Inspired Fabrication of Complex 3D Hydrogel Constructs.

    PubMed

    Li, Moxiao; Yang, Qingzhen; Liu, Hao; Qiu, Mushu; Lu, Tian Jian; Xu, Feng

    2016-09-01

    Hydrogels have found broad applications in various engineering and biomedical fields, where the shape and size of hydrogels can profoundly influence their functions. Although numerous methods have been developed to tailor 3D hydrogel structures, it is still challenging to fabricate complex 3D hydrogel constructs. Inspired by the capillary origami phenomenon where surface tension of a droplet on an elastic membrane can induce spontaneous folding of the membrane into 3D structures along with droplet evaporation, a facile strategy is established for the fabrication of complex 3D hydrogel constructs with programmable shapes and sizes by crosslinking hydrogels during the folding process. A mathematical model is further proposed to predict the temporal structure evolution of the folded 3D hydrogel constructs. Using this model, precise control is achieved over the 3D shapes (e.g., pyramid, pentahedron, and cube) and sizes (ranging from hundreds of micrometers to millimeters) through tuning membrane shape, dimensionless parameter of the process (elastocapillary number Ce ), and evaporation time. This work would be favorable to multiple areas, such as flexible electronics, tissue regeneration, and drug delivery.

  3. Sequential assembly of 3D perfusable microfluidic hydrogels.

    PubMed

    He, Jiankang; Zhu, Lin; Liu, Yaxiong; Li, Dichen; Jin, Zhongmin

    2014-11-01

    Bottom-up tissue engineering provides a promising way to recreate complex structural organizations of native organs in artificial constructs by assembling functional repeating modules. However, it is challenging for current bottom-up strategies to simultaneously produce a controllable and immediately perfusable microfluidic network in modularly assembled 3D constructs. Here we presented a bottom-up strategy to produce perfusable microchannels in 3D hydrogels by sequentially assembling microfluidic modules. The effects of agarose-collagen composition on microchannel replication and 3D assembly of hydrogel modules were investigated. The unique property of predefined microchannels in transporting fluids within 3D assemblies was evaluated. Endothelial cells were incorporated into the microfluidic network of 3D hydrogels for dynamic culture in a house-made bioreactor system. The results indicated that the sequential assembly method could produce interconnected 3D predefined microfluidic networks in optimized agarose-collagen hydrogels, which were fully perfusable and successfully functioned as fluid pathways to facilitate the spreading of endothelial cells. We envision that the presented method could be potentially used to engineer 3D vascularized parenchymal constructs by encapsulating primary cells in bulk hydrogels and incorporating endothelial cells in predefined microchannels.

  4. Programming Mechanical and Physicochemical Properties of 3D Hydrogel Cellular Microcultures via Direct Ink Writing.

    PubMed

    McCracken, Joselle M; Badea, Adina; Kandel, Mikhail E; Gladman, A Sydney; Wetzel, David J; Popescu, Gabriel; Lewis, Jennifer A; Nuzzo, Ralph G

    2016-05-01

    3D hydrogel scaffolds are widely used in cellular microcultures and tissue engineering. Using direct ink writing, microperiodic poly(2-hydroxyethyl-methacrylate) (pHEMA) scaffolds are created that are then printed, cured, and modified by absorbing 30 kDa protein poly-l-lysine (PLL) to render them biocompliant in model NIH/3T3 fibroblast and MC3T3-E1 preosteoblast cell cultures. Spatial light interference microscopy (SLIM) live cell imaging studies are carried out to quantify cellular motilities for each cell type, substrate, and surface treatment of interest. 3D scaffold mechanics is investigated using atomic force microscopy (AFM), while their absorption kinetics are determined by confocal fluorescence microscopy (CFM) for a series of hydrated hydrogel films prepared from prepolymers with different homopolymer-to-monomer (Mr ) ratios. The observations reveal that the inks with higher Mr values yield relatively more open-mesh gels due to a lower degree of entanglement. The biocompatibility of printed hydrogel scaffolds can be controlled by both PLL content and hydrogel mesh properties.

  5. Hydrogels and scaffolds for immunomodulation.

    PubMed

    Singh, Ankur; Peppas, Nicholas A

    2014-10-01

    For over two decades, immunologists and biomaterials scientists have co-existed in parallel world with the rationale of understanding the molecular profile of immune responses to vaccination, implantation, and treating incurable diseases. Much of the field of biomaterial-based immunotherapy has relied on evaluating model antigens such as chicken egg ovalbumin in mouse models but their relevance to humans has been point of much discussion. Nevertheless, such model antigens have provided important insights into the mechanisms of immune regulation and served as a proof-of-concept for plethora of biomaterial-based vaccines. After years of extensive development of numerous biomaterials for immunomodulation, it is only recently that an experimental scaffold vaccine implanted beneath the skin has begun to use the human model to study the immune responses to cancer vaccination by co-delivering patient-derived tumor lysates and immunomodulatory proteins. If successful, this scaffold vaccine will change the way we approached untreatable cancers, but more importantly, will allow a faster and more rational translation of therapeutic regimes to other cancers, chronic infections, and autoimmune diseases. Most materials reviews have focused on immunomodulatory adjuvants and micro-nano-particles. Here we provide an insight into emerging hydrogel and scaffold based immunomodulatory approaches that continue to demonstrate efficacy against immune associated diseases.

  6. Influence of scaffold design on 3D printed cell constructs.

    PubMed

    Souness, Auryn; Zamboni, Fernanda; Walker, Gavin M; Collins, Maurice N

    2017-02-14

    Additive manufacturing is currently receiving significant attention in the field of tissue engineering and biomaterial science. The development of precise, affordable 3D printing technologies has provided a new platform for novel research to be undertaken in 3D scaffold design and fabrication. In the past, a number of 3D scaffold designs have been fabricated to investigate the potential of a 3D printed scaffold as a construct which could support cellular life. These studies have shown promising results; however, few studies have utilized a low-cost desktop 3D printing technology as a potential rapid manufacturing route for different scaffold designs. Here six scaffold designs were manufactured using a Fused deposition modeling, a "bottom-up" solid freeform fabrication approach, to determine optimal scaffold architecture for three-dimensional cell growth. The scaffolds, produced from PLA, are coated using pullulan and hyaluronic acid to assess the coating influence on cell proliferation and metabolic rate. Scaffolds are characterized both pre- and postprocessing using water uptake analysis, mechanical testing, and morphological evaluation to study the inter-relationships between the printing process, scaffold design, and scaffold properties. It was found that there were key differences between each scaffold design in terms of porosity, diffusivity, swellability, and compressive strength. An optimal design was chosen based on these physical measurements which were then weighted in accordance to design importance based on literature and utilizing a design matrix technique. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2017.

  7. Automated quantitative assessment of three-dimensional bioprinted hydrogel scaffolds using optical coherence tomography

    PubMed Central

    Wang, Ling; Xu, Mingen; Zhang, LieLie; Zhou, QingQing; Luo, Li

    2016-01-01

    Reconstructing and quantitatively assessing the internal architecture of opaque three-dimensional (3D) bioprinted hydrogel scaffolds is difficult but vital to the improvement of 3D bioprinting techniques and to the fabrication of functional engineered tissues. In this study, swept-source optical coherence tomography was applied to acquire high-resolution images of hydrogel scaffolds. Novel 3D gelatin/alginate hydrogel scaffolds with six different representative architectures were fabricated using our 3D bioprinting system. Both the scaffold material networks and the interconnected flow channel networks were reconstructed through volume rendering and binarisation processing to provide a 3D volumetric view. An image analysis algorithm was developed based on the automatic selection of the spatially-isolated region-of–interest. Via this algorithm, the spatially-resolved morphological parameters including pore size, pore shape, strut size, surface area, porosity, and interconnectivity were quantified precisely. Fabrication defects and differences between the designed and as-produced scaffolds were clearly identified in both 2D and 3D; the locations and dimensions of each of the fabrication defects were also defined. It concludes that this method will be a key tool for non-destructive and quantitative characterization, design optimisation and fabrication refinement of 3D bioprinted hydrogel scaffolds. Furthermore, this method enables investigation into the quantitative relationship between scaffold structure and biological outcome. PMID:27231597

  8. 3D Printing of Ultratough Polyion Complex Hydrogels.

    PubMed

    Zhu, Fengbo; Cheng, Libo; Yin, Jun; Wu, Zi Liang; Qian, Jin; Fu, Jianzhong; Zheng, Qiang

    2016-11-16

    Polyion complex (PIC) hydrogels have been proposed as promising engineered soft materials due to their high toughness and good processability. In this work, we reported manufacturing of complex structures with tough PIC hydrogels based on three-dimensional (3D) printing technology. The strategy relies on the distinct strength of ionic bonding in PIC hydrogels at different stages of printing. In concentrated saline solution, PIC forms viscous solution, which can be directly extruded out of a nozzle into water, where dialyzing out of salt and counterions results in sol-gel transition to form tough physical PIC gel with intricate structures. The printability of PIC solutions was systematically investigated by adjusting the PIC material formula and printing parameters in which proper viscosity and gelation rate were found to be key factors for successful 3D printing. Uniaxial tensile tests were performed to printed single fibers and multilayer grids, both exhibiting distinct yet controllable strength and toughness. More complex 3D structures with negative Poisson's ratio, gradient grid, and material anisotropy were constructed as well, demonstrating the flexible printability of PIC hydrogels. The methodology and capability here provide a versatile platform to fabricate complex structures with tough PIC hydrogels, which should broaden the use of such materials in applications such as biomedical devices and artificial tissues.

  9. Design of novel 3D gene activated PEG scaffolds with ordered pore structure.

    PubMed

    Orsi, Silvia; Guarnieri, Daniela; Netti, Paolo A

    2010-03-01

    The ability to genetically modify cells seeded inside synthetic hydrogel scaffolds offers a suitable approach to induce and control tissue repair and regeneration guiding cell fate. In fact the transfected cells can act as local in vivo bioreactor, secreting plasmid encoded proteins that augment tissue regeneration processes. We have realized a DNA bioactivated high porous poly(ethylene glycol) (PEG) matrix by polyethyleneimine (PEI)/DNA complexes adsorption. As the design of the microarchitectural features of a scaffold also contributes to promote and influence cell fate, we appropriately designed the inner structure of gene activated PEG hydrogels by gelatine microparticles templating. Microarchitectural properties of the scaffold were analysed by scanning electron microscopy. 3D cell migration and transfection were monitored through time-lapse videomicroscopy and confocal laser scanning microscopy.

  10. Cell proliferation and migration in silk fibroin 3D scaffolds.

    PubMed

    Mandal, Biman B; Kundu, Subhas C

    2009-05-01

    Pore architecture in 3D polymeric scaffolds is known to play a critical role in tissue engineering as it provides the vital framework for the seeded cells to organize into a functioning tissue. In this report, we investigated the effects of different freezing temperature regimes on silk fibroin protein 3D scaffold pore microstructure. The fabricated scaffolds using freeze-dry technique were used as a 3D model to monitor cell proliferation and migration. Pores of 200-250microm diameter were formed by slow cooling at temperatures of -20 and -80 degrees C but were found to be limited in porosity and pore interconnectivity as observed through scanning electron microscopic images. In contrast, highly interconnected pores with 96% porosity were observed when silk solutions were rapidly frozen at -196 degrees C. A detailed study was conducted to assess the affect of pore size, porosity and interconnectivity on human dermal fibroblast cell proliferation and migration on these 3D scaffolds using confocal microscopy. The cells were observed to migrate within the scaffold interconnectivities and were found to reach scaffold periphery within 28 days of culture. Confocal images further confirmed normal cell attachment and alignment of actin filaments within the porous scaffold matrix with well-developed nuclei. This study indicates rapid freeze-drying technique as an alternative method to fabricate highly interconnected porous scaffolds for developing functional 3D silk fibroin matrices for potential tissue engineering, biomedical and biotechnological applications.

  11. 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

    2017-01-11

    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.

  12. 3D Printed Trileaflet Valve Conduits Using Biological Hydrogels and Human Valve Interstitial Cells

    PubMed Central

    Duan, Bin; Kapetanovic, Edi; Hockaday, Laura A.; Butcher, Jonathan T.

    2014-01-01

    Tissue engineering has great potential to provide a functional de novo living valve replacement capable of integration with host tissue and growth. Among various valve conduit fabrication techniques, 3D bioprinting enables deposition of cells and hydrogels into 3D constructs with anatomical geometry and heterogeneous mechanical properties. Successful translation of this approach is however constrained by the dearth of printable and biocompatible hydrogel materials. Furthermore, it is not known how human valve cells respond to these printed environments. In this study, we develop 3D printable formulations of hybrid hydrogels based on methacrylated hyaluronic acid (Me-HA) and methacrylated gelatin (Me-Gel), and utilize them to bioprint heart valve conduits containing encapsulated human aortic valvular interstitial cells (HAVIC). Increasing Me-Gel concentration resulted in lower stiffness and higher viscosity, facilitated cell spreading, and better maintained HAVIC fibroblastic phenotype. Bioprinting accuracy was dependent upon the relative concentrations of Me-Gel and Me-HA, but when optimized enabled the fabrication of a trileaflet valve shape accurate to the original design. HAVIC encapsulated within bioprinted heart valves maintained high viability, and remodeled the initial matrix by depositing collagen and glyosaminoglycans. These findings represent the first rational design of bioprinted trileaflet valve hydrogels that regulate encapsulated human VIC behavior. The use of anatomically accurate living valve scaffolds through bioprinting may accelerate our understanding of physiological valve cell interactions and our progress towards de novo living valve replacements. PMID:24334142

  13. 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.

  14. Multifunctional 3D printing of heterogeneous hydrogel structures.

    PubMed

    Nadernezhad, Ali; Khani, Navid; Skvortsov, Gözde Akdeniz; Toprakhisar, Burak; Bakirci, Ezgi; Menceloglu, Yusuf; Unal, Serkan; Koc, Bahattin

    2016-09-15

    Multimaterial additive manufacturing or three-dimensional (3D) printing of hydrogel structures provides the opportunity to engineer geometrically dependent functionalities. However, current fabrication methods are mostly limited to one type of material or only provide one type of functionality. In this paper, we report a novel method of multimaterial deposition of hydrogel structures based on an aspiration-on-demand protocol, in which the constitutive multimaterial segments of extruded filaments were first assembled in liquid state by sequential aspiration of inks into a glass capillary, followed by in situ gel formation. We printed different patterned objects with varying chemical, electrical, mechanical, and biological properties by tuning process and material related parameters, to demonstrate the abilities of this method in producing heterogeneous and multi-functional hydrogel structures. Our results show the potential of proposed method in producing heterogeneous objects with spatially controlled functionalities while preserving structural integrity at the switching interface between different segments. We anticipate that this method would introduce new opportunities in multimaterial additive manufacturing of hydrogels for diverse applications such as biosensors, flexible electronics, tissue engineering and organ printing.

  15. Multifunctional 3D printing of heterogeneous hydrogel structures

    NASA Astrophysics Data System (ADS)

    Nadernezhad, Ali; Khani, Navid; Skvortsov, Gözde Akdeniz; Toprakhisar, Burak; Bakirci, Ezgi; Menceloglu, Yusuf; Unal, Serkan; Koc, Bahattin

    2016-09-01

    Multimaterial additive manufacturing or three-dimensional (3D) printing of hydrogel structures provides the opportunity to engineer geometrically dependent functionalities. However, current fabrication methods are mostly limited to one type of material or only provide one type of functionality. In this paper, we report a novel method of multimaterial deposition of hydrogel structures based on an aspiration-on-demand protocol, in which the constitutive multimaterial segments of extruded filaments were first assembled in liquid state by sequential aspiration of inks into a glass capillary, followed by in situ gel formation. We printed different patterned objects with varying chemical, electrical, mechanical, and biological properties by tuning process and material related parameters, to demonstrate the abilities of this method in producing heterogeneous and multi-functional hydrogel structures. Our results show the potential of proposed method in producing heterogeneous objects with spatially controlled functionalities while preserving structural integrity at the switching interface between different segments. We anticipate that this method would introduce new opportunities in multimaterial additive manufacturing of hydrogels for diverse applications such as biosensors, flexible electronics, tissue engineering and organ printing.

  16. Multifunctional 3D printing of heterogeneous hydrogel structures

    PubMed Central

    Nadernezhad, Ali; Khani, Navid; Skvortsov, Gözde Akdeniz; Toprakhisar, Burak; Bakirci, Ezgi; Menceloglu, Yusuf; Unal, Serkan; Koc, Bahattin

    2016-01-01

    Multimaterial additive manufacturing or three-dimensional (3D) printing of hydrogel structures provides the opportunity to engineer geometrically dependent functionalities. However, current fabrication methods are mostly limited to one type of material or only provide one type of functionality. In this paper, we report a novel method of multimaterial deposition of hydrogel structures based on an aspiration-on-demand protocol, in which the constitutive multimaterial segments of extruded filaments were first assembled in liquid state by sequential aspiration of inks into a glass capillary, followed by in situ gel formation. We printed different patterned objects with varying chemical, electrical, mechanical, and biological properties by tuning process and material related parameters, to demonstrate the abilities of this method in producing heterogeneous and multi-functional hydrogel structures. Our results show the potential of proposed method in producing heterogeneous objects with spatially controlled functionalities while preserving structural integrity at the switching interface between different segments. We anticipate that this method would introduce new opportunities in multimaterial additive manufacturing of hydrogels for diverse applications such as biosensors, flexible electronics, tissue engineering and organ printing. PMID:27630079

  17. Hybrid 3D-2D printing for bone scaffolds fabrication

    NASA Astrophysics Data System (ADS)

    Seleznev, V. A.; Prinz, V. Ya

    2017-02-01

    It is a well-known fact that bone scaffold topography on micro- and nanometer scale influences the cellular behavior. Nano-scale surface modification of scaffolds allows the modulation of biological activity for enhanced cell differentiation. To date, there has been only a limited success in printing scaffolds with micro- and nano-scale features exposed on the surface. To improve on the currently available imperfect technologies, in our paper we introduce new hybrid technologies based on a combination of 2D (nano imprint) and 3D printing methods. The first method is based on using light projection 3D printing and simultaneous 2D nanostructuring of each of the layers during the formation of the 3D structure. The second method is based on the sequential integration of preliminarily created 2D nanostructured films into a 3D printed structure. The capabilities of the developed hybrid technologies are demonstrated with the example of forming 3D bone scaffolds. The proposed technologies can be used to fabricate complex 3D micro- and nanostructured products for various fields.

  18. 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

  19. 3D Cell Entrapment as a Function of the Weight Percent of Peptide-Amphiphile Hydrogels

    PubMed Central

    Scott, Carolyn M.; Forster, Colleen L.; Kokkoli, Efrosini

    2015-01-01

    The design of scaffolds which mimic the stiffness, nanofiber structure, and biochemistry of the native extra-cellular matrix (ECM) has been a major objective for the tissue engineering field. Furthermore, mimicking the innate three dimensional (3D) environment of the ECM has been shown to significantly alter cellular response compared to traditional two dimensional (2D) culture. We report the development of a self-assembling, fibronectin-mimetic, peptide-amphiphile nanofiber scaffold for 3D cell culture. To form such a scaffold, 5 mol% of a bioactive PR_g fibronectin-mimetic peptide-amphiphile was mixed with 95 mol% of a diluent peptide-amphiphile (E2) whose purpose was to neutralize electrostatic interactions, increase the gelation kinetics and promote cell survival. Atomic force microscopy verified the fibrilar structure of the gels and the mechanical properties were characterized for various weight percent (wt%) formulations of the 5 mol% PR_g - 95 mol% E2 peptide-amphiphile mixture. The 0.5 wt% formulations had an elastic modulus of 429.0 ± 21.3 Pa while the 1.0 wt% peptide-amphiphile hydrogels had an elastic modulus of 808.6 ± 38.1 Pa. The presence of entrapped cells in the gels decreased the elastic modulus and the decrease was a function of the cell loading. While both formulations supported cell proliferation, the 0.5 wt% gels supported significantly greater NIH3T3/GFP fibroblast cell proliferation throughout the gels than the 1.0 wt% gels. However, compared to the 0.5 wt% formulations, the 1.0 wt% hydrogels promoted greater increase in mRNA expression and production of fibronectin and type IV collagen ECM proteins. This study suggests that this fibronectin-mimetic scaffold holds great promise in the advance of 3D culture applications and cell therapies. PMID:25970351

  20. 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.

  1. 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.

  2. Hybrid 3D-2D printing of bone scaffolds Hybrid 3D-2D printing methods for bone scaffolds fabrication.

    PubMed

    Prinz, V Ya; Seleznev, Vladimir

    2016-12-13

    It is a well-known fact that bone scaffold topography on micro- and nanometer scale influences the cellular behavior. Nano-scale surface modification of scaffolds allows the modulation of biological activity for enhanced cell differentiation. To date, there has been only a limited success in printing scaffolds with micro- and nano-scale features exposed on the surface. To improve on the currently available imperfect technologies, in our paper we introduce new hybrid technologies based on a combination of 2D (nano imprint) and 3D printing methods. The first method is based on using light projection 3D printing and simultaneous 2D nanostructuring of each of the layers during the formation of the 3D structure. The second method is based on the sequential integration of preliminarily created 2D nanostructured films into a 3D printed structure. The capabilities of the developed hybrid technologies are demonstrated with the example of forming 3D bone scaffolds. The proposed technologies can be used to fabricate complex 3D micro- and nanostructured products for various fields.

  3. 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.

  4. 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

  5. Hydrogel scaffolds for tissue engineering: Progress and challenges

    PubMed Central

    El-Sherbiny, Ibrahim M.; Yacoub, Magdi H.

    2013-01-01

    Designing of biologically active scaffolds with optimal characteristics is one of the key factors for successful tissue engineering. Recently, hydrogels have received a considerable interest as leading candidates for engineered tissue scaffolds due to their unique compositional and structural similarities to the natural extracellular matrix, in addition to their desirable framework for cellular proliferation and survival. More recently, the ability to control the shape, porosity, surface morphology, and size of hydrogel scaffolds has created new opportunities to overcome various challenges in tissue engineering such as vascularization, tissue architecture and simultaneous seeding of multiple cells. This review provides an overview of the different types of hydrogels, the approaches that can be used to fabricate hydrogel matrices with specific features and the recent applications of hydrogels in tissue engineering. Special attention was given to the various design considerations for an efficient hydrogel scaffold in tissue engineering. Also, the challenges associated with the use of hydrogel scaffolds were described. PMID:24689032

  6. 3D bioprinting of neural stem cell-laden thermoresponsive biodegradable polyurethane hydrogel and potential in central nervous system repair.

    PubMed

    Hsieh, Fu-Yu; Lin, Hsin-Hua; Hsu, Shan-Hui

    2015-12-01

    The 3D bioprinting technology serves as a powerful tool for building tissue in the field of tissue engineering. Traditional 3D printing methods involve the use of heat, toxic organic solvents, or toxic photoinitiators for fabrication of synthetic scaffolds. In this study, two thermoresponsive water-based biodegradable polyurethane dispersions (PU1 and PU2) were synthesized which may form gel near 37 °C without any crosslinker. The stiffness of the hydrogel could be easily fine-tuned by the solid content of the dispersion. Neural stem cells (NSCs) were embedded into the polyurethane dispersions before gelation. The dispersions containing NSCs were subsequently printed and maintained at 37 °C. The NSCs in 25-30% PU2 hydrogels (∼680-2400 Pa) had excellent proliferation and differentiation but not in 25-30% PU1 hydrogels. Moreover, NSC-laden 25-30% PU2 hydrogels injected into the zebrafish embryo neural injury model could rescue the function of impaired nervous system. However, NSC-laden 25-30% PU1 hydrogels only showed a minor repair effect in the zebrafish model. In addition, the function of adult zebrafish with traumatic brain injury was rescued after implantation of the 3D-printed NSC-laden 25% PU2 constructs. Therefore, the newly developed 3D bioprinting technique involving NSCs embedded in the thermoresponsive biodegradable polyurethane ink offers new possibilities for future applications of 3D bioprinting in neural tissue engineering.

  7. 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.

  8. 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.

  9. The medial scaffold of 3D unorganized point clouds.

    PubMed

    Leymarie, Frederic F; Kimia, Benjamin B

    2007-02-01

    We introduce the notion of the medial scaffold, a hierarchical organization of the medial axis of a 3D shape in the form of a graph constructed from special medial curves connecting special medial points. A key advantage of the scaffold is that it captures the qualitative aspects of shape in a hierarchical and tightly condensed representation. We propose an efficient and exact method for computing the medial scaffold based on a notion of propagation along the scaffold itself, starting from initial sources of the flow and constructing the scaffold during the propagation. We examine this method specifically in the context of an unorganized cloud of points in 3D, e.g., as obtained from laser range finders, which typically involve hundreds of thousands of points, but the ideas are generalizable to data arising from geometrically described surface patches. The computational bottleneck in the propagation-based scheme is in finding the initial sources of the flow. We thus present several ideas to avoid the unnecessary consideration of pairs of points which cannot possibly form a medial point source, such as the "visibility" of a point from another given a third point and the interaction of clusters of points. An application of using the medial scaffold for the representation of point samplings of real-life objects is also illustrated.

  10. 3D Printing of Highly Stretchable and Tough Hydrogels into Complex, Cellularized Structures.

    PubMed

    Hong, Sungmin; Sycks, Dalton; Chan, Hon Fai; Lin, Shaoting; Lopez, Gabriel P; Guilak, Farshid; Leong, Kam W; Zhao, Xuanhe

    2015-07-15

    A 3D printable and highly stretchable tough hydrogel is developed by combining poly(ethylene glycol) and sodium alginate, which synergize to form a hydrogel tougher than natural cartilage. Encapsulated cells maintain high viability over a 7 d culture period and are highly deformed together with the hydrogel. By adding biocompatible nanoclay, the tough hydrogel is 3D printed in various shapes without requiring support material.

  11. 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.

  12. Hydrogel-based reinforcement of 3D bioprinted constructs

    PubMed Central

    Levato, R; Peiffer, Q C; de Ruijter, M; Hennink, W E; Vermonden, T; Malda, J

    2016-01-01

    Progress within the field of biofabrication is hindered by a lack of suitable hydrogel formulations. Here, we present a novel approach based on a hybrid printing technique to create cellularized 3D printed constructs. The hybrid bioprinting strategy combines a reinforcing gel for mechanical support with a bioink to provide a cytocompatible environment. In comparison with thermoplastics such as є-polycaprolactone, the hydrogel-based reinforcing gel platform enables printing at cell-friendly temperatures, targets the bioprinting of softer tissues and allows for improved control over degradation kinetics. We prepared amphiphilic macromonomers based on poloxamer that form hydrolysable, covalently cross-linked polymer networks. Dissolved at a concentration of 28.6%w/w in water, it functions as reinforcing gel, while a 5%w/w gelatin-methacryloyl based gel is utilized as bioink. This strategy allows for the creation of complex structures, where the bioink provides a cytocompatible environment for encapsulated cells. Cell viability of equine chondrocytes encapsulated within printed constructs remained largely unaffected by the printing process. The versatility of the system is further demonstrated by the ability to tune the stiffness of printed constructs between 138 and 263 kPa, as well as to tailor the degradation kinetics of the reinforcing gel from several weeks up to more than a year. PMID:27431861

  13. 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.

  14. 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.

  15. 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.

  16. Reinforcement of mono- and bi-layer poly(ethylene glycol) hydrogels with a fibrous collagen scaffold

    PubMed Central

    Kinneberg, K. R. C.; Nelson, A.; Stender, M.; Aziz, A. H.; Mozdzen, L. C.; Harley, B. A. C.; Bryant, S. J.; Ferguson, V. L.

    2015-01-01

    Biomaterial-based tissue engineering strategies hold great promise for osteochondral tissue repair. Yet significant challenges remain in joining highly dissimilar materials to achieve a biomimetic, mechanically robust design for repairing interfaces between soft tissue and bone. This study sought to improve interfacial properties and function in a bilayer, multi-phase hydrogel interpenetrated with a fibrous collagen scaffold. ‘Soft’ 10% (w/w) and ‘stiff’ 30% (w/w) PEGDM was formed into mono- or bilayer hydrogels possessing a sharp diffusional interface. Hydrogels were evaluated as single- (hydrogel only) or multi-phase (hydrogel+fibrous scaffold penetrating throughout the stiff layer and extending >500μm into the soft layer). Including a fibrous scaffold into both soft and stiff single-phase hydrogels significantly increased tangent modulus and toughness and decreased lateral expansion under compressive loading. In multi-phase hydrogels, finite element simulations predict substantially reduced stress and strain gradients across the soft—stiff hydrogel interface. When combining two low moduli constituent material, composites theory poorly predicts the observed, large modulus increases. These results suggest material structure associated with the fibrous scaffold penetrating within the PEG hydrogel as the major contributor to improved properties and function – the hydrogel bore compressive loads and the 3D fibrous scaffold was loaded in tension thus resisting lateral expansion. PMID:26001970

  17. Inorganic-organic hydrogel scaffolds for tissue engineering

    NASA Astrophysics Data System (ADS)

    Bailey, Brennan Margaret

    Analogous to the extracellular matrix (ECM) of natural tissues, properties of a tissue engineering scaffold direct cell behavior and thus regenerated tissue properties. These include both physical properties (e.g. morphology and modulus) and chemical properties (e.g. hydrophobicity, hydration and bioactivity). Notably, recent studies suggest that scaffold properties (e.g. modulus) may be as potent as growth factors in terms of directing stem cell fate. Thus, 3D scaffolds possessing specific properties modified for optimal cell regeneration have the potential to regenerate native-like tissues. Photopolymerizable poly(ethylene glycol) diacrylate (PEG-DA)-based hydrogels are frequently used as scaffolds for tissue engineering. They are ideal for controlled studies of cell-material interactions due to their poor protein adsorption in the absence of adhesive ligands thereby making them "biological blank slates". However, their range of physical and chemical properties is limited. Thus, hydrogel scaffolds which maintain the benefits of PEG-DA but possess a broader set of tunable properties would allow the establishment of predictive relationships between scaffold properties, cell behavior and regenerated tissue properties. Towards this goal, this work describes a series of unique hybrid inorganic-organic hydrogel scaffolds prepared using different solvents and also in the form of continuous gradients. Properties relevant to tissue regeneration were investigated including: swelling, morphology, modulus, degradation rates, bioactivity, cytocompatibility, and protein adhesion. These scaffolds were based on the incorporation of hydrophobic, bioactive and osteoinductive methacrylated star polydimethylsiloxane (PDMSstar-MA) ["inorganic component"] into hydrophilic PEG-DA ["organic component"]. The following parameters were varied: molecular weight (Mn) of PEG-DA (Mn = 3k & 6k g/mol) and PDMSstar-MA (Mn = 1.8k, 7k, 14k), ratio of PDMSstar-MA to PEG-DA (0:100 to 20:80), total

  18. 3D Printing of Human Tissue Mimics via Layer-by-Layer Assembly of Polymer/Hydrogel Biopapers

    NASA Astrophysics Data System (ADS)

    Ringeisen, Bradley

    2015-03-01

    The foundations of tissue engineering were built on two fundamental areas of research: cells and scaffolds. Multipotent cells and their derivatives are traditionally randomly seeded into sophisticated polymer or hydrogel scaffolds, ultimately with the goal of forming a tissue-like material through cell differentiation and cell-material interactions. One problem with this approach is that no matter how complex or biomimetic the scaffold is, the cells are still homogeneously distributed throughout this three dimensional (3D) material. Natural tissue is inherently heterogeneous on both a microscopic and macroscopic level. It also contains different types of cells in close proximity, extracellular matrix, voids, and a complex vascularized network. Recently developed 3D cell and organ printers may be able to enhance traditional tissue engineering experiments by building scaffolds layer-by-layer that are crafted to mimic the microscopic and macroscopic structure of natural tissue or organs. Over the past decade, my laboratory has developed a capillary-free, live cell printer termed biological laser printing, or BioLP. We find that printed cells do not express heat shock protein and retain >99% viability. Printed cells also incur no DNA strand fracture and preserve their ability to differentiate. Recent work has used a layer-by-layer approach, stacking sheets of hybrid polymer/hydrogel biopapers in conjunction with live cell printing to create 3D tissue structures. Our specific work is now focused on the blood-brain-barrier and air-lung interface and will be described during the presentation.

  19. In Vivo Chondrogenesis in 3D Bioprinted Human Cell-laden Hydrogel Constructs

    PubMed Central

    Möller, Thomas; Hägg, Daniel; Brantsing, Camilla; Rotter, Nicole; Apelgren, Peter; Lindahl, Anders; Kölby, Lars; Gatenholm, Paul

    2017-01-01

    Background: The three-dimensional (3D) bioprinting technology allows creation of 3D constructs in a layer-by-layer fashion utilizing biologically relevant materials such as biopolymers and cells. The aim of this study is to investigate the use of 3D bioprinting in a clinically relevant setting to evaluate the potential of this technique for in vivo chondrogenesis. Methods: Thirty-six nude mice (Balb-C, female) received a 5- × 5- × 1-mm piece of bioprinted cell-laden nanofibrillated cellulose/alginate construct in a subcutaneous pocket. Four groups of printed constructs were used: (1) human (male) nasal chondrocytes (hNCs), (2) human (female) bone marrow–derived mesenchymal stem cells (hBMSCs), (3) coculture of hNCs and hBMSCs in a 20/80 ratio, and (4) Cell-free scaffolds (blank). After 14, 30, and 60 days, the scaffolds were harvested for histological, immunohistochemical, and mechanical analysis. Results: The constructs had good mechanical properties and keep their structural integrity after 60 days of implantation. For both the hNC constructs and the cocultured constructs, a gradual increase of glycosaminoglycan production and hNC proliferation was observed. However, the cocultured group showed a more pronounced cell proliferation and enhanced deposition of human collagen II demonstrated by immunohistochemical analysis. Conclusions: In vivo chondrogenesis in a 3D bioprinted human cell-laden hydrogel construct has been demonstrated. The trophic role of the hBMSCs in stimulating hNC proliferation and matrix deposition in the coculture group suggests the potential of 3D bioprinting of human cartilage for future application in reconstructive surgery. PMID:28280669

  20. Anticancer Drug Camptothecin Test in 3D Hydrogel Networks with HeLa cells

    PubMed Central

    Liang, Jun; Susan Sun, Xiuzhi; Yang, Zhilong; Cao, Shuai

    2017-01-01

    Development of a biomimetic 3D culture system for drug screening is necessary to fully understand the in vivo environment. Previously, a self-assembling peptide hydrogel has been reported; the hydrogel exhibited physiological properties superior to a 3D cell culture matrix. In this work, further research using H9e hydrogel with HeLa cells was carried out considering H9e hydrogel’s interaction with camptothecin, a hydrophobic drug. According to AFM images, a PGworks solution triggered H9e hydrogel fiber aggregation and forms a 3D matrix suitable for cell culture. Dynamic rheological studies showed that camptothecin was encapsulated within the hydrogel network concurrently with peptide self-assembly without permanently destroying the hydrogel’s architecture and remodeling ability. Fluorescence measurement indicated negligible interaction between the fluorophore part of camptothecin and the hydrogel, especially at concentration 0.25 and 0.5 wt%. Using a dialysis method, we found that H9e hydrogel could not significantly inhibit the diffusion of camptothecin encapsulated inside the hydrogel matrix. In the cell culture experiment, HeLa cells were simultaneously embedded in the H9e hydrogel with the initialization of hydrogelation. Most importantly, cell viability data after camptothecin treatment showed responses that were drug-dose dependent but unaffected by the H9e hydrogel concentration, indicating that the hydrogel did not inhibit the drug. PMID:28145436

  1. 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.

  2. Living nano-micro fibrous woven fabric/hydrogel composite scaffolds for heart valve engineering.

    PubMed

    Wu, Shaohua; Duan, Bin; Qin, Xiaohong; Butcher, Jonathan T

    2017-03-15

    Regeneration and repair of injured or diseased heart valves remains a clinical challenge. Tissue engineering provides a promising treatment approach to facilitate living heart valve repair and regeneration. Three-dimensional (3D) biomimetic scaffolds that possess heterogeneous and anisotropic features that approximate those of native heart valve tissue are beneficial to the successful in vitro development of tissue engineered heart valves (TEHV). Here we report the development and characterization of a novel composite scaffold consisting of nano- and micro-scale fibrous woven fabrics and 3D hydrogels by using textile techniques combined with bioactive hydrogel formation. Embedded nano-micro fibrous scaffolds within hydrogel enhanced mechanical strength and physical structural anisotropy of the composite scaffold (similar to native aortic valve leaflets) and also reduced its compaction. We determined that the composite scaffolds supported the growth of human aortic valve interstitial cells (HAVIC), balanced the remodeling of heart valve ECM against shrinkage, and maintained better physiological fibroblastic phenotype in both normal and diseased HAVIC over single materials. These fabricated composite scaffolds enable the engineering of a living heart valve graft with improved anisotropic structure and tissue biomechanics important for maintaining valve cell phenotypes.

  3. 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.

  4. Design properties of hydrogel tissue-engineering scaffolds

    PubMed Central

    Zhu, Junmin; Marchant, Roger E

    2011-01-01

    This article summarizes the recent progress in the design and synthesis of hydrogels as tissue-engineering scaffolds. Hydrogels are attractive scaffolding materials owing to their highly swollen network structure, ability to encapsulate cells and bioactive molecules, and efficient mass transfer. Various polymers, including natural, synthetic and natural/synthetic hybrid polymers, have been used to make hydrogels via chemical or physical crosslinking. Recently, bioactive synthetic hydrogels have emerged as promising scaffolds because they can provide molecularly tailored biofunctions and adjustable mechanical properties, as well as an extracellular matrix-like microenvironment for cell growth and tissue formation. This article addresses various strategies that have been explored to design synthetic hydrogels with extracellular matrix-mimetic bioactive properties, such as cell adhesion, proteolytic degradation and growth factor-binding. PMID:22026626

  5. The effect of interface microstructure on interfacial shear strength for osteochondral scaffolds based on biomimetic design and 3D printing.

    PubMed

    Zhang, Weijie; Lian, Qin; Li, Dichen; Wang, Kunzheng; Hao, Dingjun; Bian, Weiguo; Jin, Zhongmin

    2015-01-01

    Interface integration between chondral phase and osseous phase is crucial in engineered osteochondral scaffolds. However, the integration was poorly understood and commonly failed to meet the need of osteochondral scaffolds. In this paper, a biphasic polyethylene glycol (PEG)/β-tricalcium phosphate (β-TCP) scaffold with enhanced interfacial integration was developed. The chondral phase was a PEG hydrogel. The osseous phase was a β-TCP ceramic scaffold. The PEG hydrogel was directly cured on the ceramic interface layer by layer to fabricate osteochondral scaffolds by 3D printing technology. Meanwhile, a series of interface structure were designed with different interface pore area percentages (0/10/20/30/40/50/60%), and interfacial shear test was applied for interface structure optimization (n=6 samples/group). The interfacial shear strength of 30% pore area group was nearly three folds improved compared with that of 0% pore area percentage group, and more than fifty folds improved compared with that of traditional integration (5.91±0.59 kPa). In conclusion, the biomimetic PEG/β-TCP scaffolds with interface structure enhanced integration show promising potential application for osteochondral tissue engineering.

  6. Enhanced bone tissue regeneration using a 3D printed microstructure incorporated with a hybrid nano hydrogel.

    PubMed

    Heo, Dong Nyoung; Castro, Nathan J; Lee, Se-Jun; Noh, Hanaul; Zhu, Wei; Zhang, Lijie Grace

    2017-02-17

    Three-dimensional (3D) functional constructs with biomimetic mechanical and chemical properties are ideal for various regenerative medicine applications. These properties of 3D fabricated constructs mainly depend on the intrinsic characteristics of the materials and fabrication method. In this respect, the current use of hydrogels for musculoskeletal tissue repair is not ideal due to the lack of suitable mechanical properties, as well as the high biomimetic requirement for success. To overcome this limitation, we developed a novel functionalized hydrogel with bioactive gold nanoparticles (GNPs), reinforcing a 3D printed microstructure via fused deposition modeling (FDM) for bone tissue regeneration. We used biodegradable thermoplastic polylactic acid (PLA) as the 3D printed microstructure in combination with photo-curable gelatin hydrogels as the encapsulation matrix for the incorporation of cyclic RGD conjugated GNPs (RGNP), and investigated their mechanical properties. In addition, human adipose-derived stem cells (ADSCs) were encapsulated within the gelatin hydrogel and examined for viability, morphology, and osteogenic differentiation in vitro. The results showed that the stiffness of the composite hydrogel on reinforcing a 3D printed microstructure can be readily modulated to simulate the stiffness of the human mandibular condyle. ADSCs encapsulated in the composite structures remained viable within the hydrogel and showed excellent spreading on the 3D printed PLA microstructure. More importantly, osteogenic differentiation with incorporated RGNPs promoted significantly higher gene expression of osteogenic specific factors. Therefore, reinforced composite hydrogels are suitable for stem cell differentiation control and bone tissue regeneration.

  7. Dextran-based hydrogel formed by thiol-Michael addition reaction for 3D cell encapsulation.

    PubMed

    Liu, Zhen Qi; Wei, Zhao; Zhu, Xv Long; Huang, Guo You; Xu, Feng; Yang, Jian Hai; Osada, Yoshihito; Zrínyi, Miklós; Li, Jian Hui; Chen, Yong Mei

    2015-04-01

    Cell encapsulation in three-dimensional (3D) hydrogels can mimic native cell microenvironment and plays a major role in cell-based transplantation therapies. In this contribution, a novel in situ-forming hydrogel, Dex-l-DTT hydrogel ("l" means "linked-by"), by cross-linking glycidyl methacrylate derivatized dextran (Dex-GMA) and dithiothreitol (DTT) under physiological conditions, has been developed using thiol-Michael addition reaction. The mechanical properties, gelation process and degree of swelling of the hydrogel can be easily adjusted by changing the pH of phosphate buffer saline. The 3D cell encapsulation ability is demonstrated by encapsulating rat bone marrow mesenchymal stem cells (BMSCs) and NIH/3T3 fibroblasts into the in situ-forming hydrogel with maintained high viability. The BMSCs also maintain their differentiation potential after encapsulation. These results demonstrate that the Dex-l-DTT hydrogel holds great potential for biomedical field.

  8. Construction of Modular Hydrogel Sheets for Micropatterned Macro-scaled 3D Cellular Architecture.

    PubMed

    Son, Jaejung; Bae, Chae Yun; Park, Je-Kyun

    2016-01-11

    Hydrogels can be patterned at the micro-scale using microfluidic or micropatterning technologies to provide an in vivo-like three-dimensional (3D) tissue geometry. The resulting 3D hydrogel-based cellular constructs have been introduced as an alternative to animal experiments for advanced biological studies, pharmacological assays and organ transplant applications. Although hydrogel-based particles and fibers can be easily fabricated, it is difficult to manipulate them for tissue reconstruction. In this video, we describe a fabrication method for micropatterned alginate hydrogel sheets, together with their assembly to form a macro-scale 3D cell culture system with a controlled cellular microenvironment. Using a mist form of the calcium gelling agent, thin hydrogel sheets are easily generated with a thickness in the range of 100 - 200 µm, and with precise micropatterns. Cells can then be cultured with the geometric guidance of the hydrogel sheets in freestanding conditions. Furthermore, the hydrogel sheets can be readily manipulated using a micropipette with an end-cut tip, and can be assembled into multi-layered structures by stacking them using a patterned polydimethylsiloxane (PDMS) frame. These modular hydrogel sheets, which can be fabricated using a facile process, have potential applications of in vitro drug assays and biological studies, including functional studies of micro- and macrostructure and tissue reconstruction.

  9. In Situ "Clickable" Zwitterionic Starch-Based Hydrogel for 3D Cell Encapsulation.

    PubMed

    Dong, Dianyu; Li, Junjie; Cui, Man; Wang, Jinmei; Zhou, Yuhang; Luo, Liu; Wei, Yufei; Ye, Lei; Sun, Hong; Yao, Fanglian

    2016-02-01

    Three-dimensional (3D) cell encapsulation in hydrogel provides superb methods to investigate the biochemical cues in directing cellular fate and behaviors outside the organism, the primary step of which is to establish suitable "blank platform" to mimic and simplify native ECM microenvironment. In this study, zwitterionic starch-based "clickable" hydrogels were fabricated via a "copper- and light- free" Michael-type "thiol-ene" addition reaction between acylated-modified sulfobetaine-derived starch (SB-ST-A) and dithiol-functionalized poly(ethylene glycol) (PEG-SH). By incorporating antifouling SB-ST and PEG, the hydrogel system would be excellently protected from nontarget protein adsorption to act as a "blank platform". The hydrogels could rapidly gel under physiological conditions in less than 7 min. Dynamic rheology experiments suggested the stiffness of the hydrogel was close to the native tissues, and the mechanical properties as well as the gelation times and swelling behaviors could be easily tuned by varying the precursor proportions. The protein and cell adhesion assays demonstrated that the hydrogel surface could effectively resist nonspecific protein and cell adhesion. The degradation study in vitro confirmed that the hydrogel was biodegradable. A549 cells encapsulated in the hydrogel maintained high viability (up to 93%) and started to proliferate in number and extend in morphology after 2 days' culture. These results indicated the hydrogel presented here could be a potential candidate as "blank platform" for 3D cell encapsulation and biochemical cues induced cellular behavior investigation in vitro.

  10. 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

  11. 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.

  12. Hydrogel-laden paper scaffold system for origami-based tissue engineering

    PubMed Central

    Kim, Su-Hwan; Lee, Hak Rae; Yu, Seung Jung; Han, Min-Eui; Lee, Doh Young; Kim, Soo Yeon; Ahn, Hee-Jin; Han, Mi-Jung; Lee, Tae-Ik; Kim, Taek-Soo; Kwon, Seong Keun; Im, Sung Gap; Hwang, Nathaniel S.

    2015-01-01

    In this study, we present a method for assembling biofunctionalized paper into a multiform structured scaffold system for reliable tissue regeneration using an origami-based approach. The surface of a paper was conformally modified with a poly(styrene-co-maleic anhydride) layer via initiated chemical vapor deposition followed by the immobilization of poly-l-lysine (PLL) and deposition of Ca2+. This procedure ensures the formation of alginate hydrogel on the paper due to Ca2+ diffusion. Furthermore, strong adhesion of the alginate hydrogel on the paper onto the paper substrate was achieved due to an electrostatic interaction between the alginate and PLL. The developed scaffold system was versatile and allowed area-selective cell seeding. Also, the hydrogel-laden paper could be folded freely into 3D tissue-like structures using a simple origami-based method. The cylindrically constructed paper scaffold system with chondrocytes was applied into a three-ring defect trachea in rabbits. The transplanted engineered tissues replaced the native trachea without stenosis after 4 wks. As for the custom-built scaffold system, the hydrogel-laden paper system will provide a robust and facile method for the formation of tissues mimicking native tissue constructs. PMID:26621717

  13. Hydrogel-laden paper scaffold system for origami-based tissue engineering.

    PubMed

    Kim, Su-Hwan; Lee, Hak Rae; Yu, Seung Jung; Han, Min-Eui; Lee, Doh Young; Kim, Soo Yeon; Ahn, Hee-Jin; Han, Mi-Jung; Lee, Tae-Ik; Kim, Taek-Soo; Kwon, Seong Keun; Im, Sung Gap; Hwang, Nathaniel S

    2015-12-15

    In this study, we present a method for assembling biofunctionalized paper into a multiform structured scaffold system for reliable tissue regeneration using an origami-based approach. The surface of a paper was conformally modified with a poly(styrene-co-maleic anhydride) layer via initiated chemical vapor deposition followed by the immobilization of poly-l-lysine (PLL) and deposition of Ca(2+). This procedure ensures the formation of alginate hydrogel on the paper due to Ca(2+) diffusion. Furthermore, strong adhesion of the alginate hydrogel on the paper onto the paper substrate was achieved due to an electrostatic interaction between the alginate and PLL. The developed scaffold system was versatile and allowed area-selective cell seeding. Also, the hydrogel-laden paper could be folded freely into 3D tissue-like structures using a simple origami-based method. The cylindrically constructed paper scaffold system with chondrocytes was applied into a three-ring defect trachea in rabbits. The transplanted engineered tissues replaced the native trachea without stenosis after 4 wks. As for the custom-built scaffold system, the hydrogel-laden paper system will provide a robust and facile method for the formation of tissues mimicking native tissue constructs.

  14. The cavity-to-cavity migration of leukaemic cells through 3D honey-combed hydrogels with adjustable internal dimension and stiffness.

    PubMed

    da Silva, Joakim; Lautenschläger, Franziska; Sivaniah, Easan; Guck, Jochen R

    2010-03-01

    Whilst rigid, planar surfaces are often used to study cell migration, a physiological scenario requires three-dimensional (3D) scaffolds with tissue-like stiffness. This paper presents a method for fabricating periodic hydrogel scaffolds with a 3D honeycomb-like structure from colloidal crystal templates. The scaffolds, made of hydrogel-walled cavities interconnected by pores, have separately tuneable internal dimensions and adjustable gel stiffness down to that of soft tissues. In conjunction with confocal microscopy, these scaffolds were used to study the importance of cell compliance on invasive potential. Acute promyelocytic leukaemia (APL) cells were differentiated with all-trans retinoic acid (ATRA) and treated with paclitaxel. Their migration ability into the scaffolds' size-restricted pores, enabled by cell softening during ATRA differentiation, was significantly reduced by paclitaxel treatment, which interferes with cell shape recovery. These findings demonstrate the usability of the scaffolds for investigating factors that affect cell migration, and potentially other cell functions, in a realistic 3D tissue model.

  15. Measuring dynamic cell-material interactions and remodeling during 3D human mesenchymal stem cell migration in hydrogels.

    PubMed

    Schultz, Kelly M; Kyburz, Kyle A; Anseth, Kristi S

    2015-07-21

    Biomaterials that mimic aspects of the extracellular matrix by presenting a 3D microenvironment that cells can locally degrade and remodel are finding increased applications as wound-healing matrices, tissue engineering scaffolds, and even substrates for stem cell expansion. In vivo, cells do not simply reside in a static microenvironment, but instead, they dynamically reengineer their surroundings. For example, cells secrete proteases that degrade extracellular components, attach to the matrix through adhesive sites, and can exert traction forces on the local matrix, causing its spatial reorganization. Although biomaterials scaffolds provide initially well-defined microenvironments for 3D culture of cells, less is known about the changes that occur over time, especially local matrix remodeling that can play an integral role in directing cell behavior. Here, we use microrheology as a quantitative tool to characterize dynamic cellular remodeling of peptide-functionalized poly(ethylene glycol) (PEG) hydrogels that degrade in response to cell-secreted matrix metalloproteinases (MMPs). This technique allows measurement of spatial changes in material properties during migration of encapsulated cells and has a sensitivity that identifies regions where cells simply adhere to the matrix, as well as the extent of local cell remodeling of the material through MMP-mediated degradation. Collectively, these microrheological measurements provide insight into microscopic, cellular manipulation of the pericellular region that gives rise to macroscopic tracks created in scaffolds by migrating cells. This quantitative and predictable information should benefit the design of improved biomaterial scaffolds for medically relevant applications.

  16. Bio-inspired detoxification using 3D-printed hydrogel nanocomposites

    NASA Astrophysics Data System (ADS)

    Gou, Maling; Qu, Xin; Zhu, Wei; Xiang, Mingli; Yang, Jun; Zhang, Kang; Wei, Yuquan; Chen, Shaochen

    2014-05-01

    Rationally designed nanoparticles that can bind toxins show great promise for detoxification. However, the conventional intravenous administration of nanoparticles for detoxification often leads to nanoparticle accumulation in the liver, posing a risk of secondary poisoning especially in liver-failure patients. Here we present a liver-inspired three-dimensional (3D) detoxification device. This device is created by 3D printing of designer hydrogels with functional polydiacetylene nanoparticles installed in the hydrogel matrix. The nanoparticles can attract, capture and sense toxins, while the 3D matrix with a modified liver lobule microstructure allows toxins to be trapped efficiently. Our results show that the toxin solution completely loses its virulence after treatment using this biomimetic detoxification device. This work provides a proof-of-concept of detoxification by a 3D-printed biomimetic nanocomposite construct in hydrogel, and could lead to the development of alternative detoxification platforms.

  17. Bio-inspired detoxification using 3D-printed hydrogel nanocomposites.

    PubMed

    Gou, Maling; Qu, Xin; Zhu, Wei; Xiang, Mingli; Yang, Jun; Zhang, Kang; Wei, Yuquan; Chen, Shaochen

    2014-05-08

    Rationally designed nanoparticles that can bind toxins show great promise for detoxification. However, the conventional intravenous administration of nanoparticles for detoxification often leads to nanoparticle accumulation in the liver, posing a risk of secondary poisoning especially in liver-failure patients. Here we present a liver-inspired three-dimensional (3D) detoxification device. This device is created by 3D printing of designer hydrogels with functional polydiacetylene nanoparticles installed in the hydrogel matrix. The nanoparticles can attract, capture and sense toxins, while the 3D matrix with a modified liver lobule microstructure allows toxins to be trapped efficiently. Our results show that the toxin solution completely loses its virulence after treatment using this biomimetic detoxification device. This work provides a proof-of-concept of detoxification by a 3D-printed biomimetic nanocomposite construct in hydrogel, and could lead to the development of alternative detoxification platforms.

  18. Bio-inspired detoxification using 3D-printed hydrogel nanocomposites

    PubMed Central

    Gou, Maling; Qu, Xin; Zhu, Wei; Xiang, Mingli; Yang, Jun; Zhang, Kang; Wei, Yuquan; Chen, Shaochen

    2014-01-01

    Rationally designed nanoparticles that can bind toxins show great promise for detoxification. However, the conventional intravenous administration of nanoparticles for detoxification often leads to nanoparticle accumulation in the liver, posing a risk of secondary poisoning especially in liver-failure patients. Here we present a liver-inspired three-dimensional (3D) detoxification device. This device is created by 3D printing of designer hydrogels with functional polydiacetylene nanoparticles installed in the hydrogel matrix. The nanoparticles can attract, capture and sense toxins, while the 3D matrix with a modified liver lobule microstructure allows toxins to be trapped efficiently. Our results show that the toxin solution completely loses its virulence after treatment using this biomimetic detoxification device. This work provides a proof-of-concept of detoxification by a 3D-printed biomimetic nanocomposite construct in hydrogel, and could lead to the development of alternative detoxification platforms. PMID:24805923

  19. A thermo-responsive and photo-polymerizable chondroitin sulfate-based hydrogel for 3D printing applications.

    PubMed

    Abbadessa, A; Blokzijl, M M; Mouser, V H M; Marica, P; Malda, J; Hennink, W E; Vermonden, T

    2016-09-20

    The aim of this study was to design a hydrogel system based on methacrylated chondroitin sulfate (CSMA) and a thermo-sensitive poly(N-(2-hydroxypropyl) methacrylamide-mono/dilactate)-polyethylene glycol triblock copolymer (M15P10) as a suitable material for additive manufacturing of scaffolds. CSMA was synthesized by reaction of chondroitin sulfate with glycidyl methacrylate (GMA) in dimethylsulfoxide at 50°C and its degree of methacrylation was tunable up to 48.5%, by changing reaction time and GMA feed. Unlike polymer solutions composed of CSMA alone (20% w/w), mixtures based on 2% w/w of CSMA and 18% of M15P10 showed strain-softening, thermo-sensitive and shear-thinning properties more pronounced than those found for polymer solutions based on M15P10 alone. Additionally, they displayed a yield stress of 19.2±7.0Pa. The 3D printing of this hydrogel resulted in the generation of constructs with tailorable porosity and good handling properties. Finally, embedded chondrogenic cells remained viable and proliferating over a culture period of 6days. The hydrogel described herein represents a promising biomaterial for cartilage 3D printing applications.

  20. Bio-inspired mineralization of hydroxyapatite in 3D silk fibroin hydrogel for bone tissue engineering.

    PubMed

    Jin, Yashi; Kundu, Banani; Cai, Yurong; Kundu, Subhas C; Yao, Juming

    2015-10-01

    To fabricate hard tissue implants with bone-like structure using a biomimetic mineralization method is drawing much more attentions in bone tissue engineering. The present work focuses in designing 3D silk fibroin hydrogel to modulate the nucleation and growth of hydroxyapatite crystals via a simple ion diffusion method. The study indicates that Ca(2+) incorporation within the hydrogel provides the nucleation sites for hydroxyapatite crystals and subsequently regulates their oriented growth. The mineralization process is regulated in a Ca(2+) concentration- and minerlization time-dependent way. Further, the compressive strength of the mineralized hydrogels is directly proportional with the mineral content in hydrogel. The orchestrated organic/inorganic composite supports well the viability and proliferation of human osteoblast cells; improved cyto-compatibility with increased mineral content. Together, the present investigation reports a simple and biomimetic process to fabricate 3D bone-like biomaterial with desired efficacy to repair bone defects.

  1. Composite hydrogel scaffolds with controlled pore opening via biodegradable hydrogel porogen degradation.

    PubMed

    Hawkins, Ashley M; Milbrandt, Todd A; Puleo, David A; Hilt, J Zach

    2014-02-01

    Poly(β-amino ester) (PBAE) biodegradable hydrogel systems have garnered much attention in recent years due to their appealing properties for biomedical applications. These hydrogel systems exhibit properties similar to natural soft tissue, degrade in aqueous environments, and have easily tunable properties that have been well studied and understood. In most cases, tissue engineering scaffolds must possess a three-dimensional interconnected porous network for tissue ingrowth and construct vascularization. Here, PBAE properties were explored and systems were selected to serve as both the pore-forming agent and the outer matrix of a scaffold that exhibits controlled pore opening upon degradation. To our knowledge, this is the first demonstration of a biodegradable hydrogel porogen system entrapped in a degradable hydrogel outer matrix. Scaffolds were prepared, and the degradation, compressive moduli, and porosity were analyzed. An added advantage of a degradable porogen is the potential for controlled drug release, and a model protein was released from the porogen particles to demonstrate this application. Finally, pluripotent cells seeded onto predegraded scaffolds were viable during the first 24 h of exposure, and furthermore, cell tracking confirmed the presence of cells within the pores of the scaffold. Overall, these present studies demonstrate the possibility of using these biodegradable hydrogel porogen-matrix systems as tissue engineering scaffolding materials.

  2. Constructing 3D heterogeneous hydrogels from electrically manipulated prepolymer droplets and crosslinked microgels

    PubMed Central

    Chiang, Min-Yu; Hsu, Yao-Wen; Hsieh, Hsin-Yi; Chen, San-Yuan; Fan, Shih-Kang

    2016-01-01

    Formation of multifunctional, heterogeneous, and encoded hydrogel building blocks, or microgels, by crosslinking and assembly of microgels are two essential steps in establishing hierarchical, complicated, and three-dimensional (3D) hydrogel architectures that recapitulate natural and biological structures or originate new materials by design. However, for the variety of the hydrogel materials crosslinked differently and for the varied scales of microgels and architectures, the formation and assembly processes are usually performed separately, which increases the manufacturing complexity of designed hydrogel materials. We show the construction of hydrogel architectures through programmable formation and assembly on an electromicrofluidic platform, adopting two reciprocal electric manipulations (electrowetting and dielectrophoresis) to manipulate varied objects (i) in multiple phases, including prepolymer liquid droplets and crosslinked microgels, (ii) on a wide range of scales from micrometer functional particles or cells to millimeter-assembled hydrogel architectures, and (iii) with diverse properties, such as conductive and dielectric droplets that are photocrosslinkable, chemically crosslinkable, or thermally crosslinkable. Prepolymer droplets, particles, and dissolved molecules are electrically addressable to adjust the properties of the microgel building blocks in liquid phase that subsequently undergo crosslinking and assembly in a flexible sequence to accomplish heterogeneous and seamless hydrogel architectures. We expect the electromicrofluidic platform to become a general technique to obtain 3D complex architectures. PMID:27819046

  3. 3D bioprinting of GelMA scaffolds triggers mineral deposition by primary human osteoblasts.

    PubMed

    McBeth, Christine; Lauer, Jasmin; Ottersbach, Michael; Campbell, Jennifer; Sharon, Andre; Sauer-Budge, Alexis F

    2017-01-10

    Due to its relatively low level of antigenicity and high durability, titanium has successfully been used as the major material for biological implants. However, because the typical interface between titanium and tissue precludes adequate transmission of load into the surrounding bone, over time, load-bearing implants tend to loosen and revision surgeries are required. Osseointegration of titanium implants requires presentation of both biological and mechanical cues that promote attachment of and trigger mineral deposition by osteoblasts. While many factors contribute to differentiation, the relative importance of the various cues is unclear. To substantially improve osseointegration of titanium implants, we generated a gelatin methacryloyl (GelMA) scaffold, using an extrusion-based 3D bioprinter, which can be directly printed on and grafted to the titanium implant surface. We demonstrate that this scaffold is able to trigger mineral deposition of both MG63 osteoblasts and primary normal human osteoblasts in the absence of any exogenous osteogenic factors. Films of the same formulation failed to promote mineral deposition suggesting that the three dimensional scaffold was able to tip the balance in favor of differentiation despite other potentially unfavorable differentiation cues of the material. We further show that these GelMA lattices can be directly grafted to titanium alloy and are secure in vitro over a period of seven weeks. When grafted within a groove system, the GelMA hydrogel is protected from shearing forces in a marrow implantation model. This prepares the way for osteogenic coatings to be directly manufactured on the implant surface and packaged for surgery.

  4. 3D bioprinting of GelMA scaffolds triggers mineral deposition by primary human osteoblasts.

    PubMed

    McBeth, Christine; Lauer, Jasmin; Ottersbach, Michael; Campbell, Jennifer; Sharon, Andre; Sauer-Budge, Alexis

    2016-12-14

    Due to its relatively low level of antigenicity and high durability, titanium has successfully been used as the major material for biological implants. However, because the typical interface between titanium and tissue precludes adequate transmission of load into the surrounding bone, over time, load-bearing implants tend to loosen and revision surgeries are required. Osseointegration of titanium implants requires presentation of both biological and mechanical cues that promote attachment of and trigger mineral deposition by osteoblasts. While many factors contribute to differentiation, the relative importance of the various cues is unclear. To substantially improve osseointegration of titanium implants, we generated a gelatin methacryloyl (GelMA) scaffold, using an extrusion-based 3D bioprinter, which can be directly printed on and grafted to the titanium implant surface. We demonstrate that this scaffold is able to trigger mineral deposition of both MG63 osteoblasts and normal human primary osteoblasts in the absence of any exogenous osteogenic factors. Films of the same formulation failed to promote mineral deposition suggesting that the three dimensional scaffold was able to tip the balance in favor of differentiation despite other potentially unfavorable differentiation cues of the material. We further show that these GelMA lattices can be directly grafted to titanium alloy and are secure in vitro over a period of seven weeks. When grafted within a groove system, the GelMA hydrogel is protected from shearing forces in a marrow implantation model. This prepares the way for osteogenic coatings to be directly manufactured on the implant surface and packaged for surgery.

  5. Development of Hydrogels and Biomimetic Regulators as Tissue Engineering Scaffolds

    PubMed Central

    Shi, Junbin; Xing, Malcolm M. Q.; Zhong, Wen

    2012-01-01

    This paper reviews major research and development issues relating to hydrogels as scaffolds for tissue engineering, the article starts with a brief introduction of tissue engineering and hydrogels as extracellular matrix mimics, followed by a description of the various types of hydrogels and preparation methods, before a discussion of the physical and chemical properties that are important to their application. There follows a short comment on the trends of future research and development. Throughout the discussion there is an emphasis on the genetic understanding of bone tissue engineering application. PMID:24957963

  6. Cellular Microcultures: Programming Mechanical and Physicochemical Properties of 3D Hydrogel Cellular Microcultures via Direct Ink Writing (Adv. Healthcare Mater. 9/2016).

    PubMed

    McCracken, Joselle M; Badea, Adina; Kandel, Mikhail E; Gladman, A Sydney; Wetzel, David J; Popescu, Gabriel; Lewis, Jennifer A; Nuzzo, Ralph G

    2016-05-01

    R. Nuzzo and co-workers show on page 1025 how compositional differences in hydrogels are used to tune their cellular compliance by controlling their polymer mesh properties and subsequent uptake of the protein poly-l-lysine (green spheres in circled inset). The cover image shows pyramid micro-scaffolds prepared using direct ink writing (DIW) that differentially direct fibroblast and preosteoblast growth in 3D, depending on cell motility and surface treatment.

  7. 3D patterned stem cell differentiation using thermo-responsive methylcellulose hydrogel molds.

    PubMed

    Lee, Wonjae; Park, Jon

    2016-07-06

    Tissue-specific patterned stem cell differentiation serves as the basis for the development, remodeling, and regeneration of the multicellular structure of the native tissues. We herein proposed a cytocompatible 3D casting process to recapitulate this patterned stem cell differentiation for reconstructing multicellular tissues in vitro. We first reconstituted the 2D culture conditions for stem cell fate control within 3D hydrogel by incorporating the sets of the diffusible signal molecules delivered through drug-releasing microparticles. Then, utilizing thermo-responsivity of methylcellulose (MC), we developed a cytocompatible casting process to mold these hydrogels into specific 3D configurations, generating the targeted spatial gradients of diffusible signal molecules. The liquid phase of the MC solution was viscous enough to adopt the shapes of 3D impression patterns, while the gelated MC served as a reliable mold for patterning the hydrogel prepolymers. When these patterned hydrogels were integrated together, the stem cells in each hydrogel distinctly differentiated toward individually defined fates, resulting in the formation of the multicellular tissue structure bearing the very structural integrity and characteristics as seen in vascularized bones and osteochondral tissues.

  8. Dual-stage growth factor release within 3D protein-engineered hydrogel niches promotes adipogenesis

    PubMed Central

    Greenwood-Goodwin, Midori; Teasley, Eric S.; Heilshorn, Sarah C.

    2014-01-01

    Engineered biomimetic microenvironments from hydrogels are an emerging strategy to achieve lineage-specific differentiation in vitro. In addition to recapitulating critical matrix cues found in the native three-dimensional (3D) niche, the hydrogel can also be designed to deliver soluble factors that are present within the native inductive microenvironment. We demonstrate a versatile materials approach for the dual-stage delivery of multiple soluble factors within a 3D hydrogel to induce adipogenesis. We use a Mixing-Induced Two-Component Hydrogel (MITCH) embedded with alginate microgels to deliver two pro-adipogenic soluble factors, fibroblast growth factor 1 (FGF-1) and bone morphogenetic protein 4 (BMP-4) with two distinct delivery profiles. We show that dual-stage delivery of FGF-1 and BMP-4 to human adipose-derived stromal cells (hADSCs) significantly increases lipid accumulation compared with the simultaneous delivery of both growth factors together. Furthermore, dual-stage growth factor delivery within a 3D hydrogel resulted in substantially more lipid accumulation compared to identical delivery profiles in 2D cultures. Gene expression analysis shows upregulation of key adipogenic markers indicative of brown-like adipocytes. These data suggest that dual-stage release of FGF-1 and BMP-4 within 3D microenvironments can promote the in vitro development of mature adipocytes. PMID:25309741

  9. 3D patterned stem cell differentiation using thermo-responsive methylcellulose hydrogel molds

    PubMed Central

    Lee, Wonjae; Park, Jon

    2016-01-01

    Tissue-specific patterned stem cell differentiation serves as the basis for the development, remodeling, and regeneration of the multicellular structure of the native tissues. We herein proposed a cytocompatible 3D casting process to recapitulate this patterned stem cell differentiation for reconstructing multicellular tissues in vitro. We first reconstituted the 2D culture conditions for stem cell fate control within 3D hydrogel by incorporating the sets of the diffusible signal molecules delivered through drug-releasing microparticles. Then, utilizing thermo-responsivity of methylcellulose (MC), we developed a cytocompatible casting process to mold these hydrogels into specific 3D configurations, generating the targeted spatial gradients of diffusible signal molecules. The liquid phase of the MC solution was viscous enough to adopt the shapes of 3D impression patterns, while the gelated MC served as a reliable mold for patterning the hydrogel prepolymers. When these patterned hydrogels were integrated together, the stem cells in each hydrogel distinctly differentiated toward individually defined fates, resulting in the formation of the multicellular tissue structure bearing the very structural integrity and characteristics as seen in vascularized bones and osteochondral tissues. PMID:27381562

  10. 3D Bioprinting of Heterogeneous Aortic Valve Conduits with Alginate/Gelatin Hydrogels

    PubMed Central

    Duan, Bin; Hockaday, Laura A.; Kang, Kevin H.; Butcher, Jonathan T.

    2013-01-01

    Heart valve disease is a serious and growing public health problem for which prosthetic replacement is most commonly indicated. Current prosthetic devices are inadequate for younger adults and growing children. Tissue engineered living aortic valve conduits have potential for remodeling, regeneration, and growth, but fabricating natural anatomical complexity with cellular heterogeneity remain challenging. In the current study, we implement 3D bioprinting to fabricate living alginate/gelatin hydrogel valve conduits with anatomical architecture and direct incorporation of dual cell types in a regionally constrained manner. Encapsulated aortic root sinus smooth muscle cells (SMC) and aortic valve leaflet interstitial cells (VIC) were viable within alginate/gelatin hydrogel discs over 7 days in culture. Acellular 3D printed hydrogels exhibited reduced modulus, ultimate strength, and peak strain reducing slightly over 7-day culture, while the tensile biomechanics of cell-laden hydrogels were maintained. Aortic valve conduits were successfully bioprinted with direct encapsulation of SMC in the valve root and VIC in the leaflets. Both cell types were viable (81.4±3.4% for SMC and 83.2±4.0% for VIC) within 3D printed tissues. Encapsulated SMC expressed elevated alpha-smooth muscle actin when printed in stiff matrix, while VIC expressed elevated vimentin in soft matrix. These results demonstrate that anatomically complex, heterogeneously encapsulated aortic valve hydrogel conduits can be fabricated with 3D bioprinting. PMID:23015540

  11. 3D patterned stem cell differentiation using thermo-responsive methylcellulose hydrogel molds

    NASA Astrophysics Data System (ADS)

    Lee, Wonjae; Park, Jon

    2016-07-01

    Tissue-specific patterned stem cell differentiation serves as the basis for the development, remodeling, and regeneration of the multicellular structure of the native tissues. We herein proposed a cytocompatible 3D casting process to recapitulate this patterned stem cell differentiation for reconstructing multicellular tissues in vitro. We first reconstituted the 2D culture conditions for stem cell fate control within 3D hydrogel by incorporating the sets of the diffusible signal molecules delivered through drug-releasing microparticles. Then, utilizing thermo-responsivity of methylcellulose (MC), we developed a cytocompatible casting process to mold these hydrogels into specific 3D configurations, generating the targeted spatial gradients of diffusible signal molecules. The liquid phase of the MC solution was viscous enough to adopt the shapes of 3D impression patterns, while the gelated MC served as a reliable mold for patterning the hydrogel prepolymers. When these patterned hydrogels were integrated together, the stem cells in each hydrogel distinctly differentiated toward individually defined fates, resulting in the formation of the multicellular tissue structure bearing the very structural integrity and characteristics as seen in vascularized bones and osteochondral tissues.

  12. Characterisation of the surface structure of 3D printed scaffolds for cell infiltration and surgical suturing.

    PubMed

    Ruiz-Cantu, Laura; Gleadall, Andrew; Faris, Callum; Segal, Joel; Shakesheff, Kevin; Yang, Jing

    2016-03-01

    3D printing is of great interest for tissue engineering scaffolds due to the ability to form complex geometries and control internal structures, including porosity and pore size. The porous structure of scaffolds plays an important role in cell ingrowth and nutrition infusion. Although the internal porosity and pore size of 3D printed scaffolds have been frequently studied, the surface porosity and pore size, which are critical for cell infiltration and mass transport, have not been investigated. The surface geometry can differ considerably from the internal scaffold structure depending on the 3D printing process. It is vital to be able to control the surface geometry of scaffolds as well as the internal structure to fabricate optimal architectures. This work presents a method to control the surface porosity and pore size of 3D printed scaffolds. Six scaffold designs have been printed with surface porosities ranging from 3% to 21%. We have characterised the overall scaffold porosity and surface porosity using optical microscopy and microCT. It has been found that surface porosity has a significant impact on cell infiltration and proliferation. In addition, the porosity of the surface has been found to have an effect on mechanical properties and on the forces required to penetrate the scaffold with a surgical suturing needle. To the authors' knowledge, this study is the first to investigate the surface geometry of extrusion-based 3D printed scaffolds and demonstrates the importance of surface geometry in cell infiltration and clinical manipulation.

  13. A phase diagram for microfabrication of geometrically controlled hydrogel scaffolds.

    PubMed

    Tirella, A; Orsini, A; Vozzi, G; Ahluwalia, A

    2009-12-01

    Hydrogels are considered as excellent candidates for tissue substitutes by virtue of their high water content and biphasic nature. However, the fact that they are soft, wet and floppy renders them difficult to process and use as custom-designed scaffolds. To address this problem alginate hydrogels were modeled and characterized by measuring stress-strain and creep behavior as well as viscosity as a function of sodium alginate concentration, cross-linking time and calcium ion concentration. The gels were then microfabricated into scaffolds using the pressure-assisted microsyringe. The mechanical and viscous characteristics were used to generate a processing window in the form of a phase diagram which describes the fidelity of the scaffolds as a function of the material and machine parameters. The approach can be applied to a variety of microfabrication methods and biomaterials in order to design well-controlled custom scaffolds.

  14. 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.

  15. A Genetically Modified Protein-Based Hydrogel for 3D Culture of AD293 Cells

    PubMed Central

    Du, Xiao; Wang, Jingyu; Diao, Wentao; Wang, Ling; Long, Jiafu; Zhou, Hao

    2014-01-01

    Hydrogels have strong application prospects for drug delivery, tissue engineering and cell therapy because of their excellent biocompatibility and abundant availability as scaffolds for drugs and cells. In this study, we created hybrid hydrogels based on a genetically modified tax interactive protein-1 (TIP1) by introducing two or four cysteine residues in the primary structure of TIP1. The introduced cysteine residues were crosslinked with a four-armed poly (ethylene glycol) having their arm ends capped with maleimide residues (4-armed-PEG-Mal) to form hydrogels. In one form of the genetically modification, we incorporated a peptide sequence ‘GRGDSP’ to introduce bioactivity to the protein, and the resultant hydrogel could provide an excellent environment for a three dimensional cell culture of AD293 cells. The AD293 cells continued to divide and displayed a polyhedron or spindle-shape during the 3-day culture period. Besides, AD293 cells could be easily separated from the cell-gel constructs for future large-scale culture after being cultured for 3 days and treating hydrogel with trypsinase. This work significantly expands the toolbox of recombinant proteins for hydrogel formation, and we believe that our hydrogel will be of considerable interest to those working in cell therapy and controlled drug delivery. PMID:25233088

  16. User-friendly 3D bioassays with cell-containing hydrogel modules: narrowing the gap between microfluidic bioassays and clinical end-users' needs.

    PubMed

    Lee, Do-Hyun; Bae, Chae Yun; Kwon, Seyong; Park, Je-Kyun

    2015-06-07

    Cell-containing hydrogel modules as cell-hydrogel microunits for creating a physiologically relevant 3D in vivo-like microenvironment with multiple cell types and unique extracellular matrix (ECM) compositions facilitate long-term cell maintenance and bioassays. To date, there have been many important advances in microfluidic bioassays, which incorporate hydrogel scaffolds into surface-accessible microchambers, driven by the strong demand for the application of spatiotemporally defined biochemical stimuli to construct in vivo-like conditions and perform real-time imaging of cell-matrix interactions. In keeping with the trend of fostering collaborations among biologists, clinicians, and microfluidic engineers, it is essential to create a simpler approach for coupling cell-containing hydrogel modules and an automated bioassay platform in a user-friendly format. In this article, we review recent progress in hydrogel-incorporated microfluidics for long-term cell maintenance and discuss some of the simpler and user-friendly 3D bioassay techniques combined with cell-containing hydrogel modules that can be applied to mutually beneficial collaborations with non-engineers. We anticipate that this modular and user-friendly format interfaced with existing laboratory infrastructure will help address several clinical questions in ways that extend well beyond the current 2D cell-culture systems.

  17. 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.

  18. Superabsorbent 3D Scaffold Based on Electrospun Nanofibers for Cartilage Tissue Engineering.

    PubMed

    Chen, Weiming; Chen, Shuai; Morsi, Yosry; El-Hamshary, Hany; El-Newhy, Mohamed; Fan, Cunyi; Mo, Xiumei

    2016-09-21

    Electrospun nanofibers have been used for various biomedical applications. However, electrospinning commonly produces two-dimensional (2D) membranes, which limits the application of nanofibers for the 3D tissue engineering scaffold. In the present study, a porous 3D scaffold (3DS-1) based on electrospun gelatin/PLA nanofibers has been prepared for cartilage tissue regeneration. To further improve the repairing effect of cartilage, a modified scaffold (3DS-2) cross-linked with hyaluronic acid (HA) was also successfully fabricated. The nanofibrous structure, water absorption, and compressive mechanical properties of 3D scaffold were studied. Chondrocytes were cultured on 3D scaffold, and their viability and morphology were examined. 3D scaffolds were also subjected to an in vivo cartilage regeneration study on rabbits using an articular cartilage injury model. The results indicated that 3DS-1 and 3DS-2 exhibited superabsorbent property and excellent cytocompatibility. Both these scaffolds present elastic property in the wet state. An in vivo study showed that 3DS-2 could enhance the repair of cartilage. The present 3D nanofibrous scaffold (3DS-2) would be promising for cartilage tissue engineering application.

  19. Ultrafine fibrous gelatin scaffolds with deep cell infiltration mimicking 3D ECMs for soft tissue repair.

    PubMed

    Jiang, Qiuran; Xu, Helan; Cai, Shaobo; Yang, Yiqi

    2014-07-01

    In this research, ultrafine fibrous scaffolds with deep cell infiltration and sufficient water stability have been developed from gelatin, aiming to mimic the extracellular matrices (ECMs) as three dimensional (3D) stromas for soft tissue repair. The ultrafine fibrous scaffolds produced from the current technologies of electrospinning and phase separation are either lack of 3D oriented fibrous structure or too compact to be penetrated by cells. Whilst electrospun scaffolds are able to emulate two dimensional (2D) ECMs, they cannot mimic the 3D ECM stroma. In this work, ultralow concentration phase separation (ULCPS) has been developed to fabricate gelatin scaffolds with 3D randomly oriented ultrafine fibers and loose structures. Besides, a non-toxic citric acid crosslinking system has been established for the ULCPS method. This system could endow the scaffolds with sufficient water stability, while maintain the fibrous structures of scaffolds. Comparing with electrospun scaffolds, the ULCPS scaffolds showed improved cytocompatibility and more importantly, cell infiltration. This research has proved the possibility of using gelatin ULCPS scaffolds as the substitutes of 3D ECMs.

  20. Fabrication and Mechanical Characterization of Hydrogel Infused Network Silk Scaffolds

    PubMed Central

    Kundanati, Lakshminath; Singh, Saket K.; Mandal, Biman B.; Murthy, Tejas G.; Gundiah, Namrata; Pugno, Nicola M.

    2016-01-01

    Development and characterization of porous scaffolds for tissue engineering and regenerative medicine is of great importance. In recent times, silk scaffolds were developed and successfully tested in tissue engineering and drug release applications. We developed a novel composite scaffold by mechanical infusion of silk hydrogel matrix into a highly porous network silk scaffold. The mechanical behaviour of these scaffolds was thoroughly examined for their possible use in load bearing applications. Firstly, unconfined compression experiments show that the denser composite scaffolds displayed significant enhancement in the elastic modulus as compared to either of the components. This effect was examined and further explained with the help of foam mechanics principles. Secondly, results from confined compression experiments that resemble loading of cartilage in confinement, showed nonlinear material responses for all scaffolds. Finally, the confined creep experiments were performed to calculate the hydraulic permeability of the scaffolds using soil mechanics principles. Our results show that composite scaffolds with some modifications can be a potential candidate for use of cartilage like applications. We hope such approaches help in developing novel scaffolds for tissue engineering by providing an understanding of the mechanics and can further be used to develop graded scaffolds by targeted infusion in specific regions. PMID:27681725

  1. Microfabrication of complex porous tissue engineering scaffolds using 3D projection stereolithography

    PubMed Central

    Gauvin, Robert; Chen, Ying-Chieh; Lee, Jin Woo; Soman, Pranav; Zorlutuna, Pinar; Nichol, Jason W.; Bae, Hojae; Chen, Shaochen; Khademhosseini, Ali

    2013-01-01

    The success of tissue engineering will rely on the ability to generate complex, cell seeded three-dimensional (3D) structures. Therefore, methods that can be used to precisely engineer the architecture and topography of scaffolding materials will represent a critical aspect of functional tissue engineering. Previous approaches for 3D scaffold fabrication based on top-down and process driven methods are often not adequate to produce complex structures due to the lack of control on scaffold architecture, porosity, and cellular interactions. The proposed projection stereolithography (PSL) platform can be used to design intricate 3D tissue scaffolds that can be engineered to mimic the microarchitecture of tissues, based on computer aided design (CAD). The PSL system was developed, programmed and optimized to fabricate 3D scaffolds using gelatin methacrylate (GelMA). Variation of the structure and prepolymer concentration enabled tailoring the mechanical properties of the scaffolds. A dynamic cell seeding method was utilized to improve the coverage of the scaffold throughout its thickness. The results demonstrated that the interconnectivity of pores allowed for uniform human umbilical vein endothelial cells (HUVECs) distribution and proliferation in the scaffolds, leading to high cell density and confluency at the end of the culture period. Moreover, immunohistochemistry results showed that cells seeded on the scaffold maintained their endothelial phenotype, demonstrating the biological functionality of the microfabricated GelMA scaffolds. PMID:22365811

  2. Engineering Cellular Microenvironments with Photo- and Enzymatically Responsive Hydrogels: Toward Biomimetic 3D Cell Culture Models.

    PubMed

    Tam, Roger Y; Smith, Laura J; Shoichet, Molly S

    2017-03-27

    Conventional cell culture techniques using 2D polystyrene or glass have provided great insight into key biochemical mechanisms responsible for cellular events such as cell proliferation, differentiation, and cell-cell interactions. However, the physical and chemical properties of 2D culture in vitro are dramatically different than those found in the native cellular microenvironment in vivo. Cells grown on 2D substrates differ significantly from those grown in vivo, and this explains, in part, why many promising drug candidates discovered through in vitro drug screening assays fail when they are translated to in vivo animal or human models. To overcome this obstacle, 3D cell culture using biomimetic hydrogels has emerged as an alternative strategy to recapitulate native cell growth in vitro. Hydrogels, which are water-swollen polymers, can be synthetic or naturally derived. Many methods have been developed to control the physical and chemical properties of the hydrogels to match those found in specific tissues. Compared to 2D culture, cells cultured in 3D gels with the appropriate physicochemical cues can behave more like they naturally do in vivo. While conventional hydrogels involve modifications to the bulk material to mimic the static aspects of the cellular microenvironment, recent progress has focused on using more dynamic hydrogels, the chemical and physical properties of which can be altered with external stimuli to better mimic the dynamics of the native cellular microenvironment found in vivo. In this Account, we describe our progress in designing stimuli-responsive, optically transparent hydrogels that can be used as biomimetic extracellular matrices (ECMs) to study cell differentiation and migration in the context of modeling the nervous system and cancer. Specifically, we developed photosensitive agarose and hyaluronic acid hydrogels that are activated by single or two-photon irradiation for biomolecule immobilization at specific volumes within the 3D

  3. Development of hybrid scaffolds using ceramic and hydrogel for articular cartilage tissue regeneration.

    PubMed

    Seol, Young-Joon; Park, Ju Young; Jeong, Wonju; Kim, Tae-Ho; Kim, Shin-Yoon; Cho, Dong-Woo

    2015-04-01

    The regeneration of articular cartilage consisting of hyaline cartilage and hydrogel scaffolds has been generally used in tissue engineering. However, success in in vivo studies has been rarely reported. The hydrogel scaffolds implanted into articular cartilage defects are mechanically unstable and it is difficult for them to integrate with the surrounding native cartilage tissue. Therefore, it is needed to regenerate cartilage and bone tissue simultaneously. We developed hybrid scaffolds with hydrogel scaffolds for cartilage tissue and with ceramic scaffolds for bone tissue. For in vivo study, hybrid scaffolds were press-fitted into osteochondral tissue defects in a rabbit knee joints and the cartilage tissue regeneration in blank, hydrogel scaffolds, and hybrid scaffolds was compared. In 12th week after implantation, the histological and immunohistochemical analyses were conducted to evaluate the cartilage tissue regeneration. In the blank and hydrogel scaffold groups, the defects were filled with fibrous tissues and the implanted hydrogel scaffolds could not maintain their initial position; in the hybrid scaffold group, newly generated cartilage tissues were morphologically similar to native cartilage tissues and were smoothly connected to the surrounding native tissues. This study demonstrates hybrid scaffolds containing hydrogel and ceramic scaffolds can provide mechanical stability to hydrogel scaffolds and enhance cartilage tissue regeneration at the defect site.

  4. A multimaterial bioink method for 3D printing tunable, cell-compatible hydrogels.

    PubMed

    Rutz, Alexandra L; Hyland, Kelly E; Jakus, Adam E; Burghardt, Wesley R; Shah, Ramille N

    2015-03-04

    A multimaterial bio-ink method using polyethylene glycol crosslinking is presented for expanding the biomaterial palette required for 3D bioprinting of more mimetic and customizable tissue and organ constructs. Lightly crosslinked, soft hydrogels are produced from precursor solutions of various materials and 3D printed. Rheological and biological characterizations are presented, and the promise of this new bio-ink synthesis strategy is discussed.

  5. Photoactive Self-Shaping Hydrogels as Noncontact 3D Macro/Microscopic Photoprinting Platforms.

    PubMed

    Liao, Yue; An, Ning; Wang, Ning; Zhang, Yinyu; Song, Junfei; Zhou, Jinxiong; Liu, Wenguang

    2015-12-01

    A photocleavable terpolymer hydrogel cross-linked with o-nitrobenzyl derivative cross-linker is shown to be capable of self-shaping without losing its physical integrity and robustness due to spontaneous asymmetric swelling of network caused by UV-light-induced gradient cleavage of chemical cross-linkages. The continuum model and finite element method are used to elucidate the curling mechanism underlying. Remarkably, based on the self-changing principle, the photosensitive hydrogels can be developed as photoprinting soft and wet platforms onto which specific 3D characters and images are faithfully duplicated in macro/microscale without contact by UV light irradiation under the cover of customized photomasks. Importantly, a quick response (QR) code is accurately printed on the photoactive hydrogel for the first time. Scanning QR code with a smartphone can quickly connect to a web page. This photoactive hydrogel is promising to be a new printing or recording material.

  6. A Periosteum-Inspired 3D Hydrogel-Bioceramic Composite for Enhanced Bone Regeneration .

    PubMed

    Chun, Yong Yao; Wang, Jun Kit; Tan, Nguan Soon; Chan, Peggy Puk Yik; Tan, Timothy Thatt Yang; Choong, Cleo

    2016-02-01

    A 3D injectable hydrogel-bioceramic composite consisting of gelatin-3-(4-hydroxyphenyl) propionic acid (Gtn-HPA) and carboxymethyl cellulose-tyramine (CMC-Tyr), incorporated with fish scale-derived calcium phosphate (CaP), is developed for bone applications. The hydrogel-bioceramic composite has significantly improved the elastic modulus compared to the non-filled hydrogel, of which the addition of 10 w/v% CaP showed zero order fluorescein isothiocyanate (FITC)-dextran release profile and a significantly higher proliferation rate of encapsulated cells. All the samples promote the nucleation and growth of CaP minerals when exposed to 1× SBF. Overall, the hydrogel-bioceramic composite with 10 w/v% CaP can potentially be used as a periosteum-mimicking membrane to facilitate bone regeneration.

  7. Enzymatically triggered peptide hydrogels for 3D cell encapsulation and culture.

    PubMed

    Szkolar, Laura; Guilbaud, Jean-Baptiste; Miller, Aline F; Gough, Julie E; Saiani, Alberto

    2014-07-01

    We have investigated the possibility of using enzymatically triggered peptide hydrogels for the encapsulation and culture of cells. Based on recent work done on the enzymatically triggered gelation of FEFK (F, phenylalanine; E, glutamic acid; K, lysine) using thermolysin, a protease enzyme from Bacillus Thermoproteolyticus Rokko, we have investigated the possibility of using this gelation triggering mechanism to encapsulate cells within a 3D hydrogel matrix. First, the properties of enzymatically triggered hydrogels prepared in phosphate buffer solution were investigated and compared with the properties of hydrogels prepared in HPLC grade water from our previous work. We showed that the use of phosphate buffer solution allowed the production of hydrogels with very high shear moduli (>1 MPa). The gelation kinetics was also investigated, and the mechanical properties of the system were shown to closely follow the synthesis of the octapeptide by the enzyme through reverse hydrolysis. In a second phase, we developed, on the basis of information acquired, a facile protocol for the encapsulation of cells and plating of the hydrogel. Human dermal fibroblasts were then used to exemplify the use of these materials. FEFEFKFK octapeptide hydrogels prepared under the same conditions and with the same mechanical properties were used as a control. We showed that no significant differences were observed between the two systems and that after a decrease in cell number on day 1, cells start to proliferate. After 5 days of culture, the cells can be seen to start to adopt a stretched morphology typical of fibroblasts. The results clearly show that the protocol developed minimises the potential detrimental effect that thermolysin can have on the cells and that these enzymatically triggered hydrogels can be used for the 3D encapsulation and culture of cells.

  8. Scaffolds for 3D in vitro culture of neural lineage cells.

    PubMed

    Murphy, Ashley R; Laslett, Andrew; O'Brien, Carmel M; Cameron, Neil R

    2017-03-01

    Understanding how neurodegenerative disorders develop is not only a key challenge for researchers but also for the wider society, given the rapidly aging populations in developed countries. Advances in this field require new tools with which to recreate neural tissue in vitro and produce realistic disease models. This in turn requires robust and reliable systems for performing 3D in vitro culture of neural lineage cells. This review provides a state of the art update on three-dimensional culture systems for in vitro development of neural tissue, employing a wide range of scaffold types including hydrogels, solid porous polymers, fibrous materials and decellularised tissues as well as microfluidic devices and lab-on-a-chip systems. To provide some context with in vivo development of the central nervous system (CNS), we also provide a brief overview of the neural stem cell niche, neural development and neural differentiation in vitro. We conclude with a discussion of future directions for this exciting and important field of biomaterials research.

  9. Degradation behaviors of geometric cues and mechanical properties in a 3D scaffold for tendon repair.

    PubMed

    Wu, Yang; Wong, Yoke San; Fuh, Jerry Ying Hsi

    2017-04-01

    A three-dimensional (3D) scaffold fabricated via electrohydrodynamic jet printing (E-jetting) and thermally uniaxial stretching, has been developed for tendon tissue regeneration in our previous study. In this study, more in-depth biological test showed that the aligned cell morphology guided by the anisotropic geometries of the 3D tendon scaffolds, leading to up-regulated tendious gene expression including collagen type I, decorin, tenascin-C, and biglycan, as compared to the electrospun scaffolds. Given the importance of geometric cues to the biological function of the scaffolds, the degradation behaviors of the 3D scaffolds were investigated. Results from accelerated hydrolysis showed that the E-jetted portion followed bulk-controlled erosion, while the unaixially stretched portion followed surface-controlled erosion. The 3D tendon scaffold exhibited consistency between the weight loss and the decline of mechanical properties, which indicated by a 65% decrease in mass with a corresponding 56% loss in ultimate tensile strength after degradation. This study not only reveals that the anisotropic geometries of 3D tendon scaffold could affect cell morphology and lead to desired gene expression toward tendon tissue but also gives an insight into how the degradation impacts geometric cues and mechanical properties of the as-fabricated scaffold. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1138-1149, 2017.

  10. Ornamenting 3D printed scaffolds with cell-laid extracellular matrix for bone tissue regeneration.

    PubMed

    Pati, Falguni; Song, Tae-Ha; Rijal, Girdhari; Jang, Jinah; Kim, Sung Won; Cho, Dong-Woo

    2015-01-01

    3D printing technique is the most sophisticated technique to produce scaffolds with tailorable physical properties. But, these scaffolds often suffer from limited biological functionality as they are typically made from synthetic materials. Cell-laid mineralized ECM was shown to be potential for improving the cellular responses and drive osteogenesis of stem cells. Here, we intend to improve the biological functionality of 3D-printed synthetic scaffolds by ornamenting them with cell-laid mineralized extracellular matrix (ECM) that mimics a bony microenvironment. We developed bone graft substitutes by using 3D printed scaffolds made from a composite of polycaprolactone (PCL), poly(lactic-co-glycolic acid) (PLGA), and β-tricalcium phosphate (β-TCP) and mineralized ECM laid by human nasal inferior turbinate tissue-derived mesenchymal stromal cells (hTMSCs). A rotary flask bioreactor was used to culture hTMSCs on the scaffolds to foster formation of mineralized ECM. A freeze/thaw cycle in hypotonic buffer was used to efficiently decellularize (97% DNA reduction) the ECM-ornamented scaffolds while preserving its main organic and inorganic components. The ECM-ornamented 3D printed scaffolds supported osteoblastic differentiation of newly-seeded hTMSCs by upregulating four typical osteoblastic genes (4-fold higher RUNX2; 3-fold higher ALP; 4-fold higher osteocalcin; and 4-fold higher osteopontin) and increasing calcium deposition compared to bare 3D printed scaffolds. In vivo, in ectopic and orthotopic models in rats, ECM-ornamented scaffolds induced greater bone formation than that of bare scaffolds. These results suggest a valuable method to produce ECM-ornamented 3D printed scaffolds as off-the-shelf bone graft substitutes that combine tunable physical properties with physiological presentation of biological signals.

  11. 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.

  12. 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.

  13. Modulus-regulated 3D-cell proliferation in an injectable self-healing hydrogel.

    PubMed

    Li, Yongsan; Zhang, Yingwei; Shi, Feng; Tao, Lei; Wei, Yen; Wang, Xing

    2017-01-01

    Cell therapy has attracted wide attention among researchers in biomaterial and medical areas. As a carrier, hydrogels that could keep high viability of the embedded cells have been developed. However, few researches were conducted on 3D cell proliferation, a key factor for cell therapy, especially after injection. In this study, we demonstrated for the first time the proliferation regulation of the 3D-embedded L929 cells in a modulus-tunable and injectable self-healing hydrogel before and after injection without adding specific growth factor. The cells showed a stiffness-dependent proliferation to grow faster in higher stiffness hydrogels. The proliferating rates of the encapsulated cells before and after injection were quantified, and the shearing force as a possible negative influence factor was discussed, suggesting the both internal property of the hydrogel and injection process are critical for further practical applications. Due to the high operability and good biocompatibility, this injectable self-healing hydrogel can be a promising carrier for cell therapy.

  14. VA-086 methacrylate gelatine photopolymerizable hydrogels: A parametric study for highly biocompatible 3D cell embedding.

    PubMed

    Occhetta, Paola; Visone, Roberta; Russo, Laura; Cipolla, Laura; Moretti, Matteo; Rasponi, Marco

    2015-06-01

    The ability to replicate in vitro the native extracellular matrix (ECM) features and to control the three-dimensional (3D) cell organization plays a fundamental role in obtaining functional engineered bioconstructs. In tissue engineering (TE) applications, hydrogels have been successfully implied as biomatrices for 3D cell embedding, exhibiting high similarities to the natural ECM and holding easily tunable mechanical properties. In the present study, we characterized a promising photocrosslinking process to generate cell-laden methacrylate gelatin (GelMA) hydrogels in the presence of VA-086 photoinitiator using a ultraviolet LED source. We investigated the influence of prepolymer concentration and light irradiance on mechanical and biomimetic properties of resulting hydrogels. In details, the increasing of gelatin concentration resulted in enhanced rheological properties and shorter polymerization time. We then defined and validated a reliable photopolymerization protocol for cell embedding (1.5% VA-086, LED 2 mW/cm2) within GelMA hydrogels, which demonstrated to support bone marrow stromal cells viability when cultured up to 7 days. Moreover, we showed how different mechanical properties, derived from different crosslinking parameters, strongly influence cell behavior. In conclusion, this protocol can be considered a versatile tool to obtain biocompatible cell-laden hydrogels with properties easily adaptable for different TE applications.

  15. 3D fibre deposition and stereolithography techniques for the design of multifunctional nanocomposite magnetic scaffolds.

    PubMed

    De Santis, Roberto; D'Amora, Ugo; Russo, Teresa; Ronca, Alfredo; Gloria, Antonio; Ambrosio, Luigi

    2015-10-01

    Magnetic nanocomposite scaffolds based on poly(ε-caprolactone) and poly(ethylene glycol) were fabricated by 3D fibre deposition modelling (FDM) and stereolithography techniques. In addition, hybrid coaxial and bilayer magnetic scaffolds were produced by combining such techniques. The aim of the current research was to analyse some structural and functional features of 3D magnetic scaffolds obtained by the 3D fibre deposition technique and by stereolithography as well as features of multimaterial scaffolds in the form of coaxial and bilayer structures obtained by the proper integration of such methods. The compressive mechanical behaviour of these scaffolds was investigated in a wet environment at 37 °C, and the morphological features were analysed through scanning electron microscopy (SEM) and X-ray micro-computed tomography. The capability of a magnetic scaffold to absorb magnetic nanoparticles (MNPs) in water solution was also assessed. confocal laser scanning microscopy was used to assess the in vitro biological behaviour of human mesenchymal stem cells (hMSCs) seeded on 3D structures. Results showed that a wide range of mechanical properties, covering those spanning hard and soft tissues, can be obtained by 3D FDM and stereolithography techniques. 3D virtual reconstruction and SEM showed the precision with which the scaffolds were fabricated, and a good-quality interface between poly(ε-caprolactone) and poly(ethylene glycol) based scaffolds was observed for bilayer and coaxial scaffolds. Magnetised scaffolds are capable of absorbing water solution of MNPs, and a preliminary information on cell adhesion and spreading of hMSCs was obtained without the application of an external magnetic field.

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

    SciTech Connect

    Chen, Chih-Hao; Kuo, Shyh Ming; Liu, Guei-Sheung; Chen, Wan-Nan U.; Chuang, Chin-Wen; Liu, Li-Feng

    2012-11-09

    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 porous 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.

  17. Design of 3D engineered protein hydrogels for tailored control of neurite growth

    PubMed Central

    Lampe, Kyle J.; Antaris, Alexander L.; Heilshorn, Sarah C.

    2013-01-01

    The design of bioactive materials allows for tailored studies probing cell-biomaterial interactions; however, relatively few studies have examined effects of ligand density and material stiffness on neurite growth in 3D. Elastin-like proteins (ELPs) have been designed with modular bioactive and structural regions to enable the systematic characterization of design parameters within 3D materials. To promote neurite outgrowth and better understand the effects of common biomaterial design parameters on neuronal cultures, we here focused on cell-adhesive ligand density and hydrogel stiffness as design variables for ELP hydrogels. With the inherent design freedom of engineered proteins, these 3D ELP hydrogels enabled decoupled investigation into the effects of biomechanics and biochemistry on neurite outgrowth from dorsal root ganglia (DRG). Increasing the cell-adhesive RGD ligand density from 0 to 1.9 × 107 ligands/μm3 led to a significant increase in the rate, length, and density of neurite outgrowth, as quantified by a high-throughput algorithm developed for dense neurite analysis. An approximately two-fold improvement in total neurite outgrowth was observed in materials with the higher ligand density at all time-points through 7 days. ELP hydrogels with initial elastic moduli of 0.5, 1.5, or 2.1 kPa and identical RGD ligand densities revealed that the most compliant materials led to the greatest outgrowth, with some neurites extending over 1800 μm by day 7. Given the ability of ELP hydrogels to efficiently promote neurite outgrowth within defined and tunable 3D microenvironments, these materials may be useful in developing therapeutic nerve guides and the further study of basic neuron-biomaterial interactions. PMID:23128159

  18. Optimizing Photo-Encapsulation Viability of Heart Valve Cell Types in 3D Printable Composite Hydrogels.

    PubMed

    Kang, Laura Hockaday; Armstrong, Patrick A; Lee, Lauren Julia; Duan, Bin; Kang, Kevin Heeyong; Butcher, Jonathan Talbot

    2017-02-01

    Photocrosslinking hydrogel technologies are attractive for the biofabrication of cardiovascular soft tissues, but 3D printing success is dependent on multiple variables. In this study we systematically test variables associated with photocrosslinking hydrogels (photoinitiator type, photoinitiator concentration, and light intensity) for their effects on encapsulated cells in an extrusion 3D printable mixture of methacrylated gelatin/poly-ethylene glycol diacrylate/alginate (MEGEL/PEGDA3350/alginate). The fabrication conditions that produced desired hydrogel mechanical properties were compared against those that optimize aortic valve or mesenchymal stem cell viability. In the 3D hydrogel culture environment and fabrication setting studied, Irgacure can increase hydrogel stiffness with a lower proportional decrease in encapsulated cell viability compared to VA086. Human adipose derived mesenchymal stem cells (HADMSC) survived increasing photoinitiator concentrations in photo-encapsulation conditions better than aortic valve interstitial cells (HAVIC) and aortic valve sinus smooth muscle cells (HASSMC). Within the range of photo-encapsulation fabrication conditions tested with MEGEL/PEGDA/alginate (0.25-1.0% w/v VA086, 0.025-0.1% w/v Irgacure 2959, and 365 nm light intensity 2-136 mW/cm(2)), the highest viabilities achieved were 95, 93, and 93% live for HASSMC, HAVIC, and HADMSC respectively. These results identify parameter combinations that optimize cell viability during 3D printing for multiple cell types. These results also indicate that general oxidative stress is higher in photocrosslinking conditions that induce lower cell viability. However, suppressing this increase in intracellular oxidative stress did not improve cell viability, which suggests that other stress mechanisms also contribute.

  19. Control of the pore architecture in three-dimensional hydroxyapatite-reinforced hydrogel scaffolds

    PubMed Central

    Román, Jesús; Cabañas, María Victoria; Peña, Juan; Vallet-Regí, María

    2011-01-01

    Hydrogels (gellan or agarose) reinforced with nanocrystalline carbonated hydroxyapatite (nCHA) were prepared by the GELPOR3D technique. This simple method is characterized by compositional flexibility; it does not require expensive equipment, thermal treatment, or aggressive or toxic solvents, and yields a three-dimensional (3D) network of interconnected pores 300–900 μm in size. In addition, an interconnected porosity is generated, yielding a hierarchical porous architecture from the macro to the molecular scale. This porosity depends on both the drying/preservation technology (freeze drying or oven drying at 37 ○C) and on the content and microstructure of the reinforcing ceramic. For freeze-dried samples, the porosities were approximately 30, 66 and below 3% for pore sizes of 600–900 μm, 100–200 μm and 50–100 nm, respectively. The pore structure depends much on the ceramic content, so that higher contents lead to the disappearance of the characteristic honeycomb structure observed in low-ceramic scaffolds and to a lower fraction of the 100–200-μm-sized pores. The nature of the hydrogel did not affect the pore size distribution but was crucial for the behavior of the scaffolds in a hydrated medium: gellan-containing scaffolds showed a higher swelling degree owing to the presence of more hydrophilic groups. PMID:27877422

  20. Control of the pore architecture in three-dimensional hydroxyapatite-reinforced hydrogel scaffolds.

    PubMed

    Román, Jesús; Cabañas, María Victoria; Peña, Juan; Vallet-Regí, María

    2011-08-01

    Hydrogels (gellan or agarose) reinforced with nanocrystalline carbonated hydroxyapatite (nCHA) were prepared by the GELPOR3D technique. This simple method is characterized by compositional flexibility; it does not require expensive equipment, thermal treatment, or aggressive or toxic solvents, and yields a three-dimensional (3D) network of interconnected pores 300-900 μm in size. In addition, an interconnected porosity is generated, yielding a hierarchical porous architecture from the macro to the molecular scale. This porosity depends on both the drying/preservation technology (freeze drying or oven drying at 37 (○)C) and on the content and microstructure of the reinforcing ceramic. For freeze-dried samples, the porosities were approximately 30, 66 and below 3% for pore sizes of 600-900 μm, 100-200 μm and 50-100 nm, respectively. The pore structure depends much on the ceramic content, so that higher contents lead to the disappearance of the characteristic honeycomb structure observed in low-ceramic scaffolds and to a lower fraction of the 100-200-μm-sized pores. The nature of the hydrogel did not affect the pore size distribution but was crucial for the behavior of the scaffolds in a hydrated medium: gellan-containing scaffolds showed a higher swelling degree owing to the presence of more hydrophilic groups.

  1. Self-assembling hydrogel scaffolds for photocatalytic hydrogen production

    NASA Astrophysics Data System (ADS)

    Weingarten, Adam S.; Kazantsev, Roman V.; Palmer, Liam C.; McClendon, Mark; Koltonow, Andrew R.; Samuel, Amanda P. S.; Kiebala, Derek J.; Wasielewski, Michael R.; Stupp, Samuel I.

    2014-11-01

    Integration into a soft material of all the molecular components necessary to generate storable fuels is an interesting target in supramolecular chemistry. The concept is inspired by the internal structure of photosynthetic organelles, such as plant chloroplasts, which colocalize molecules involved in light absorption, charge transport and catalysis to create chemical bonds using light energy. We report here on the light-driven production of hydrogen inside a hydrogel scaffold built by the supramolecular self-assembly of a perylene monoimide amphiphile. The charged ribbons formed can electrostatically attract a nickel-based catalyst, and electrolyte screening promotes gelation. We found the emergent phenomenon that screening by the catalyst or the electrolytes led to two-dimensional crystallization of the chromophore assemblies and enhanced the electronic coupling among the molecules. Photocatalytic production of hydrogen is observed in the three-dimensional environment of the hydrogel scaffold and the material is easily placed on surfaces or in the pores of solid supports.

  2. Concentrated hydroxyapatite inks for direct-write assembly of 3-D periodic scaffolds.

    PubMed

    Michna, Sarah; Wu, Willie; Lewis, Jennifer A

    2005-10-01

    Hydroxyapatite (HA) scaffolds with a 3-D periodic architecture and multiscale porosity have been fabricated by direct-write assembly. Concentrated HA inks with tailored viscoelastic properties were developed to enable the construction of complex 3-D architectures comprised of self-supporting cylindrical rods in a layer-by-layer patterning sequence. By controlling their lattice constant and sintering conditions, 3-D periodic HA scaffolds were produced with a bimodal pore size distribution. Mercury intrusion porosimetry (MIP) was used to determine the characteristic pore size and volume associated with the interconnected pore channels between HA rods and the finer pores within the partially sintered HA rods.

  3. 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.

  4. 3D in vitro bioengineered tumors based on collagen I hydrogels

    PubMed Central

    Szot, Christopher S.; Buchanan, Cara F.; Freeman, Joseph W.; Rylander, Marissa N.

    2011-01-01

    Cells cultured within a three-dimensional (3D) in vitro environment have the ability to acquire phenotypes and respond to stimuli analogous to in vivo biological systems. This approach has been utilized in tissue engineering and can also be applied to the development of a physiologically relevant in vitro tumor model. In this study, collagen I hydrogels cultured with MDA-MB-231 human breast cancer cells were bioengineered as a platform for in vitro solid tumor development. The cell–cell and cell-matrix interactions present during in vivo tissue progression were encouraged within the 3D hydrogel architecture, and the biocompatibility of collagen I supported unconfined cellular proliferation. The development of necrosis beyond a depth of ~150–200 μm and the expression of hypoxia-inducible factor (HIF)-1α were demonstrated in the in vitro bioengineered tumors. Oxygen and nutrient diffusion limitations through the collagen I matrix as well as competition for available nutrients resulted in growing levels of intra-cellular hypoxia, quantified by a statistically significant (p < 0.01) upregulation of HIF-1α gene expression. The bioengineered tumors also demonstrated promising angiogenic potential with a statistically significant (p < 0.001) upregulation of vascular endothelial growth factor (VEGF)-A gene expression. In addition, comparable gene expression analysis demonstrated a statistically significant increase of HIF-1α (p < 0.05) and VEGF-A (p < 0.001) by MDA-MB-231 cells cultured in the 3D collagen I hydrogels compared to cells cultured in a monolayer on two-dimensional tissue culture polystyrene. The results presented in this study demonstrate the capacity of collagen I hydrogels to facilitate the development of 3D in vitro bioengineered tumors that are representative of the pre-vascularized stages of in vivo solid tumor progression. PMID:21782234

  5. Natural assembly of platelet lysate-loaded nanocarriers into enriched 3D hydrogels for cartilage regeneration.

    PubMed

    Santo, Vítor E; Popa, Elena G; Mano, João F; Gomes, Manuela E; Reis, Rui L

    2015-06-01

    The role of Platelet Lysates (PLs) as a source of growth factors (GFs) and as main element of three-dimensional (3D) hydrogels has been previously described. However, the resulting hydrogels usually suffer from high degree of contraction, limiting their usefulness. This work describes the development of a stable biomimetic 3D hydrogel structure based on PLs, through the spontaneous assembling of a high concentration of chitosan-chondroitin sulfate nanoparticles (CH/CS NPs) with PLs loaded by adsorption. The interactions between the NPs and the lysates resemble the ones observed in the extracellular matrix (ECM) native environment between glycosaminoglycans and ECM proteins. In vitro release studies were carried out focusing on the quantification of PDGF-BB and TGF-β1 GFs. Human adipose derived stem cells (hASCs) were entrapped in these 3D hydrogels and cultured in vitro under chondrogenic stimulus, in order to assess their potential use for cartilage regeneration. Histological, immunohistological and gene expression analysis demonstrated that the PL-assembled constructs entrapping hASCs exhibited results similar to the positive control (hASCS cultured in pellets), concerning the levels of collagen II expression and immunolocalization of collagen type I and II and aggrecan. Moreover, the deposition of new cartilage ECM was detected by alcian blue and safranin-O positive stainings. This work demonstrates the potential of PLs to act simultaneously as a source/carrier of GFs and as a 3D structure of support, through the application of a "bottom-up" approach involving the assembly of NPs, resulting in an enriched construct for cartilage regeneration applications.

  6. Pluripotent stem cell expansion and neural differentiation in 3-D scaffolds of tunable Poisson's ratio.

    PubMed

    Yan, Yuanwei; Li, Yan; Song, Liqing; Zeng, Changchun; Li, Yan

    2017-02-01

    Biophysical properties of the scaffolds such as the elastic modulus, have been recently shown to impact stem cell lineage commitment. On the other hand, the contribution of the Poisson's ratio, another important biophysical property, to the stem cell fate decision, has not been studied. Scaffolds with tunable Poisson's ratio (ν) (termed as auxetic scaffolds when Poisson's ratio is zero or negative) are anticipated to provide a spectrum of unique biophysical 3-D microenvironments to influence stem cell fate. To test this hypothesis, in the present work we fabricated auxetic polyurethane scaffolds (ν=0 to -0.45) and evaluated their effects on neural differentiation of mouse embryonic stem cells (ESCs) and human induced pluripotent stem cells (hiPSCs). Compared to the regular scaffolds (ν=+0.30) before auxetic conversion, the auxetic scaffolds supported smaller aggregate formation and higher expression of β-tubulin III upon neural differentiation. The influences of pore structure, Poisson's ratio, and elastic modulus on neural lineage commitment were further evaluated using a series of auxetic scaffolds. The results indicate that Poisson's ratio may confound the effects of elastic modulus, and auxetic scaffolds with proper pore structure and Poisson's ratio enhance neural differentiation. This study demonstrates that tuning the Poisson's ratio of the scaffolds together with elastic modulus and microstructure would enhance the capability to generate broader, more diversified ranges of biophysical 3-D microenvironments for the modulation of cellular differentiation.

  7. 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.

  8. Development of nanocellulose scaffolds with tunable structures to support 3D cell culture.

    PubMed

    Liu, Jun; Cheng, Fang; Grénman, Henrik; Spoljaric, Steven; Seppälä, Jukka; E Eriksson, John; Willför, Stefan; Xu, Chunlin

    2016-09-05

    Swollen three-dimensional nanocellulose films and their resultant aerogels were prepared as scaffolds towards tissue engineering application. The nanocellulose hydrogels with various swelling degree (up to 500 times) and the resultant aerogels with desired porosity (porosity up to 99.7% and specific surface area up to 308m(2)/g) were prepared by tuning the nanocellulose charge density, the swelling media conditions, and the material processing approach. Representative cell-based assays were applied to assess the material biocompatibility and efficacy of the human extracellular matrix (ECM)-mimicking nanocellulose scaffolds. The effects of charge density and porosity of the scaffolds on the biological tests were investigated for the first time. The results reveal that the nanocellulose scaffolds could promote the survival and proliferation of tumor cells, and enhance the transfection of exogenous DNA into the cells. These results suggest the usefulness of the nanocellulose-based matrices in supporting crucial cellular processes during cell growth and proliferation.

  9. Rapid Fabrication of Cell-Laden Alginate Hydrogel 3D Structures by Micro Dip-Coating

    PubMed Central

    Ghanizadeh Tabriz, Atabak; Mills, Christopher G.; Mullins, John J.; Davies, Jamie A.; Shu, Wenmiao

    2017-01-01

    Development of a simple, straightforward 3D fabrication method to culture cells in 3D, without relying on any complex fabrication methods, remains a challenge. In this paper, we describe a new technique that allows fabrication of scalable 3D cell-laden hydrogel structures easily, without complex machinery: the technique can be done using only apparatus already available in a typical cell biology laboratory. The fabrication method involves micro dip-coating of cell-laden hydrogels covering the surface of a metal bar, into the cross-linking reagents calcium chloride or barium chloride to form hollow tubular structures. This method can be used to form single layers with thickness ranging from 126 to 220 µm or multilayered tubular structures. This fabrication method uses alginate hydrogel as the primary biomaterial and a secondary biomaterial can be added depending on the desired application. We demonstrate the feasibility of this method, with survival rate over 75% immediately after fabrication and normal responsiveness of cells within these tubular structures using mouse dermal embryonic fibroblast cells and human embryonic kidney 293 cells containing a tetracycline-responsive, red fluorescent protein (tHEK cells). PMID:28286747

  10. Development of 3-D Hydrogel Culture Systems With On-Demand Cell Separation

    PubMed Central

    Hamilton, Sharon K.; Bloodworth, Nathaniel C.; Massad, Christopher S.; Hammoudi, Taymour M.; Suri, Shalu; Yang, Peter J.; Lu, Hang; Temenoff, Johnna S.

    2013-01-01

    Recently there has been an increased interest in the effects of paracrine signaling between groups of cells, particularly in the context of better understanding how stem cells contribute to tissue repair. Most current 3-D co-culture methods lack the ability to effectively separate 2 cell populations after the culture period, which is important for simultaneously analyzing the reciprocal effects of each cell type on the other. Here, we detail the development of a 3-D hydrogel co-culture system that allows us to culture different cell types for up to 7 days and subsequently separate and isolate the different cell populations using enzyme-sensitive glues. Separable 3-D co-culture laminates were prepared by laminating PEG-based hydrogels with enzyme-degradable hydrogel adhesives. Encapsulated cell populations exhibited good segregation with well-defined interfaces. Furthermore, constructs can be separated on-demand upon addition of the appropriate enzyme and cell viability remains high throughout the culture period, even after laminate separation. This platform offers great potential for a variety of basic cell signaling studies as the incorporation of an enzyme-sensitive adhesive interface allows the on-demand separation of individual cell populations for immediate analysis or further culture to examine persistence of co-culture effects and paracrine signaling on cell populations. PMID:23447378

  11. High-performance 3D printing of hydrogels by water-dispersible photoinitiator nanoparticles

    PubMed Central

    Pawar, Amol A.; Saada, Gabriel; Cooperstein, Ido; Larush, Liraz; Jackman, Joshua A.; Tabaei, Seyed R.; Cho, Nam-Joon; Magdassi, Shlomo

    2016-01-01

    In the absence of water-soluble photoinitiators with high absorbance in the ultraviolet (UV)–visible range, rapid three-dimensional (3D) printing of hydrogels for tissue engineering is challenging. A new approach enabling rapid 3D printing of hydrogels in aqueous solutions is presented on the basis of UV-curable inks containing nanoparticles of highly efficient but water-insoluble photoinitiators. The extinction coefficient of the new water-dispersible nanoparticles of 2,4,6-trimethylbenzoyl-diphenylphosphine oxide (TPO) is more than 300 times larger than the best and most used commercially available water-soluble photoinitiator. The TPO nanoparticles absorb significantly in the range from 385 to 420 nm, making them suitable for use in commercially available, low-cost, light-emitting diode–based 3D printers using digital light processing. The polymerization rate at this range is very fast and enables 3D printing that otherwise is impossible to perform without adding solvents. The TPO nanoparticles were prepared by rapid conversion of volatile microemulsions into water-dispersible powder, a process that can be used for a variety of photoinitiators. Such water-dispersible photoinitiator nanoparticles open many opportunities to enable rapid 3D printing of structures prepared in aqueous solutions while bringing environmental advantages by using low-energy curing systems and avoiding the need for solvents. PMID:27051877

  12. High-performance 3D printing of hydrogels by water-dispersible photoinitiator nanoparticles.

    PubMed

    Pawar, Amol A; Saada, Gabriel; Cooperstein, Ido; Larush, Liraz; Jackman, Joshua A; Tabaei, Seyed R; Cho, Nam-Joon; Magdassi, Shlomo

    2016-04-01

    In the absence of water-soluble photoinitiators with high absorbance in the ultraviolet (UV)-visible range, rapid three-dimensional (3D) printing of hydrogels for tissue engineering is challenging. A new approach enabling rapid 3D printing of hydrogels in aqueous solutions is presented on the basis of UV-curable inks containing nanoparticles of highly efficient but water-insoluble photoinitiators. The extinction coefficient of the new water-dispersible nanoparticles of 2,4,6-trimethylbenzoyl-diphenylphosphine oxide (TPO) is more than 300 times larger than the best and most used commercially available water-soluble photoinitiator. The TPO nanoparticles absorb significantly in the range from 385 to 420 nm, making them suitable for use in commercially available, low-cost, light-emitting diode-based 3D printers using digital light processing. The polymerization rate at this range is very fast and enables 3D printing that otherwise is impossible to perform without adding solvents. The TPO nanoparticles were prepared by rapid conversion of volatile microemulsions into water-dispersible powder, a process that can be used for a variety of photoinitiators. Such water-dispersible photoinitiator nanoparticles open many opportunities to enable rapid 3D printing of structures prepared in aqueous solutions while bringing environmental advantages by using low-energy curing systems and avoiding the need for solvents.

  13. Mechanical properties and shape memory effect of 3D-printed PLA-based porous scaffolds.

    PubMed

    Senatov, F S; Niaza, K V; Zadorozhnyy, M Yu; Maksimkin, A V; Kaloshkin, S D; Estrin, Y Z

    2016-04-01

    In the present work polylactide (PLA)/15wt% hydroxyapatite (HA) porous scaffolds with pre-modeled structure were obtained by 3D-printing by fused filament fabrication. Composite filament was obtained by extrusion. Mechanical properties, structural characteristics and shape memory effect (SME) were studied. Direct heating was used for activation of SME. The average pore size and porosity of the scaffolds were 700μm and 30vol%, respectively. Dispersed particles of HA acted as nucleation centers during the ordering of PLA molecular chains and formed an additional rigid fixed phase that reduced molecular mobility, which led to a shift of the onset of recovery stress growth from 53 to 57°C. A more rapid development of stresses was observed for PLA/HA composites with the maximum recovery stress of 3.0MPa at 70°C. Ceramic particles inhibited the growth of cracks during compression-heating-compression cycles when porous PLA/HA 3D-scaffolds recovered their initial shape. Shape recovery at the last cycle was about 96%. SME during heating may have resulted in "self-healing" of scaffold by narrowing the cracks. PLA/HA 3D-scaffolds were found to withstand up to three compression-heating-compression cycles without delamination. It was shown that PLA/15%HA porous scaffolds obtained by 3D-printing with shape recovery of 98% may be used as self-fitting implant for small bone defect replacement owing to SME.

  14. Thermoforming techniques for manufacturing porous scaffolds for application in 3D cell cultivation.

    PubMed

    Borowiec, Justyna; Hampl, Jörg; Gebinoga, Michael; Elsarnagawy, Tarek; Elnakady, Yasser A; Fouad, Hassan; Almajhadi, Fahd; Fernekorn, Uta; Weise, Frank; Singh, Sukhdeep; Elsarnagawy, Dief; Schober, Andreas

    2015-04-01

    Within the scientific community, there is an increasing demand to apply advanced cell cultivation substrates with increased physiological functionalities for studying spatially defined cellular interactions. Porous polymeric scaffolds are utilized for mimicking an organ-like structure or engineering complex tissues and have become a key element for three-dimensional (3D) cell cultivation in the meantime. As a consequence, efficient 3D scaffold fabrication methods play an important role in modern biotechnology. Here, we present a novel thermoforming procedure for manufacturing porous 3D scaffolds from permeable materials. We address the issue of precise thermoforming of porous polymer foils by using multilayer polymer thermoforming technology. This technology offers a new method for structuring porous polymer foils that are otherwise available for non-porous polymers only. We successfully manufactured 3D scaffolds from solvent casted and phase separated polylactic acid (PLA) foils and investigated their biocompatibility and basic cellular performance. The HepG2 cell culture in PLA scaffold has shown enhanced albumin secretion rate in comparison to a previously reported polycarbonate based scaffold with similar geometry.

  15. Photocrosslinked hyaluronic acid hydrogels: natural, biodegradable tissue engineering scaffolds.

    PubMed

    Baier Leach, Jennie; Bivens, Kathryn A; Patrick, Charles W; Schmidt, Christine E

    2003-06-05

    Ideally, rationally designed tissue engineering scaffolds promote natural wound healing and regeneration. Therefore, we sought to synthesize a biomimetic hydrogel specifically designed to promote tissue repair and chose hyaluronic acid (HA; also called hyaluronan) as our initial material. Hyaluronic acid is a naturally occurring polymer associated with various cellular processes involved in wound healing, such as angiogenesis. Hyaluronic acid also presents unique advantages: it is easy to produce and modify, hydrophilic and nonadhesive, and naturally biodegradable. We prepared a range of glycidyl methacrylate-HA (GMHA) conjugates, which were subsequently photopolymerized to form crosslinked GMHA hydrogels. A range of hydrogel degradation rates was achieved as well as a corresponding, modest range of material properties (e.g., swelling, mesh size). Increased amounts of conjugated methacrylate groups corresponded with increased crosslink densities and decreased degradation rates and yet had an insignificant effect on human aortic endothelial cell cytocompatibility and proliferation. Rat subcutaneous implants of the GMHA hydrogels showed good biocompatibility, little inflammatory response, and similar levels of vascularization at the implant edge compared with those of fibrin positive controls. Therefore, these novel GMHA hydrogels are suitable for modification with adhesive peptide sequences (e.g., RGD) and use in a variety of wound-healing applications.

  16. 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.

  17. 3D-Printed ABS and PLA Scaffolds for Cartilage and Nucleus Pulposus Tissue Regeneration

    PubMed Central

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

    2015-01-01

    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. PMID:26151846

  18. Thermo-responsive non-woven scaffolds for "smart" 3D cell culture.

    PubMed

    Rossouw, Claire L; Chetty, Avashnee; Moolman, Francis Sean; Birkholtz, Lyn-Marie; Hoppe, Heinrich; Mancama, Dalu T

    2012-08-01

    The thermo-responsive polymer poly(N-isopropylacrylamide) has received widespread attention for its in vitro application in the non-invasive, non-destructive release of adherent cells on two dimensional surfaces. In this study, 3D non-woven scaffolds fabricated from poly(propylene) (PP), poly(ethylene terephthalate) (PET), and nylon that had been grafted with PNIPAAm were tested for their ability to support the proliferation and subsequent thermal release of HC04 and HepG2 hepatocytes. Hepatocyte viability and proliferation were estimated using the Alamar Blue assay and Hoechst 33258 total DNA quantification. The assays revealed that the pure and grafted non-woven scaffolds maintained the hepatocytes within the matrix and promoted 3D proliferation comparable to that of the commercially available Algimatrix™ alginate scaffold. Albumin production and selected cytochrome P450 genes expression was found to be superior in cells growing on pure and grafted non-woven PP scaffolds as compared to cells grown as a 2D monolayer. Two scaffolds, namely, PP-g-PNIPAAm-A and PP-g-PNIPAAm-B were identified as having far superior thermal release capabilities; releasing the majority of the cells from the matrices within 2 h. This is the first report for the development of 3D non-woven, thermo-responsive scaffolds able to release cells from the matrix without the use of any enzymatic assistance or scaffold degradation.

  19. Ceramic scaffolds produced by computer-assisted 3D printing and sintering: characterization and biocompatibility investigations.

    PubMed

    Warnke, Patrick H; Seitz, Hermann; Warnke, Frauke; Becker, Stephan T; Sivananthan, Sureshan; Sherry, Eugene; Liu, Qin; Wiltfang, Jörg; Douglas, Timothy

    2010-04-01

    Hydroxyapatite (HAP) and tricalcium phosphate (TCP) are two very common ceramic materials for bone replacement. However, in general HAP and TCP scaffolds are not tailored to the exact dimensions of the defect site and are mainly used as granules or beads. Some scaffolds are available as ordinary blocks, but cannot be customized for individual perfect fit. Using computer-assisted 3D printing, an emerging rapid prototyping technique, individual three-dimensional ceramic scaffolds can be built up from TCP or HAP powder layer by layer with subsequent sintering. These scaffolds have precise dimensions and highly defined and regular internal characteristics such as pore size. External shape and internal characteristics such as pore size can be fabricated using Computer Assisted Design (CAD) based on individual patient data. Thus, these scaffolds could be designed as perfect fit replacements to reconstruct the patient's skeleton. Before their use as bone replacement materials in vivo, in vitro testing of these scaffolds is necessary. In this study, the behavior of human osteoblasts on HAP and TCP scaffolds was investigated. The commonly used bone replacement material BioOss(R) served as control. Biocompatibility was assessed by scanning electron microscopy (SEM), fluorescence microscopy after staining for cell vitality with fluorescin diacetate (FDA) and propidium iodide (PI) and the MTT, LDH, and WST biocompatibility tests. Both versions were colonised by human osteoblasts, however more cells were seen on HAP scaffolds than TCP scaffolds. Cell vitality staining and MTT, LDH, and WST tests showed superior biocompatibility of HAP scaffolds to BioOss, while BioOss was more compatible than TCP. Further experiments are necessary to determine biocompatibility in vivo. Future modifications of 3D printed scaffolds offer advantageous features for Tissue Engineering. The integration of channels could allow for vascular and nerve ingrowth into the scaffold. Also the complex shapes

  20. Partitioning microfluidic channels with hydrogel to construct tunable 3-D cellular microenvironments.

    PubMed

    Wong, Amy P; Perez-Castillejos, Raquel; Christopher Love, J; Whitesides, George M

    2008-04-01

    Accurate modeling of the cellular microenvironment is important for improving studies of cell biology in vitro. Here, we demonstrate a flexible method for creating a cellular microenvironment in vitro that allows (i) controlled spatial distribution (patterning) of multiple types of cells within three-dimensional (3-D) matrices of a biologically derived, thermally curable hydrogel (Matrigel) and (ii) application of gradients of soluble factors, such as cytokines, across the hydrogel. The technique uses laminar flow to divide a microchannel into multiple subchannels separated by microslabs of hydrogel. It does not require the use of UV light or photoinitiators and is compatible with cell culture in the hydrogel. This technique makes it possible to design model systems to study cellular communication mediated by the diffusion of soluble factors within 3-D matrices. Such factors can originate either from secretions of neighboring cells patterned within the microchannel, or from an external source -- e.g., a solution of growth factors injected into a subchannel. This method is particularly useful for studying cells such as those of the immune system, which are often weakly adherent and difficult to position precisely with standard systems for cell culture. We demonstrated this application by co-culturing two types of macrophage-like cells (BAC1.2F5 and LADMAC cell lines) within spatially separated regions of a slab of hydrogel. This pair of cell lines represents a simple model system for intercellular communication: the LADMAC cells produce colony-stimulating factor 1 (CSF-1), which is required by the BAC cells for survival.

  1. Partitioning Microfluidic Channels with Hydrogel to Construct Tunable 3-D Cellular Microenvironments

    PubMed Central

    Wong, Amy P.; Perez-Castillejos, Raquel; Love, J. Christopher; Whitesides, George M.

    2008-01-01

    Accurate modeling of the cellular microenvironment is important for improving studies of cell biology in vitro. Here, we demonstrate a flexible method for creating a cellular microenvironment in vitro that allows i) controlled spatial distribution (patterning) of multiple types of cells within three-dimensional (3-D) matrices of a biologically-derived, thermally-curable hydrogel (Matrigel) and ii) application of gradients of soluble factors, such as cytokines, across the hydrogel. The technique uses laminar flow to divide a microchannel into multiple subchannels separated by microslabs of hydrogel. It does not require the use of UV light or photoinitiators, and is compatible with cell culture in the hydrogel. This technique makes it possible to design model systems to study cellular communication mediated by the diffusion of soluble factors within 3-D matrices. Such factors can originate either from secretions of neighboring cells patterned within the microchannel, or from an external source—e.g., a solution of growth factors injected into a subchannel. This method is particularly useful for studying cells such as those of the immune system, which are often weakly adherent and difficult to position precisely with standard systems for cell culture. We demonstrated this application by co-culturing two types of macrophage-like cells (BAC1.2F5 and LADMAC cell lines) within spatially separated regions of a slab of hydrogel. This pair of cell lines represents a simple model system for intercellular communication: the LADMAC cells produce colony-stimulating factor 1 (CSF-1), which is required by the BAC cells for survival. PMID:18243301

  2. Peptide-directed self-assembly of functionalized polymeric nanoparticles part I: design and self-assembly of peptide-copolymer conjugates into nanoparticle fibers and 3D scaffolds.

    PubMed

    Ding, Xiaochu; Janjanam, Jagadeesh; Tiwari, Ashutosh; Thompson, Martin; Heiden, Patricia A

    2014-06-01

    A robust self-assembly of nanoparticles into fibers and 3D scaffolds is designed and fabricated by functionalizing a RAFT-polymerized amphiphilic triblock copolymer with designer ionic complementary peptides so that the assembled core-shell polymeric nanoparticles are directed by peptide assembly into continuous "nanoparticle fibers," ultimately leading to 3D fiber scaffolds. The assembled nanostructure is confirmed by FESEM and optical microscopy. The assembly is not hindered when a protein (insulin) is incorporated within the nanoparticles as an active ingredient. MTS cytotoxicity tests on SW-620 cell lines show that the peptides, copolymers, and peptide-copolymer conjugates are biocompatible. The methodology of self-assembled nanoparticle fibers and 3D scaffolds is intended to combine the advantages of a flexible hydrogel scaffold with the versatility of controlled release nanoparticles to offer unprecedented ability to incorporate desired drug(s) within a self-assembled scaffold system with individual control over the release of each drug.

  3. Human Skin 3D Bioprinting Using Scaffold-Free Approach.

    PubMed

    Pourchet, Léa J; Thepot, Amélie; Albouy, Marion; Courtial, Edwin J; Boher, Aurélie; Blum, Loïc J; Marquette, Christophe A

    2017-02-01

    Organ in vitro synthesis is one of the last bottlenecks between tissue engineering and transplantation of synthetic organs. Bioprinting has proven its capacity to produce 3D objects composed of living cells but highly organized tissues such as full thickness skin (dermis + epidermis) are rarely attained. The focus of the present study is to demonstrate the capability of a newly developed ink formulation and the use of an open source printer, for the production of a really complete skin model. Proofs are given through immunostaining and electronic microscopy that the bioprinted skin presents all characteristics of human skin, both at the molecular and macromolecular level. Finally, the printability of large skin objects is demonstrated with the printing of an adult-size ear.

  4. Design of a Novel 3D Printed Bioactive Nanocomposite Scaffold for Improved Osteochondral Regeneration.

    PubMed

    Castro, Nathan J; Patel, Romil; Zhang, Lijie Grace

    2015-09-01

    Chronic and acute osteochondral defects as a result of osteoarthritis and trauma present a common and serious clinical problem due to the tissue's inherent complexity and poor regenerative capacity. In addition, cells within the osteochondral tissue are in intimate contact with a 3D nanostructured extracellular matrix composed of numerous bioactive organic and inorganic components. As an emerging manufacturing technique, 3D printing offers great precision and control over the microarchitecture, shape and composition of tissue scaffolds. Therefore, the objective of this study is to develop a biomimetic 3D printed nanocomposite scaffold with integrated differentiation cues for improved osteochondral tissue regeneration. Through the combination of novel nano-inks composed of organic and inorganic bioactive factors and advanced 3D printing, we have successfully fabricated a series of novel constructs which closely mimic the native 3D extracellular environment with hierarchical nanoroughness, microstructure and spatiotemporal bioactive cues. Our results illustrate several key characteristics of the 3D printed nanocomposite scaffold to include improved mechanical properties as well as excellent cytocompatibility for enhanced human bone marrow-derived mesenchymal stem cell adhesion, proliferation, and osteochondral differentiation in vitro. The present work further illustrates the effectiveness of the scaffolds developed here as a promising and highly tunable platform for osteochondral tissue regeneration.

  5. Design of a Novel 3D Printed Bioactive Nanocomposite Scaffold for Improved Osteochondral Regeneration

    PubMed Central

    Castro, Nathan J.; Patel, Romil; Zhang, Lijie Grace

    2015-01-01

    Chronic and acute osteochondral defects as a result of osteoarthritis and trauma present a common and serious clinical problem due to the tissue's inherent complexity and poor regenerative capacity. In addition, cells within the osteochondral tissue are in intimate contact with a 3D nanostructured extracellular matrix composed of numerous bioactive organic and inorganic components. As an emerging manufacturing technique, 3D printing offers great precision and control over the microarchitecture, shape and composition of tissue scaffolds. Therefore, the objective of this study is to develop a biomimetic 3D printed nanocomposite scaffold with integrated differentiation cues for improved osteochondral tissue regeneration. Through the combination of novel nano-inks composed of organic and inorganic bioactive factors and advanced 3D printing, we have successfully fabricated a series of novel constructs which closely mimic the native 3D extracellular environment with hierarchical nanoroughness, microstructure and spatiotemporal bioactive cues. Our results illustrate several key characteristics of the 3D printed nanocomposite scaffold to include improved mechanical properties as well as excellent cytocompatibility for enhanced human bone marrow-derived mesenchymal stem cell adhesion, proliferation, and osteochondral differentiation in vitro. The present work further illustrates the effectiveness of the scaffolds developed here as a promising and highly tunable platform for osteochondral tissue regeneration. PMID:26366231

  6. 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

  7. Mechanical evaluation of gradient electrospun scaffolds with 3D printed ring reinforcements for tracheal defect repair.

    PubMed

    Ott, Lindsey M; Zabel, Taylor A; Walker, Natalie K; Farris, Ashley L; Chakroff, Jason T; Ohst, Devan G; Johnson, Jed K; Gehrke, Steven H; Weatherly, Robert A; Detamore, Michael S

    2016-04-21

    Tracheal stenosis can become a fatal condition, and current treatments include augmentation of the airway with autologous tissue. A tissue-engineered approach would not require a donor source, while providing an implant that meets both surgeons' and patients' needs. A fibrous, polymeric scaffold organized in gradient bilayers of polycaprolactone (PCL) and poly-lactic-co-glycolic acid (PLGA) with 3D printed structural ring supports, inspired by the native trachea rings, could meet this need. The purpose of the current study was to characterize the tracheal scaffolds with mechanical testing models to determine the design most suitable for maintaining a patent airway. Degradation over 12 weeks revealed that scaffolds with the 3D printed rings had superior properties in tensile and radial compression, with at least a three fold improvement and 8.5-fold improvement, respectively, relative to the other scaffold groups. The ringed scaffolds produced tensile moduli, radial compressive forces, and burst pressures similar to or exceeding physiological forces and native tissue data. Scaffolds with a thicker PCL component had better suture retention and tube flattening recovery properties, with the monolayer of PCL (PCL-only group) exhibiting a 2.3-fold increase in suture retention strength (SRS). Tracheal scaffolds with ring reinforcements have improved mechanical properties, while the fibrous component increased porosity and cell infiltration potential. These scaffolds may be used to treat various trachea defects (patch or circumferential) and have the potential to be employed in other tissue engineering applications.

  8. In-vivo behavior of Si-hydroxyapatite/polycaprolactone/DMB scaffolds fabricated by 3D printing.

    PubMed

    Meseguer-Olmo, Luis; Vicente-Ortega, Vicente; Alcaraz-Baños, Miguel; Calvo-Guirado, José Luis; Vallet-Regí, María; Arcos, Daniel; Baeza, Alejandro

    2013-07-01

    Scaffolds made of polycaprolactone and nanocrystalline silicon-substituted hydroxyapatite have been fabricated by 3D printing rapid prototyping technique. To asses that the scaffolds fulfill the requirements to be considered for bone grafting applications, they were implanted in New Zealand rabbits. Histological and radiological studies have demonstrated that the scaffolds implanted in bone exhibited an excellent osteointegration without the interposition of fibrous tissue between bone and implants and without immune response after 4 months of implantation. In addition, we have evaluated the possibility of improving the scaffolds efficiency by incorporating demineralized bone matrix during the preparation by 3D printing. When demineralized bone matrix (DBM) is incorporated, the efficacy of the scaffolds is enhanced, as new bone formation occurs not only in the peripheral portions of the scaffolds but also within its pores after 4 months of implantation. This enhanced performance can be explained in terms of the osteoinductive properties of the DBM in the scaffolds, which have been assessed through the new bone tissue formation when the scaffolds are ectopically implanted.

  9. A photo-degradable supramolecular hydrogel for selective delivery of microRNA into 3D-cultured cells.

    PubMed

    Zhou, Zhengquan; Yi, Qikun; Xia, Tingting; Yin, Wencui; Kadi, Adnan A; Li, Jinbo; Zhang, Yan

    2017-03-08

    Multi-functional supramolecular hydrogels have emerged as smart biomaterials for diverse biomedical applications. Here we report a multi-functional supramolecular hydrogel formed by the conjugate of the bioactive GRGDS peptide with biaryltetrazole that is the substrate of photo-click reaction. The hydrogel was used as a biocompatible matrix to encapsulate live cells for 3D culture. The presence of the RGD epitope in the hydrogelator enhanced the interaction of the nanofiber with integrin over-expressing cells, which resulted in the selective enhancement in the miRNA delivery into the encapsulated U87 cells. The intramolecular photo-click reaction of the biaryltetrazole moiety in the hydrogelator leads to a sensitive photo-response of the hydrogel, which allowed photo-degradation of the hydrogel for release of the encapsulated live cells for further bio-assay of the intracellular species.

  10. Controlled release of TGF-beta 1 from RADA self-assembling peptide hydrogel scaffolds

    PubMed Central

    Zhou, Ao; Chen, Shuo; He, Bin; Zhao, Weikang; Chen, Xiaojun; Jiang, Dianming

    2016-01-01

    Bioactive mediators, cytokines, and chemokines have an important role in regulating and optimizing the synergistic action of materials, cells, and cellular microenvironments for tissue engineering. RADA self-assembling peptide hydrogels have been proved to have an excellent ability to promote cell proliferation, wound healing, tissue repair, and drug delivery. Here, we report that D-RADA16 and L-RADA16-RGD self-assembling peptides can form stable second structure and hydrogel scaffolds, affording the slow release of growth factor (transforming growth factor cytokine-beta 1 [TGF-beta 1]). In vitro tests demonstrated that the plateau release amount can be obtained till 72 hours. Moreover, L-RADA16, D-RADA16, and L-RADA16-RGD self-assembling peptide hydrogels containing TGF-beta 1 were used for 3D cell culture of bone mesenchymal stem cells of rats for 2 weeks. The results revealed that these three RADA16 peptide hydrogels had a significantly favorable influence on proliferation of bone mesenchymal stem cells and hold some promise in slow and sustained release of growth factor. PMID:27703332

  11. Preparation and mechanical property of a novel 3D porous magnesium scaffold for bone tissue engineering.

    PubMed

    Zhang, Xue; Li, Xiao-Wu; Li, Ji-Guang; Sun, Xu-Dong

    2014-09-01

    Porous magnesium has been recently recognized as a biodegradable metal for bone substitute applications. A novel porous Mg scaffold with three-dimensional (3D) interconnected pores and with a porosity of 33-54% was produced by the fiber deposition hot pressing (FDHP) technology. The microstructure and morphologies of the porous Mg scaffold were characterized by scanning electron microscopy (SEM), and the effects of porosities on the microstructure and mechanical properties of the porous Mg were investigated. Experimental results indicate that the measured Young's modulus and compressive strength of the Mg scaffold are ranged in 0.10-0.37 GPa, and 11.1-30.3 MPa, respectively, which are fairly comparable to those of cancellous bone. Such a porous Mg scaffold having a 3D interconnected network structure has the potential to be used in bone tissue engineering.

  12. Towards the Development of Smart 3D "gated scaffolds" for on-command delivery.

    PubMed

    Mas, Núria; Arcos, Daniel; Polo, Lorena; Aznar, Elena; Sánchez-Salcedo, Sandra; Sancenón, Félix; García, Ana; Marcos, M Dolores; Baeza, Alejandro; Vallet-Regí, María; Martínez-Máñez, Ramón

    2014-12-10

    A new approach towards the design of "gated scaffolds" based on the combination of capped mesoporous silica nanoparticles (MSNs) with porous biomaterials is reported. Using this approach, a 3D gelatin-based scaffold able to selectively deliver cargo in the presence of an APase enzyme is prepared and tested. This new design opens up the possibility of developing new smart biomaterials with advanced drug-delivery features.

  13. 3D printed PLA-based scaffolds: a versatile tool in regenerative medicine.

    PubMed

    Serra, Tiziano; Mateos-Timoneda, Miguel A; Planell, Josep A; Navarro, Melba

    2013-10-01

    Rapid prototyping (RP), also known as additive manufacturing (AM), has been well received and adopted in the biomedical field. The capacity of this family of techniques to fabricate customized 3D structures with complex geometries and excellent reproducibility has revolutionized implantology and regenerative medicine. In particular, nozzle-based systems allow the fabrication of high-resolution polylactic acid (PLA) structures that are of interest in regenerative medicine. These 3D structures find interesting applications in the regenerative medicine field where promising applications including biodegradable templates for tissue regeneration purposes, 3D in vitro platforms for studying cell response to different scaffolds conditions and for drug screening are considered among others. Scaffolds functionality depends not only on the fabrication technique, but also on the material used to build the 3D structure, the geometry and inner architecture of the structure, and the final surface properties. All being crucial parameters affecting scaffolds success. This Commentary emphasizes the importance of these parameters in scaffolds' fabrication and also draws the attention toward the versatility of these PLA scaffolds as a potential tool in regenerative medicine and other medical fields.

  14. 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.

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

    NASA Astrophysics Data System (ADS)

    Farkas, Balázs; Dante, Silvia; Brandi, Fernando

    2017-01-01

    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.

  16. Stem cells catalyze cartilage formation by neonatal articular chondrocytes in 3D biomimetic hydrogels

    NASA Astrophysics Data System (ADS)

    Lai, Janice H.; Kajiyama, Glen; Smith, Robert Lane; Maloney, William; Yang, Fan

    2013-12-01

    Cartilage loss is a leading cause of disability among adults and effective therapy remains elusive. Neonatal chondrocytes (NChons) are an attractive allogeneic cell source for cartilage repair, but their clinical translation has been hindered by scarce donor availability. Here we examine the potential for catalyzing cartilage tissue formation using a minimal number of NChons by co-culturing them with adipose-derived stem cells (ADSCs) in 3D hydrogels. Using three different co-culture models, we demonstrated that the effects of co-culture on cartilage tissue formation are dependent on the intercellular distance and cell distribution in 3D. Unexpectedly, increasing ADSC ratio in mixed co-culture led to increased synergy between NChons and ADSCs, and resulted in the formation of large neocartilage nodules. This work raises the potential of utilizing stem cells to catalyze tissue formation by neonatal chondrocytes via paracrine signaling, and highlights the importance of controlling cell distribution in 3D matrices to achieve optimal synergy.

  17. Development of a 3D polymer reinforced calcium phosphate cement scaffold for cranial bone tissue engineering

    NASA Astrophysics Data System (ADS)

    Alge, Daniel L.

    The repair of critical-sized cranial bone defects represents an important clinical challenge. The limitations of autografts and alloplastic materials make a bone tissue engineering strategy desirable, but success depends on the development of an appropriate scaffold. Key scaffold properties include biocompatibility, osteoconductivity, sufficient strength to maintain its structure, and resorbability. Furthermore, amenability to rapid prototyping fabrication methods is desirable, as these approaches offer precise control over scaffold architecture and have the potential for customization. While calcium phosphate cements meet many of these criteria due to their composition and their injectability, which can be leveraged for scaffold fabrication via indirect casting, their mechanical properties are a major limitation. Thus, the overall goal of this work was to develop a 3D polymer reinforced calcium phosphate cement scaffold for use in cranial bone tissue engineering. Dicalcium phosphate dihydrate (DCPD) setting cements are of particular interest because of their excellent resorbability. We demonstrated for the first time that DCPD cement can be prepared from monocalcium phosphate monohydrate (MCPM)/hydroxyapatite (HA) mixtures. However, subsequent characterization revealed that MCPM/HA cements rapidly convert to HA during degradation, which is undesirable and led us to choose a more conventional formulation for scaffold fabrication. In addition, we developed a novel method for calcium phosphate cement reinforcement that is based on infiltrating a pre-set cement structure with a polymer, and then crosslinking the polymer in situ. Unlike prior methods of cement reinforcement, this method can be applied to the reinforcement of 3D scaffolds fabricated by indirect casting. Using our novel method, composites of poly(propylene fumarate) (PPF) reinforced DCPD were prepared and demonstrated as excellent candidate scaffold materials, as they had increased strength and ductility

  18. 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.

  19. Relevance of PEG in PLA-based blends for tissue engineering 3D-printed scaffolds.

    PubMed

    Serra, Tiziano; Ortiz-Hernandez, Monica; Engel, Elisabeth; Planell, Josep A; Navarro, Melba

    2014-05-01

    Achieving high quality 3D-printed structures requires establishing the right printing conditions. Finding processing conditions that satisfy both the fabrication process and the final required scaffold properties is crucial. This work stresses the importance of studying the outcome of the plasticizing effect of PEG on PLA-based blends used for the fabrication of 3D-direct-printed scaffolds for tissue engineering applications. For this, PLA/PEG blends with 5, 10 and 20% (w/w) of PEG and PLA/PEG/bioactive CaP glass composites were processed in the form of 3D rapid prototyping scaffolds. Surface analysis and differential scanning calorimetry revealed a rearrangement of polymer chains and a topography, wettability and elastic modulus increase of the studied surfaces as PEG was incorporated. Moreover, addition of 10 and 20% PEG led to non-uniform 3D structures with lower mechanical properties. In vitro degradation studies showed that the inclusion of PEG significantly accelerated the degradation rate of the material. Results indicated that the presence of PEG not only improves PLA processing but also leads to relevant surface, geometrical and structural changes including modulation of the degradation rate of PLA-based 3D printed scaffolds.

  20. 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.

  1. 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.

  2. 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.

  3. 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

  4. 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.

  5. 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

  6. Role of bioactive 3D hybrid fibrous scaffolds on mechanical behavior and spatiotemporal osteoblast gene expression.

    PubMed

    Allo, Bedilu A; Lin, Shigang; Mequanint, Kibret; Rizkalla, Amin S

    2013-08-14

    Three-dimensional (3D) bioactive organic-inorganic (O/I) hybrid fibrous scaffolds are attractive extracellular matrix (ECM) surrogates for bone tissue engineering. With the aim of regulating osteoblast gene expression in 3D, a new class of hybrid fibrous scaffolds with two distinct fiber diameters (260 and 600 nm) and excellent physico-mechanical properties were fabricated from tertiary (SiO2-CaO-P2O5) bioactive glass (BG) and poly (ε-caprolactone) (PCL) by in situ sol-gel and electrospinning process. The PCL/BG hybrid fibrous scaffolds exhibited accelerated wetting properties, enhanced pore sizes and porosity, and superior mechanical properties that were dependent on fiber diameter. Contrary to control PCL fibrous scaffolds that were devoid of bonelike apatite particles, incubating PCL/BG hybrid fibrous scaffolds in simulated body fluid (SBF) revealed bonelike apatite deposition. Osteoblast cells cultured on PCL/BG hybrid fibrous scaffolds spread with multiple attachments and actively proliferated suggesting that the low temperature in situ sol-gel and electrospinning process did not have a detrimental effect. Targeted bone-associated gene expressions by rat calvarial osteoblasts seeded on these hybrid scaffolds demonstrated remarkable spatiotemporal gene activation. Transcriptional-level gene expressions for alkaline phosphatase (ALP), osteopontin (OPN), bone sialoprotein (BSP), and osteocalcin (OCN) were significantly higher on the hybrid fibrous scaffolds (p < 0.001) that were largely dependent on fiber diameter compared. Taken together, our results suggest that PCL/BG fibrous scaffolds may accelerate bone formation by providing a favorable microenvironment.

  7. 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.

  8. Engineering multi-layered skeletal muscle tissue by using 3D microgrooved collagen scaffolds.

    PubMed

    Chen, Shangwu; Nakamoto, Tomoko; Kawazoe, Naoki; Chen, Guoping

    2015-12-01

    Preparation of three-dimensional (3D) micropatterned porous scaffolds remains a great challenge for engineering of highly organized tissues such as skeletal muscle tissue and cardiac tissue. Two-dimensional (2D) micropatterned surfaces with periodic features (several nanometers to less than 100 μm) are commonly used to guide the alignment of muscle myoblasts and myotubes and lead to formation of pre-patterned cell sheets. However, cell sheets from 2D patterned surfaces have limited thickness, and harvesting the cell sheets for implantation is inconvenient and can lead to less alignment of myotubes. 3D micropatterned scaffolds can promote cell alignment and muscle tissue formation. In this study, we developed a novel type of 3D porous collagen scaffolds with concave microgrooves that mimic muscle basement membrane to engineer skeletal muscle tissue. Highly aligned and multi-layered muscle bundle tissues were engineered by controlling the size of microgrooves and cell seeding concentration. Myoblasts in the engineered muscle tissue were well-aligned and had high expression of myosin heavy chain and synthesis of muscle extracellular matrix. The microgrooved collagen scaffolds could be used to engineer organized multi-layered muscle tissue for implantation to repair/restore the function of diseased tissues or be used to investigate the cell-cell interaction in 3D microscale topography.

  9. Hydroxyapatite scaffolds for bone tissue engineering made by 3D printing.

    PubMed

    Leukers, Barbara; Gülkan, Hülya; Irsen, Stephan H; Milz, Stefan; Tille, Carsten; Schieker, Matthias; Seitz, Hermann

    2005-12-01

    Nowadays, there is a significant need for synthetic bone replacement materials used in bone tissue engineering (BTE). Rapid prototyping and especially 3D printing is a suitable technique to create custom implants based on medical data sets. 3D printing allows to fabricate scaffolds based on Hydroxyapatite with complex internal structures and high resolution. To determine the in vitro behaviour of cells cultivated on the scaffolds, we designed a special test-part. MC3T3-E1 cells were seeded on the scaffolds and cultivated under static and dynamic setups. Histological evaluation was carried out to characterise the cell ingrowth. In summary, the dynamic cultivation method lead to a stronger population compared to the static cultivation method. The cells proliferated deep into the structure forming close contact to Hydroxyapatite granules.

  10. Investigation of osteoblast cells behavior in polymeric 3D micropatterned scaffolds using digital holographic microscopy.

    PubMed

    Mihailescu, M; Popescu, R C; Matei, A; Acasandrei, A; Paun, I A; Dinescu, M

    2014-08-01

    The effect of micropatterned polymeric scaffolds on the features of the cultured cells at different time intervals after seeding was investigated by digital holographic microscopy. Both parallel and perpendicular walls, with different heights, were fabricated using two-photon lithography on photopolymers. The walls were subsequently coated with polypyrrole-based thin films using the matrix assisted pulsed laser evaporation technique. Osteoblast-like cells, MG-63 line, were cultured on these polymeric 3D micropatterned scaffolds. To analyze these scaffolds with/without cultured cells, an inverted digital holographic microscope, which provides 3D images, was used. Information about the samples' refractive indices and heights was obtained from the phase shift introduced in the optical path. Characteristics of cell adhesion, alignment, orientation, and morphology as a function of the wall heights and time from seeding were highlighted.

  11. Engineering EMT using 3D micro-scaffold to promote hepatic functions for drug hepatotoxicity evaluation.

    PubMed

    Wang, Jingyu; Chen, Fengling; Liu, Longwei; Qi, Chunxiao; Wang, Bingjie; Yan, Xiaojun; Huang, Chenyu; Hou, Wei; Zhang, Michael Q; Chen, Yang; Du, Yanan

    2016-06-01

    Accompanied by decreased hepatic functions, epithelial-mesenchymal transition (EMT) was observed in two dimensional (2D) cultured hepatocytes with elongated morphology, loss of polarity and weakened cell-cell interaction, while upgrading to 3D culture has been considered as significant improvement of its 2D counterpart for hepatocyte maintenance. Here we hypothesize that 3D culture enhances hepatic functions through regulating the EMT status. Biomaterial-engineered EMT was achieved by culturing HepaRG as 3D spheroids (SP-3D) or 3D stretched cells (ST-3D) in non-adherent and adherent micro-scaffold respectively. In SP-3D, constrained EMT of HepaRG, a hepatic stem cell line, as represented by increased epithelial markers and decreased mesenchymal markers, was echoed by improved hepatic functions. To investigate the relationship between EMT status and hepatic functions, time-series RNA-Seq and gene network analysis were used for comparing different cell culture models, which identified histone deacetylases (HDACs) as key mediating factors. Protein analysis confirmed that high HDAC activity was correlated with high expression of Cadherin-1 (CDH1) and hepatic function genes, which were decreased upon HDAC inhibitor treatment in SP-3D, suggesting HDACs may play positive role in regulating EMT and hepatic functions. To illustrate the application of 3D micro-scaffold culture in drug safety evaluation, hepatotoxicity and metabolism assays of two hepatotoxins (i.e. N-acetyl-p-aminophenol and Doxorubicin) were performed and SP-3D showed more biomimetic toxicity response, indicating regulation of EMT as a vital consideration in designing 3D hepatocyte culture configuration.

  12. 3D scaffold with effective multidrug sequential release against bacteria biofilm.

    PubMed

    García-Alvarez, Rafaela; Izquierdo-Barba, Isabel; Vallet-Regí, María

    2017-02-01

    Bone infection is a feared complication following surgery or trauma that remains as an extremely difficult disease to deal with. So far, the outcome of therapy could be improved with the design of 3D implants, which combine the merits of osseous regeneration and local multidrug therapy so as to avoid bacterial growth, drug resistance and the feared side effects. Herein, hierarchical 3D multidrug scaffolds based on nanocomposite bioceramic and polyvinyl alcohol (PVA) prepared by rapid prototyping with an external coating of gelatin-glutaraldehyde (Gel-Glu) have been fabricated. These 3D scaffolds contain three antimicrobial agents (rifampin, levofloxacin and vancomycin), which have been localized in different compartments of the scaffold to obtain different release kinetics and more effective combined therapy. Levofloxacin was loaded into the mesopores of nanocomposite bioceramic part, vancomycin was localized into PVA biopolymer part and rifampin was loaded in the external coating of Gel-Glu. The obtained results show an early and fast release of rifampin followed by sustained and prolonged release of vancomycin and levofloxacin, respectively, which are mainly governed by the progressive in vitro degradability rate of these scaffolds. This combined therapy is able to destroy Gram-positive and Gram-negative bacteria biofilms as well as inhibit the bacteria growth. In addition, these multifunctional scaffolds exhibit excellent bioactivity as well as good biocompatibility with complete cell colonization of preosteoblast in the entire surface, ensuring good bone regeneration. These findings suggest that these hierarchical 3D multidrug scaffolds are promising candidates as platforms for local bone infection therapy.

  13. Hydrogel based injectable scaffolds for cardiac tissue regeneration.

    PubMed

    Radhakrishnan, Janani; Krishnan, Uma Maheswari; Sethuraman, Swaminathan

    2014-01-01

    Tissue engineering promises to be an effective strategy that can overcome the lacuna existing in the current pharmacological and interventional therapies and heart transplantation. Heart failure continues to be a major contributor to the morbidity and mortality across the globe. This may be attributed to the limited regeneration capacity after the adult cardiomyocytes are terminally differentiated or injured. Various strategies involving acellular scaffolds, stem cells, and combinations of stem cells, scaffolds and growth factors have been investigated for effective cardiac tissue regeneration. Recently, injectable hydrogels have emerged as a potential candidate among various categories of biomaterials for cardiac tissue regeneration due to improved patient compliance and facile administration via minimal invasive mode that treats complex infarction. This review discusses in detail on the advances made in the field of injectable materials for cardiac tissue engineering highlighting their merits over their preformed counterparts.

  14. In vivo bone response to 3D periodic hydroxyapatite scaffolds assembled by direct ink writing.

    PubMed

    Simon, Joshua L; Michna, Sarah; Lewis, Jennifer A; Rekow, E Dianne; Thompson, Van P; Smay, James E; Yampolsky, Andrew; Parsons, J Russell; Ricci, John L

    2007-12-01

    The in vivo bone response of 3D periodic hydroxyapatite (HA) scaffolds is investigated. Two groups of HA scaffolds (11 mm diameter x 3.5 mm thick) are fabricated by direct-write assembly of a concentrated HA ink. The scaffolds consist of cylindrical rods periodically arranged into four quadrants with varying separation distances between rods. In the first group, HA rods (250 microm in diameter) are patterned to create pore channels, whose areal dimensions are 250 x 250 microm(2) in quadrant 1, 250 x 500 microm(2) in quadrants 2 and 4, and 500 x 500 microm(2) in quadrant 3. In the second group, HA rods (400 microm in diameter) are patterned to create pore channels, whose areal dimensions of 500 x 500 microm(2) in quadrant 1, 500 x 750 microm(2) in quadrants 2 and 4, and 750 x 750 microm(2) in quadrant 3. Each group of scaffolds is partially densified by sintering at 1200 degrees C prior to being implanted bilaterally in trephine defects of skeletally mature New Zealand White rabbits. Their tissue response is evaluated at 8 and 16 weeks using micro-computed tomography, histology, and scanning electron microscopy. New trabecular bone is conducted rapidly and efficiently across substantial distances within these patterned 3D HA scaffolds. Our observations suggest that HA rods are first coated with a layer of new bone followed by subsequent scaffold infilling via outward and inward radial growth of the coated regions. Direct-write assembly of 3D periodic scaffolds composed of micro-porous HA rods arrayed to produce macro-pores that are size-matched to trabecular bone may represent an optimal strategy for bone repair and replacement structures.

  15. Bioinspired Tuning of Hydrogel Permeability-Rigidity Dependency for 3D Cell Culture

    NASA Astrophysics Data System (ADS)

    Lee, Min Kyung; Rich, Max H.; Baek, Kwanghyun; Lee, Jonghwi; Kong, Hyunjoon

    2015-03-01

    Hydrogels are being extensively used for three-dimensional immobilization and culture of cells in fundamental biological studies, biochemical processes, and clinical treatments. However, it is still a challenge to support viability and regulate phenotypic activities of cells in a structurally stable gel, because the gel becomes less permeable with increasing rigidity. To resolve this challenge, this study demonstrates a unique method to enhance the permeability of a cell-laden hydrogel while avoiding a significant change in rigidity of the gel. Inspired by the grooved skin textures of marine organisms, a hydrogel is assembled to present computationally optimized micro-sized grooves on the surface. Separately, a gel is engineered to preset aligned microchannels similar to a plant's vascular bundles through a uniaxial freeze-drying process. The resulting gel displays significantly increased water diffusivity with reduced changes of gel stiffness, exclusively when the microgrooves and microchannels are aligned together. No significant enhancement of rehydration is achieved when the microgrooves and microchannels are not aligned. Such material design greatly enhances viability and neural differentiation of stem cells and 3D neural network formation within the gel.

  16. 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

  17. MCF-7 Human Breast Cancer Cells Form Differentiated Microtissues in Scaffold-Free Hydrogels

    PubMed Central

    Vantangoli, Marguerite M.; Madnick, Samantha J.; Huse, Susan M.; Weston, Paula; Boekelheide, Kim

    2015-01-01

    Three-dimensional (3D) cultures are increasing in use because of their ability to represent in vivo human physiology when compared to monolayer two-dimensional (2D) cultures. When grown in 3D using scaffold-free agarose hydrogels, MCF-7 human breast cancer cells self-organize to form directionally-oriented microtissues that contain a luminal space, reminiscent of the in vivo structure of the mammary gland. When compared to MCF-7 cells cultured in 2D monolayer culture, MCF-7 microtissues exhibit increased mRNA expression of luminal epithelial markers keratin 8 and keratin 19 and decreased expression of basal marker keratin 14 and the mesenchymal marker vimentin. These 3D MCF-7 microtissues remain responsive to estrogens, as demonstrated by induction of known estrogen target mRNAs following exposure to 17β-estradiol. Culture of MCF-7 cells in scaffold-free conditions allows for the formation of more differentiated, estrogen-responsive structures that are a more relevant system for evaluation of estrogenic compounds than traditional 2D models. PMID:26267486

  18. Fabrication of 3D cell-laden hydrogel microstructures through photo-mold patterning.

    PubMed

    Occhetta, P; Sadr, N; Piraino, F; Redaelli, A; Moretti, M; Rasponi, M

    2013-09-01

    Native tissues are characterized by spatially organized three-dimensional (3D) microscaled units which functionally define cells-cells and cells-extracellular matrix interactions. The ability to engineer biomimetic constructs mimicking these 3D microarchitectures is subject to the control over cell distribution and organization. In the present study we introduce a novel protocol to generate 3D cell laden hydrogel micropatterns with defined size and shape. The method, named photo-mold patterning (PMP), combines hydrogel micromolding within polydimethylsiloxane (PDMS) stamps and photopolymerization through a recently introduced biocompatible ultraviolet (UVA) activated photoinitiator (VA-086). Exploiting PDMS micromolds as geometrical constraints for two methacrylated prepolymers (polyethylene glycol diacrylate and gelatin methacrylate), micrometrically resolved structures were obtained within a 3 min exposure to a low cost and commercially available UVA LED. The PMP was validated both on a continuous cell line (human umbilical vein endothelial cells expressing green fluorescent protein, HUVEC GFP) and on primary human bone marrow stromal cells (BMSCs). HUVEC GFP and BMSCs were exposed to 1.5% w/v VA-086 and UVA light (1 W, 385 nm, distance from sample = 5 cm). Photocrosslinking conditions applied during the PMP did not negatively affect cells viability or specific metabolic activity. Quantitative analyses demonstrated the potentiality of PMP to uniformly embed viable cells within 3D microgels, creating biocompatible and favorable environments for cell proliferation and spreading during a seven days' culture. PMP can thus be considered as a promising and cost effective tool for designing spatially accurate in vitro models and, in perspective, functional constructs.

  19. 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

    2017-03-22

    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.

  20. Synthesis and Characterization of Carboxymethylcellulose-Methacrylate Hydrogel Cell Scaffolds

    PubMed Central

    Reeves, Robert; Ribeiro, Andreia; Lombardo, Leonard; Boyer, Richard; Leach, Jennie B.

    2012-01-01

    Many carbohydrates pose advantages for tissue engineering applications due to their hydrophilicity, degradability, and availability of chemical groups for modification. For example, carboxymethylcellulose (CMC) is a water-soluble cellulose derivative that is degradable by cellulase. Though this enzyme is not synthesized by mammalian cells, cellulase and the fragments derived from CMC degradation are biocompatible. With this in mind, we created biocompatible, selectively degradable CMC-based hydrogels that are stable in routine culture, but degrade when exposed to exogenous cellulase. Solutions of CMC-methacrylate and polyethylene glycol dimethacrylate (PEG-DM) were co-crosslinked to form stable hydrogels; we found that greater CMC-methacrylate content resulted in increased gel swelling, protein diffusion and rates of degradation by cellulase, as well as decreased gel shear modulus. CMC-methacrylate/PEG-DM gels modified with the adhesive peptide RGD supported fibroblast adhesion and viability. We conclude that hydrogels based on CMC-methacrylate are suitable for bioengineering applications where selective degradability may be favorable, such as cell scaffolds or controlled release devices. PMID:22708058

  1. 3D printing of composite calcium phosphate and collagen scaffolds for bone regeneration.

    PubMed

    Inzana, Jason A; Olvera, Diana; Fuller, Seth M; Kelly, James P; Graeve, Olivia A; Schwarz, Edward M; Kates, Stephen L; Awad, Hani A

    2014-04-01

    Low temperature 3D printing of calcium phosphate scaffolds holds great promise for fabricating synthetic bone graft substitutes with enhanced performance over traditional techniques. Many design parameters, such as the binder solution properties, have yet to be optimized to ensure maximal biocompatibility and osteoconductivity with sufficient mechanical properties. This study tailored the phosphoric acid-based binder solution concentration to 8.75 wt% to maximize cytocompatibility and mechanical strength, with a supplementation of Tween 80 to improve printing. To further enhance the formulation, collagen was dissolved into the binder solution to fabricate collagen-calcium phosphate composites. Reducing the viscosity and surface tension through a physiologic heat treatment and Tween 80, respectively, enabled reliable thermal inkjet printing of the collagen solutions. Supplementing the binder solution with 1-2 wt% collagen significantly improved maximum flexural strength and cell viability. To assess the bone healing performance, we implanted 3D printed scaffolds into a critically sized murine femoral defect for 9 weeks. The implants were confirmed to be osteoconductive, with new bone growth incorporating the degrading scaffold materials. In conclusion, this study demonstrates optimization of material parameters for 3D printed calcium phosphate scaffolds and enhancement of material properties by volumetric collagen incorporation via inkjet printing.

  2. 3D Printing of Composite Calcium Phosphate and Collagen Scaffolds for Bone Regeneration

    PubMed Central

    Inzana, Jason A.; Olvera, Diana; Fuller, Seth M.; Kelly, James P.; Graeve, Olivia A.; Schwarz, Edward M.; Kates, Stephen L.; Awad, Hani A.

    2014-01-01

    Low temperature 3D printing of calcium phosphate scaffolds holds great promise for fabricating synthetic bone graft substitutes with enhanced performance over traditional techniques. Many design parameters, such as the binder solution properties, have yet to be optimized to ensure maximal biocompatibility and osteoconductivity with sufficient mechanical properties. This study tailored the phosphoric acid-based binder solution concentration to 8.75 wt% to maximize cytocompatibility and mechanical strength, with a supplementation of Tween 80 to improve printing. To further enhance the formulation, collagen was dissolved into the binder solution to fabricate collagen-calcium phosphate composites. Reducing the viscosity and surface tension through a physiologic heat treatment and Tween 80, respectively, enabled reliable thermal inkjet printing of the collagen solutions. Supplementing the binder solution with 1–2 wt% collagen significantly improved maximum flexural strength and cell viability. To assess the bone healing performance, we implanted 3D printed scaffolds into a critically sized murine femoral defect for 9 weeks. The implants were confirmed to be osteoconductive, with new bone growth incorporating the degrading scaffold materials. In conclusion, this study demonstrates optimization of material parameters for 3D printed calcium phosphate scaffolds and enhancement of material properties by volumetric collagen incorporation via inkjet printing. PMID:24529628

  3. 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).

  4. Two-photon polymerization microfabrication of hydrogels: an advanced 3D printing technology for tissue engineering and drug delivery.

    PubMed

    Xing, Jin-Feng; Zheng, Mei-Ling; Duan, Xuan-Ming

    2015-08-07

    3D printing technology has attracted much attention due to its high potential in scientific and industrial applications. As an outstanding 3D printing technology, two-photon polymerization (TPP) microfabrication has been applied in the fields of micro/nanophotonics, micro-electromechanical systems, microfluidics, biomedical implants and microdevices. In particular, TPP microfabrication is very useful in tissue engineering and drug delivery due to its powerful fabrication capability for precise microstructures with high spatial resolution on both the microscopic and the nanometric scale. The design and fabrication of 3D hydrogels widely used in tissue engineering and drug delivery has been an important research area of TPP microfabrication. The resolution is a key parameter for 3D hydrogels to simulate the native 3D environment in which the cells reside and the drug is controlled to release with optimal temporal and spatial distribution in vitro and in vivo. The resolution of 3D hydrogels largely depends on the efficiency of TPP initiators. In this paper, we will review the widely used photoresists, the development of TPP photoinitiators, the strategies for improving the resolution and the microfabrication of 3D hydrogels.

  5. 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.

  6. A 3D Fibrous Scaffold Inducing Tumoroids: A Platform for Anticancer Drug Development

    PubMed Central

    Girard, Yvonne K.; Wang, Chunyan; Ravi, Sowndharya; Howell, Mark C.; Mallela, Jaya; Alibrahim, Mahmoud; Green, Ryan; Hellermann, Gary; Mohapatra, Shyam S.; Mohapatra, Subhra

    2013-01-01

    The development of a suitable three dimensional (3D) culture system for anticancer drug development remains an unmet need. Despite progress, a simple, rapid, scalable and inexpensive 3D-tumor model that recapitulates in vivo tumorigenesis is lacking. Herein, we report on the development and characterization of a 3D nanofibrous scaffold produced by electrospinning a mixture of poly(lactic-co-glycolic acid) (PLGA) and a block copolymer of polylactic acid (PLA) and mono-methoxypolyethylene glycol (mPEG) designated as 3P. Cancer cells cultured on the 3P scaffold formed tight irregular aggregates similar to in vivo tumors, referred to as tumoroids that depended on the topography and net charge of the scaffold. 3P scaffolds induced tumor cells to undergo the epithelial-to-mesenchymal transition (EMT) as demonstrated by up-regulation of vimentin and loss of E-cadherin expression. 3P tumoroids showed higher resistance to anticancer drugs than the same tumor cells grown as monolayers. Inhibition of ERK and PI3K signal pathways prevented EMT and reduced tumoroid formation, diameter and number. Fine needle aspirates, collected from tumor cells implanted in mice when cultured on 3P scaffolds formed tumoroids, but showed decreased sensitivity to anticancer drugs, compared to tumoroids formed by direct seeding. These results show that 3P scaffolds provide an excellent platform for producing tumoroids from tumor cell lines and from biopsies and that the platform can be used to culture patient biopsies, test for anticancer compounds and tailor a personalized cancer treatment. PMID:24146752

  7. 3D printing scaffold coupled with low level light therapy for neural tissue regeneration.

    PubMed

    Zhu, Wei; George, Jonathan; Sorger, Volker; Zhang, Lijie

    2017-03-28

    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 day culture. Over culturing time of 14 day 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.

  8. 3-D Intestinal Scaffolds for Evaluating the Therapeutic Potential of Probiotics

    PubMed Central

    2015-01-01

    Biomimetic in vitro intestinal models are becoming useful tools for studying host–microbial interactions. In the past, these models have typically been limited to simple cultures on 2-D scaffolds or Transwell inserts, but it is widely understood that epithelial cells cultured in 3-D environments exhibit different phenotypes that are more reflective of native tissue, and that different microbial species will preferentially adhere to select locations along the intestinal villi. We used a synthetic 3-D tissue scaffold with villous features that could support the coculture of epithelial cell types with select bacterial populations. Our end goal was to establish microbial niches along the crypt–villus axis in order to mimic the natural microenvironment of the small intestine, which could potentially provide new insights into microbe-induced intestinal disorders, as well as enabling targeted probiotic therapies. We recreated the surface topography of the small intestine by fabricating a biodegradable and biocompatible villous scaffold using poly lactic-glycolic acid to enable the culture of Caco-2 with differentiation along the crypt–villus axis in a similar manner to native intestines. This was then used as a platform to mimic the adhesion and invasion profiles of both Salmonella and Pseudomonas, and assess the therapeutic potential of Lactobacillus and commensal Escherichia coli in a 3-D setting. We found that, in a 3-D environment, Lactobacillus is more successful at displacing pathogens, whereas Nissle is more effective at inhibiting pathogen adhesion. PMID:24798584

  9. A biomimetic hydrogel based on methacrylated dextran-graft-lysine and gelatin for 3D smooth muscle cell culture.

    PubMed

    Liu, Yunxiao; Chan-Park, Mary B

    2010-02-01

    Many synthetic hydrogels for cell encapsulation have hitherto been based on polyethylene glycol which is non-natural, non-biodegradable and only terminal-functionalizable, all of which are drawbacks for tissue engineering or cell delivery. The polysaccharide dextran is also highly hydrophilic but biodegradable and pendant-functionalizable and more closely resembles glycosaminoglycans to mimic the natural extracellular matrix. This study reports synthesis of a methacrylate and lysine functionalized dextran and development of hydrogel composite systems based on this material and methacrylamide modified gelatin. The mechanical stiffness and degree of swelling of the hydrogels were varied by manipulation of the degree of functionalization of dextran and gelatin and concentration/composition of precursor solution. Human umbilical artery smooth muscle cells (SMCs) were encapsulated inside hydrogels during gel hardening with photopolymerization. Rapid cell spreading, extensive cellular network formation and high SMC proliferation occurred within softer hydrogels (with shear storage moduli ranging from 898 to 3124Pa). The encapsulated SMCs appear to be relatively contractile in the initial culture than on tissue culture polystyrene dish due to physical constraint imposed by the hydrogels but they become more synthetic with time possibly due to the inability of cells to reach confluence inside these cell-mediated degradable hydrogels. From the impressive cell proliferation and network formation, these new hydrogels combining polysaccharide and protein derivatives appear to be excellent candidates for further development as bioactive scaffolds for use in vascular tissue engineering and regeneration.

  10. 3D scaffold alters cellular response to graphene in a polymer composite for orthopedic applications.

    PubMed

    Kumar, Sachin; Azam, Dilkash; Raj, Shammy; Kolanthai, Elayaraja; Vasu, K S; Sood, A K; Chatterjee, Kaushik

    2016-05-01

    Graphene-based polymer nanocomposites are being studied for biomedical applications. Polymer nanocomposites can be processed differently to generate planar two-dimensional (2D) substrates and porous three-dimensional (3D) scaffolds. The objective of this work was to investigate potential differences in biological response to graphene in polymer composites in the form of 2D substrates and 3D scaffolds. Polycaprolactone (PCL) nanocomposites were prepared by incorporating 1% of graphene oxide (GO) and reduced graphene oxide (RGO). GO increased modulus and strength of PCL by 44 and 22% respectively, whereas RGO increased modulus and strength by 22 and 16%, respectively. RGO increased the water contact angle of PCL from 81° to 87° whereas GO decreased it to 77°. In 2D, osteoblast proliferated 15% more on GO composites than on PCL whereas RGO composite showed 17% decrease in cell proliferation, which may be attributed to differences in water wettability. In 3D, initial cell proliferation was markedly retarded in both GO (36% lower) and RGO (55% lower) composites owing to increased roughness due to the presence of the protruding nanoparticles. Cells organized into aggregates in 3D in contrast to spread and randomly distributed cells on 2D discs due to the macro-porous architecture of the scaffolds. Increased cell-cell contact and altered cellular morphology led to significantly higher mineralization in 3D. This study demonstrates that the cellular response to nanoparticles in composites can change markedly by varying the processing route and has implications for designing orthopedic implants such as resorbable fracture fixation devices and tissue scaffolds using such nanocomposites.

  11. The potential of encapsulating "raw materials" in 3D osteochondral gradient scaffolds.

    PubMed

    Mohan, Neethu; Gupta, Vineet; Sridharan, Banupriya; Sutherland, Amanda; Detamore, Michael S

    2014-04-01

    Scaffolds with continuous gradients in material composition and bioactive signals enable a smooth transition of properties at the interface. Components like chondroitin sulfate (CS) and bioactive glass (BG) in 3D scaffolds may serve as "raw materials" for synthesis of new extracellular matrix (ECM), and may have the potential to completely or partially replace expensive growth factors. We hypothesized that scaffolds with gradients of ECM components would enable superior performance of engineered constructs. Raw material encapsulation altered the appearance, structure, porosity, and degradation of the scaffolds. They allowed the scaffolds to better retain their 3D structure during culture and provided a buffering effect to the cells in culture. Following seeding of rat mesenchymal stem cells, there were several instances where glycosaminoglycan (GAG), collagen, or calcium contents were higher with the scaffolds containing raw materials (CS or BG) than with those containing transforming growth factor (TGF)-β3 or bone morphogenetic protein (BMP)-2. It was also noteworthy that a combination of both CS and TGF-β3 increased the secretion of collagen type II. Moreover, cells seeded in scaffolds containing opposing gradients of CS/TGF-β3 and BG/BMP-2 produced clear regional variations in the secretion of tissue-specific ECM. The study demonstrated raw materials have the potential to create a favorable microenvironment for cells; they can significantly enhance the synthesis of certain extracellular matrix (ECM) components when compared to expensive growth factors; either alone or in combination with growth factors they can enhance the secretion of tissue specific matrix proteins. Raw materials are promising candidates that can be used to either replace or be used in combination with growth factors. Success with raw materials in lieu of growth factors could have profound implications in terms of lower cost and faster regulatory approval for more rapid translation of

  12. 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.

  13. The effect of conjugating RGD into 3D alginate hydrogels on adipogenic differentiation of human adipose-derived stromal cells.

    PubMed

    Kang, Sun-Woong; Cha, Byung-Hyun; Park, Honghyun; Park, Kwang-Sook; Lee, Kuen Yong; Lee, Soo-Hong

    2011-05-12

    The effects of RGD peptide conjugation to alginate hydrogel on the adipogenic differentiation of ASCs was investigated. After 3 d of culture, RGD-modified alginate hydrogels significantly stimulated FAK and integrin α1 gene expressions and vinculin expression in ASCs. In addition, RGD-modified alginate hydrogels significantly enhanced the adipogenic differentiation of human ASCs to exhibit higher expression levels of oil red O staining and adipogenic genes compared to those of the control group (unmodified gels). These results suggest potential applications of RGD-modified alginate gels for adipose tissue regeneration.

  14. Fibrous hydrogel scaffolds with cells embedded in the fibers as a potential tissue scaffold for skin repair.

    PubMed

    Lin, Hsin-Yi; Peng, Chih-Wei; Wu, Wei-Wen

    2014-01-01

    A novel approach was undertaken to create a potential skin wound dressing. L929 fibroblast cells and alginate solution were simultaneously dispensed into a calcium chloride solution using a three-dimensional plotting system to manufacture a fibrous alginate scaffold with interconnected pores. These cells were then embedded in the alginate hydrogel fibers of the scaffold. A conventional scaffold with cells directly seeded on the fiber surface was used as a control. The encapsulated fibroblasts made using the co-dispensing method distributed homogeneously within the scaffold and showed the delayed formation of large cell aggregates compared to the control. The cells embedded in the hydrogel fibers also deposited more type I collagen in the extracellular matrix and expressed higher levels of fgf11 and fn1 than the control, indicating increased cellular proliferation and attachment. The results indicate that the novel co-dispensing alginate scaffold may promote skin regeneration better than the conventional directly-seeded scaffold.

  15. Impact of 3-D printed PLA- and chitosan-based scaffolds on human monocyte/macrophage responses: unraveling the effect of 3-D structures on inflammation.

    PubMed

    Almeida, Catarina R; Serra, Tiziano; Oliveira, Marta I; Planell, Josep A; Barbosa, Mário A; Navarro, Melba

    2014-02-01

    Recent studies have pointed towards a decisive role of inflammation in triggering tissue repair and regeneration, while at the same time it is accepted that an exacerbated inflammatory response may lead to rejection of an implant. Within this context, understanding and having the capacity to regulate the inflammatory response elicited by 3-D scaffolds aimed for tissue regeneration is crucial. This work reports on the analysis of the cytokine profile of human monocytes/macrophages in contact with biodegradable 3-D scaffolds with different surface properties, architecture and controlled pore geometry, fabricated by 3-D printing technology. Fabrication processes were optimized to create four different 3-D platforms based on polylactic acid (PLA), PLA/calcium phosphate glass or chitosan. Cytokine secretion and cell morphology of human peripheral blood monocytes allowed to differentiate on the different matrices were analyzed. While all scaffolds supported monocyte/macrophage adhesion and stimulated cytokine production, striking differences between PLA-based and chitosan scaffolds were found, with chitosan eliciting increased secretion of tumor necrosis factor (TNF)-α, while PLA-based scaffolds induced higher production of interleukin (IL)-6, IL-12/23 and IL-10. Even though the material itself induced the biggest differences, the scaffold geometry also impacted on TNF-α and IL-12/23 production, with chitosan scaffolds having larger pores and wider angles leading to a higher secretion of these pro-inflammatory cytokines. These findings strengthen the appropriateness of these 3-D platforms to study modulation of macrophage responses by specific parameters (chemistry, topography, scaffold architecture).

  16. Scaffold-free and scaffold-assisted 3D culture enhances differentiation of bone marrow stromal cells.

    PubMed

    Vidyasekar, Prasanna; Shyamsunder, Pavithra; Sahoo, Sanjeeb Kumar; Verma, Rama Shanker

    2016-02-01

    3D cultures of stem cells can preserve differentiation potential or increase the efficiency of methods that induce differentiation. Mouse bone marrow-derived stromal cells (BMSCs) were cultured in 3D as scaffold-free spheroids or "mesoid bodies" (MBs) and as aggregates on poly(lactic) acid microspheres (MB/MS). 3D cultures demonstrated viable cells, interaction on multiple planes, altered cell morphology, and the formation of structures similar to epithelial cell bridges. Cell proliferation was limited in suspension cultures of MB and MB/MS; however, cells regained proliferative capacity when transferred to flat substrates of tissue culture plates (TCPs). Expanded as monolayer, cells retained expression of Sca-1 and CD44 stem cell markers. 3D cultures demonstrated enhanced potential for adipogenic and osteogenic differentiation showing higher triglyceride accumulation and robust mineralization in comparison with TCP cultures. Enhanced and efficient adipogenesis was also observed in 3D cultures generated in a rotating cell culture system. Preservation of multilineage potential of BMSC was demonstrated in 5-azacytidine treatment of 3D cultures and TCP by expression of cardiac markers GATA4 and ACTA1 although functioning cardiomyocytes were not derived.

  17. Modulating mechanical behaviour of 3D-printed cartilage-mimetic PCL scaffolds: influence of molecular weight and pore geometry.

    PubMed

    Olubamiji, Adeola D; Izadifar, Zohreh; Si, Jennifer L; Cooper, David M L; Eames, B Frank; Chen, Daniel X B

    2016-06-22

    Three-dimensional (3D)-printed poly(ε)-caprolactone (PCL)-based scaffolds are increasingly being explored for cartilage tissue engineering (CTE) applications. However, ensuring that the mechanical properties of these PCL-based constructs are comparable to that of articular cartilage that they are meant to regenerate is an area that has been under-explored. This paper presents the effects of PCL's molecular weight (MW) and scaffold's pore geometric configurations; strand size (SZ), strand spacing (SS), and strand orientation (SO), on mechanical properties of 3D-printed PCL scaffolds. The results illustrate that MW has significant effect on compressive moduli and yield strength of 3D-printed PCL scaffolds. Specifically, PCL with MW of 45 K was a more feasible choice for fabrication of visco-elastic, flexible and load-bearing PCL scaffolds. Furthermore, pore geometric configurations; SZ, SS, and SO, all significantly affect on tensile moduli of scaffolds. However, only SZ and SS have statistically significant effects on compressive moduli and porosity of these scaffolds. That said, inverse linear relationship was observed between porosity and mechanical properties of 3D-printed PCL scaffolds in Pearson's correlation test. Altogether, this study illustrates that modulating MW of PCL and pore geometrical configurations of the scaffolds enabled design and fabrication of PCL scaffolds with mechanical and biomimetic properties that better mimic mechanical behaviour of human articular cartilage. Thus, the modulated PCL scaffold proposed in this study is a framework that offers great potentials for CTE applications.

  18. A 3D Chemically Modified Graphene Hydrogel for Fast, Highly Sensitive, and Selective Gas Sensor

    PubMed Central

    Wu, Jin; Tao, Kai; Guo, Yuanyuan; Li, Zhong; Wang, Xiaotian; Luo, Zhongzhen; Du, Chunlei; Chen, Di; Norford, Leslie K.

    2016-01-01

    Reduced graphene oxide (RGO) has proved to be a promising candidate in high‐performance gas sensing in ambient conditions. However, trace detection of different kinds of gases with simultaneously high sensitivity and selectivity is challenging. Here, a chemiresistor‐type sensor based on 3D sulfonated RGO hydrogel (S‐RGOH) is reported, which can detect a variety of important gases with high sensitivity, boosted selectivity, fast response, and good reversibility. The NaHSO3 functionalized RGOH displays remarkable 118.6 and 58.9 times higher responses to NO2 and NH3, respectively, compared with its unmodified RGOH counterpart. In addition, the S‐RGOH sensor is highly responsive to volatile organic compounds. More importantly, the characteristic patterns on the linearly fitted response–temperature curves are employed to distinguish various gases for the first time. The temperature of the sensor is elevated rapidly by an imbedded microheater with little power consumption. The 3D S‐RGOH is characterized and the sensing mechanisms are proposed. This work gains new insights into boosting the sensitivity of detecting various gases by combining chemical modification and 3D structural engineering of RGO, and improving the selectivity of gas sensing by employing temperature dependent response characteristics of RGO for different gases. PMID:28331786

  19. Biomimetic Hydrogels Incorporating Polymeric Cell-Adhesive Peptide to Promote the 3D Assembly of Tumoroids

    PubMed Central

    Hao, Ying; Zerdoum, Aidan B.; Stuffer, Alexander J.; Rajasekaran, Ayyappan K.; Jia, Xinqiao

    2016-01-01

    Towards the goal of establishing physiologically relevant in vitro tumor models, we synthesized and characterized a biomimetic hydrogel using thiolated hyaluronic acid (HA-SH) and an acrylated copolymer carrying multiple copies of cell adhesive peptide (PolyRGD-AC). PolyRGD-AC was derived from a random copolymer of tert-butyl methacrylate (tBMA) and oligomeric (ethylene glycol) methacrylate (OEGMA), synthesized via atom transfer radical polymerization (ATRP). Acid hydrolysis of tert-butyl moieties revealed the carboxylates, through which acrylate groups were installed. Partial modification of the acrylate groups with a cysteine-containing RGD peptide generated PolyRGD-AC. When PolyRGD-AC was mixed with HA-SH under physiological conditions, a macroscopic hydrogel with an average elastic modulus of 630 Pa was produced. LNCaP prostate cancer cells encapsulated in HA-PolyRGD gels as dispersed single cells formed multicellular tumoroids by day 4 and reached an average diameter of ~95 μm by day 28. Cells in these structures were viable, formed cell-cell contacts through E-cadherin (E-CAD and displayed cortical organization of F-actin. Compared to the control gels prepared using PolyRDG, multivalent presentation of the RGD signal in the HA matrix increased cellular metabolism, promoted the development of larger tumoroids and enhanced the expression of E-CAD and integrins. Overall, hydrogels with multivalently immobilized RGD is a promising 3D culture platform for dissecting principles of tumorigenesis and for screening anticancer drugs. PMID:27723964

  20. Exploring natural silk protein sericin for regenerative medicine: an injectable, photoluminescent, cell-adhesive 3D hydrogel

    NASA Astrophysics Data System (ADS)

    Wang, Zheng; Zhang, Yeshun; Zhang, Jinxiang; Huang, Lei; Liu, Jia; Li, Yongkui; Zhang, Guozheng; Kundu, Subhas C.; Wang, Lin

    2014-11-01

    Sericin, a major component of silk, has a long history of being discarded as a waste during silk processing. The value of sericin for tissue engineering is underestimated and its potential application in regenerative medicine has just begun to be explored. Here we report the successful fabrication and characterization of a covalently-crosslinked 3D pure sericin hydrogel for delivery of cells and drugs. This hydrogel is injectable, permitting its implantation through minimally invasive approaches. Notably, this hydrogel is found to exhibit photoluminescence, enabling bioimaging and in vivo tracking. Moreover, this hydrogel system possesses excellent cell-adhesive capability, effectively promoting cell attachment, proliferation and long-term survival of various types of cells. Further, the sericin hydrogel releases bioactive reagents in a sustained manner. Additionally, this hydrogel demonstrates good elasticity, high porosity, and pH-dependent degradation dynamics, which are advantageous for this sericin hydrogel to serve as a delivery vehicle for cells and therapeutic drugs. With all these unique features, it is expected that this sericin hydrogel will have wide utility in the areas of tissue engineering and regenerative medicine.

  1. Exploring natural silk protein sericin for regenerative medicine: an injectable, photoluminescent, cell-adhesive 3D hydrogel.

    PubMed

    Wang, Zheng; Zhang, Yeshun; Zhang, Jinxiang; Huang, Lei; Liu, Jia; Li, Yongkui; Zhang, Guozheng; Kundu, Subhas C; Wang, Lin

    2014-11-20

    Sericin, a major component of silk, has a long history of being discarded as a waste during silk processing. The value of sericin for tissue engineering is underestimated and its potential application in regenerative medicine has just begun to be explored. Here we report the successful fabrication and characterization of a covalently-crosslinked 3D pure sericin hydrogel for delivery of cells and drugs. This hydrogel is injectable, permitting its implantation through minimally invasive approaches. Notably, this hydrogel is found to exhibit photoluminescence, enabling bioimaging and in vivo tracking. Moreover, this hydrogel system possesses excellent cell-adhesive capability, effectively promoting cell attachment, proliferation and long-term survival of various types of cells. Further, the sericin hydrogel releases bioactive reagents in a sustained manner. Additionally, this hydrogel demonstrates good elasticity, high porosity, and pH-dependent degradation dynamics, which are advantageous for this sericin hydrogel to serve as a delivery vehicle for cells and therapeutic drugs. With all these unique features, it is expected that this sericin hydrogel will have wide utility in the areas of tissue engineering and regenerative medicine.

  2. 3D Printing Bioceramic Porous Scaffolds with Good Mechanical Property and Cell Affinity

    PubMed Central

    Chang, Chih-Hao; Lin, Chih-Yang; Liu, Fwu-Hsing; Chen, Mark Hung-Chih; Lin, Chun-Pin; Ho, Hong-Nerng; Liao, Yunn-Shiuan

    2015-01-01

    Artificial bone grafting is widely used in current orthopedic surgery for bone defect problems. Unfortunately, surgeons remain unsatisfied with the current commercially available products. One of the major complaints is that these products cannot provide sufficient mechanical strength to support the human skeletal structure. In this study, we aimed to develop a bone scaffold with better mechanical property and good cell affinity by 3D printing (3DP) techniques. A self-developed 3D printer with laser-aided gelling (LAG) process was used to fabricate bioceramic scaffolds with inter-porous structures. To improve the mechanical property of the bioceramic parts after heating, CaCO3 was added to the silica ceramic slurry. CaCO3 was blended into a homogenous SiO2-sol dispersion at weight ratios varying from 0/100 to 5/95 to 9/91 (w/w). Bi-component CaCO3/SiO2-sol was prepared as a biocomposite for the 3DP scaffold. The well-mixed biocomposite was used to fabricate the bioceramic green part using the LAG method. The varied scaffolds were sintered at different temperatures ranging from 900 to 1500°C, and the mechanical property was subsequently analyzed. The scaffolds showed good property with the composite ratio of 5:95 CaCO3:SiO2 at a sintering temperature of 1300°C. The compressive strength was 47 MPa, and the porosity was 34%. The topography of the sintered 3DP bioceramic scaffold was examined by SEM, EDS and XRD. The silica bioceramic presented no cytotoxicity and good MG-63 osteoblast-like cell affinity, demonstrating good biocompatibility. Therefore, the new silica biocomposite is viable for fabricating 3DP bone bioceramics with improved mechanical property and good cell affinity. PMID:26618362

  3. 3D Printing Bioceramic Porous Scaffolds with Good Mechanical Property and Cell Affinity.

    PubMed

    Chang, Chih-Hao; Lin, Chih-Yang; Liu, Fwu-Hsing; Chen, Mark Hung-Chih; Lin, Chun-Pin; Ho, Hong-Nerng; Liao, Yunn-Shiuan

    2015-01-01

    Artificial bone grafting is widely used in current orthopedic surgery for bone defect problems. Unfortunately, surgeons remain unsatisfied with the current commercially available products. One of the major complaints is that these products cannot provide sufficient mechanical strength to support the human skeletal structure. In this study, we aimed to develop a bone scaffold with better mechanical property and good cell affinity by 3D printing (3DP) techniques. A self-developed 3D printer with laser-aided gelling (LAG) process was used to fabricate bioceramic scaffolds with inter-porous structures. To improve the mechanical property of the bioceramic parts after heating, CaCO3 was added to the silica ceramic slurry. CaCO3 was blended into a homogenous SiO2-sol dispersion at weight ratios varying from 0/100 to 5/95 to 9/91 (w/w). Bi-component CaCO3/SiO2-sol was prepared as a biocomposite for the 3DP scaffold. The well-mixed biocomposite was used to fabricate the bioceramic green part using the LAG method. The varied scaffolds were sintered at different temperatures ranging from 900 to 1500°C, and the mechanical property was subsequently analyzed. The scaffolds showed good property with the composite ratio of 5:95 CaCO3:SiO2 at a sintering temperature of 1300°C. The compressive strength was 47 MPa, and the porosity was 34%. The topography of the sintered 3DP bioceramic scaffold was examined by SEM, EDS and XRD. The silica bioceramic presented no cytotoxicity and good MG-63 osteoblast-like cell affinity, demonstrating good biocompatibility. Therefore, the new silica biocomposite is viable for fabricating 3DP bone bioceramics with improved mechanical property and good cell affinity.

  4. Patterned and functionalized nanofiber scaffolds in three-dimensional hydrogel constructs enhance neurite outgrowth and directional control

    NASA Astrophysics Data System (ADS)

    McMurtrey, Richard J.

    2014-12-01

    Objective. Neural tissue engineering holds incredible potential to restore functional capabilities to damaged neural tissue. It was hypothesized that patterned and functionalized nanofiber scaffolds could control neurite direction and enhance neurite outgrowth. Approach. A method of creating aligned electrospun nanofibers was implemented and fiber characteristics were analyzed using environmental scanning electron microscopy. Nanofibers were composed of polycaprolactone (PCL) polymer, PCL mixed with gelatin, or PCL with a laminin coating. Three-dimensional hydrogels were then integrated with embedded aligned nanofibers to support neuronal cell cultures. Microscopic images were captured at high-resolution in single and multi-focal planes with eGFP-expressing neuronal SH-SY5Y cells in a fluorescent channel and nanofiber scaffolding in another channel. Neuronal morphology and neurite tracking of nanofibers were then analyzed in detail. Main results. Aligned nanofibers were shown to enable significant control over the direction of neurite outgrowth in both two-dimensional (2D) and three-dimensional (3D) neuronal cultures. Laminin-functionalized nanofibers in 3D hyaluronic acid (HA) hydrogels enabled significant alignment of neurites with nanofibers, enabled significant neurite tracking of nanofibers, and significantly increased the distance over which neurites could extend. Specifically, the average length of neurites per cell in 3D HA constructs with laminin-functionalized nanofibers increased by 66% compared to the same laminin fibers on 2D laminin surfaces, increased by 59% compared to 2D laminin-coated surface without fibers, and increased by 1052% compared to HA constructs without fibers. Laminin functionalization of fibers also doubled average neurite length over plain PCL fibers in the same 3D HA constructs. In addition, neurites also demonstrated tracking directly along the fibers, with 66% of neurite lengths directly tracking laminin-coated fibers in 3D HA

  5. Laser 3D printing with sub-microscale resolution of porous elastomeric scaffolds for supporting human bone stem cells.

    PubMed

    Petrochenko, Peter E; Torgersen, Jan; Gruber, Peter; Hicks, Lucas A; Zheng, Jiwen; Kumar, Girish; Narayan, Roger J; Goering, Peter L; Liska, Robert; Stampfl, Jürgen; Ovsianikov, Aleksandr

    2015-04-02

    A reproducible method is needed to fabricate 3D scaffold constructs that results in periodic and uniform structures with precise control at sub-micrometer and micrometer length scales. In this study, fabrication of scaffolds by two-photon polymerization (2PP) of a biodegradable urethane and acrylate-based photoelastomer is demonstrated. This material supports 2PP processing with sub-micrometer spatial resolution. The high photoreactivity of the biophotoelastomer permits 2PP processing at a scanning speed of 1000 mm s(-1), facilitating rapid fabrication of relatively large structures (>5 mm(3)). These structures are custom printed for in vitro assay screening in 96-well plates and are sufficiently flexible to enable facile handling and transplantation. These results indicate that stable scaffolds with porosities of greater than 60% can be produced using 2PP. Human bone marrow stromal cells grown on 3D scaffolds exhibit increased growth and proliferation compared to smooth 2D scaffold controls. 3D scaffolds adsorb larger amounts of protein than smooth 2D scaffolds due to their larger surface area; the scaffolds also allow cells to attach in multiple planes and to completely infiltrate the porous scaffolds. The flexible photoelastomer material is biocompatible in vitro and is associated with facile handling, making it a viable candidate for further study of complex 3D-printed scaffolds.

  6. Differentiation of liver progenitor cell line to functional organotypic cultures in 3D nanofibrillar cellulose and hyaluronan-gelatin hydrogels.

    PubMed

    Malinen, Melina M; Kanninen, Liisa K; Corlu, Anne; Isoniemi, Helena M; Lou, Yan-Ru; Yliperttula, Marjo L; Urtti, Arto O

    2014-06-01

    Physiologically relevant hepatic cell culture models must be based on three-dimensional (3D) culture of human cells. However, liver cells are generally cultured in two-dimensional (2D) format that deviates from the normal in vivo morphology. We generated 3D culture environment for HepaRG liver progenitor cells using wood-derived nanofibrillar cellulose (NFC) and hyaluronan-gelatin (HG) hydrogels. Culture of undifferentiated HepaRG cells in NFC and HG hydrogels induced formation of 3D multicellular spheroids with apicobasal polarity and functional bile canaliculi-like structures, structural hallmarks of the liver tissue. Furthermore, hepatobiliary drug transporters, MRP2 and MDR1, were localized on the canalicular membranes of the spheroids and vectorial transport of fluorescent probes towards the biliary compartment was demonstrated. Cell culture in 3D hydrogel supported the mRNA expression of hepatocyte markers (albumin and CYP3A4), and metabolic activity of CYP3A4 in the HepaRG cell cultures. On the contrary, the 3D hydrogel cultures with pre-differentiated HepaRG cells showed decreasing expression of albumin and CYP3A4 transcripts as well as CYP3A4 activity. It is concluded that NFC and HG hydrogels expedite the hepatic differentiation of HepaRG liver progenitor cells better than the standard 2D culture environment. This was shown as improved cell morphology, expression and localization of hepatic markers, metabolic activity and vectorial transport. The NFC and HG hydrogels are promising materials for hepatic cell culture and tissue engineering.

  7. Interfacing polymeric scaffolds with primary pancreatic ductal adenocarcinoma cells to develop 3D cancer models

    PubMed Central

    Ricci, Claudio; Mota, Carlos; Moscato, Stefania; D’Alessandro, Delfo; Ugel, Stefano; Sartoris, Silvia; Bronte, Vincenzo; Boggi, Ugo; Campani, Daniela; Funel, Niccola; Moroni, Lorenzo; Danti, Serena

    2014-01-01

    We analyzed the interactions between human primary cells from pancreatic ductal adenocarcinoma (PDAC) and polymeric scaffolds to develop 3D cancer models useful for mimicking the biology of this tumor. Three scaffold types based on two biocompatible polymeric formulations, such as poly(vinyl alcohol)/gelatin (PVA/G) mixture and poly(ethylene oxide terephthalate)/poly(butylene terephthalate) (PEOT/PBT) copolymer, were obtained via different techniques, namely, emulsion and freeze-drying, compression molding followed by salt leaching, and electrospinning. In this way, primary PDAC cells interfaced with different pore topographies, such as sponge-like pores of different shape and size or nanofiber interspaces. The aim of this study was to investigate the influence played by the scaffold architecture over cancerous cell growth and function. In all scaffolds, primary PDAC cells showed good viability and synthesized tumor-specific metalloproteinases (MMPs) such as MMP-2, and MMP-9. However, only sponge-like pores, obtained via emulsion-based and salt leaching-based techniques allowed for an organized cellular aggregation very similar to the native PDAC morphological structure. Differently, these cell clusters were not observed on PEOT/PBT electrospun scaffolds. MMP-2 and MMP-9, as active enzymes, resulted to be increased in PVA/G and PEOT/PBT sponges, respectively. These findings suggested that spongy scaffolds supported the generation of pancreatic tumor models with enhanced aggressiveness. In conclusion, primary PDAC cells showed diverse behaviors while interacting with different scaffold types that can be potentially exploited to create stage-specific pancreatic cancer models likely to provide new knowledge on the modulation and drug susceptibility of MMPs. PMID:25482337

  8. Interfacing polymeric scaffolds with primary pancreatic ductal adenocarcinoma cells to develop 3D cancer models.

    PubMed

    Ricci, Claudio; Mota, Carlos; Moscato, Stefania; D'Alessandro, Delfo; Ugel, Stefano; Sartoris, Silvia; Bronte, Vincenzo; Boggi, Ugo; Campani, Daniela; Funel, Niccola; Moroni, Lorenzo; Danti, Serena

    2014-01-01

    We analyzed the interactions between human primary cells from pancreatic ductal adenocarcinoma (PDAC) and polymeric scaffolds to develop 3D cancer models useful for mimicking the biology of this tumor. Three scaffold types based on two biocompatible polymeric formulations, such as poly(vinyl alcohol)/gelatin (PVA/G) mixture and poly(ethylene oxide terephthalate)/poly(butylene terephthalate) (PEOT/PBT) copolymer, were obtained via different techniques, namely, emulsion and freeze-drying, compression molding followed by salt leaching, and electrospinning. In this way, primary PDAC cells interfaced with different pore topographies, such as sponge-like pores of different shape and size or nanofiber interspaces. The aim of this study was to investigate the influence played by the scaffold architecture over cancerous cell growth and function. In all scaffolds, primary PDAC cells showed good viability and synthesized tumor-specific metalloproteinases (MMPs) such as MMP-2, and MMP-9. However, only sponge-like pores, obtained via emulsion-based and salt leaching-based techniques allowed for an organized cellular aggregation very similar to the native PDAC morphological structure. Differently, these cell clusters were not observed on PEOT/PBT electrospun scaffolds. MMP-2 and MMP-9, as active enzymes, resulted to be increased in PVA/G and PEOT/PBT sponges, respectively. These findings suggested that spongy scaffolds supported the generation of pancreatic tumor models with enhanced aggressiveness. In conclusion, primary PDAC cells showed diverse behaviors while interacting with different scaffold types that can be potentially exploited to create stage-specific pancreatic cancer models likely to provide new knowledge on the modulation and drug susceptibility of MMPs.

  9. Hierarchical bioceramic scaffolds with 3D-plotted macropores and mussel-inspired surface nanolayers for stimulating osteogenesis.

    PubMed

    Xu, Mengchi; Zhai, Dong; Xia, Lunguo; Li, Hong; Chen, Shiyi; Fang, Bing; Chang, Jiang; Wu, Chengtie

    2016-07-14

    The hierarchical structure of biomaterials plays an important role in the process of tissue reconstruction and regeneration. 3D-plotted scaffolds have been widely used for bone tissue engineering due to their controlled macropore structure and mechanical properties. However, the lack of micro- or nano-structures on the strut surface of 3D-plotted scaffolds, especially for bioceramic scaffolds, limits their biological activity. Inspired by the adhesive versatility of mussels and the active ion-chelating capacity of polydopamine, we set out to prepare a hierarchical bioceramic scaffold with controlled macropores and mussel-inspired surface nanolayers by combining the 3D-plotting technique with the polydopamine/apatite hybrid strategy in order to synergistically accelerate the osteogenesis and angiogenesis. β-Tricalcium phosphate (TCP) scaffolds were firstly 3D-plotted and then treated in dopamine-Tris/HCl and dopamine-SBF solutions to obtain TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds, respectively. It was found that polydopamine/apatite hybrid nanolayers were formed on the surface of both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds and TCP-DOPA-SBF scaffolds induced apatite mineralization for the second time during the cell culture. As compared to TCP scaffolds, both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds significantly promoted the osteogenesis of bone marrow stromal cells (BMSCs) as well as the angiogenesis of human umbilical vein endothelial cells (HUVECs), and the TCP-DOPA-SBF group presented the highest in vitro osteogenic/angiogenic activity among the three groups. Furthermore, both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds significantly improved the formation of new bone in vivo as compared to TCP scaffolds without a nanostructured surface. Our results suggest that the utilization of a mussel-inspired Ca, P-chelated polydopamine nanolayer on 3D-plotted bioceramic scaffolds is a viable and effective strategy to construct a hierarchical structure for synergistically

  10. 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.

  11. Microfluidic assay of endothelial cell migration in 3D interpenetrating polymer semi-network HA-Collagen hydrogel.

    PubMed

    Jeong, Gi Seok; Kwon, Gu Han; Kang, Ah Ran; Jung, Bo Young; Park, Yongdoo; Chung, Seok; Lee, Sang-Hoon

    2011-08-01

    Cell migration through the extracellular matrix (ECM) is one of the key features for physiological and pathological processes such as angiogenesis, cancer metastasis, and wound healing. In particular, the quantitative assay of endothelial cell migration under the well-defined three dimensional (3D) microenvironment is important to analyze the angiogenesis mechanism. In this study, we report a microfluidic assay of endothelial cell sprouting and migration into an interpenetrating polymer semi-network HA-Collagen (SIPNs CH) hydrogel as ECM providing an enhanced in vivo mimicking 3D microenvironment to cells. The microfluidic chip could provide a well-controlled gradient of growth factor to cells, whereas the hydrogel could mimic a well-defined 3D microenvironment in vivo. (In addition/Furthermore, the microfluidic chip gives a well-controlled gradient of growth factor to cells) For this reason, three types of hydrogel, composed of semi-interpenetrating networks of collagen and hyaluronic acid were prepared, and firstly we proved the role of the hydrogel in endothelial cell migration. The diffusion property and swelling ratio of the hydrogel were characterized. It modulated the migration of endothelial cells in quantified manner, also being influenced by additional synthesis of Matrix metalloproteinase(MMP)-sensitive remodeling peptides and Arginine-glycine-lycinee (RGD) cell adhesion peptides. We successfully established a novel cell migration platform by changing major determinants such as ECM material under biochemical synthesis and under growth factor gradients in a microfluidic manner.

  12. Assessment of Photodynamic Inactivation against Periodontal Bacteria Mediated by a Chitosan Hydrogel in a 3D Gingival Model

    PubMed Central

    Peng, Po-Chun; Hsieh, Chien-Ming; Chen, Chueh-Pin; Tsai, Tsuimin; Chen, Chin-Tin

    2016-01-01

    Chitosan hydrogels containing hydroxypropyl methylcellulose (HPMC) and toluidine blue O were prepared and assessed for their mucoadhesive property and antimicrobial efficacy of photodynamic inactivation (PDI). Increased HPMC content in the hydrogels resulted in increased mucoadhesiveness. Furthermore, we developed a simple In Vitro 3D gingival model resembling the oral periodontal pocket to culture the biofilms of Staphylococcus aureus (S. aureus), Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans), and Porphyromonas gingivalis (P. gingivalis). The PDI efficacy of chitosan hydrogel was examined against periodontal biofilms cultured in this 3D gingival model. We found that the PDI effectiveness was limited due to leaving some of the innermost bacteria alive at the non-illuminated site. Using this 3D gingival model, we further optimized PDI procedures with various adjustments of light energy and irradiation sites. The PDI efficacy of the chitosan hydrogel against periodontal biofilms can significantly improve via four sides of irradiation. In conclusion, this study not only showed the clinical applicability of this chitosan hydrogel but also the importance of the light irradiation pattern in performing PDI for periodontal disease. PMID:27809278

  13. Direct 3D powder printing of biphasic calcium phosphate scaffolds for substitution of complex bone defects.

    PubMed

    Castilho, Miguel; Moseke, Claus; Ewald, Andrea; Gbureck, Uwe; Groll, Jürgen; Pires, Inês; Teßmar, Jörg; Vorndran, Elke

    2014-03-01

    The 3D printing technique based on cement powders is an excellent method for the fabrication of individual and complex bone substitutes even in the case of large defects. The outstanding bone remodeling capacity of biphasic calcium phosphates (BCPs) containing hydroxyapatite (HA) as well as tricalcium phosphate (TCP) in varying ratios makes the adaption of powder systems resulting in BCP materials to this fabrication technique a desirable aim. This study presents the synthesis and characterization of a novel powder system for the 3D printing process, intended for the production of complexly shaped BCP scaffolds by a hydraulic setting reaction of calcium carbonate and TCP with phosphoric acid. The HA/TCP ratio in the specimens could be tailored by the calcium/phosphate ratio of the starting powder. The scaffolds could be fabricated with a dimensional accuracy of >96.5% and a minimal macro pore size of 300 µm. Independent of the phase composition the printed specimens showed a microporosity of approximately 68%, while the compressive strength strongly depended on the chemical composition and increased with rising TCP content in the scaffolds to a maximum of 1.81 MPa. Post-treatment of the scaffolds with a polylactic-co-glycolic acid-solution enhanced the mechanical properties by a factor of 8. In vitro studies showed that all BCP scaffolds were cytocompatible and enhanced the cell viability as well as the cell proliferation, as compared with pure TCP. Cell proliferation is even better on BCP when compared to HA and cell viability is in a similar range on these materials.

  14. Additive manufactured polymeric 3D scaffolds with tailored surface topography influence mesenchymal stromal cells activity.

    PubMed

    Neves, Sara C; Mota, Carlos; Longoni, Alessia; Barrias, Cristina C; Granja, Pedro L; Moroni, Lorenzo

    2016-05-24

    Additive manufactured three-dimensional (3D) scaffolds with tailored surface topography constitute a clear advantage in tissue regeneration strategies to steer cell behavior. 3D fibrous scaffolds of poly(ethylene oxide terephthalate)/poly(butylene terephthalate) block copolymer presenting different fiber surface features were successfully fabricated by additive manufacturing combined with wet-spinning, in a single step, without any post-processing. The optimization of the processing parameters, mainly driven by different solvent/non-solvent combinations, led to four distinct scaffold types, with average surface roughness values ranging from 0.071 ± 0.012 μm to 1.950 ± 0.553 μm, average pore sizes in the x- and y-axis between 351.1 ± 33.6 μm and 396.1 ± 32.3 μm, in the z-axis between 36.5 ± 5.3 μm and 70.7 ± 8.8 μm, average fiber diameters between 69.4 ± 6.1 μm and 99.0 ± 9.4 μm, and porosity values ranging from 60.2 ± 0.8% to 71.7 ± 2.6%. Human mesenchymal stromal cells (hMSCs) cultured on these scaffolds adhered, proliferated, and produced endogenous extracellular matrix. The effect of surface roughness and topography on hMSCs differentiation was more evident for cells seeded at lower density, where the percentage of cells in direct contact with the surface was higher compared to more densely seeded scaffolds. Under osteogenic conditions, lower surface roughness values (0.227 ± 0.035 μm) had a synergistic effect on hMSCs behavior, while chondrogenesis was favored on rougher surfaces (1.950 ± 0.553 μm).

  15. Novel enzymatically cross-linked hyaluronan hydrogels support the formation of 3D neuronal networks.

    PubMed

    Broguiere, Nicolas; Isenmann, Luca; Zenobi-Wong, Marcy

    2016-08-01

    Hyaluronan (HA) is an essential component of the central nervous system's extracellular matrix and its high molecular weight (MW) form has anti-inflammatory and anti-fibrotic properties relevant for regenerative medicine. Here, we introduce a new hydrogel based on high MW HA which is cross-linked using the transglutaminase (TG) activity of the activated blood coagulation factor XIII (FXIIIa). These HA-TG gels have significant advantages for neural tissue engineering compared to previous HA gels. Due to their chemical inertness in the absence of FXIIIa, the material can be stored long-term, is stable in solution, and shows no cytotoxicity. The gelation is completely cell-friendly due to the specificity of the enzyme and the gelation rate can be tuned from seconds to hours at physiological pH and independently of stiffness. The gels are injectable, and attach covalently to fibrinogen and fibrin, two common bioactive components in in vitro tissue engineering, as well as proteins present in vivo, allowing the gels to covalently bind to brain or spinal cord defects. These optimal chemical and bioactive properties of HA-TG gels enabled the formation of 3D neuronal cultures of unprecedented performance, showing fast neurite outgrowth, axonal and dendritic speciation, strong synaptic connectivity in 3D networks, and rapidly-occurring and long-lasting coordinated electrical activity.

  16. Lipid Coated Microbubbles and Low Intensity Pulsed Ultrasound Enhance Chondrogenesis of Human Mesenchymal Stem Cells in 3D Printed Scaffolds

    PubMed Central

    Aliabouzar, Mitra; Zhang, Lijie Grace; Sarkar, Kausik

    2016-01-01

    Lipid-coated microbubbles are used to enhance ultrasound imaging and drug delivery. Here we apply these microbubbles along with low intensity pulsed ultrasound (LIPUS) for the first time to enhance proliferation and chondrogenic differentiation of human mesenchymal stem cells (hMSCs) in a 3D printed poly-(ethylene glycol)-diacrylate (PEG-DA) hydrogel scaffold. The hMSC proliferation increased up to 40% after 5 days of culture in the presence of 0.5% (v/v) microbubbles and LIPUS in contrast to 18% with LIPUS alone. We systematically varied the acoustic excitation parameters—excitation intensity, frequency and duty cycle—to find 30 mW/cm2, 1.5 MHz and 20% duty cycle to be optimal for hMSC proliferation. A 3-week chondrogenic differentiation results demonstrated that combining LIPUS with microbubbles enhanced glycosaminoglycan (GAG) production by 17% (5% with LIPUS alone), and type II collagen production by 78% (44% by LIPUS alone). Therefore, integrating LIPUS and microbubbles appears to be a promising strategy for enhanced hMSC growth and chondrogenic differentiation, which are critical components for cartilage regeneration. The results offer possibilities of novel applications of microbubbles, already clinically approved for contrast enhanced ultrasound imaging, in tissue engineering. PMID:27883051

  17. 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.

  18. Generation and transplantation of reprogrammed human neurons in the brain using 3D microtopographic scaffolds

    PubMed Central

    Carlson, Aaron L.; Bennett, Neal K.; Francis, Nicola L.; Halikere, Apoorva; Clarke, Stephen; Moore, Jennifer C.; Hart, Ronald P.; Paradiso, Kenneth; Wernig, Marius; Kohn, Joachim; Pang, Zhiping P.; Moghe, Prabhas V.

    2016-01-01

    Cell replacement therapy with human pluripotent stem cell-derived neurons has the potential to ameliorate neurodegenerative dysfunction and central nervous system injuries, but reprogrammed neurons are dissociated and spatially disorganized during transplantation, rendering poor cell survival, functionality and engraftment in vivo. Here, we present the design of three-dimensional (3D) microtopographic scaffolds, using tunable electrospun microfibrous polymeric substrates that promote in situ stem cell neuronal reprogramming, neural network establishment and support neuronal engraftment into the brain. Scaffold-supported, reprogrammed neuronal networks were successfully grafted into organotypic hippocampal brain slices, showing an ∼3.5-fold improvement in neurite outgrowth and increased action potential firing relative to injected isolated cells. Transplantation of scaffold-supported neuronal networks into mouse brain striatum improved survival ∼38-fold at the injection site relative to injected isolated cells, and allowed delivery of multiple neuronal subtypes. Thus, 3D microscale biomaterials represent a promising platform for the transplantation of therapeutic human neurons with broad neuro-regenerative relevance. PMID:26983594

  19. Materials and scaffolds in medical 3D printing and bioprinting in the context of bone regeneration.

    PubMed

    Heller, Martin; Bauer, Heide-Katharina; Goetze, Elisabeth; Gielisch, Matthias; Ozbolat, Ibrahim T; Moncal, Kazim K; Rizk, Elias; Seitz, Hermann; Gelinsky, Michael; Schröder, Heinz C; Wang, Xiaohong H; Müller, Werner E G; Al-Nawas, Bilal

    The structural and functional repair of lost bone is still one of the biggest challenges in regenerative medicine. In many cases, autologous bone is used for the reconstruction of bone tissue; however, the availability of autologous material is limited, which always means additional stress to the patient. Due to this, more and more frequently various biocompatible materials are being used instead for bone augmentation. In this context, in order to ensure the structural function of the bone, scaffolds are implanted and fixed into the bone defect, depending on the medical indication. Nevertheless, for the surgeon, every individual clinical condition in which standardized scaffolds have to be aligned is challenging, and in many cases the alignment is not possible without limitations. Therefore, in the last decades, 3D printing (3DP) or additive manufacturing (AM) of scaffolds has become one of the most innovative approaches in surgery to individualize and improve the treatment of patients. Numerous biocompatible materials are available for 3DP, and various printing techniques can be applied, depending on the process conditions of these materials. Besides these conventional printing techniques, another promising approach in the context of medical AM is 3D bioprinting, a technique which makes it possible to print human cells embedded in special carrier substances to generate functional tissues. Even the direct printing into bone defects or lesions becomes possible. 3DP is already improving the treatment of patients, and has the potential to revolutionize regenerative medicine in future.

  20. 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.

  1. Effects of Novel 3D Antibiotic-Containing Electrospun Scaffolds on Dentin Discoloration

    PubMed Central

    Porter, Margaret Louise A.; Münchow, Eliseu A.; Albuquerque, Maria T. P.; Spolnik, Kenneth J.; Hara, Anderson T.; Bottino, Marco C.

    2016-01-01

    Introduction Although intracanal application of the triple antibiotic paste (TAP) may offer advantages (e.g., disinfection), this practice has been associated with significant drawbacks, including tooth discoloration. In this study, the color change of dentin was monitored during treatment with distinct TAP pastes and novel tubular-shaped three-dimensional (3D) electrospun scaffolds containing minocycline-MINO or doxycycline-DOX. Methods Two TAP pastes (TAPMINO–MINO, metronidazole/MET, and ciprofloxacin/CIP; and TAPDOX–DOX, MET, and CIP), four scaffold-based groups containing MINO or DOX, at distinct concentrations; one antibiotic-free scaffold (Scaffold); and one untreated group (Control) were investigated. Human canines were sectioned at the cemento-enamel junction (CEJ) and tubular-shaped scaffolds or paste were placed into the root canals and sealed. Color measurements (CIEL*a*b* parameters) were performed at baseline and after 1, 3, 7, 14, 21, and 28 days. Color changes were expressed as ΔE* values. In addition, scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS) were also performed on the specimens after treatment. Data were analyzed using Repeated-measures ANOVA (alpha=0.05). Results All antibiotic-containing groups led to greater discoloration than the antibiotic-free groups. A severe discoloration occurred after 1 day. At the end of the experiment, antibiotic-treated samples exhibited crusts/agglomerates over the dentin surface, which totally or partially obliterated the dentinal tubules. The presence of MINO resulted in greater color change than DOX. Conclusion Scaffolds containing MINO or DOX produced similar color change to dentin when compared to their respective TAP systems, although DOX-related discoloration was less pronounced. PMID:26602451

  2. Dextran hydrogel scaffolds enhance angiogenic responses and promote complete skin regeneration during burn wound healing.

    PubMed

    Sun, Guoming; Zhang, Xianjie; Shen, Yu-I; Sebastian, Raul; Dickinson, Laura E; Fox-Talbot, Karen; Reinblatt, Maura; Steenbergen, Charles; Harmon, John W; Gerecht, Sharon

    2011-12-27

    Neovascularization is a critical determinant of wound-healing outcomes for deep burn injuries. We hypothesize that dextran-based hydrogels can serve as instructive scaffolds to promote neovascularization and skin regeneration in third-degree burn wounds. Dextran hydrogels are soft and pliable, offering opportunities to improve the management of burn wound treatment. We first developed a procedure to treat burn wounds on mice with dextran hydrogels. In this procedure, we followed clinical practice of wound excision to remove full-thickness burned skin, and then covered the wound with the dextran hydrogel and a dressing layer. Our procedure allows the hydrogel to remain intact and securely in place during the entire healing period, thus offering opportunities to simplify the management of burn wound treatment. A 3-week comparative study indicated that dextran hydrogel promoted dermal regeneration with complete skin appendages. The hydrogel scaffold facilitated early inflammatory cell infiltration that led to its rapid degradation, promoting the infiltration of angiogenic cells into the healing wounds. Endothelial cells homed into the hydrogel scaffolds to enable neovascularization by day 7, resulting in an increased blood flow significantly greater than treated and untreated controls. By day 21, burn wounds treated with hydrogel developed a mature epithelial structure with hair follicles and sebaceous glands. After 5 weeks of treatment, the hydrogel scaffolds promoted new hair growth and epidermal morphology and thickness similar to normal mouse skin. Collectively, our evidence shows that customized dextran-based hydrogel alone, with no additional growth factors, cytokines, or cells, promoted remarkable neovascularization and skin regeneration and may lead to novel treatments for dermal wounds.

  3. Biologically Inspired Smart Release System Based on 3D Bioprinted Perfused Scaffold for Vascularized Tissue Regeneration

    PubMed Central

    Cui, Haitao; Zhu, Wei; Holmes, Benjamin

    2016-01-01

    A critical challenge to the development of large‐scale artificial tissue grafts for defect reconstruction is vascularization of the tissue construct. As an emerging tissue/organ manufacturing technique, 3D bioprinting offers great precision in controlling the internal architecture of a scaffold with preferable mechanical strength and printing complicated microstructures comparable to native tissue. However, current bioprinting techniques still exhibit difficulty in achieving biomimetic nano resolution and cooperating with bioactive spatiotemporal signals. In this study, a comprehensive design of engineered vascularized bone construct is presented for the first time by integrating biomimetic 3D bioprinted fluid perfused microstructure with biologically inspired smart release nanocoating, which is regarded as an aspiring concept combining engineering, biological, and material science. In this biologically inspired design, angiogenesis and osteogenesis are successively induced through a matrix metalloprotease 2 regulative mechanism by delivering dual growth factors with sequential release in spatiotemporal coordination. Availability of this system is evaluated in dynamic culture condition, which is similar to fluid surrounding in vivo, as an alternative animal model study. Results, particularly from co‐cultured dynamically samples demonstrate excellent bioactivity and vascularized bone forming potential of nanocoating modified 3D bioprinted scaffolds for human bone marrow mesenchymal stem cells and human umbilical vein endothelial cells. PMID:27818910

  4. Biologically Inspired Smart Release System Based on 3D Bioprinted Perfused Scaffold for Vascularized Tissue Regeneration.

    PubMed

    Cui, Haitao; Zhu, Wei; Holmes, Benjamin; Zhang, Lijie Grace

    2016-08-01

    A critical challenge to the development of large-scale artificial tissue grafts for defect reconstruction is vascularization of the tissue construct. As an emerging tissue/organ manufacturing technique, 3D bioprinting offers great precision in controlling the internal architecture of a scaffold with preferable mechanical strength and printing complicated microstructures comparable to native tissue. However, current bioprinting techniques still exhibit difficulty in achieving biomimetic nano resolution and cooperating with bioactive spatiotemporal signals. In this study, a comprehensive design of engineered vascularized bone construct is presented for the first time by integrating biomimetic 3D bioprinted fluid perfused microstructure with biologically inspired smart release nanocoating, which is regarded as an aspiring concept combining engineering, biological, and material science. In this biologically inspired design, angiogenesis and osteogenesis are successively induced through a matrix metalloprotease 2 regulative mechanism by delivering dual growth factors with sequential release in spatiotemporal coordination. Availability of this system is evaluated in dynamic culture condition, which is similar to fluid surrounding in vivo, as an alternative animal model study. Results, particularly from co-cultured dynamically samples demonstrate excellent bioactivity and vascularized bone forming potential of nanocoating modified 3D bioprinted scaffolds for human bone marrow mesenchymal stem cells and human umbilical vein endothelial cells.

  5. Surface functionalization of 3D glass-ceramic porous scaffolds for enhanced mineralization in vitro

    NASA Astrophysics Data System (ADS)

    Ferraris, Sara; Vitale-Brovarone, Chiara; Bretcanu, Oana; Cassinelli, Clara; Vernè, Enrica

    2013-04-01

    Bone reconstruction after tissue loosening due to traumatic, pathological or surgical causes is in increasing demand. 3D scaffolds are a widely studied solution for supporting new bone growth. Bioactive glass-ceramic porous materials can offer a three-dimensional structure that is able to chemically bond to bone. The ability to surface modify these devices by grafting biologically active molecules represents a challenge, with the aim of stimulating physiological bone regeneration with both inorganic and organic signals. In this research work glass ceramic scaffolds with very high mechanical properties and moderate bioactivity have been functionalized with the enzyme alkaline phosphatase (ALP). The material surface was activated in order to expose hydroxyl groups. The activated surface was further grafted with ALP both via silanization and also via direct grafting to the surface active hydroxyl groups. Enzymatic activity of grafted samples were measured by means of UV-vis spectroscopy before and after ultrasonic washing in TRIS-HCl buffer solution. In vitro inorganic bioactivity was investigated by soaking the scaffolds after the different steps of functionalization in a simulated body fluid (SBF). SEM observations allowed the monitoring of the scaffold morphology and surface chemical composition after soaking in SBF. The presence of ALP enhanced the in vitro inorganic bioactivity of the tested material.

  6. Highly Porous Gelatin Reinforced 3D Scaffolds for Articular Cartilage Regeneration.

    PubMed

    Amadori, Sofia; Torricelli, Paola; Panzavolta, Silvia; Parrilli, Annapaola; Fini, Milena; Bigi, Adriana

    2015-07-01

    3D highly porous (93% total porosity) gelatin scaffolds were prepared according to a novel, simple method, which implies gelatin foaming, gelification, soaking into ethanol and successive freeze-drying. Reinforcement of the as-prepared scaffolds (GEL) was performed through immersion in aqueous solutions at different gelatin concentrations. Reinforcement solutions with and without genipin addition allowed to prepare two series of samples:cross-linked and uncross-linked samples, respectively. The amount of gelatin adsorbed onto the reinforced samples increases as a function of gelatin concentration in solution and provokes a drastic improvement of the compressive modulus and collapse strength up to values of about 30 and 4 MPa, respectively. The open and interconnected porosity, although slightly reduced, is still of the order of 80% in the samples reinforced with the highest concentration of gelatin. Water uptake ability evaluated after immersion in PBS for 20 s decreases with gelatin reinforcement. The presence of genipin in cross-linked samples reduces gelatin release and stabilizes the scaffolds in solution. Chondrocytes from human articular cartilage adhere, proliferate, and penetrate into the scaffolds. The evaluation of differentiation markers both on the supernatants of cell culture and by means of quantitative polymerase chain reaction (qPCR) indicates a dose-dependent promotion of cell differentiation.

  7. Increased lipid accumulation and adipogenic gene expression of adipocytes in 3D bioprinted nanocellulose scaffolds.

    PubMed

    Henriksson, I; Gatenholm, P; Hägg, D A

    2017-02-21

    Compared to standard 2D culture systems, new methods for 3D cell culture of adipocytes could provide more physiologically accurate data and a deeper understanding of metabolic diseases such as diabetes. By resuspending living cells in a bioink of nanocellulose and hyaluronic acid, we were able to print 3D scaffolds with uniform cell distribution. After one week in culture, cell viability was 95%, and after two weeks the cells displayed a more mature phenotype with larger lipid droplets than standard 2D cultured cells. Unlike cells in 2D culture, the 3D bioprinted cells did not detach upon lipid accumulation. After two weeks, the gene expression of the adipogenic marker genes PPARγ and FABP4 was increased 2.0- and 2.2-fold, respectively, for cells in 3D bioprinted constructs compared with 2D cultured cells. Our 3D bioprinted culture system produces better adipogenic differentiation of mesenchymal stem cells and a more mature cell phenotype than conventional 2D culture systems.

  8. 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.

  9. A composite scaffold of MSC affinity peptide-modified demineralized bone matrix particles and chitosan hydrogel for cartilage regeneration

    PubMed Central

    Meng, Qingyang; Man, Zhentao; Dai, Linghui; Huang, Hongjie; Zhang, Xin; Hu, Xiaoqing; Shao, Zhenxing; Zhu, Jingxian; Zhang, Jiying; Fu, Xin; Duan, Xiaoning; Ao, Yingfang

    2015-01-01

    Articular cartilage injury is still a significant challenge because of the poor intrinsic healing potential of cartilage. Stem cell-based tissue engineering is a promising technique for cartilage repair. As cartilage defects are usually irregular in clinical settings, scaffolds with moldability that can fill any shape of cartilage defects and closely integrate with the host cartilage are desirable. In this study, we constructed a composite scaffold combining mesenchymal stem cells (MSCs) E7 affinity peptide-modified demineralized bone matrix (DBM) particles and chitosan (CS) hydrogel for cartilage engineering. This solid-supported composite scaffold exhibited appropriate porosity, which provided a 3D microenvironment that supports cell adhesion and proliferation. Cell proliferation and DNA content analysis indicated that the DBM-E7/CS scaffold promoted better rat bone marrow-derived MSCs (BMMSCs) survival than the CS or DBM/CS groups. Meanwhile, the DBM-E7/CS scaffold increased matrix production and improved chondrogenic differentiation ability of BMMSCs in vitro. Furthermore, after implantation in vivo for four weeks, compared to those in control groups, the regenerated issue in the DBM-E7/CS group exhibited translucent and superior cartilage-like structures, as indicated by gross observation, histological examination, and assessment of matrix staining. Overall, the functional composite scaffold of DBM-E7/CS is a promising option for repairing irregularly shaped cartilage defects. PMID:26632447

  10. A composite scaffold of MSC affinity peptide-modified demineralized bone matrix particles and chitosan hydrogel for cartilage regeneration

    NASA Astrophysics Data System (ADS)

    Meng, Qingyang; Man, Zhentao; Dai, Linghui; Huang, Hongjie; Zhang, Xin; Hu, Xiaoqing; Shao, Zhenxing; Zhu, Jingxian; Zhang, Jiying; Fu, Xin; Duan, Xiaoning; Ao, Yingfang

    2015-12-01

    Articular cartilage injury is still a significant challenge because of the poor intrinsic healing potential of cartilage. Stem cell-based tissue engineering is a promising technique for cartilage repair. As cartilage defects are usually irregular in clinical settings, scaffolds with moldability that can fill any shape of cartilage defects and closely integrate with the host cartilage are desirable. In this study, we constructed a composite scaffold combining mesenchymal stem cells (MSCs) E7 affinity peptide-modified demineralized bone matrix (DBM) particles and chitosan (CS) hydrogel for cartilage engineering. This solid-supported composite scaffold exhibited appropriate porosity, which provided a 3D microenvironment that supports cell adhesion and proliferation. Cell proliferation and DNA content analysis indicated that the DBM-E7/CS scaffold promoted better rat bone marrow-derived MSCs (BMMSCs) survival than the CS or DBM/CS groups. Meanwhile, the DBM-E7/CS scaffold increased matrix production and improved chondrogenic differentiation ability of BMMSCs in vitro. Furthermore, after implantation in vivo for four weeks, compared to those in control groups, the regenerated issue in the DBM-E7/CS group exhibited translucent and superior cartilage-like structures, as indicated by gross observation, histological examination, and assessment of matrix staining. Overall, the functional composite scaffold of DBM-E7/CS is a promising option for repairing irregularly shaped cartilage defects.

  11. Combinatorial screening of osteoblast response to 3D calcium phosphate/poly(ε-caprolactone) scaffolds using gradients and arrays

    PubMed Central

    Chatterjee, Kaushik; Sun, Limin; Chow, Laurence C.; Young, Marian F.; Simon, Carl G.

    2012-01-01

    There is a need for combinatorial and high-throughput methods for screening cell–biomaterial interactions to maximize tissue generation in scaffolds. Current methods employ a flat two-dimensional (2D) format even though three-dimensional (3D) scaffolds are more representative of the tissue environment in vivo and cells are responsive to topographical differences of 2D substrates and 3D scaffolds. Thus, combinatorial libraries of 3D porous scaffolds were developed and used to screen the effect of nano-amorphous calcium phosphate (nACP) particles on osteoblast response. Increasing nACP content in poly (ε-caprolactone) (PCL) scaffolds promoted osteoblast adhesion and proliferation. The nACP-containing scaffolds released calcium and phosphate ions which are known to activate osteoblast function. Scaffold libraries were fabricated in two formats, gradients and arrays, and the magnitude of the effect of nACP on osteoblast proliferation was greater for arrays than gradients. The enhanced response in arrays can be explained by differences in cell culture designs, diffusional effects and differences in the ratio of “scaffold mass to culture medium”. These results introduce a gradient library approach for screening large pore 3D scaffolds and demonstrate that inclusion of the nACP particles enhances osteoblast proliferation in 3D scaffolds. Further, comparison of gradients and arrays suggests that gradients were more sensitive for detecting effects of scaffold composition on cell adhesion (short time points, 1 day) whereas arrays were more sensitive at detecting effects on cell proliferation (longer time points, 14 day). PMID:21074846

  12. Bicomponent electrospinning to fabricate three-dimensional hydrogel-hybrid nanofibrous scaffolds with spatial fiber tortuosity.

    PubMed

    Jin, Gyuhyung; Lee, Slgirim; Kim, Seung-Hyun; Kim, Minhee; Jang, Jae-Hyung

    2014-12-01

    Electrospun fibrous mats have emerged as powerful tissue engineering scaffolds capable of providing highly effective and versatile physical guidance, mimicking the extracellular environment. However, electrospinning typically produces a sheet-like structure, which is a major limitation associated with current electrospinning technologies. To address this challenge, highly porous, volumetric hydrogel-hybrid fibrous scaffolds were fabricated by one Taylor cone-based side-by-side dual electrospinning of poly (ε-caprolactone) (PCL) and poly (vinyl pyrrolidone) (PVP), which possess distinct properties (i.e., hydrophobic and hydrogel properties, respectively). Immersion of the resulting scaffolds in water induced spatial tortuosity of the hydrogel PVP fibers while maintaining their aligned fibrous structures in parallel with the PCL fibers. The resulting conformational changes in the entire bicomponent fibers upon immersion in water led to volumetric expansion of the fibrous scaffolds. The spatial fiber tortuosity significantly increased the pore volumes of electrospun fibrous mats and dramatically promoted cellular infiltration into the scaffold interior both in vitro and in vivo. Harmonizing the flexible PCL fibers with the soft PVP-hydrogel layers produced highly ductile fibrous structures that could mechanically resist cellular contractile forces upon in vivo implantation. This facile dual electrospinning followed by the spatial fiber tortuosity for fabricating three-dimensional hydrogel-hybrid fibrous scaffolds will extend the use of electrospun fibers toward various tissue engineering applications.

  13. A Tunable Scaffold of Microtubular Graphite for 3D Cell Growth

    PubMed Central

    2016-01-01

    Aerographite (AG) is a novel carbon-based material that exists as a self-supportive 3D network of interconnected hollow microtubules. It can be synthesized in a variety of architectures tailored by the growth conditions. This flexibility in creating structures presents interesting bioengineering possibilities such as the generation of an artificial extracellular matrix. Here we have explored the feasibility and potential of AG as a scaffold for 3D cell growth employing cyclic RGD (cRGD) peptides coupled to poly(ethylene glycol) (PEG) conjugated phospholipids for surface functionalization to promote specific adhesion of fibroblast cells. Successful growth and invasion of the bulk material was followed over a period of 4 days. PMID:27258400

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

    PubMed

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

    2016-09-06

    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/cm(2) 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.

  15. 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

  16. Adipose tissue-derived stem cells display a proangiogenic phenotype on 3D scaffolds.

    PubMed

    Neofytou, Evgenios A; Chang, Edwin; Patlola, Bhagat; Joubert, Lydia-Marie; Rajadas, Jayakumar; Gambhir, Sanjiv S; Cheng, Zhen; Robbins, Robert C; Beygui, Ramin E

    2011-09-01

    Ischemic heart disease is the leading cause of death worldwide. Recent studies suggest that adipose tissue-derived stem cells (ASCs) can be used as a potential source for cardiovascular tissue engineering due to their ability to differentiate along the cardiovascular lineage and to adopt a proangiogenic phenotype. To understand better ASCs' biology, we used a novel 3D culture device. ASCs' and b.END-3 endothelial cell proliferation, migration, and vessel morphogenesis were significantly enhanced compared to 2D culturing techniques. ASCs were isolated from inguinal fat pads of 6-week-old GFP+/BLI+ mice. Early passage ASCs cells (P3-P4), PKH26-labeled murine b.END-3 cells or a co-culture of ASCs and b.END-3 cells were seeded at a density of 1 × 10(5) on three different surface configurations: (a) a 2D surface of tissue culture plastic, (b) Matrigel, and (c) a highly porous 3D scaffold fabricated from inert polystyrene. VEGF expression, cell proliferation, and tubulization, were assessed using optical microscopy, fluorescence microscopy, 3D confocal microscopy, and SEM imaging (n = 6). Increased VEGF levels were seen in conditioned media harvested from co-cultures of ASCs and b.END-3 on either Matrigel or a 3D matrix. Fluorescence, confocal, SEM, bioluminescence revealed improved cell, proliferation, and tubule formation for cells seeded on the 3D polystyrene matrix. Collectively, these data demonstrate that co-culturing ASCs with endothelial cells in a 3D matrix environment enable us to generate prevascularized tissue-engineered constructs. This can potentially help us to surpass the tissue thickness limitations faced by the tissue engineering community today.

  17. Hydrogel with chains functionalized with carboxyl groups as universal 3D platform in DNA biosensors.

    PubMed

    Kowalczyk, Agata; Fau, Michal; Karbarz, Marcin; Donten, Mikolaj; Stojek, Zbigniew; Nowicka, Anna M

    2014-04-15

    Application of hydrogel based on N-isopropylacrylamide with carboxyl groups grafted to the chains enabled the immobilization of DNA at an extent exceeding that for flat surfaces by at least one order of magnitude. The probe DNA strands in the 3D platform were fully available for the hybridization process. The examination of the gels containing different amounts of grafted carboxyl groups (1-10%) was done using quartz crystal microbalance, electrochemical impedance spectroscopy, chronoamperometry and ionic coupled plasma with laser ablation. The optimal carboxyl group content was determined to be 5%. A very good agreement of the data obtained with independent techniques on content of DNA in the gel was obtained. In comparison to the other methods of immobilization of DNA the new platform enabled complete removal of DNA after the measurements and analysis and, therefore, could be used many times. After a 10-fold exchange of the DNA-sensing layer the efficiency of hybridization and analytical signal did not change by more than 5%. The sensor response increased linearly with logarithm of concentration of target DNA in the range 1×10(-13)-1×10(-6) M. The obtained detection limit was circa 8×10(-13) M of target DNA in the sample which is a substantial improvement over the planar sensing layers.

  18. Fabrication and development of artificial osteochondral constructs based on cancellous bone/hydrogel hybrid scaffold.

    PubMed

    Song, Kedong; Li, Liying; Yan, Xinyu; Zhang, Yu; Li, Ruipeng; Wang, Yiwei; Wang, Ling; Wang, Hong; Liu, Tianqing

    2016-06-01

    Using tissue engineering techniques, an artificial osteochondral construct was successfully fabricated to treat large osteochondral defects. In this study, porcine cancellous bones and chitosan/gelatin hydrogel scaffolds were used as substitutes to mimic bone and cartilage, respectively. The porosity and distribution of pore size in porcine bone was measured and the degradation ratio and swelling ratio for chitosan/gelatin hydrogel scaffolds was also determined in vitro. Surface morphology was analyzed with the scanning electron microscope (SEM). The physicochemical properties and the composition were tested by using an infrared instrument. A double layer composite scaffold was constructed via seeding adipose-derived stem cells (ADSCs) induced to chondrocytes and osteoblasts, followed by inoculation in cancellous bones and hydrogel scaffolds. Cell proliferation was assessed through Dead/Live staining and cellular activity was analyzed with IpWin5 software. Cell growth, adhesion and formation of extracellular matrix in composite scaffolds blank cancellous bones or hydrogel scaffolds were also analyzed. SEM analysis revealed a super porous internal structure of cancellous bone scaffolds and pore size was measured at an average of 410 ± 59 μm while porosity was recorded at 70.6 ± 1.7 %. In the hydrogel scaffold, the average pore size was measured at 117 ± 21 μm and the porosity and swelling rate were recorded at 83.4 ± 0.8 % and 362.0 ± 2.4 %, respectively. Furthermore, the remaining hydrogel weighed 80.76 ± 1.6 % of the original dry weight after hydration in PBS for 6 weeks. In summary, the cancellous bone and hydrogel composite scaffold is a promising biomaterial which shows an essential physical performance and strength with excellent osteochondral tissue interaction in situ. ADSCs are a suitable cell source for osteochondral composite reconstruction. Moreover, the bi-layered scaffold significantly enhanced cell proliferation compared to the cells seeded on

  19. Generation of Multi-Scale Vascular Network System within 3D Hydrogel using 3D Bio-Printing Technology.

    PubMed

    Lee, Vivian K; Lanzi, Alison M; Haygan, Ngo; Yoo, Seung-Schik; Vincent, Peter A; Dai, Guohao

    2014-09-01

    Although 3D bio-printing technology has great potential in creating complex tissues with multiple cell types and matrices, maintaining the viability of thick tissue construct for tissue growth and maturation after the printing is challenging due to lack of vascular perfusion. Perfused capillary network can be a solution for this issue; however, construction of a complete capillary network at single cell level using the existing technology is nearly impossible due to limitations in time and spatial resolution of the dispensing technology. To address the vascularization issue, we developed a 3D printing method to construct larger (lumen size of ~1mm) fluidic vascular channels and to create adjacent capillary network through a natural maturation process, thus providing a feasible solution to connect the capillary network to the large perfused vascular channels. In our model, microvascular bed was formed in between two large fluidic vessels, and then connected to the vessels by angiogenic sprouting from the large channel edge. Our bio-printing technology has a great potential in engineering vascularized thick tissues and vascular niches, as the vascular channels are simultaneously created while cells and matrices are printed around the channels in desired 3D patterns.

  20. Hyperbranched poly(glycidol)/poly(ethylene oxide) crosslinked hydrogel for tissue engineering scaffold using e-beams.

    PubMed

    Haryanto; Singh, Deepti; Huh, Pil Ho; Kim, Seong Cheol

    2016-01-01

    A microporous hydrogel scaffold was developed from hyperbranched poly(glycidol) (HPG) and poly(ethylene oxide) (PEO) using electron beam (e-beam) induced cross-linking for tissue engineering applications. In this study, HPG was synthesized from glycidol using trimethylol propane as a core initiator and cross-linked hydrogels were made using 0, 10, 20, and 30% HPG with respect to PEO. The effects of %-HPG on the swelling ratio, cross-linking density, mechanical properties, morphology, degradation, and cytotoxicity of the hydrogel scaffolds were then investigated. Increasing the HPG content increased the pore size of the hydrogel scaffold, as well as the porosity, elongation at break, degree of degradation and swelling ratio. In contrast, the presence of HPG decreased the cross-linking density of the hydrogel. There was no significant difference in compressive modulus and tensile strength of all compositions. The pore size of hydrogel scaffolds could be easily tailored by controlling the content of HPG in the polymer blend. Evaluation of the cytotoxicity demonstrated that HPG/PEO hydrogel scaffold has potential for use as a matrix for cellular attachment and proliferation. These results indicate that cross-linked HPG/PEO hydrogel can function as a potential material for tissue engineering scaffolds. Moreover, a facile method to prepare hydrogel microporous scaffolds for tissue engineering by e-beam irradiation was developed.

  1. 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

  2. Graphene-based 3D composite hydrogel by anchoring Co3O4 nanoparticles with enhanced electrochemical properties.

    PubMed

    Yuan, Jingjing; Zhu, Junwu; Bi, Huiping; Meng, Xiaoqian; Liang, Shiming; Zhang, Lili; Wang, Xin

    2013-08-21

    Three-dimensional (3D) graphene-based composite materials have attracted increasing attention, owing to their specific surface area, high conductivity and electronic interactions. Here, we report a convenient route to fabricate a 3D Co3O4/Graphene Hydrogel (CGH) composite as an electrode material for supercapacitors. Utilizing the gelation of a graphene oxide dispersion enables the anchoring of Co3O4 nanoparticles on the graphene sheet surfaces and formation of the hydrogel simultaneously. Remarkably, the spherical Co3O4 particles can serve as spacers to keep the neighboring graphene sheets separated. The CGH exhibits a high specific capacitance (Cs) of 757.5 F g(-1) at a current density of 0.5 A g(-1), indicating its potential application as an electrode material for supercapacitors.

  3. Differentiation of neural stem cells in three-dimensional growth factor-immobilized chitosan hydrogel scaffolds.

    PubMed

    Leipzig, Nic D; Wylie, Ryan G; Kim, Howard; Shoichet, Molly S

    2011-01-01

    The adult central nervous system (CNS) contains adult neural stem/progenitor cells (NSPCs) that possess the ability to differentiate into the primary cell types found in the CNS and to regenerate lost or damaged tissue. The ability to specifically and spatially control differentiation is vital to enable cell-based CNS regenerative strategies. Here we describe the development of a protein-biomaterial system that allows rapid, stable and homogenous linking of a growth factor to a photocrosslinkable material. A bioactive recombinant fusion protein incorporating pro-neural rat interferon-γ (rIFN-γ) and the AviTag for biotinylation was successfully expressed in Escherichia coli and purified. The photocrosslinkable biopolymer, methacrylamide chitosan (MAC), was thiolated, allowing conjugation of maleimide-strepatavidin via Michael-type addition. We demonstrated that biotin-rIFN-γ binds specifically to MAC-streptavidin in stoichiometric yields at 100 and 200 ng/mL in photocrosslinked hydrogels. For cell studies, NSPCs were photo-encapsulated in 100 ng/mL biotin-rIFN-γ immobilized MAC based scaffolds and compared to similar NSPC-seeded scaffolds combining 100 ng/mL soluble biotin-rIFN-γ vs. no growth factor. Cells were cultured for 8 days after which differentiation was assayed using immunohistochemistry for lineage specific markers. Quantification showed that immobilized biotin-rIFN-γ promoted neuronal differentiation (72.8 ± 16.0%) similar to soluble biotin-rIFN-γ (71.8 ± 13.2%). The percentage of nestin-positive (stem/progenitor) cells as well as RIP-positive (oligodendrocyte) cells were significantly higher in scaffolds with soluble vs. immobilized biotin-rIFN-γ suggesting that 3-D immobilization results in a more committed lineage specification.

  4. Bioactive gyroid scaffolds formed by sacrificial templating of nanocellulose and nanochitin hydrogels as instructive platforms for biomimetic tissue engineering.

    PubMed

    Torres-Rendon, Jose Guillermo; Femmer, Tim; De Laporte, Laura; Tigges, Thomas; Rahimi, Khosrow; Gremse, Felix; Zafarnia, Sara; Lederle, Wiltrud; Ifuku, Shinsuke; Wessling, Matthias; Hardy, John G; Walther, Andreas

    2015-05-20

    A sacrificial templating process using lithographically printed minimal surface structures allows complex de novo geo-metries of delicate hydrogel materials. The hydrogel scaffolds based on cellulose and chitin nanofibrils show differences in terms of attachment of human mesenchymal stem cells, and allow their differentiation into osteogenic outcomes. The approach here serves as a first example toward designer hydrogel scaffolds viable for biomimetic tissue engineering.

  5. Solid state synthesis of chitosan and its unsaturated derivatives for laser microfabrication of 3D scaffolds

    NASA Astrophysics Data System (ADS)

    Akopova, T. A.; Demina, T. S.; Bagratashvili, V. N.; Bardakova, K. N.; Novikov, M. M.; Selezneva, I. I.; Istomin, A. V.; Svidchenko, E. A.; Cherkaev, G. V.; Surin, N. M.; Timashev, P. S.

    2015-07-01

    Chitosans with various degrees of deacetylation and molecular weights and their allyl substituted derivatives were obtained through a solvent-free reaction under shear deformation in an extruder. Structure and physical-chemical analysis of the samples were carried out using nuclear magnetic resonance (NMR), ultraviolet (UV) and infrared radiation (IR) spectroscopy. Photosensitive materials based on the synthesized polymers were successfully used for microfabrication of 3D well-defined architectonic structures by laser stereolithography. Study on the metabolic activity of NCTC L929 cultured in the presence of the cured chitosan extracts indicates that the engineered biomaterials could support adhesion, spreading and growth of adherent-dependent cells, and thus could be considered as biocompatible scaffolds.

  6. Cancer Cell Migration within 3D Layer-By-Layer Microfabricated Photocrosslinked PEG Scaffolds with Tunable Stiffness

    PubMed Central

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

    2012-01-01

    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. PMID:22809641

  7. 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.

  8. Collagen Hydrogel Scaffold and Fibroblast Growth Factor-2 Accelerate Periodontal Healing of Class II Furcation Defects in Dog

    PubMed Central

    Momose, Takehito; Miyaji, Hirofumi; Kato, Akihito; Ogawa, Kosuke; Yoshida, Takashi; Nishida, Erika; Murakami, Syusuke; Kosen, Yuta; Sugaya, Tsutomu; Kawanami, Masamitsu

    2016-01-01

    Objective: Collagen hydrogel scaffold exhibits bio-safe properties and facilitates periodontal wound healing. However, regenerated tissue volume is insufficient. Fibroblast growth factor-2 (FGF2) up-regulates cell behaviors and subsequent wound healing. We evaluated whether periodontal wound healing is promoted by application of collagen hydrogel scaffold in combination with FGF2 in furcation defects in beagle dogs. Methods: Collagen hydrogel was fabricated from bovine type I collagen with an ascorbate-copper ion cross-linking system. Collagen hydrogel was mingled with FGF2 and injected into sponge-form collagen. Subsequently, FGF2 (50 µg)/collagen hydrogel scaffold and collagen hydrogel scaffold alone were implanted into class II furcation defects in dogs. In addition, no implantation was performed as a control. Histometric parameters were assessed at 10 days and 4 weeks after surgery. Result: FGF2 application to scaffold promoted considerable cell and tissue ingrowth containing numerous cells and blood vessel-like structure at day 10. At 4 weeks, reconstruction of alveolar bone was stimulated by implantation of scaffold loaded with FGF2. Furthermore, periodontal attachment, consisting of cementum-like tissue, periodontal ligament-like tissue and Sharpey’s fibers, was also repaired, indicating that FGF2-loaded scaffold guided self-assembly and then re-established the function of periodontal organs. Aberrant healing, such as ankylosis and root resorption, was not observed. Conclusion: FGF2-loaded collagen hydrogel scaffold possessed excellent biocompatibility and strongly promoted periodontal tissue engineering, including periodontal attachment re-organization. PMID:27583044

  9. Tough and elastic hydrogel of hyaluronic acid and chondroitin sulfate as potential cell scaffold materials.

    PubMed

    Ni, Yilu; Tang, Zhurong; Cao, Wanxu; Lin, Hai; Fan, Yujiang; Guo, Likun; Zhang, Xingdong

    2015-03-01

    Natural polysaccharides are extensively investigated as cell scaffold materials for cellular adhesion, proliferation, and differentiation due to their excellent biocompatibility, biodegradability, and biofunctions. However, their application is often severely limited by their mechanical behavior. In this study, a tough and elastic hydrogel scaffold was prepared with hyaluronic acid (HA) and chondroitin sulfate (CS). HA and CS were conjugated with tyramine (TA) and the degree of substitution (DS) was 10.7% and 11.3%, respectively, as calculated by (1)H NMR spectra. The hydrogel was prepared by mixing HA-TA and CS-TA in presence of H2O2 and HRP. The sectional morphology of hydrogels was observed by SEM, static and dynamic mechanical properties were analyzed by Shimadzu electromechanical testing machine and dynamic mechanical thermal analyzer Q800. All samples showed good ability to recover their appearances after deformation, the storage modulus (E') of hydrogels became higher as the testing frequency went up. Hydrogels also showed fatigue resistance to cyclic compression. Mesenchymal stem cells encapsulated in hydrogels showed good cell viability as detected by CLSM. This study suggests that the hydrogels have both good mechanical properties and biocompatibility, and may serve as model systems to explore mechanisms of deformation and energy dissipation or find some applications in tissue engineering.

  10. Preparation of collagen/hydroxyapatite/alendronate hybrid hydrogels as potential scaffolds for bone regeneration.

    PubMed

    Ma, Xin; He, Zhiwei; Han, Fengxuan; Zhong, Zhiyuan; Chen, Liang; Li, Bin

    2016-07-01

    Development of biomimetic scaffolds represents a promising direction in bone tissue engineering. In this study, we designed a two-step process to prepare a type of biomimetic hybrid hydrogels that were composed of collagen, hydroxyapatite (HAP) and alendronate (ALN), an anti-osteoporosis drug. First, water-soluble ALN-conjugated HAP (HAP-ALN) containing 4.0wt.% of ALN was synthesized by treating HAP particles with ALN. Hydrogels were then formed from HAP-ALN conjugate and collagen under physiological conditions using genipin (GNP) as the crosslinker. Depending on the ALN/collagen molar ratio and GNP concentration, the gelation time of hydrogels ranged from 5 to 37min. Notably, these hybrid hydrogels exhibited markedly improved mechanical property (storage modulus G'=38-187kPa), higher gel contents, and lower swelling ratios compared to the hydrogels prepared from collagen alone under similar conditions. Moreover, they showed tunable degradation behaviors against collagenase. The collagen/HAP-ALN hybrid hydrogels supported the adhesion and growth of murine MC3T3-E1 osteoblastic cells well. Such tough yet enzymatically degradable hybrid hydrogels hold potential as scaffolds for bone tissue engineering.

  11. Flyweight 3D Graphene Scaffolds with Microinterface Barrier-Derived Tunable Thermal Insulation and Flame Retardancy.

    PubMed

    Zhang, Qiangqiang; Hao, Menglong; Xu, Xiang; Xiong, Guoping; Li, Hui; Fisher, Timothy S

    2017-04-14

    In this article, flyweight three-dimensional (3D) graphene scaffolds (GSs) have been demonstrated with a microinterface barrier-derived thermal insulation and flame retardancy characteristics. Such 3D GSs were fabricated by a modified hydrothermal method and a unidirectional freeze-casting process with hierarchical porous microstructures. Because of high porosity (99.9%), significant phonon scattering, and strong π-π interaction at the interface barriers of multilayer graphene cellular walls, the GSs demonstrate a sequence of multifunctional properties simultaneously, such as lightweight density, thermal insulating characteristics, and outstanding mechanical robustness. At 100 °C, oxidized GSs exhibit a thermal conductivity of 0.0126 ± 0.0010 W/(m K) in vacuum. The thermal conductivity of oxidized GSs remains relatively unaffected despite large-scale deformation-induced densification of the microstructures, as compared to the behavior of reduced GSs (rGSs) whose thermal conductivity increases dramatically under compression. The contrasting behavior of oxidized GSs and rGSs appears to derive from large differences in the intersheet contact resistance and varying intrinsic thermal conductivity between reduced and oxidized graphene sheets. The oxidized GSs also exhibit excellent flame retardant behavior and mechanical robustness, with only 2% strength decay after flame treatment. In a broader context, this work demonstrates a useful strategy to design porous nanomaterials with a tunable heat conduction behavior through interface engineering at the nanoscale.

  12. Effect of bioglass on growth and biomineralization of SaOS-2 cells in hydrogel after 3D cell bioprinting.

    PubMed

    Wang, Xiaohong; Tolba, Emad; Schröder, Heinz C; Neufurth, Meik; Feng, Qingling; Diehl-Seifert, Bärbel; Müller, Werner E G

    2014-01-01

    We investigated the effect of bioglass (bioactive glass) on growth and mineralization of bone-related SaOS-2 cells, encapsulated into a printable and biodegradable alginate/gelatine hydrogel. The hydrogel was supplemented either with polyphosphate (polyP), administered as polyP • Ca2+-complex, or silica, or as biosilica that had been enzymatically prepared from ortho-silicate by silicatein. These hydrogels, together with SaOS-2 cells, were bioprinted to computer-designed scaffolds. The results revealed that bioglass (nano)particles, with a size of 55 nm and a molar ratio of SiO2 : CaO : P2O5 of 55 : 40 : 5, did not affect the growth of the encapsulated cells. If silica, biosilica, or polyP • Ca2+-complex is co-added to the cell-containing alginate/gelatin hydrogel the growth behavior of the cells is not changed. Addition of 5 mg/ml of bioglass particles to this hydrogel significantly enhanced the potency of the entrapped SaOS-2 cells to mineralize. If compared with the extent of the cells to form mineral deposits in the absence of bioglass, the cells exposed to bioglass together with 100 µmoles/L polyP • Ca2+-complex increased their mineralization activity from 2.1- to 3.9-fold, or with 50 µmoles/L silica from 1.8- to 2.9-fold, or with 50 µmoles/L biosilica from 2.7- to 4.8-fold or with the two components together (100 µmoles/L polyP • Ca2+-complex and 50 µmoles/L biosilica) from 4.1- to 6.8-fold. Element analysis by EDX spectrometry of the mineral nodules formed by SaOS-2 revealed an accumulation of O, P, Ca and C, indicating that the mineral deposits contain, besides Ca-phosphate also Ca-carbonate. The results show that bioglass added to alginate/gelatin hydrogel increases the proliferation and mineralization of bioprinted SaOS-2 cells. We conclude that the development of cell-containing scaffolds consisting of a bioprintable, solid and cell-compatible inner matrix surrounded by a printable hard and flexible outer matrix containing bioglass, provide

  13. Effect of Bioglass on Growth and Biomineralization of SaOS-2 Cells in Hydrogel after 3D Cell Bioprinting

    PubMed Central

    Wang, Xiaohong; Tolba, Emad; Schröder, Heinz C.; Neufurth, Meik; Feng, Qingling; Diehl-Seifert, Bärbel; Müller, Werner E. G.

    2014-01-01

    We investigated the effect of bioglass (bioactive glass) on growth and mineralization of bone-related SaOS-2 cells, encapsulated into a printable and biodegradable alginate/gelatine hydrogel. The hydrogel was supplemented either with polyphosphate (polyP), administered as polyP•Ca2+-complex, or silica, or as biosilica that had been enzymatically prepared from ortho-silicate by silicatein. These hydrogels, together with SaOS-2 cells, were bioprinted to computer-designed scaffolds. The results revealed that bioglass (nano)particles, with a size of 55 nm and a molar ratio of SiO2∶CaO∶P2O5 of 55∶40∶5, did not affect the growth of the encapsulated cells. If silica, biosilica, or polyP•Ca2+-complex is co-added to the cell-containing alginate/gelatin hydrogel the growth behavior of the cells is not changed. Addition of 5 mg/ml of bioglass particles to this hydrogel significantly enhanced the potency of the entrapped SaOS-2 cells to mineralize. If compared with the extent of the cells to form mineral deposits in the absence of bioglass, the cells exposed to bioglass together with 100 µmoles/L polyP•Ca2+-complex increased their mineralization activity from 2.1- to 3.9-fold, or with 50 µmoles/L silica from 1.8- to 2.9-fold, or with 50 µmoles/L biosilica from 2.7- to 4.8-fold or with the two components together (100 µmoles/L polyP•Ca2+-complex and 50 µmoles/L biosilica) from 4.1- to 6.8-fold. Element analysis by EDX spectrometry of the mineral nodules formed by SaOS-2 revealed an accumulation of O, P, Ca and C, indicating that the mineral deposits contain, besides Ca-phosphate also Ca-carbonate. The results show that bioglass added to alginate/gelatin hydrogel increases the proliferation and mineralization of bioprinted SaOS-2 cells. We conclude that the development of cell-containing scaffolds consisting of a bioprintable, solid and cell-compatible inner matrix surrounded by a printable hard and flexible outer matrix containing bioglass, provide a

  14. Production of Uniform 3D Microtumors in Hydrogel Microwell Arrays for Measurement of Viability, Morphology, and Signaling Pathway Activation

    PubMed Central

    Singh, Manjulata; Close, David A.; Mukundan, Shilpaa; Johnston, Paul A.

    2015-01-01

    Abstract Despite significant investments in cancer research and drug discovery/development, the rate of new cancer drug approval is ≤5% and most cases of metastatic cancer remain incurable. Ninety-five percent of new cancer drugs fail in clinical development because of a lack of therapeutic efficacy and/or unacceptable toxicity. One of the major factors responsible for the low success rate of anticancer drug development is the failure of preclinical models to adequately recapitulate the complexity and heterogeneity of human cancer. For throughput and capacity reasons, high-throughput screening growth inhibition assays almost exclusively use two-dimensional (2D) monolayers of tumor cell lines cultured on tissue culture-treated plastic/glass surfaces in serum-containing medium. However, these 2D tumor cell line cultures fail to recapitulate the three-dimensional (3D) context of cells in solid tumors even though the tumor microenvironment has been shown to have a profound effect on anticancer drug responses. Tumor spheroids remain the best characterized and most widely used 3D models; however, spheroid sizes tend to be nonuniform, making them unsuitable for high-throughput drug testing. To circumvent this challenge, we have developed defined size microwell arrays using nonadhesive hydrogels that are applicable to a wide variety of cancer cell lines to fabricate size-controlled 3D microtumors. We demonstrate that the hydrogel microwell array platform can be applied successfully to generate hundreds of uniform microtumors within 3–6 days from many cervical and breast, as well as head and neck squamous cell carcinoma (HNSCC) cells. Moreover, controlling size of the microwells in the hydrogel array allows precise control over the size of the microtumors. Finally, we demonstrate the application of this platform technology to probe activation as well as inhibition of epidermal growth factor receptor (EGFR) signaling in 3D HNSCC microtumors in response to EGF and

  15. A hydrogel pen for electrochemical reaction and its applications for 3D printing

    NASA Astrophysics Data System (ADS)

    Kang, Hosuk; Hwang, Seongpil; Kwak, Juhyoun

    2014-12-01

    A hydrogel pen consisting of a microscopic pyramid containing an electrolyte offers a localized electroactive area on the nanometer scale via controlled contact of the apex with a working electrode. The hydrogel pen merges the fine control of atomic force microscopy with non-linear diffusion of an ultramicroelectrode, producing a faradaic current that depends on the small electroactive area. The theoretical and experimental investigations of the mass transport behavior within the hydrogel reveal that the steady-state current from the faradaic reaction is linearly proportional to the deformed length of the hydrogel pen by contact, i.e. signal transduction of deformation to an electrochemical signal, which enables the fine control of the electroactive area in the nanometer-scale regime. Combined with electrodeposition, localized electrochemistry of the hydrogel pen results in the ability to fabricate small sizes (110 nm in diameter), tall heights (up to 30 μm), and arbitrary structures, thereby indicating an additive process in 3 dimensions by localized electrodeposition.A hydrogel pen consisting of a microscopic pyramid containing an electrolyte offers a localized electroactive area on the nanometer scale via controlled contact of the apex with a working electrode. The hydrogel pen merges the fine control of atomic force microscopy with non-linear diffusion of an ultramicroelectrode, producing a faradaic current that depends on the small electroactive area. The theoretical and experimental investigations of the mass transport behavior within the hydrogel reveal that the steady-state current from the faradaic reaction is linearly proportional to the deformed length of the hydrogel pen by contact, i.e. signal transduction of deformation to an electrochemical signal, which enables the fine control of the electroactive area in the nanometer-scale regime. Combined with electrodeposition, localized electrochemistry of the hydrogel pen results in the ability to fabricate

  16. 3D Printing of Transparent and Conductive Heterogeneous Hydrogel-Elastomer Systems.

    PubMed

    Tian, Kevin; Bae, Jinhye; Bakarich, Shannon E; Yang, Canhui; Gately, Reece D; Spinks, Geoffrey M; In Het Panhuis, Marc; Suo, Zhigang; Vlassak, Joost J

    2017-03-01

    A hydrogel-dielectric-elastomer system, polyacrylamide and poly(dimethylsiloxane) (PDMS), is adapted for extrusion printing for integrated device fabrication. A lithium-chloride-containing hydrogel printing ink is developed and printed onto treated PDMS with no visible signs of delamination and geometrically scaling resistance under moderate uniaxial tension and fatigue. A variety of designs are demonstrated, including a resistive strain gauge and an ionic cable.

  17. A chitosan-hyaluronic acid hydrogel scaffold for periodontal tissue engineering.

    PubMed

    Miranda, Diego G; Malmonge, Sônia M; Campos, Doris M; Attik, Nina G; Grosgogeat, Brigitte; Gritsch, Kerstin

    2016-11-01

    The current challenge in treating periodontitis is regenerating the periodontium. This motivates tissue-engineering researchers to develop scaffolds as artificial matrices that give mechanical support for osteoblasts, cementoblasts, gingival and periodontal ligament fibroblast cells. In this study, modified hyaluronic acid (HA) and chitosan (CS) were employed to create a hybrid CS-HA hydrogel scaffold for periodontal regeneration. CS, HA, and CS-HA scaffolds were obtained by freeze-drying technique, resulting in porous structures suitable for use in tissue engineering. Scaffolds were submitted to gamma and UV-sterilization without significant morphology changes. The ATR-FTIR spectra of CS-HA hydrogels showed peaks at 377 cm(-1) , 1566 cm(-1) , and 1614 cm(-1) , representing secondary amide, primary amine, and carboxyl acid respectively, and it was also observed the emergence of peaks at 886 cm(-1) , which probably represents the Schiff base formed in the case of hybrid CS-HA hydrogels. The scaffolds presented a high rate of PBS uptake, reaching values higher than 95%. Thermal degradation of HA scaffolds was around 225°C and CS was around 285°C. The ATR-FTIR spectra and swelling degree were slightly disturbed mainly after gamma sterilization, but degradation temperature did not change after sterilization. The performance of the CS-HA hydrogel scaffolds for in vitro cell culture was tested using NIH3T3 and MG63 cell lines. The Alamar Blue test showed a significant increase in cellular viability and high CD44 expression, suggesting that the cells migrated more when seeded onto the scaffolds. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 1691-1702, 2016.

  18. Hierarchical bioceramic scaffolds with 3D-plotted macropores and mussel-inspired surface nanolayers for stimulating osteogenesis

    NASA Astrophysics Data System (ADS)

    Xu, Mengchi; Zhai, Dong; Xia, Lunguo; Li, Hong; Chen, Shiyi; Fang, Bing; Chang, Jiang; Wu, Chengtie

    2016-07-01

    The hierarchical structure of biomaterials plays an important role in the process of tissue reconstruction and regeneration. 3D-plotted scaffolds have been widely used for bone tissue engineering due to their controlled macropore structure and mechanical properties. However, the lack of micro- or nano-structures on the strut surface of 3D-plotted scaffolds, especially for bioceramic scaffolds, limits their biological activity. Inspired by the adhesive versatility of mussels and the active ion-chelating capacity of polydopamine, we set out to prepare a hierarchical bioceramic scaffold with controlled macropores and mussel-inspired surface nanolayers by combining the 3D-plotting technique with the polydopamine/apatite hybrid strategy in order to synergistically accelerate the osteogenesis and angiogenesis. β-Tricalcium phosphate (TCP) scaffolds were firstly 3D-plotted and then treated in dopamine-Tris/HCl and dopamine-SBF solutions to obtain TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds, respectively. It was found that polydopamine/apatite hybrid nanolayers were formed on the surface of both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds and TCP-DOPA-SBF scaffolds induced apatite mineralization for the second time during the cell culture. As compared to TCP scaffolds, both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds significantly promoted the osteogenesis of bone marrow stromal cells (BMSCs) as well as the angiogenesis of human umbilical vein endothelial cells (HUVECs), and the TCP-DOPA-SBF group presented the highest in vitro osteogenic/angiogenic activity among the three groups. Furthermore, both TCP-DOPA-Tris and TCP-DOPA-SBF scaffolds significantly improved the formation of new bone in vivo as compared to TCP scaffolds without a nanostructured surface. Our results suggest that the utilization of a mussel-inspired Ca, P-chelated polydopamine nanolayer on 3D-plotted bioceramic scaffolds is a viable and effective strategy to construct a hierarchical structure for synergistically

  19. Combination of thermal extrusion printing and ultrafast laser fabrication for the manufacturing of 3D composite scaffolds

    NASA Astrophysics Data System (ADS)

    Balčiūnas, Evaldas; Lukoševičius, Laurynas; Mackevičiūtė, Dovilė; Rekštytė, Sima; Rutkūnas, Vygandas; Paipulas, Domas; Stankevičiūtė, Karolina; Baltriukienė, Daiva; Bukelskienė, Virginija; Piskarskas, Algis P.; Malinauskas, Mangirdas

    2014-03-01

    We present a novel approach to manufacturing 3D microstructured composite scaffolds for tissue engineering applications. A thermal extrusion 3D printer - a simple, low-cost tabletop device enabling rapid materialization of CAD models in plastics - was used to produce cm-scale microporous scaffolds out of polylactic acid (PLA). The fabricated objects were subsequently immersed in a photosensitive monomer solution and direct laser writing technique (DLW) was used to refine its inner structure by fabricating a fine mesh inside the previously produced scaffold. In addition, a composite material structure out of four different materials fabricated via DLW is presented. This technique, empowered by ultrafast lasers allows 3D structuring with high spatial resolution in a great variety of photosensitive materials. A composite scaffold made of distinct materials and periodicities is acquired after the development process used to wash out non-linked monomers. Another way to modify the 3D printed PLA surfaces was also demonstrated - ablation with femtosecond laser beam. Structure geometry on macro- to micro- scales could be finely tuned by combining these fabrication techniques. Such artificial 3D substrates could be used for cell growth or as biocompatible-biodegradable implants. To our best knowledge, this is the first experimental demonstration showing the creation of composite 3D scaffolds using convenient 3D printing combined with DLW. This combination of distinct material processing techniques enables rapid fabrication of diverse functional micro-featured and integrated devices. Hopefully, the proposed approach will find numerous applications in the field of tissue engineering, as well as in microelectromechanical systems, microfluidics, microoptics and others.

  20. Collagen-coated polylactide microcarriers/chitosan hydrogel composite: injectable scaffold for cartilage regeneration.

    PubMed

    Hong, Yi; Gong, Yihong; Gao, Changyou; Shen, Jiacong

    2008-06-01

    A novel structure of injectable scaffold is designed and fabricated by combining collagen-coated polylactide (PLA) microcarriers and crosslinkable chitosan hydrogel. The collagen-coated PLA microcarriers were firstly mixed with the hydrogel precursor, a thickening agent of konjac glucomannan (KGM), and redox initiators of ammonium persulfate and tetramethylethylenediamine (TMEDA). The mixture was then injected into a mold and incubated at 37 degrees C to obtain the composite scaffold. The hydrogel can deliver the collagen-coated PLA microcarriers to the desired site and, after gelation, will prevent them from uncontrolled movement. On the other hand, the collagen-coated PLA microcarriers can substantially enhance the mechanical properties of the composite system. It was found that the microcarriers suspended stably in 0.6% KGM/1% chitosan derivative (CML) solution at 37 degrees C at least for 15 min. The dynamic elastic modulus (G') of the composite scaffold increased along with the increase of the microcarrier content. G' of the composite scaffold with 10% microcarriers was measured as 0.87-2.15 MPa at a frequency range of 0.1-100 rad/s, which was 120-90 times higher than that of its hydrogel system alone (12.1-24.4 kPa). In vitro culture of chondrocytes/composite scaffold showed that the cell metabolic activity increased rapidly before day 9, then leveled off. Cells in the hydrogel could attach and grow on the surface of the collagen-coated PLA microcarriers to form confluent cell layers after days 9-12. These features make the composite scaffold to be injectable and applicable in either tissue engineering, or regenerative medicine, and in particular, in orthopaedics.

  1. Resilin-PEG Hybrid Hydrogels Yield Degradable Elastomeric Scaffolds with Heterogeneous Microstructure.

    PubMed

    McGann, Christopher L; Akins, Robert E; Kiick, Kristi L

    2016-01-11

    Hydrogels derived from resilin-like polypeptides (RLPs) have shown outstanding mechanical resilience and cytocompatibility; expanding the versatility of RLP-based materials via conjugation with other polypeptides and polymers would offer great promise in the design of a range of materials. Here, we present an investigation of the biochemical and mechanical properties of hybrid hydrogels composed of a recombinant RLP and a multiarm PEG macromer. These hybrid hydrogels can be rapidly cross-linked through a Michael-type addition reaction between the thiols of cysteine residues on the RLP and vinyl sulfone groups on the multiarm PEG. Oscillatory rheology and tensile testing confirmed the formation of elastomeric hydrogels with mechanical resilience comparable to aortic elastin; hydrogel stiffness was easily modulated through the cross-linking ratio. Macromolecular phase separation of the RLP-PEG hydrogels offers the unique advantage of imparting a heterogeneous microstructure, which can be used to localize cells, through simple mixing and cross-linking. Assessment of degradation of the RLP by matrix metalloproteinases (MMPs) illustrated the specific proteolysis of the polypeptide in both its soluble form and when cross-linked into hydrogels. Finally, the successful encapsulation and viable three-dimensional culture of human mesenchymal stem cells (hMSCs) demonstrated the cytocompatibility of the RLP-PEG gels. Overall, the cytocompatibility, elastomeric mechanical properties, microheterogeneity, and degradability of the RLP-PEG hybrid hydrogels offer a suite of promising properties for the development of cell-instructive, structured tissue engineering scaffolds.

  2. Maintaining dimensions and mechanical properties of ionically crosslinked alginate hydrogel scaffolds in vitro.

    PubMed

    Kuo, Catherine K; Ma, Peter X

    2008-03-15

    Ionically crosslinked alginate hydrogels are attractive scaffolds because of their biocompatibility and mild gelation reaction that allows for gentle cell incorporation. However, the instability of ionically crosslinked hydrogels in an aqueous environment is a challenge that limits their application. This report presents a novel method to control the dimensions and mechanical properties of ionically crosslinked hydrogels via control of the ionic concentration of the medium. Homogeneous calcium-alginate gels were incubated in physiological saline baths adjusted to specific calcium ion concentrations. Swelling and shrinking occurred at low and high ionic concentrations of the medium, respectively, while an "optimal" intermediate calcium ion concentration of the medium was found to maintain original size and shape of the hydrogel. This optimal calcium ion concentration was found to be a function of crosslinking density and polymer concentration of the hydrogel and chemical composition of the alginate. The effects of optimal and high calcium ion concentrations of the medium on swelling behavior, calcium content, dry weight, and mechanical properties of the immersed hydrogels were investigated. It was found that the resulting hydrogel composition and mechanical properties depended on not only the calcium concentration of the medium, but also the crosslinking density and polymer concentration of the gel. In an 8-week experiment, controlled dimensions and mechanical properties of alginate gels in an aqueous environment were demonstrated. This new technique significantly enhances the potential of alginate hydrogels for tissue engineering and other biomedical applications.

  3. Fabrication and performance characteristics of tough hydrogel scaffolds based on biocompatible polymers.

    PubMed

    Islam, Atif; Yasin, Tariq; Gull, Nafisa; Khan, Shahzad Maqsood; Sabir, Aneela; Munawwar, Muhammad Azeem; Shafiq, Muhammad; Jamil, Tahir; Raza, Muhammad Hamid

    2016-11-01

    Novel silane crosslinked tough hydrogel scaffolds were prepared using chitosan (CS) and polyvinyl alcohol (PVA) to give network structure and scaffolds properties. The influence of crosslinking and PVA concentration on scaffolds were studied. Fourier transform infrared spectroscopy (FTIR) spectroscopy confirmed the presence of incorporated components. Tensile strength (TS) and fracture strain analysis of scaffolds were detected owing to the mutual effect of chemically and physically crosslinked network. Tough hydrogel scaffolds having 90% CS and 10% PVA exhibited TS of 49.18MPa and 10.15% elongation at break. The contact angle is less than 90(°) exhibited the hydrophilic nature of the scaffold. X-ray diffraction analysis (XRD) indicated the characteristics peaks fitting to CS and PVA and increase in the crystallinity of scaffolds. Cytotoxicity of scaffolds with different human fibroblast cell lines (F121, F192 and F84) for indirect method and human dermal fibroblast cell lines (F121) for direct method was evaluated. This indicated that these biomaterials were non-toxic, viable to the used cell lines, helpful in the growth of these cells and did not discharge toxic material damaging to the living cells.

  4. Self-Assembling Hydrogel Scaffolds for Photocatalytic Hydrogen Production

    PubMed Central

    Weingarten, Adam S.; Kazantsev, Roman V.; Palmer, Liam C.; McClendon, Mark; Koltonow, Andrew R.; Samuel, Amanda P. S.; Kiebala, Derek J.; Wasielewski, Michael R.; Stupp, Samuel I.

    2015-01-01

    Integration in a soft material of all molecular components necessary to generate storable fuels is an interesting target in supramolecular chemistry. The concept is inspired by the internal structure of photosynthetic organelles such as plant chloroplasts which co-localize molecules involved in light absorption, charge transport, and catalysis to create chemical bonds with light energy. We report here on the light-driven production of hydrogen inside a hydrogel scaffold built by the supramolecular self-assembly of a perylene monoimide amphiphile. The charged ribbons formed can electrostatically attract a nickel-based catalyst, and electrolyte screening promotes gelation. We found the emergent phenomenon that screening by the catalyst or the electrolytes led to two-dimensional crystallization of the chromophore assemblies and enhanced the electronic coupling among the molecules. Photocatalytic production of hydrogen is observed in the three-dimensional environment of the hydrogel scaffold and the material is easily placed on surfaces or in the pores of solid supports. The development of soft materials that integrate all necessary molecular components to generate storable fuels in the presence of sunlight is an unexplored area of chemistry with potential impact in renewable energy. Such systems could have advantages over the use of large volumes of liquids, dispersions of expensive or toxic inorganic particles, or complex devices. The use of such soft materials with integrated functions and high water content is bioinspired by the internal structure of chloroplasts in plants. These photosynthetic organelles have evolved to co-localize within stacked lipid bilayers in their stroma the protein machinery which integrates light-absorption, charge transport, and the catalytic functions necessary to convert light energy into chemical bonds1,2. Efforts to emulate natural photosynthetic systems over the past several decades have concentrated on the development of efficient

  5. Interpenetrated Si-HPMC/alginate hydrogels as a potential scaffold for human tissue regeneration.

    PubMed

    Viguier, Alexia; Boyer, Cecile; Chassenieux, Christophe; Benyahia, Lazhar; Guicheux, Jérôme; Weiss, Pierre; Rethore, Gildas; Nicolai, Taco

    2016-05-01

    Interpenetrated gels of biocompatible polysaccharides alginate and silanized hydroxypropyl methyl cellulose (Si-HPMC) have been studied in order to assess their potential as scaffolds for the regeneration of human tissues. Si-HPMC networks were formed by reduction of the pH to neutral and alginate networks were formed by progressive in situ release of Ca(2+). Linear and non-linear mechanical properties of the mixed gels at different polymer and calcium concentrations were compared with those of the corresponding single gels. The alginate/Si-HPMC gels were found to be stiffer than pure Si-HPMC gels, but weaker and more deformable than pure alginate gels. No significant difference was found for the maximum stress at rupture measured during compression for all these gels. The degrees of swelling or contraction in excess water at pH 7 as well as the release of Ca(2+) was measured as a function of time. Pure alginate gels contracted by as much as 50 % and showed syneresis, which was much reduced or even eliminated for mixed gels. The important release of Ca(2+) upon ageing for pure alginate gels was much reduced for the mixed gels. Furthermore, results of cytocompatibility assays indicated that there was no cytotoxicity of Si-HPMC/alginate hydrogels in 2D and 3D culture of human SW1353 cells. The results show that using interpenetrated Si-HPMC/alginate gels has clear advantages over the use of single gels for application in tissue regeneration.

  6. The fabrication of double layer tubular vascular tissue engineering scaffold via coaxial electrospinning and its 3D cell coculture.

    PubMed

    Ye, Lin; Cao, Jie; Chen, Lamei; Geng, Xue; Zhang, Ai-Ying; Guo, Lian-Rui; Gu, Yong-Quan; Feng, Zeng-Guo

    2015-12-01

    A continuous electrospinning technique was applied to fabricate double layer tubular tissue engineering vascular graft (TEVG) scaffold. The luminal layer was made from poly(ɛ-caprolac-tone)(PCL) ultrafine fibers via common single axial electrospinning followed by the outer layer of core-shell structured nanofibers via coaxial electrospinning. For preparing the outer layernano-fibers, the PCL was electrospun into the shell and both bovine serum albumin (BSA) and tetrapeptide val-gal-pro-gly (VAPG) were encapsulated into the core. The core-shell structure in the outer layer fibers was observed by transmission electron microscope (TEM). The in vitro release tests exhibited the sustainable release behavior of BSA and VAPG so that they provided a better cell growth environment in the interior of tubular scaffold wall. The in vitro culture of smooth muscle cells (SMCs) demonstrated their potential to penetrate into the scaffold wall for the 3D cell culture. Subsequently, 3D cell coculture was conducted. First, SMCs were seeded on the luminal surface of the scaffold and cultured for 5 days, and then endothelial cells (ECs) were also seeded on the luminal surface and cocultured with SMCs for another 2 days. After stained with antibodies, 3D cell distribution on the scaffold was revealed by confocal laser scanning microscopy (CLSM) where ECs were mainly located on the luminal surface whereas SMCs penetrated into the surface and distributed inside the scaffold wall. This double layer tubular scaffold with 3D cell distribution showed the promise to develop it into a novel TEVG for clinical trials in the near future.

  7. Development of bioartificial myocardium by electrostimulation of 3D collagen scaffolds seeded with stem cells.

    PubMed

    Haneef, Kanwal; Lila, Nermine; Benadda, Samira; Legrand, Fabien; Carpentier, Alain; Chachques, Juan C

    2012-06-05

    Electrostimulation (ES) can be defined as a safe physical method to induce stem cell differentiation. The aim of this study is to evaluate the effectiveness of ES on bone marrow mesenchymal stem cells (BMSCs) seeded in collagen scaffolds in terms of proliferation and differentiation into cardiomyocytes. BMSCs were isolated from Wistar rats and seeded into 3D collagen type 1 templates measuring 25 × 25 × 6 mm. Bipolar in vitro ES was performed during 21 days. Electrical impedance and cell proliferation were measured. Expression of cardiac markers was assessed by immunocytochemistry. Viscoelasticity of collagen matrix was evaluated. Electrical impedance assessments showed a low resistance of 234±41 Ohms which indicates good electrical conductivity of collagen matrix. Cell proliferation at 570 nm as significantly increased in ES groups after seven day (ES 0.129±0.03 vs non-stimulated control matrix 0.06±0.01, P=0.002) and after 21 days, (ES 0.22±0.04 vs control 0.13±0.01, P=0.01). Immunocytoche mistry of BMSCs after 21 days ES showed positive staining of cardiac markers, troponin I, connexin 43, sarcomeric alpha-actinin, slow myosin, fast myosin and desmin. Staining for BMSCs marker CD29 after 21 days was negative. Electrostimulation of cell-seeded collagen matrix changed stem cell morphology and biochemical characteristics, increasing the expression of cardiac markers. Thus, MSC-derived differentiated cells by electrostimulation grafted in biological scaffolds might result in a convenient tissue engineering source for myocardial diseases.

  8. Nanofibrillar hydrogel scaffolds from recombinant protein-based polymers with integrin- and proteoglycan-binding domains.

    PubMed

    Włodarczyk-Biegun, Małgorzata K; Werten, Marc W T; Posadowska, Urszula; Storm, Ingeborg M; de Wolf, Frits A; van den Beucken, Jeroen J J P; Leeuwenburgh, Sander C G; Cohen Stuart, Martien A; Kamperman, Marleen

    2016-12-01

    This study describes the design, production, and testing of functionalized variants of a recombinant protein-based polymer that forms nanofibrillar hydrogels with self-healing properties. With a view to bone tissue engineering applications, we equipped these variants with N-terminal extensions containing either (1) integrin-binding (RGD) or (2) less commonly studied proteoglycan-binding (KRSR) cell-adhesive motifs. The polymers were efficiently produced as secreted proteins using the yeast Pichia pastoris and were essentially monodisperse. The pH-responsive protein-based polymers are soluble at low pH and self-assemble into supramolecular fibrils and hydrogels at physiological pH. By mixing functionalized and nonfunctionalized proteins in different ratios, and adjusting pH, hydrogel scaffolds with the same protein concentration but varying content of the two types of cell-adhesive motifs were readily obtained. The scaffolds were used for the two-dimensional culture of MG-63 osteoblastic cells. RGD domains had a slightly stronger effect than KRSR domains on adhesion, activity, and spreading. However, scaffolds featuring both functional domains revealed a clear synergistic effect on cell metabolic activity and spreading, and provided the highest final degree of cell confluency. The mixed functionalized hydrogels presented here thus allowed to tailor the osteoblastic cell response, offering prospects for their further development as scaffolds for bone regeneration. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 3082-3092, 2016.

  9. Nanofibrillar hydrogel scaffolds from recombinant protein‐based polymers with integrin‐ and proteoglycan‐binding domains

    PubMed Central

    Włodarczyk‐Biegun, Małgorzata K.; Posadowska, Urszula; Storm, Ingeborg M.; de Wolf, Frits A.; van den Beucken, Jeroen J. J. P.; Leeuwenburgh, Sander C. G.; Cohen Stuart, Martien A.; Kamperman, Marleen

    2016-01-01

    Abstract This study describes the design, production, and testing of functionalized variants of a recombinant protein‐based polymer that forms nanofibrillar hydrogels with self‐healing properties. With a view to bone tissue engineering applications, we equipped these variants with N‐terminal extensions containing either (1) integrin‐binding (RGD) or (2) less commonly studied proteoglycan‐binding (KRSR) cell‐adhesive motifs. The polymers were efficiently produced as secreted proteins using the yeast Pichia pastoris and were essentially monodisperse. The pH‐responsive protein‐based polymers are soluble at low pH and self‐assemble into supramolecular fibrils and hydrogels at physiological pH. By mixing functionalized and nonfunctionalized proteins in different ratios, and adjusting pH, hydrogel scaffolds with the same protein concentration but varying content of the two types of cell‐adhesive motifs were readily obtained. The scaffolds were used for the two‐dimensional culture of MG‐63 osteoblastic cells. RGD domains had a slightly stronger effect than KRSR domains on adhesion, activity, and spreading. However, scaffolds featuring both functional domains revealed a clear synergistic effect on cell metabolic activity and spreading, and provided the highest final degree of cell confluency. The mixed functionalized hydrogels presented here thus allowed to tailor the osteoblastic cell response, offering prospects for their further development as scaffolds for bone regeneration. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 3082–3092, 2016. PMID:27449385

  10. Laser sintered porous polycaprolacone scaffolds loaded with hyaluronic acid and gelatin-grafted thermoresponsive hydrogel for cartilage tissue engineering.

    PubMed

    Lee, Ming-Yih; Tsai, Wen-Wei; Chen, His-Jung; Chen, Jyh-Ping; Chen, Chih-Hao; Yeh, Wen-Lin; An, Jia

    2013-01-01

    The aim of this study is to evaluate a soft/hard bi-phase scaffold for cartilage tissue engineering. Chondrocyte proliferation, glycoaminoglycan production and total collagen content are compared between laser-sintered porous polycaprolactone (PCL) scaffolds with and without a thermoresponsive hydrogel grafted with hyaluronic acid and gelatin. The in vitro results show that scaffolds loaded with hydrogel have a higher initial chondrocyte attachment than PCL scaffolds. At day 21 and 28, scaffolds loaded with hydrogel have a significantly higher glycosaminoglycan (GAG) production than PCL scaffolds alone, and total collagen content including collagen type II in the hydrogel-loaded group is three times higher than the group without hydrogel. It is concluded that the laser-sintered porous PCL scaffold has good cytocompatibility, and that the hydrogel phase is able to enhance initial chondrocytes attachment as well as GAG and collagen production of chondrocytes. This study suggests that a soft/hard bi-phase scaffold may be used for cartilage tissue engineering to enhance in vitro chondrogenesis.

  11. 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.

  12. Engineering Photocrosslinkable Bicomponent Hydrogel Constructs for Creating 3D Vascularized Bone.

    PubMed

    Kazemzadeh-Narbat, Mehdi; Rouwkema, Jeroen; Annabi, Nasim; Cheng, Hao; Ghaderi, Masoumeh; Cha, Byung-Hyun; Aparnathi, Mansi; Khalilpour, Akbar; Byambaa, Batzaya; Jabbari, Esmaiel; Tamayol, Ali; Khademhosseini, Ali

    2017-02-27

    Engineering bone tissue requires the generation of a highly organized vasculature. Cellular behavior is affected by the respective niche. Directing cellular behavior and differentiation for creating mineralized regions surrounded by vasculature can be achieved by controlling the pattern of osteogenic and angiogenic niches. This manuscript reports on engineering vascularized bone tissues by incorporating osteogenic and angiogenic cell-laden niches in a photocrosslinkable hydrogel construct. Two-step photolithography process is used to control the stiffness of the hydrogel and distribution of cells in the patterned hydrogel. In addittion, osteoinductive nanoparticles are utilized to induce osteogenesis. The size of microfabricated constructs has a pronounced effect on cellular organization and function. It is shown that the simultaneous presence of both osteogenic and angiogenic niches in one construct results in formation of mineralized regions surrounded by organized vasculature. In addition, the presence of angiogenic niche improves bone formation. This approach can be used for engineered constructs that can be used for treatment of bone defects.

  13. 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

  14. 3D-printed microfluidic chips with patterned, cell-laden hydrogel constructs.

    PubMed

    Knowlton, Stephanie; Yu, Chu Hsiang; Ersoy, Fulya; Emadi, Sharareh; Khademhosseini, Ali; Tasoglu, Savas

    2016-06-20

    Three-dimensional (3D) printing offers potential to fabricate high-throughput and low-cost fabrication of microfluidic devices as a promising alternative to traditional techniques which enables efficient design iterations in the development stage. In this study, we demonstrate a single-step fabrication of a 3D transparent microfluidic chip using two alternative techniques: a stereolithography-based desktop 3D printer and a two-step fabrication using an industrial 3D printer based on polyjet technology. This method, compared to conventional fabrication using relatively expensive materials and labor-intensive processes, presents a low-cost, rapid prototyping technique to print functional 3D microfluidic chips. We enhance the capabilities of 3D-printed microfluidic devices by coupling 3D cell encapsulation and spatial patterning within photocrosslinkable gelatin methacryloyl (GelMA). The platform presented here serves as a 3D culture environment for long-term cell culture and growth. Furthermore, we have demonstrated the ability to print complex 3D microfluidic channels to create predictable and controllable fluid flow regimes. Here, we demonstrate the novel use of 3D-printed microfluidic chips as controllable 3D cell culture environments, advancing the applicability of 3D printing to engineering physiological systems for future applications in bioengineering.

  15. 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.

  16. The effect of porosity on cell ingrowth into accurately defined, laser-made, polylactide-based 3D scaffolds

    NASA Astrophysics Data System (ADS)

    Danilevicius, Paulius; Georgiadi, Leoni; Pateman, Christopher J.; Claeyssens, Frederik; Chatzinikolaidou, Maria; Farsari, Maria

    2015-05-01

    The aim of this study is to demonstrate the accuracy required for the investigation of the role of solid scaffolds' porosity in cell proliferation. We therefore present a qualitative investigation into the effect of porosity on MC3T3-E1 pre-osteoblastic cell ingrowth of three-dimensional (3D) scaffolds fabricated by direct femtosecond laser writing. The material we used is a purpose made photosensitive pre-polymer based on polylactide. We designed and fabricated complex, geometry-controlled 3D scaffolds with pore sizes ranging from 25 to 110 μm, representing porosities 70%, 82%, 86%, and 90%. The 70% porosity scaffolds did not support cell growth initially and in the long term. For the other porosities, we found a strong adhesion of the pre-osteoblastic cells from the first hours after seeding and a remarkable proliferation increase after 3 weeks and up to 8 weeks. The 86% porosity scaffolds exhibited a higher efficiency compared to 82% and 90%. In addition, bulk material degradation studies showed that the employed, highly-acrylated polylactide is degradable. These findings support the potential use of the proposed material and the scaffold fabrication technique in bone tissue engineering.

  17. Soft Hydrogels Featuring In-Depth Surface Density Gradients for the Simple Establishment of 3D Tissue Models for Screening Applications.

    PubMed

    Zhang, Ning; Milleret, Vincent; Thompson-Steckel, Greta; Huang, Ning-Ping; Vörös, János; Simona, Benjamin R; Ehrbar, Martin

    2017-03-01

    Three-dimensional (3D) cell culture models are gaining increasing interest for use in drug development pipelines due to their closer resemblance to human tissues. Hydrogels are the first-choice class of materials to recreate in vitro the 3D extra-cellular matrix (ECM) environment, important in studying cell-ECM interactions and 3D cellular organization and leading to physiologically relevant in vitro tissue models. Here we propose a novel hydrogel platform consisting of a 96-well plate containing pre-cast synthetic PEG-based hydrogels for the simple establishment of 3D (co-)culture systems without the need for the standard encapsulation method. The in-depth density gradient at the surface of the hydrogel promotes the infiltration of cells deposited on top of it. The ability to decouple hydrogel production and cell seeding is intended to simplify the use of hydrogel-based platforms and thus increase their accessibility. Using this platform, we established 3D cultures relevant for studying stem cell differentiation, angiogenesis, and neural and cancer models.

  18. Subacute Tissue Response to 3D Graphene Oxide Scaffolds Implanted in the Injured Rat Spinal Cord.

    PubMed

    López-Dolado, Elisa; González-Mayorga, Ankor; Portolés, María Teresa; Feito, María José; Ferrer, María Luisa; Del Monte, Francisco; Gutiérrez, María Concepción; Serrano, María Concepción

    2015-08-26

    The increasing prevalence and high sanitary costs of lesions affecting the central nervous system (CNS) at the spinal cord are encouraging experts in different fields to explore new avenues for neural repair. In this context, graphene and its derivatives are attracting significant attention, although their toxicity and performance in the CNS in vivo remains unclear. Here, the subacute tissue response to 3D flexible and porous scaffolds composed of partially reduced graphene oxide is investigated when implanted in the injured rat spinal cord. The interest of these structures as potentially useful platforms for CNS regeneration mainly relies on their mechanical compliance with neural tissues, adequate biocompatibility with neural cells in vitro and versatility to carry topographical and biological guidance cues. Early tissue responses are thoroughly investigated locally (spinal cord at C6 level) and in the major organs (i.e., kidney, liver, lung, and spleen). The absence of local and systemic toxic responses, along with the positive signs found at the lesion site (e.g., filler effect, soft interface for no additional scaring, preservation of cell populations at the perilesional area, presence of M2 macrophages), encourages further investigation of these materials as promising components of more efficient material-based platforms for CNS repair.

  19. Using Polymer Confinement for Stem Cell Differentiation: 3D Printed vs Molded Scaffolds

    NASA Astrophysics Data System (ADS)

    Rafailovich, Miriam

    Additive manufacturing technologies are increasingly being used to replace standard extrusion or molding methods in engineering polymeric biomedical implants, which can be further seeded with cells for tissue regeneration. The principal advantage of this new technology is the ability to print directly from a scan and hence produce parts which are an ideal fit for an individual, eliminating much of the sizing and fitting associated with standard manufacturing methods. The question though arises whether devices which may be macroscopically similar, serve identical functions and are produced from the same material, interact in the same manner with cells and living tissue. Here we show that fundamental differences can exist between 3-D printed and extruded scaffolds which can impact stem cell differentiation and lineage selection. We will show how polymer confinement inherent in these methods affect the printed features on multiple length scales. We will also and how the differentiation of stem cells is affected by substrate heterogeneity in both morphological and mechanical features. NSF-Inspire award # 1344267.

  20. Bone response to 3-D periodic hydroxyapatite scaffolds with and without tailored microporosity to deliver bone morphogenetic protein 2.

    SciTech Connect

    Eurell, Jo Ann; Dellinger, Jennifer Gwynne; Cesarano, Joseph, III; Jamison, Russell D.

    2005-06-01

    The in vivo bone response of 3D periodic hydroxyapatite (HA) scaffolds is investigated. Two groups of HA scaffolds (11 mm diameter x 3.5 mm thick) are fabricated by direct-write assembly of a concentrated HA ink. The scaffolds consist of cylindrical rods periodically arranged into four quadrants with varying separation distances between rods. In the first group, HA rods (250 {micro}m in diameter) are patterned to create pore channels, whose areal dimensions are 250 x 250 {micro}m{sup 2} in quadrant 1, 250 x 500 {micro}m{sup 2} in quadrants 2 and 4, and 500 x 500 {micro}m{sup 2} in quadrant 3. In the second group, HA rods (400 {micro}m in diameter) are patterned to create pore channels, whose areal dimensions of 500 x 500 {micro}m{sup 2} in quadrant 1, 500 x 750 {micro}m{sup 2} in quadrants 2 and 4, and 750 x 750 {micro}m{sup 2} in quadrant 3. Each group of scaffolds is partially densified by sintering at 1200 C prior to being implanted bilaterally in trephine defects of skeletally mature New Zealand White rabbits. Their tissue response is evaluated at 8 and 16 weeks using micro-computed tomography, histology, and scanning electron microscopy. New trabecular bone is conducted rapidly and efficiently across substantial distances within these patterned 3D HA scaffolds. Our observations suggest that HA rods are first coated with a layer of new bone followed by subsequent scaffold infilling via outward and inward radial growth of the coated regions. Direct-write assembly of 3D periodic scaffolds composed of micro-porous HA rods arrayed to produce macro-pores that are size-matched to trabecular bone may represent an optimal strategy for bone repair and replacement structures.

  1. Artificial biomembranes stabilized over spin coated hydrogel scaffolds. Crosslinking agent nature induces wrinkled or flat surfaces on the hydrogel.

    PubMed

    González-Henríquez, C M; Pizarro-Guerra, G C; Córdova-Alarcón, E N; Sarabia-Vallejos, M A; Terraza-Inostroza, C A

    2016-03-01

    Hydrogel films possess the ability of retain water and deliver it to a phospholipid bilayer mainly composed by DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine); moisture of the medium favors the stability of an artificial biomembrane when it is subjected to repetitive heating cycles. This hypothesis is valid when the hydrogel film, used as scaffold, present a flat surface morphology and a high ability for water releasing. On the other hand, when the sample presents a wrinkle topography (periodic undulations), free lateral molecular movement of the bilayer becomes lower, disfavoring the occurrence of clear phases/phase transitions according to applied temperature. Hydrogel films were prepared using HEMA (hydroxyethylmetacrylate), different crosslinking agents and initiators. This reaction mixture was spread over hydrophilic silicon wafers using spin coating technique. Resultant films were then exposed to UV light favoring polymeric chain crosslinking and interactions between hydrogel and substrate; this process is also known to generate tensile stress mismatch between different hydrogel strata, producing out-of-plane net force that generate ordered undulations or collapsed crystals at surface level. DPPC bilayers were then placed over hydrogel using Langmuir-Blodgett technique. Surface morphology was detected in order to clarify the behavior of these films. Obtained data corroborate DPPC membrane stability making possible to detect phases/phase transitions by ellipsometric methods and Atomic Force Microscopy due to their high hydration level. This system is intended to be used as biosensor through the insertion of transmembrane proteins or peptides that detect minimal variations of some analyte in the environment; artificial biomembrane stability and behavior is fundamental for this purpose.

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

    PubMed Central

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

    2015-01-01

    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. PMID:27877821

  3. 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.

  4. 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.

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

    PubMed

    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.

  6. Hydrogel scaffolds promote neural gene expression and structural reorganization in human astrocyte cultures

    PubMed Central

    Knight, V. Bleu

    2017-01-01

    Biomaterial scaffolds have the potential to enhance neuronal development and regeneration. Understanding the genetic responses of astrocytes and neurons to biomaterials could facilitate the development of synthetic environments that enable the specification of neural tissue organization with engineered scaffolds. In this study, we used high throughput transcriptomic and imaging methods to determine the impact of a hydrogel, PuraMatrix™, on human glial cells in vitro. Parallel studies were undertaken with cells grown in a monolayer environment on tissue culture polystyrene. When the Normal Human Astrocyte (NHA) cell line is grown in a hydrogel matrix environment, the glial cells adopt a structural organization that resembles that of neuronal-glial cocultures, where neurons form clusters that are distinct from the surrounding glia. Statistical analysis of next generation RNA sequencing data uncovered a set of genes that are differentially expressed in the monolayer and matrix hydrogel environments. Functional analysis demonstrated that hydrogel-upregulated genes can be grouped into three broad categories: neuronal differentiation and/or neural plasticity, response to neural insult, and sensory perception. Our results demonstrate that hydrogel biomaterials have the potential to transform human glial cell identity, and may have applications in the repair of damaged brain tissue. PMID:28097054

  7. Engineering interconnected 3D vascular networks in hydrogels using molded sodium alginate lattice as the sacrificial template.

    PubMed

    Wang, Xue-Ying; Jin, Zi-He; Gan, Bo-Wen; Lv, Song-Wei; Xie, Min; Huang, Wei-Hua

    2014-08-07

    Engineering 3D perfusable vascular networks in vitro and reproducing the physiological environment of blood vessels is very challenging for tissue engineering and investigation of blood vessel function. Here, we engineer interconnected 3D microfluidic vascular networks in hydrogels using molded sodium alginate lattice as sacrificial templates. The sacrificial templates are rapidly replicated in polydimethylsiloxane (PDMS) microfluidic chips via Ca⁺²-crosslinking and then fully encapsulated in hydrogels. Interconnected channels with well controlled size and morphology are obtained by dissolving the monolayer or multilayer templates with EDTA solution. The human umbilical vein endothelial cells (HUVECs) are cultured on the channel linings and proliferated to form vascular lumens. The strong cell adhesion capability and adaptive response to shear stress demonstrate the excellent cytocompatibility of both the template and template-sacrificing process. Furthermore, the barrier function of the endothelial layer is characterized and the results show that a confluent endothelial monolayer is fully developed. Taken together, we develop a facile and rapid approach to engineer a vascular model that could be potentially used in physiological studies of vascular functions and vascular tissue engineering.

  8. 3D-printed hierarchical scaffold for localized isoniazid/rifampin drug delivery and osteoarticular tuberculosis therapy.

    PubMed

    Zhu, Min; Li, Kun; Zhu, Yufang; Zhang, Jianhua; Ye, Xiaojian

    2015-04-01

    After surgical treatment of osteoarticular tuberculosis (TB), it is necessary to fill the surgical defect with an implant, which combines the merits of osseous regeneration and local multi-drug therapy so as to avoid drug resistance and side effects. In this study, a 3D-printed macro/meso-porous composite scaffold is fabricated. High dosages of isoniazid (INH)/rifampin (RFP) anti-TB drugs are loaded into chemically modified mesoporous bioactive ceramics in advance, which are then bound with poly (3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) through a 3D printing procedure. The composite scaffolds show greatly prolonged drug release time compared to commercial calcium phosphate scaffolds either in vitro or in vivo. In addition, the drug concentrations on the periphery tissues of defect are maintained above INH/RFP minimal inhibitory concentrations even up to 12 weeks post-surgery, while they are extremely low in blood. Examinations of certain serum enzymes suggest no harm to hepatic or renal functions. Micro-CT evaluations and histology results also indicate partly degradation of the composite scaffolds and new bone growth in the cavity. These results suggest promising applications of our hierarchical composite scaffold in bone regeneration and local anti-TB therapy after osteoarticular TB debridement surgery.

  9. Hydrogels to Model 3D in vitro Microenvironment of Tumor Vascularization

    PubMed Central

    Song, Hyun-Ho Greco; Park, Kyung Min; Gerecht, Sharon

    2014-01-01

    A growing number of failing clinical trials for cancer therapy is substantiating the need to upgrade the current practice in culturing tumor cells and modeling tumor angiogenesis in vitro. Many attempts have been made to engineer vasculature in vitro by utilizing hydrogels, but the application of these tools in simulating in vivo tumor angiogenesis is still very new. In this review, we explore current use of hydrogels and their design parameters to engineer vasculogenesis and angiogenesis and to evaluate the angiogenic capability of cancerous cells and tissues. When coupled with other technologies such as lithography and three-dimensional printing, one can even create an advanced microvessel model as microfluidic channels to more accurately capture the native angiogenesis process. PMID:24969477

  10. 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.

  11. 3D scaffold of PLLA/pearl and PLLA/nacre powder for bone regeneration.

    PubMed

    Liu, Yuansheng; Huang, Qianli; Feng, Qingling

    2013-12-01

    Naturally occurring pearl and its derivatives have recently gained interest in bone regeneration due to their bioactive characteristics and good mechanical properties. In this study, three-dimensional scaffolds composed of poly-l-lactide (PLLA)/aragonite pearl powder, PLLA/vaterite pearl powder and PLLA/nacre powder were fabricated by freeze-drying. Scanning electron microscope (SEM) images indicated that the addition of powder made no visible difference to the morphology of the composite scaffolds. These composite scaffolds were found to have nearly twice the compressive strength and compressive modulus of the pure PLLA scaffold. X-ray diffraction patterns reveal that both PLLA/aragonite and PLLA/nacre composite scaffolds have pure aragonite crystals as their inorganic component, while PLLA/vaterite has pure vaterite crystals. The attachment and morphology of rat bone marrow-derived mesenchymal stem cells (rBMSCs) on scaffolds was observed by the SEM. The proliferation and osteogenic differentiation of rBMSCs on composite scaffolds was also investigated. The results indicate that PLLA/aragonite and PLLA/nacre scaffolds better stimulate cell proliferation and alkaline phosphatase activity than the PLLA scaffold. However, the PLLA/vaterite scaffold appears to decrease rBMSCs proliferation as well as the osteogenic differentiation, possibly due to the high pH of the solution containing PLLA/vaterite.

  12. A simple method for the production of large volume 3D macroporous hydrogels for advanced biotechnological, medical and environmental applications

    PubMed Central

    Savina, Irina N.; Ingavle, Ganesh C.; Cundy, Andrew B.; Mikhalovsky, Sergey V.

    2016-01-01

    The development of bulk, three-dimensional (3D), macroporous polymers with high permeability, large surface area and large volume is highly desirable for a range of applications in the biomedical, biotechnological and environmental areas. The experimental techniques currently used are limited to the production of small size and volume cryogel material. In this work we propose a novel, versatile, simple and reproducible method for the synthesis of large volume porous polymer hydrogels by cryogelation. By controlling the freezing process of the reagent/polymer solution, large-scale 3D macroporous gels with wide interconnected pores (up to 200 μm in diameter) and large accessible surface area have been synthesized. For the first time, macroporous gels (of up to 400 ml bulk volume) with controlled porous structure were manufactured, with potential for scale up to much larger gel dimensions. This method can be used for production of novel 3D multi-component macroporous composite materials with a uniform distribution of embedded particles. The proposed method provides better control of freezing conditions and thus overcomes existing drawbacks limiting production of large gel-based devices and matrices. The proposed method could serve as a new design concept for functional 3D macroporous gels and composites preparation for biomedical, biotechnological and environmental applications. PMID:26883390

  13. A simple method for the production of large volume 3D macroporous hydrogels for advanced biotechnological, medical and environmental applications

    NASA Astrophysics Data System (ADS)

    Savina, Irina N.; Ingavle, Ganesh C.; Cundy, Andrew B.; Mikhalovsky, Sergey V.

    2016-02-01

    The development of bulk, three-dimensional (3D), macroporous polymers with high permeability, large surface area and large volume is highly desirable for a range of applications in the biomedical, biotechnological and environmental areas. The experimental techniques currently used are limited to the production of small size and volume cryogel material. In this work we propose a novel, versatile, simple and reproducible method for the synthesis of large volume porous polymer hydrogels by cryogelation. By controlling the freezing process of the reagent/polymer solution, large-scale 3D macroporous gels with wide interconnected pores (up to 200 μm in diameter) and large accessible surface area have been synthesized. For the first time, macroporous gels (of up to 400 ml bulk volume) with controlled porous structure were manufactured, with potential for scale up to much larger gel dimensions. This method can be used for production of novel 3D multi-component macroporous composite materials with a uniform distribution of embedded particles. The proposed method provides better control of freezing conditions and thus overcomes existing drawbacks limiting production of large gel-based devices and matrices. The proposed method could serve as a new design concept for functional 3D macroporous gels and composites preparation for biomedical, biotechnological and environmental applications.

  14. Fabrication of 3D porous SF/β-TCP hybrid scaffolds for bone tissue reconstruction.

    PubMed

    Park, Hyun Jung; Min, Kyung Dan; Lee, Min Chae; Kim, Soo Hyeon; Lee, Ok Joo; Ju, Hyung Woo; Moon, Bo Mi; Lee, Jung Min; Park, Ye Ri; Kim, Dong Wook; Jeong, Ju Yeon; Park, Chan Hum

    2016-07-01

    Bio-ceramic is a biomaterial actively studied in the field of bone tissue engineering. But, only certain ceramic materials can resolve the corrosion problem and possess the biological affinity of conventional metal biomaterials. Therefore, the recent development of composites of hybrid composites and polymers has been widely studied. In this study, we aimed to select the best scaffold of silk fibroin and β-TCP hybrid for bone tissue engineering. We fabricated three groups of scaffold such as SF (silk fibroin scaffold), GS (silk fibroin/small granule size of β-TCP scaffold) and GM (silk fibroin/medium granule size of β-TCP scaffold), and we compared the characteristics of each group. During characterization of the scaffold, we used scanning electron microscopy (SEM) and a Fourier transform infrared spectroscopy (FTIR) for structural analysis. We compared the physiological properties of the scaffold regarding the swelling ratio, water uptake and porosity. To evaluate the mechanical properties, we examined the compressive strength of the scaffold. During in vitro testing, we evaluated cell attachment and cell proliferation (CCK-8). Finally, we confirmed in vivo new bone regeneration from the implanted scaffolds using histological staining and micro-CT. From these evaluations, the fabricated scaffold demonstrated high porosity with good inter-pore connectivity, showed good biocompatibility and high compressive strength and modulus. In particular, the present study indicates that the GM scaffold using β-TCP accelerates new bone regeneration of implanted scaffolds. Accordingly, our scaffold is expected to act a useful application in the field of bone tissue engineering. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1779-1787, 2016.

  15. The enhancement of cancer stem cell properties of MCF-7 cells in 3D collagen scaffolds for modeling of cancer and anti-cancer drugs.

    PubMed

    Chen, Lei; Xiao, Zhifeng; Meng, Yue; Zhao, Yannan; Han, Jin; Su, Guannan; Chen, Bing; Dai, Jianwu

    2012-02-01

    Three-dimensional (3D) culture could partially simulate in vivo conditions. In this work, we developed a 3D collagen scaffold to investigate cellular properties of MCF-7 cells. The porous scaffolds not only induced the diversification of cell morphologies but also extended cell proliferation. The expression of pro-angiogenic growth factors and the transcriptions of matrix metalloproteinases (MMPs) were significantly increased in cells cultured in 3D collagen scaffolds. In addition, 3D collagen scaffolds could generate a cell population with the properties of cancer stem cells (CSCs). The upregulation of EMT markers and the downregulation of the epithelial cell marker were observed in cells cultured in collagen scaffolds. The expression of stem cell markers, including OCT4A and SOX2, and breast cancer stem cell signatures, including SOX4, JAG1 and CD49F, was significantly unregulated in 3D collagen scaffolds. The proportion of cells with CSC-like CD44(+)/CD24(-/low) phenotype was notably increased. High-level expression of CSC-associated properties of MCF-7 cells cultured in 3D was further confirmed by high tumorigenicity in vivo. Moreover, xenografts with 3D cells formed larger tumors. The properties of MCF-7 cells in 3D may have partially simulated their in vivo behaviors. Thus, 3D collagen scaffolds might provide a useful platform for anti-cancer therapeutics and CSC research.

  16. Foamed oligo(poly(ethylene glycol)fumarate) hydrogels as versatile prefabricated scaffolds for tissue engineering.

    PubMed

    Henke, Matthias; Baumer, Julia; Blunk, Torsten; Tessmar, Joerg

    2014-03-01

    Radically cross-linked hydrogels are frequently used as cell carriers due to their excellent biocompatibility and their tissue-like mechanical properties. Through frequent investigation, PEG-based polymers such as oligo(poly(ethylene glycol)fumarate [OPF] have proven to be especially suitable as cell carriers by encapsulating cells during hydrogel formation. In some cases, NaCl or biodegradable gelatin microparticles were added prior to cross-linking in order to provide space for the proliferating cells, which would otherwise stay embedded in the hydrogel matrix. However, all of these immediate cross-linking procedures involve time consuming sample preparation and sterilization directly before cell culture and often show notable swelling after their preparation. In this study, ready to use OPF-hydrogel scaffolds were prepared by gas foaming, freeze drying, individual packing into bags and subsequent γ-sterilization. The scaffolds could be stored and used "off-the-shelf" without any need for further processing prior to cell culture. Thus the handling was simplified and the sterility of the cell carrier was assured. Further improvement of the gel system was achieved using a two component injectable system, which may be used for homogenous injection molding in order to create individually shaped three dimensional scaffolds. In order to evaluate the suitability of the scaffolds for tissue engineering, constructs were seeded with juvenile bovine chondrocytes and cultured for 28 days. Cross-sections of the respective constructs showed an intense and homogenous red staining of GAG with safranin O, indicating a homogenous cell distribution within the scaffolds and the production of substantial amounts of GAG-rich matrix.

  17. Heparin interacting protein mediated assembly of nano-fibrous hydrogel scaffolds for guided stem cell differentiation.

    PubMed

    Tan, Huaping; Zhou, Qixin; Qi, Haifeng; Zhu, Dan; Ma, Xiaoxue; Xiong, Dangsheng

    2012-05-01

    A new methodology is developed to conjugate hyaluronic acid (HA) hydrogel with novel nano-fibrous architectures via non-covalent assembly that specifically allows for targeted adipose-derived stem cells (ASCs) differentiation and soft tissue engineering. The assembly of non-covalently associated hydrogel network produced via the interaction of a low molecular weight heparin (LMWH) modified HA derivative and heparin interacting protein (HIP). The multifunctional star poly(ethylene glycol) (PEG) and HIP copolymer has the capability to mediate the non-covalent assembly of nano-fibrous HA hydrogel networks via affinity interactions with LMWH. The effect of the HIP mediation on in vitro gelation, rheological characteristics, degradation, equilibrium swelling, adipose-derived stem cells (ASCs) proliferation and differentiation of nano-fibrous hydrogel is examined. The results suggest the potential utility of this unique design of the bioactive nano-fibrous HA hydrogel in directing the differentiation of ASCs and adipogenesis in ECM-mimetic scaffolds in vitro. These studies demonstrate that this nano-fibrous HA hydrogel can render the formulation of a therapeutically effective platform for in vitro adipogenesis applications.

  18. Fabrication and characterization of strontium incorporated 3-D bioactive glass scaffolds for bone tissue from biosilica.

    PubMed

    Özarslan, Ali Can; Yücel, Sevil

    2016-11-01

    Bioactive glass scaffolds that contain silica are high viable biomaterials as bone supporters for bone tissue engineering due to their bioactive behaviour in simulated body fluid (SBF). In the human body, these materials help inorganic bone structure formation due to a combination of the particular ratio of elements such as silicon (Si), calcium (Ca), sodium (Na) and phosphorus (P), and the doping of strontium (Sr) into the scaffold structure increases their bioactive behaviour. In this study, bioactive glass scaffolds were produced by using rice hull ash (RHA) silica and commercial silica based bioactive glasses. The structural properties of scaffolds such as pore size, porosity and also the bioactive behaviour were investigated. The results showed that undoped and Sr-doped RHA silica-based bioactive glass scaffolds have better bioactivity than that of commercial silica based bioactive glass scaffolds. Moreover, undoped and Sr-doped RHA silica-based bioactive glass scaffolds will be able to be used instead of undoped and Sr-doped commercial silica based bioactive glass scaffolds for bone regeneration applications. Scaffolds that are produced from undoped or Sr-doped RHA silica have high potential to form new bone for bone defects in tissue engineering.

  19. 3D printed tricalcium phosphate scaffolds: Effect of SrO and MgO doping on in vivo osteogenesis in a rat distal femoral defect model.

    PubMed

    Tarafder, Solaiman; Davies, Neal M; Bandyopadhyay, Amit; Bose, Susmita

    2013-12-01

    The presence of interconnected macro pores is important in tissue engineering scaffolds for guided tissue regeneration. This study reports in vivo biological performance of interconnected macro porous tricalcium phosphate (TCP) scaffolds due to the addition of SrO and MgO as dopants in TCP. We have used direct three dimensional printing (3DP) technology for scaffold fabrication followed by microwave sintering. Mechanical strength was evaluated by scaffolds with 500 µm, 750 µm, and 1000 µm interconnected designed pore sizes. Maximum compressive strength of 12.01 ± 1.56 MPa was achieved for 500 µm interconnected designed pore size Sr-Mg doped scaffold. In vivo biological performance of the microwave sintered pure TCP and Sr-Mg doped TCP scaffolds was assessed by implanting 350 µm designed interconnected macro porous scaffolds in rat distal femoral defect. Sintered pore size of these 3D printed scaffolds were 311 ± 5.9 µm and 245 ± 7.5 µm for pure and SrO-MgO doped TCP scaffolds, respectively. These 3D printed scaffolds possessed multiscale porosity, i.e., 3D interconnected designed macro pores along with intrinsic micro pores. Histomorphology and histomorphometric analysis revealed a significant increase in osteoid like new bone formation, and accelerated mineralization inside SrO and MgO doped 3D printed TCP scaffolds as compared to pure TCP scaffolds. An increase in osteocalcin and type I collagen level was also observed in rat blood serum with SrO and MgO doped TCP scaffolds compared to pure TCP scaffolds. Our results show that these 3D printed SrO and MgO doped TCP scaffolds with multiscale porosity contributed to early healing through accelerated osteogenesis.

  20. 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.

  1. In vivo guided vascular regeneration with a non-porous elastin-like polypeptide hydrogel tubular scaffold.

    PubMed

    Mahara, Atsushi; Kiick, Kristi L; Yamaoka, Tetsuji

    2017-01-28

    Herein, we demonstrate a new approach for small-caliber vascular reconstruction using a non-porous elastin-like polypeptide hydrogel tubular scaffold, based on the concept of guided vascular regeneration (GVR). The scaffolds are composed of elastin-like polypeptide, (Val-Pro-Gly-Ile-Gly)n , for compliance matching and antithrombogenicity and an Arg-Gly-Asp (RGD) motif for connective tissue regeneration. When the polypeptide was mixed with an aqueous solution of β-[Tris(hydroxymethyl)phosphino]propionic acid at 37°C, the polypeptide hydrogel was rapidly formed. The elastic modulus of the hydrogel was 4.4kPa. The hydrogel tubular scaffold was formed in a mold and reinforced with poly(lactic acid) nanofibers. When tubular scaffolds with an inner diameter of 1 mm and length of 5 mm were implanted into rat abdominal aortae, connective tissue grew along the scaffold luminal surface from the flanking native tissues, resulting in new blood vessel tissue with a thickness of 200 μm in 1 month. In contrast, rats implanted with control scaffolds without the RGD motif died. These results indicate that the non-porous hydrogel tubular scaffold containing the RGD motif effectively induced rapid tissue regeneration and that GVR is a promising strategy for the regeneration of small-diameter blood vessels. This article is protected by copyright. All rights reserved.

  2. Biodegradable HA-PLA 3-D porous scaffolds: effect of nano-sized filler content on scaffold properties.

    PubMed

    Kothapalli, Chandrasekhar R; Shaw, Montgomery T; Wei, Mei

    2005-11-01

    Scaffolds comprising poly(lactic acid) and nano-hydroxyapatite (HA) were prepared using the solvent-casting/salt-leaching technique. NaCl was used as the leaching agent. Nano-sized HA was synthesized by a hydrothermal method at 170 degrees C and autogenous pressure. High-resolution TEM imaging revealed that the HA particles were ellipsoidal-shaped with needle-like morphologies. The particles had an average size of approximately 25 nm in width and 150 nm in length with aspect ratios ranging from 6 to 8. As the HA content increased in the scaffold from 0 to 50 wt%, the compression modulus of the scaffolds increased from 4.72+/-1.2 to 9.87+/-1.8 MPa, while the yield strength from 0.29+/-0.03 to 0.44+/-0.01 MPa. Such polymeric scaffolds should be suitable materials for non-load sharing tissue-engineering applications.

  3. Visible light cured thiol-vinyl hydrogels with tunable degradation for 3D cell culture

    PubMed Central

    Hao, Yiting; Shih, Han; Muňoz, Zachary; Kemp, Arika; Lin, Chien-Chi

    2013-01-01

    We report here a synthetically simple yet highly tunable and diverse visible light mediated thiol-vinyl gelation system for fabricating cell-instructive hydrogels. Gelation was achieved via a mixed-mode step-and-chain-growth photopolymerization using functionalized 4-arm poly(ethylene glycol) as backbone macromer, eosin-Y as photosensitizer, and di-thiol containing molecule as dual purpose co-initiator/cross-linker. N-vinylpyrrolidone (NVP) was used to accelerate gelation kinetics and to adjust the stiffness of the hydrogels. Visible light (wavelength: 400–700nm) was used to initiate rapid gelation (gel points: ~20 seconds) that reached completion within a few minutes. The major differences between current thiol-vinyl gelation and prior visible light mediated photopolymerization are that: (1) the co-initiator triethanolamine (TEOA) used in the previous systems was replaced with multifunctional thiols and (2) mixed-mode polymerized gels contain less network heterogeneity. The gelation kinetics and gel properties at the same PEG macromer concentration could be tuned by changing the identity of vinyl groups and di-thiol cross-linkers, as well as concentration of cross-linker and NVP. Specifically, acrylate-modified PEG afforded the fastest gelation rate, followed by acrylamide and methacrylate-functionalized PEG. Increasing NVP concentration also accelerated gelation and led to a higher network cross-linking density. Further, increasing di-thiol peptide concentration in the gel formulation increased hydrogel swelling and decreased gel stiffness. Due to the formation of thiol-ether-ester bonds following thiol-acrylate reaction, the gels degraded hydrolytically following a pseudo first order degradation kinetics. Degradation rate was controlled by adjusting thiol or NVP content in the polymer precursor solution. The cytocompatibility and utility of this hydrogel system were evaluated using in situ encapsulation of human mesenchymal stem cells (hMSC). Encapsulated h

  4. Biopolymeric hydrogels - nanostructured TiO2 hybrid materials as potential injectable scaffolds for bone regeneration.

    PubMed

    Zazakowny, Karolina; Lewandowska-Łańcucka, Joanna; Mastalska-Popławska, Joanna; Kamiński, Kamil; Kusior, Anna; Radecka, Marta; Nowakowska, Maria

    2016-12-01

    The present work aims at development of novel hybrid materials from genipin crosslinked collagen or collagen/chitosan hydrogels containing various types of TiO2 nanoparticles characterized with different anatase/rutile ratios. Collagen and chitosan were selected as hydrogel components since they are biopolymers being, like collagen, the major compound present in extracellular matrix or exhibit structural similarity to glycosaminoglycans, like chitosan. TiO2 nanoparticles were introduced to the hydrogel matrices to improve their mechanical properties as well as bioactivity. A series of twelve novel hybrid materials were prepared and their physicochemical, mechanical and biological properties were evaluated. It was found that TiO2 nanostructures introduced to the hydrogels have significant influence on the swelling properties of the synthesized hybrids and their impact is strongly dependent on the type of matrices. The surfaces of hybrid materials were found to be more hydrophilic than these of corresponding hydrogel matrix. It was also observed that, the storage modulus values of the hybrids based on collagen-chitosan hydrogel are comparable to these for plain hydrogels what indicates that the mechanical properties of the materials obtained are satisfactory for possible biomedical application. The in vitro cell culture studies have shown that prepared materials are biocompatible as they can support mitochondrial activity of MEFs as well as MG-63 cells. In vitro experiments performed under simulated body fluid (SBF) conditions have revealed that all studied TiO2 nanoparticles present in hydrogel matrices, regardless of anatase/rutile ratio, successfully induced formation of apatite-like structures. The hybrid materials developed here are promising candidates for preparation of bioactive, injectable scaffolds for tissue engineering.

  5. Effects of proanthocyanidin, a crosslinking agent, on physical and biological properties of collagen hydrogel scaffold

    PubMed Central

    Choi, Yoorina; Kim, Hee-Jin

    2016-01-01

    Objectives The purpose of the present study was to evaluate the effects of proanthocyanidin (PAC), a crosslinking agent, on the physical properties of a collagen hydrogel and the behavior of human periodontal ligament cells (hPDLCs) cultured in the scaffold. Materials and Methods Viability of hPDLCs treated with PAC was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The physical properties of PAC treated collagen hydrogel scaffold were evaluated by the measurement of setting time, surface roughness, and differential scanning calorimetry (DSC). The behavior of the hPDLCs in the collagen scaffold was evaluated by cell morphology observation and cell numbers counting. Results The setting time of the collagen scaffold was shortened in the presence of PAC (p < 0.05). The surface roughness of the PAC-treated collagen was higher compared to the untreated control group (p < 0.05). The thermogram of the crosslinked collagen exhibited a higher endothermic peak compared to the uncrosslinked one. Cells in the PAC-treated collagen were observed to attach in closer proximity to one another with more cytoplasmic extensions compared to cells in the untreated control group. The number of cells cultured in the PAC-treated collagen scaffolds was significantly increased compared to the untreated control (p < 0.05). Conclusions Our results showed that PAC enhanced the physical properties of the collagen scaffold. Furthermore, the proliferation of hPDLCs cultured in the collagen scaffold crosslinked with PAC was facilitated. Conclusively, the application of PAC to the collagen scaffold may be beneficial for engineering-based periodontal ligament regeneration in delayed replantation. PMID:27847751

  6. Toxicity and biocompatibility profile of 3D bone scaffold developed by Universitas Indonesia: A preliminary study

    NASA Astrophysics Data System (ADS)

    Rahyussalim A., J.; Kurniawati, T.; Aprilya, D.; Anggraini, R.; Ramahdita, Ghiska; Whulanza, Yudan

    2017-02-01

    Scaffold as a biomaterial must fulfill some requirements to be safely implanted to the human body. Toxicity and biocompatibility test are needed to evaluate scaffold material in mediating cell proliferation and differentiation, secreting extracelullar matrix and carrying biomolecular signals for cell communication. An in vitro study with mesenchymal stem cells consisted of direct contact test and indirect contact test using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium reduction assay was conducted on 4 scaffolds made of poly-L-lactic acid (PLA), polyvinyl alcohol (PVA), and hydroxyapatite-poly (vinyl alcohol) composite. There were cells-substrate adhesion impairment, morphological changes, cell death and reduction in cell proliferation seen at 2nd and 6th day in most tested scaffold. Cell count result at day-6 showed proliferation inhibition of more than 50% cell death (inhibition value >50) in all tested scaffold. In MTT assay, two scaffolds were proven non-toxic. In conclusion, various scaffold materials showed different toxicity effect. The toxicity and biocompatibility profile in this study is a preliminary data for further research aiming to use those local-made scaffolds to fill human bone defect in various needs.

  7. A building-block approach to 3D printing a multichannel, organ-regenerative scaffold.

    PubMed

    Wang, Xiaohong; Rijff, Boaz Lloyd; Khang, Gilson

    2015-06-29

    Multichannel scaffolds, formed by rapid prototyping technologies, retain a high potential for regenerative medicine and the manufacture of complex organs. This study aims to optimize several parameters for producing poly(lactic-co-glycolic acid) (PLGA) scaffolds by a low-temperature, deposition manufacturing, three-dimensional printing (3DP, or rapid prototyping) system. Concentration of the synthetic polymer solution, nozzle speed and extrusion rate were analysed and discussed. Polymer solution with a concentration of 12% w/v was determined as optimal for formation; large deviation of this figure failed to maintain the desired structure. The extrusion rate was also modified for better construct quality. Finally, several solid organ scaffolds, such as the liver, with proper wall thickness and intact contour were printed. This study gives basic instruction to design and fabricate scaffolds with de novo material systems, particularly by showing the approximation of variables for manufacturing multichannel PLGA scaffolds. Copyright © 2015 John Wiley & Sons, Ltd.

  8. 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.

  9. 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.

  10. Urethral reconstruction with a 3D porous bacterial cellulose scaffold seeded with lingual keratinocytes in a rabbit model.

    PubMed

    Huang, Jian-Wen; Lv, Xiang-Guo; Li, Zhe; Song, Lu-Jie; Feng, Chao; Xie, Min-Kai; Li, Chao; Li, Hong-Bin; Wang, Ji-Hong; Zhu, Wei-Dong; Chen, Shi-Yan; Wang, Hua-Ping; Xu, Yue-Min

    2015-09-11

    The goal of this study was to evaluate the effects of urethral reconstruction with a three-dimensional (3D) porous bacterial cellulose (BC) scaffold seeded with lingual keratinocytes in a rabbit model. A novel 3D porous BC scaffold was prepared by gelatin sponge interfering in the BC fermentation process. Rabbit lingual keratinocytes were isolated, expanded, and seeded onto 3D porous BC. BC alone (group 1, N  =  10), 3D porous BC alone (group 2, N  =  10), and 3D porous BC seeded with lingual keratinocytes (group 3, N  =  10) were used to repair rabbit ventral urethral defects (2.0   ×   0.8 cm). Scanning electron microscopy revealed that BC consisted of a compact laminate while 3D porous BC was composed of a porous sheet buttressed by a dense outer layer. The average pore diameter and porosity of the 3D porous BC were 4.23   ±   1.14 μm and 67.00   ±   6.80%, respectively. At 3 months postoperatively, macroscopic examinations and retrograde urethrograms of urethras revealed that all urethras maintained wide calibers in group 3. Strictures were found in all rabbits in groups 1 and 2. Histologically, at 1 month postoperatively, intact epithelium occurred in group 3, and discontinued epithelium was found in groups 1 and 2. However, groups 2 and 3 exhibited similar epithelial regeneration, which was superior to that of group 1 at 3 months (p  <  0.05). Comparisons of smooth muscle content and endothelia density among the three groups revealed a significant increase at each time point (p  <  0.05). Our results demonstrated that 3D porous BC seeded with lingual keratinocytes enhanced urethral tissue regeneration. 3D porous BC could potentially be used as an optimized scaffold for urethral reconstruction.

  11. Alginate hydrogel as a promising scaffold for dental-derived stem cells: an in vitro study.

    PubMed

    Moshaverinia, Alireza; Chen, Chider; Akiyama, Kentaro; Ansari, Sahar; Xu, Xingtian; Chee, Winston W; Schricker, Scott R; Shi, Songtao

    2012-12-01

    The objectives of this study were to: (1) develop an injectable and biodegradable scaffold based on oxidized alginate microbeads encapsulating periodontal ligament (PDLSCs) and gingival mesenchymal stem cells (GMSCs); and (2) investigate the stem cell viability, and osteogenic differentiation of the stem cells in vitro. Stem cells were encapsulated using alginate hydrogel. The stem cell viability, proliferation and differentiation to adipogenic and osteogenic tissues were studied. To investigate the expression of both adipogenesis and ontogenesis related genes, the RNA was extracted and RT-PCR was performed. The degradation behavior of hydrogel based on oxidized sodium alginate with different degrees of oxidation was studied in PBS at 37 °C as a function of time by monitoring the changes in weight loss. The swelling kinetics of alginate hydrogel was also investigated. The results showed that alginate is a promising candidate as a non-toxic scaffold for PDLSCs and GMSCs. It also has the ability to direct the differentiation of these stem cells to osteogenic and adipogenic tissues as compared to the control group in vitro. The encapsulated stem cells remained viable in vitro and both osteo-differentiated and adipo-differentiated after 4 weeks of culturing in the induction media. It was found that the degradation profile and swelling kinetics of alginate hydrogel strongly depends on the degree of oxidation showing its tunable chemistry and degradation rate. These findings demonstrate for the first time that immobilization of PDLSCs and GMSCs in the alginate microspheres provides a promising strategy for bone tissue engineering.

  12. Differentiation and Behavior of Dental Pulp Stem Cells in Hydrogel Scaffolds of Various Stiffnesses

    NASA Astrophysics Data System (ADS)

    Bhatnagar, Divya; Jurukovski, Vladimir; Rafailovich, Miriam; Simon, Marcia

    2011-03-01

    Dental Pulp Stem Cells (DPSCs) are known to differentiate in bone, dentine, or nerve tissue through different environment signals. This work investigates whether differentiation could occur in the absence of chemical induction and through mechanical stimuli only. For this study, we chose enzymatically cross-linked gelatin hydrogels as our substrates. Rheological studies carried out by oscillatory shear rheometry indicated that the modulus of the hardest hydrogel was of the order of 8kPa where as the medium and the softest hydrogel had modulus of the order of 1kPa and 100Pa respectively. DPSC were then plated on all three substrates and cultured with and without dexamethasone induction media. After 21 days of incubation, SEM analysis indicated that the cells cultured in the induction media produced biomineralized deposits on hard, medium as well as soft hydrogels. On the other hand, the cells cultured without the induction media also produced large amounts of biomineralized deposits.The modulus of the cells was also measured using AFM. En mass cell migration was also studied to determine the average velocity of migration of DPSCs. We also investigated whether stem cells that are induced to differentiate by their scaffold environment would continue to differentiate and biomineralize when removed from the inducing scaffold.

  13. Microwave-assisted fabrication of chitosan-hydroxyapatite superporous hydrogel composites as bone scaffolds.

    PubMed

    Beşkardeş, Işıl Gerçek; Demirtaş, Tuğrul Tolga; Durukan, Müge Dağlı; Gümüşderelioğlu, Menemşe

    2015-11-01

    In this study, a novel scaffold fabrication method was developed by combining microwave irradiation and gas foaming. Chitosan superporous hydrogels (SPHs) and chitosan-hydroxyapatite (HA) superporous hydrogel composites (SPHCs) were prepared by using this method in the presence of crosslinking agent, glyoxal, and a gas-blowing agent, NaHCO3. In order to examine the effect of HA on composite structure and cellular behaviour, two types of HA particles, i.e. spherical beads in 45-80 µm diameter and powder form, were used. While rapid heating with microwave irradiation enhances gas blowing, pH increment, which is accelerated by NaHCO3 decomposition, provides better crosslinking. Thus, interconnected and well-established macroporous hydrogels/hydrogel composites were produced easily and rapidly (~1 min). Cell culture studies, which were carried out under static and dynamic conditions with MC3T3-E1 pre-osteoblastic cells, indicated that chitosan-HA bead SPHCs supported cellular proliferation and osteoblastic differentiation better than chitosan SPHs and chitosan-HA powder SPHCs. In conclusion, simultaneous gas foaming and microwave crosslinking can be evaluated for the preparation of composite scaffolds which have superior properties for bone tissue engineering.

  14. Diagnostics of 3D Scaffolds by the Method of X-Ray Phase Contrast Visualization

    NASA Astrophysics Data System (ADS)

    Al'tapova, V. R.; Khlusov, I. A.; Karpov, D. A.; Chen, F.; Baumbach, T.; Pichugin, V. F.

    2014-02-01

    Polymers are one of the most interesting classes of materials for bioengineering due to their high biocompatibility and the possibility of regulating their strength and degradation. In bioengineering, the design of a polymer scaffold determines the functional possibilities of the scaffold and its possible medical applications. Traditionally, the design of polymer scaffolds is analyzed with the help of two-dimensional visualization methods, such as optical and electron microscopy, and computer tomography. However, the x-ray region of the electromagnetic spectrum is only insignificantly absorbed by polymers and soft tissue, which means that it does not support computer tomography with sufficient contrast. The present work investigates visualization with the help of an interferometer based on the Talbot effect for three-dimensional visualization of a polymer scaffold in absorption, phase, and dark-field contrasts. A comparison of images obtained by x-ray visualization with histological sections of the scaffold is made. Phase contrast has made it possible to visualize the polymer structure and growth of soft tissues in the volume of the scaffold. In the future, it will be possible to use phase contrast for three-dimensional visualization of polymer scaffolds and soft tissues in vivo as well as in vitro.

  15. In vivo tissue has non-linear rheological behavior distinct from 3D biomimetic hydrogels, as determined by AMOTIV microscopy.

    PubMed

    Blehm, Benjamin H; Devine, Alexus; Staunton, Jack R; Tanner, Kandice

    2016-03-01

    Variation in matrix elasticity has been shown to determine cell fate in both differentiation and development of malignant phenotype. The tissue microenvironment provides complex biochemical and biophysical signals in part due to the architectural heterogeneities found in extracellular matrices (ECMs). Three dimensional cell cultures can partially mimic in vivo tissue architecture, but to truly understand the role of viscoelasticity on cell fate, we must first determine in vivo tissue mechanical properties to improve in vitro models. We employed Active Microrheology by Optical Trapping InVivo (AMOTIV), using in situ calibration to measure in vivo zebrafish tissue mechanics. Previously used trap calibration methods overestimate complex moduli by ∼ 2-20 fold compared to AMOTIV. Applying differential microscale stresses and strains showed that hyaluronic acid (HA) gels display semi-flexible polymer behavior, while laminin-rich ECM hydrogels display flexible polymer behavior. In contrast, zebrafish tissues displayed different moduli at different stresses, with higher power law exponents at lower stresses, indicating that living tissue has greater stress dependence than the 3D hydrogels examined. To our knowledge, this work is the first vertebrate tissue rheological characterization performed in vivo. Our fundamental observations are important for the development and refinement of in vitro platforms.

  16. Glycerol-mediated nanostructure modification leading to improved transparency of porous polymeric scaffolds for high performance 3D cell imaging.

    PubMed

    Zhao, Shan; Shen, Zhiyuan; Wang, Jingyu; Li, Xiaokang; Zeng, Yang; Wang, Bingjie; He, Yonghong; Du, Yanan

    2014-07-14

    Glycerol is among the most commonly used optical clearing agents for tissues clearance largely due to refractive index (RI) matching between glycerol and the submerged tissues. Here we applied glycerol as structure modifier at both macroscopic (as porogen) and nanoscopic (as nanostructure ameliorant) scales to fabricate transparent porous scaffolds made from poly(ethylene glycol) (PEG) as well as other widely used biomaterials (e.g., PLGA, PA, or gelatin), whose nanostructures, in the scale of light wavelength, dominantly improved the optical transmittance of the scaffolds even when immersed in RI mismatched medium (e.g., culture medium or water). We further exploited the clearing mechanisms based on Mie scattering theory, illustrating that conformational changes of polymer chains induced by solvent effects of glycerol enhanced the anisotropy (i.e., directional alignment) of the nanostructures, leading to reduced crystallinity and scattering of the resulted PEG scaffolds. Our findings represent the first and systematic demonstration with both experimental and theoretical evidence in effectively clearing porous polymeric scaffolds by mechanisms other than RI matching, which could tackle the limitations of current optical imaging of cells cultured within three-dimensional (3D) opaque porous scaffolds, such as poor visibility, low spatial resolution, and small penetration depth.

  17. Development and characterization of novel porous 3D alginate-cockle shell powder nanobiocomposite bone scaffold.

    PubMed

    Bharatham, B Hemabarathy; Abu Bakar, Md Zuki; Perimal, Enoch Kumar; Yusof, Loqman Mohamed; Hamid, Muhajir

    2014-01-01

    A novel porous three-dimensional bone scaffold was developed using a natural polymer (alginate/Alg) in combination with a naturally obtained biomineral (nano cockle shell powder/nCP) through lyophilization techniques. The scaffold was developed in varying composition mixture of Alg-nCP and characterized using various evaluation techniques as well as preliminary in vitro studies on MG63 human osteoblast cells. Morphological observations using SEM revealed variations in structures with the use of different Alg-nCP composition ratios. All the developed scaffolds showed a porous structure with pore sizes ideal for facilitating new bone growth; however, not all combination mixtures showed subsequent favorable characteristics to be used for biological applications. Scaffolds produced using the combination mixture of 40% Alg and 60% nCP produced significantly promising results in terms of mechanical strength, degradation rate, and increased cell proliferation rates making it potentially the optimum composition mixture of Alg-nCP with future application prospects.

  18. In situ supramolecular hydrogel based on hyaluronic acid and dextran derivatives as cell scaffold.

    PubMed

    Chen, Jing-Xiao; Cao, Lu-Juan; Shi, Yu; Wang, Ping; Chen, Jing-Hua

    2016-09-01

    In this study, hyaluronic acid-β-cyclodextrin conjugate (HA-CD) and dextran-2-naphthylacetic acid conjugate (Dex-NAA) were synthesized as two gelators. The degrees of substitution (DS) of these two gelators were determined to be 15.5 and 7.4%, respectively. Taking advantages of the strong and selective host-guest interaction between β-CD and 2-NAA, the mixture of two gelators could form supramolecular hydrogel in situ. Moreover, the pore size, gelation time, swelling ratio as well as modulus of the hydrogel could be adjusted by simply varying the contents of HA-CD and Dex-NAA. NIH/3T3 cells that entrapped in hydrogel grew well as compared with that cultured in plates, indicating a favorable cytocompatibility of the hydrogel. Collectively, the results demonstrated that the HA-Dex hydrogel could potentially be applied in tissue engineering as cell scaffold. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2263-2270, 2016.

  19. Self-crosslinked oxidized alginate/gelatin hydrogel as injectable, adhesive biomimetic scaffolds for cartilage regeneration.

    PubMed

    Balakrishnan, Biji; Joshi, Nitin; Jayakrishnan, Athipettah; Banerjee, Rinti

    2014-08-01

    Biopolymeric hydrogels that mimic the properties of extracellular matrix have great potential in promoting cellular migration and proliferation for tissue regeneration. The authors reported earlier that rapidly gelling, biodegradable, injectable hydrogels can be prepared by self-crosslinking of periodate oxidized alginate and gelatin in the presence of borax, without using any toxic crosslinking agents. The present paper investigates the suitability of this hydrogel as a minimally invasive injectable, cell-attractive and adhesive scaffold for cartilage tissue engineering for the treatment of osteoarthritis. Time and frequency sweep rheology analysis confirmed gel formation within 20s. The hydrogel integrated well with the cartilage tissue, with a burst pressure of 70±3mmHg, indicating its adhesive nature. Hydrogel induced negligible inflammatory and oxidative stress responses, a prerequisite for the management and treatment of osteoarthritis. Scanning electron microscopy images of primary murine chondrocytes encapsulated within the matrix revealed attachment of cells onto the hydrogel matrix. Chondrocytes demonstrated viability, proliferation and migration within the matrix, while maintaining their phenotype, as seen by expression of collagen type II and aggrecan, and functionality, as seen by enhanced glycosoaminoglycan (GAG) deposition with time. DNA content and GAG deposition of chondrocytes within the matrix can be tuned by incorporation of bioactive signaling molecules such as dexamethasone, chondroitin sulphate, platelet derived growth factor (PDGF-BB) and combination of these three agents. The results suggest that self-crosslinked oxidized alginate/gelatin hydrogel may be a promising injectable, cell-attracting adhesive matrix for neo-cartilage formation in the management and treatment of osteoarthritis.

  20. 3D modeling of human cancer: A PEG-fibrin hydrogel system to study the role of tumor microenvironment and recapitulate the in vivo effect of oncolytic adenovirus.

    PubMed

    Del Bufalo, Francesca; Manzo, Teresa; Hoyos, Valentina; Yagyu, Shigeki; Caruana, Ignazio; Jacot, Jeffrey; Benavides, Omar; Rosen, Daniel; Brenner, Malcolm K

    2016-04-01

    Interactions between malignant and stromal cells and the 3D spatial architecture of the tumor both substantially modify tumor behavior, including the responses to small molecule drugs and biological therapies. Conventional 2D culture systems cannot replicate this complexity. To overcome these limitations and more accurately model solid tumors, we developed a highly versatile 3D PEG-fibrin hydrogel model of human lung adenocarcinoma. Our model relevantly recapitulates the effect of oncolytic adenovirus; tumor responses in this setting nearly reproduce those observed in vivo. We have also validated the use of this model for complex, long-term, 3D cultures of cancer cells and their stroma (fibroblasts and endothelial cells). Both tumor proliferation and invasiveness were enhanced in the presence of stromal components. These results validate our 3D hydrogel model as a relevant platform to study cancer biology and tumor responses to biological treatments.

  1. Cell culture and characterization of cross-linked poly(vinyl alcohol)-g-starch 3D scaffold for tissue engineering.

    PubMed

    Hsieh, Wen-Chuan; Liau, Jiun-Jia

    2013-10-15

    The research goal of this experiment is chemically to cross-link poly(vinyl alcohol) (PVA) and starch to form a 3D scaffold that is effective water absorbent, has a stable structure, and supports cell growth. PVA and starch can be chemically cross-linked to form a PVA-g-starch 3D scaffold polymer, as observed by Fourier transform infrared spectroscopy (FTIR), with an absorbency of up to 800%. Tensile testing reveals that, as the amount of starch increases, the strength of the 3D scaffold strength reaches 4×10(-2) MPa. Scanning electron microscope (SEM) observations of the material reveal that the 3D scaffold is highly porous formed using a homogenizer at 500 rpm. In an enzymatic degradation, the 3D scaffold was degraded by various enzymes at a rate of up to approximately 30-60% in 28 days. In vitro tests revealed that cells proliferate and grow in the 3D scaffold material. Energy dispersive spectrometer (EDS) analysis further verified that the bio-compatibility of this scaffold.

  2. Control of Retinal Ganglion Cell Positioning and Neurite Growth: Combining 3D Printing with Radial Electrospun Scaffolds.

    PubMed

    Kador, Karl E; Grogan, Shawn P; Dorthé, Erik W; Venugopalan, Praseeda; Malek, Monisha F; Goldberg, Jeffrey L; D'lima, Darryl D

    2016-02-01

    Retinal ganglion cells (RGCs) are responsible for the transfer of signals from the retina to the brain. As part of the central nervous system, RGCs are unable to regenerate following injury, and implanted cells have limited capacity to orient and integrate in vivo. During development, secreted guidance molecules along with signals from extracellular matrix and the vasculature guide cell positioning, for example, around the fovea, and axon outgrowth; however, these changes are temporally regulated and are not the same in the adult. Here, we combine electrospun cell transplantation scaffolds capable of RGC neurite guidance with thermal inkjet 3D cell printing techniques capable of precise positioning of RGCs on the scaffold surface. Optimal printing parameters are developed for viability, electrophysiological function and, neurite pathfinding. Different media, commonly used to promote RGC survival and growth, were tested under varying conditions. When printed in growth media containing both brain-derived neurotrophic factor (BDNF) and ciliary neurotrophic factor (CNTF), RGCs maintained survival and normal electrophysiological function, and displayed radial axon outgrowth when printed onto electrospun scaffolds. These results demonstrate that 3D printing technology may be combined with complex electrospun surfaces in the design of future retinal models or therapies.

  3. 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.

  4. Control of Retinal Ganglion Cell Positioning and Neurite Growth: Combining 3D Printing with Radial Electrospun Scaffolds

    PubMed Central

    Kador, Karl E.; Grogan, Shawn P.; Dorthé, Erik W.; Venugopalan, Praseeda; Malek, Monisha F.

    2016-01-01

    Retinal ganglion cells (RGCs) are responsible for the transfer of signals from the retina to the brain. As part of the central nervous system, RGCs are unable to regenerate following injury, and implanted cells have limited capacity to orient and integrate in vivo. During development, secreted guidance molecules along with signals from extracellular matrix and the vasculature guide cell positioning, for example, around the fovea, and axon outgrowth; however, these changes are temporally regulated and are not the same in the adult. Here, we combine electrospun cell transplantation scaffolds capable of RGC neurite guidance with thermal inkjet 3D cell printing techniques capable of precise positioning of RGCs on the scaffold surface. Optimal printing parameters are developed for viability, electrophysiological function and, neurite pathfinding. Different media, commonly used to promote RGC survival and growth, were tested under varying conditions. When printed in growth media containing both brain-derived neurotrophic factor (BDNF) and ciliary neurotrophic factor (CNTF), RGCs maintained survival and normal electrophysiological function, and displayed radial axon outgrowth when printed onto electrospun scaffolds. These results demonstrate that 3D printing technology may be combined with complex electrospun surfaces in the design of future retinal models or therapies. PMID:26729061

  5. Design and fabrication of 3D porous scaffolds to facilitate cell-based gene therapy.

    PubMed

    El-Ayoubi, Rouwayda; Eliopoulos, Nicoletta; Diraddo, Robert; Galipeau, Jacques; Yousefi, Azizeh-Mitra

    2008-06-01

    Biomaterials capable of efficient gene delivery by embedded cells provide a fundamental tool for the treatment of acquired or hereditary diseases. A major obstacle is maintaining adequate nutrient and oxygen diffusion to cells within the biomaterial. In this study, we combined the solid free-form fabrication and porogen leaching techniques to fabricate three-dimensional scaffolds, with bimodal pore size distribution, for cell-based gene delivery. The objective of this study was to design micro-/macroporous scaffolds to improve cell viability and drug delivery. Murine bone marrow-derived mesenchymal stromal cells (MSCs) genetically engineered to secrete erythropoietin (EPO) were seeded onto poly-L-lactide (PLLA) scaffolds with different microporosities. Over a period of 2 weeks in culture, an increase in cell proliferation and metabolic activity was observed with increasing scaffold microporosity. The concentration of EPO detected in supernatants also increased with increasing microporosity level. Our study shows that these constructs can promote cell viability and release of therapeutic proteins, and clearly demonstrates their capacity for a dual role as scaffolds for tissue regeneration and as delivery systems for soluble gene products.

  6. Microporous polymeric 3D scaffolds templated by the layer-by-layer self-assembly.

    PubMed

    Paulraj, Thomas; Feoktistova, Natalia; Velk, Natalia; Uhlig, Katja; Duschl, Claus; Volodkin, Dmitry

    2014-08-01

    Polymeric scaffolds serve as valuable supports for biological cells since they offer essential features for guiding cellular organization and tissue development. The main challenges for scaffold fabrication are i) to tune an internal structure and ii) to load bio-molecules such as growth factors and control their local concentration and distribution. Here, a new approach for the design of hollow polymeric scaffolds using porous CaCO3 particles (cores) as templates is presented. The cores packed into a microfluidic channel are coated with polymers employing the layer-by-layer (LbL) technique. Subsequent core elimination at mild conditions results in formation of the scaffold composed of interconnected hollow polymer microspheres. The size of the cores determines the feature dimensions and, as a consequence, governs cellular adhesion: for 3T3 fibroblasts an optimal microsphere size is 12 μm. By making use of the carrier properties of the porous CaCO3 cores, the microspheres are loaded with BSA as a model protein. The scaffolds developed here may also be well suited for the localized release of bio-molecules using external triggers such as IR-light.

  7. 3D polylactide-based scaffolds for studying human hepatocarcinoma processes in vitro.

    PubMed

    Scaffaro, Roberto; Re, Giada Lo; 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.

  8. Strontium eluting graphene hybrid nanoparticles augment osteogenesis in a 3D tissue scaffold.

    PubMed

    Kumar, Sachin; Chatterjee, Kaushik

    2015-02-07

    The objective of this work was to prepare hybrid nanoparticles of graphene sheets decorated with strontium metallic nanoparticles and demonstrate their advantages in bone tissue engineering. Strontium-decorated reduced graphene oxide (RGO_Sr) hybrid nanoparticles were synthesized by the facile reduction of graphene oxide and strontium nitrate. X-ray diffraction, transmission electron microscopy, and atomic force microscopy revealed that the hybrid particles were composed of RGO sheets decorated with 200-300 nm metallic strontium particles. Thermal gravimetric analysis further confirmed the composition of the hybrid particles as 22 wt% of strontium. Macroporous tissue scaffolds were prepared by incorporating RGO_Sr particles in poly(ε-caprolactone) (PCL). The PCL/RGO_Sr scaffolds were found to elute strontium ions in aqueous medium. Osteoblast proliferation and differentiation was significantly higher in the PCL scaffolds containing the RGO_Sr particles in contrast to neat PCL and PCL/RGO scaffolds. The increased biological activity can be attributed to the release of strontium ions from the hybrid nanoparticles. This study demonstrates that composites prepared using hybrid nanoparticles that elute strontium ions can be used to prepare multifunctional scaffolds with good mechanical and osteoinductive properties. These findings have important implications for designing the next generation of biomaterials for use in tissue regeneration.

  9. 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.

  10. Multimodal imaging of sustained drug release from 3-D poly(propylene fumarate) (PPF) scaffolds.

    PubMed

    Choi, Jonghoon; Kim, Kyobum; Kim, Taeho; Liu, Guanshu; Bar-Shir, Amnon; Hyeon, Taeghwan; McMahon, Michael T; Bulte, Jeff W M; Fisher, John P; Gilad, Assaf A

    2011-12-10

    The potential of poly(propylene fumarate) (PPF) scaffolds as drug carriers was investigated and the kinetics of the drug release quantified using magnetic resonance imaging (MRI) and optical imaging. Three different MR contrast agents were used for coating PPF scaffolds. Initially, iron oxide (IONP) or manganese oxide nanoparticles (MONP) carrying the anti-cancer drug doxorubicin were absorbed or mixed with the scaffold and their release into solution at physiological conditions was measured with MRI and optical imaging. A slow (hours to days) and functional release of the drug molecules into the surrounding solution was observed. In order to examine the release properties of proteins and polypeptides, protamine sulfate, a chemical exchange saturation transfer (CEST) MR contrast agent, was attached to the scaffold. Protamine sulfate showed a steady release rate for the first 24h. Due to its biocompatibility, versatile drug-loading capability and constant release rate, the porous PPF scaffold has potential in various biomedical applications, including MR-guided implantation of drug-dispensing materials, development of drug carrying vehicles, and drug delivery for tumor treatment.

  11. Sensitivity of Neural Stem Cell Survival, Differentiation and Neurite Outgrowth within 3D Hydrogels to Environmental Heavy Metals

    PubMed Central

    Tasneem, Sameera; Farrell, Kurt; Lee, Moo-Yeal; Kothapalli, Chandrasekhar R.

    2015-01-01

    We investigated the sensitivity of embryonic murine neural stem cells exposed to 10 pM – 10 μM concentrations of three heavy metals (Cd, Hg, Pb), continuously for 14 days within 3D collagen hydrogels. Critical endpoints for neurogenesis such as survival, differentiation and neurite outgrowth were assessed. Results suggest significant compromise in cell viability within the first four days at concentrations ≥ 10 nM, while lower concentrations induced a more delayed effect. Mercury and lead suppressed neural differentiation at as low as 10 pM concentration within 7 days, while all three metals inhibited neural and glial differentiation by day 14. Neurite outgrowth remained unaffected at lower cadmium or mercury concentrations (≤ 100 pM), but was completely repressed beyond day 1 at higher concentrations. Higher metal concentrations (≥ 100 pM) suppressed NSC differentiation to motor or dopaminergic neurons. Cytokines and chemokines released by NSCs, and the sub-cellular mechanisms by which metals induce damage to NSCs have been quantified and correlated to phenotypic data. The observed degree of toxicity in NSC cultures is in the order: lead > mercury > cadmium. Results point to the use of biomimetic 3D culture models to screen the toxic effects of heavy metals during developmental stages, and investigate their underlying mechanistic pathways. PMID:26621541

  12. Sensitivity of neural stem cell survival, differentiation and neurite outgrowth within 3D hydrogels to environmental heavy metals.

    PubMed

    Tasneem, Sameera; Farrell, Kurt; Lee, Moo-Yeal; Kothapalli, Chandrasekhar R

    2016-02-03

    We investigated the sensitivity of embryonic murine neural stem cells exposed to 10 pM-10 μM concentrations of three heavy metals (Cd, Hg, Pb), continuously for 14 days within 3D collagen hydrogels. Critical endpoints for neurogenesis such as survival, differentiation and neurite outgrowth were assessed. Results suggest significant compromise in cell viability within the first four days at concentrations ≥10 nM, while lower concentrations induced a more delayed effect. Mercury and lead suppressed neural differentiation at as low as 10 pM concentration within 7 days, while all three metals inhibited neural and glial differentiation by day 14. Neurite outgrowth remained unaffected at lower cadmium or mercury concentrations (≤100 pM), but was completely repressed beyond day 1 at higher concentrations. Higher metal concentrations (≥100 pM) suppressed NSC differentiation to motor or dopaminergic neurons. Cytokines and chemokines released by NSCs, and the sub-cellular mechanisms by which metals induce damage to NSCs have been quantified and correlated to phenotypic data. The observed degree of toxicity in NSC cultures is in the order: lead>mercury>cadmium. Results point to the use of biomimetic 3D culture models to screen the toxic effects of heavy metals during developmental stages, and investigate their underlying mechanistic pathways.

  13. Investigating the neuroglial differentiation effect of neuroblastoma conditioned medium in human endometrial stem cells cultured on 3D nanofibrous scaffold.

    PubMed

    Ebrahimi-Barough, Somayeh; Hoveizi, Elham; Norouzi Javidan, Abbas; Ai, Jafar

    2015-08-01

    Neural tissue engineering is an important area of research in the field of tissue-engineering especially for neurodegenerative disease such as spinal cord injury. The differentiation capacity of human endometrial stem cells (hEnSCs) into neuronal cells has yet to be elucidated. Here, the major aim of the present study was to investigate the differentiation ability of hEnSCs cultured on polylactic acid/chitosan (PLA/CS) nanofibrous scaffold into neuroglial cells in response to conditioned medium of BE(2)-C human neuroblastoma cells and growth factors. Here we investigated the use PLA/CS scaffold as a three dimensional (3D) system that increased neuro-glial cells differentiation. Human EnSCs after three passages were differentiated in neuro-glial like cells under neuroblastoma conditioned medium with FGF2/PDGF-AA on PLA/CS scaffold. By day 18, differentiated cells were analyzed for expression of neuroglial markers by qRT-PCR and immunofluorescence. The results revealed that hEnSCs attach, grow and differentiation on the nanofibrous PLA/CS scaffold. Additionally, our study showed the expression of neural and glial lineage markers such as Nestin, NF-L, MAP2, PDGFRa, CNP, Olig2, MBP, and GFAP in the level of mRNA and MAP2, Tuj-1, and NF-L in the protein level after 18 days. Our results demonstrate that hEnSCs cultured on PLA/CS nanofibrous scaffold have the potential to differentiate in neuronal and glial cells in presence of neuroblastoma conditioned medium on PLA/CS scaffold. The result of this study may have impact in tissue engineering and cells-base therapy of neurodegenerative diseases and have a great potential for wide application.

  14. Laser-ignited frontal polymerization of shape-controllable poly(VI-co-AM) hydrogels based on 3D templates toward adsorption of heavy metal ions

    NASA Astrophysics Data System (ADS)

    Fan, Suzhen; Liu, Sisi; Wang, Xiao-Qiao; Wang, Cai-Feng; Chen, Su

    2016-06-01

    Given the increasing heavy metal pollution issue, fast preparation of polymeric hydrogels with excellent adsorption property toward heavy metal ions is very attractive. In this work, a series of poly( N-vinylimidazole-co-acrylamide) (poly(VI-co-AM)) hydrogels were synthesized via laser-ignited frontal polymerization (LIFP) for the first time. The dependence of frontal velocity and temperature on two factors monomer ratios and initiator concentrations was systematically investigated. Poly(VI-co-AM) hydrogels with any self-supporting shapes can be synthesized by a one-step LIFP in seconds through the application of 3D templates. These shape-persistent hydrogels are pH-responsive and exhibit excellent adsorption/desorption characteristics toward Mn(II), Zn(II), Cd(II), Ni(II), Cu(II) and Co(II) ions, and the adsorption conformed to the pseudo-second-order kinetic model. The reusability of the hydrogels toward mental ions adsorption was further researched, which suggested that the hydrogels can be reused without serious decrease in adsorption capacity. This work might open a promising strategy to facilely prepare shape-controllable hydrogels and expand the application of LIFP.

  15. Chondrocyte calcium signaling in response to fluid flow is regulated by matrix adhesion in 3-D alginate scaffolds.

    PubMed

    Degala, Satish; Zipfel, Warren R; Bonassar, Lawrence J

    2011-01-01

    The interaction between chondrocytes and their surrounding extracellular matrix plays an important role in regulating cartilage metabolism in response to environmental cues. This study characterized the role of cell adhesion on the calcium signaling response of chondrocytes to fluid flow. Bovine chondrocytes were suspended in alginate hydrogels functionalized with RGD at concentrations of 0-400μM. The hydrogels were perfused and the calcium signaling response of the cells was measured over a range of fluid velocities from 0 to 68μm/s. Attachment to RGD-alginate doubled the sensitivity of chondrocytes to flows in the range of 8-13μm/s, but at higher fluid velocities, the contribution of cell adhesion to the observed calcium signaling response was no longer apparent. The enhanced sensitivity to flow was dependent on the density of RGD-ligand present in the scaffolds. The RGD-enhanced sensitivity to flow was completely inhibited by the addition of soluble RGD which acted as a competitive inhibitor. The results of this study indicate a role for matrix adhesion in regulating chondrocyte response to fluid flow through a calcium dependent mechanism.

  16. Decellularized human liver as a natural 3D-scaffold for liver bioengineering and transplantation

    PubMed Central

    Mazza, Giuseppe; Rombouts, Krista; Rennie Hall, Andrew; Urbani, Luca; Vinh Luong, Tu; Al-Akkad, Walid; Longato, Lisa; Brown, David; Maghsoudlou, Panagiotis; Dhillon, Amar P.; Fuller, Barry; Davidson, Brian; Moore, Kevin; Dhar, Dipok; De Coppi, Paolo; Malago, Massimo; Pinzani, Massimo

    2015-01-01

    Liver synthetic and metabolic function can only be optimised by the growth of cells within a supportive liver matrix. This can be achieved by the utilisation of decellularised human liver tissue. Here we demonstrate complete decellularization of whole human liver and lobes to form an extracellular matrix scaffold with a preserved architecture. Decellularized human liver cubic scaffolds were repopulated for up to 21 days using human cell lines hepatic stellate cells (LX2), hepatocellular carcinoma (Sk-Hep-1) and hepatoblastoma (HepG2), with excellent viability, motility and proliferation and remodelling of the extracellular matrix. Biocompatibility was demonstrated by either omental or subcutaneous xenotransplantation of liver scaffold cubes (5 × 5 × 5 mm) into immune competent mice resulting in absent foreign body responses. We demonstrate decellularization of human liver and repopulation with derived human liver cells. This is a key advance in bioartificial liver development. PMID:26248878

  17. Cell-laden 3D bioprinting hydrogel matrix depending on different compositions for soft tissue engineering: Characterization and evaluation.

    PubMed

    Park, Jisun; Lee, Sang Jin; Chung, Solchan; Lee, Jun Hee; Kim, Wan Doo; Lee, Jae Young; Park, Su A

    2017-02-01

    Cell-printing techniques that can construct three-dimensional (3D) structures with biocompatible materials and cells are of great interest for various biomedical applications, such as tissue engineering and drug-screening studies. For successful cell-printing with cells, bioinks are critical for both the processability of printing and the viability of printed cells. However, the influence of composition on 3D bio-printing with cells has not been well explored. In this study, we investigated different compositions of alginate bioinks by varying the concentrations of high molecular weight alginate (High Alg) and low molecular weight alginate (Low Alg). Bioinks of 3wt% alginate containing High Alg alone or a 1:2 (Low Alg:High Alg) composite allowed for the construction of 3D scaffolds with good processability and shapes. Cell-printing with fibroblasts and in vitro culture studies revealed good viability and growth of the printed cells after up to 7days of culture. Bioinks prepared with High and Low Alg at a 2:1 ratio exhibited better cell growth compared with those of other compositions. This study progresses the design and applications of alginate-based bioinks for cell-printing platforms in soft tissue engineering.

  18. Diarylureas and diarylamides with pyrrolo[2,3-d]pyrimidine scaffold as broad-spectrum anticancer agents.

    PubMed

    El-Gamal, Mohammed Ibrahim; Oh, Chang-Hyun

    2014-01-01

    A series of diarylureas and diarylamides possessing pyrrolo[2,3-d]pyrimidine scaffold was designed and synthesized. The in vitro antiproliferative activities of a selected group of the target compounds against NCI-60 cell line panel were tested and compared with Sorafenib and Imatinib as reference compounds. Most of the compounds showed strong and broad-spectrum antiproliferative activities. Compounds IVa, IVb, and IVd with benzamido moiety at position 4 of the pyrrolo[2,3-d]pyrimidine nucleus, para-disubstituted phenyl ring at N1-position of pyrrolo[2,3-d]pyrimidine scaffold, and urea linker showed strong and broad-spectrum anticancer results with high potencies and efficacies. In addition, the amide derivatives Vb and Vc demonstrated one-digit nanomolar IC50 values over two and one cell line(s), respectively. Amid all the target compounds, compound IVa demonstrated the best results in both one-dose and five-dose testing modes. It showed 109.18% mean % inhibition over the NCI-60 cancer cell line panel at 10 µM concentration, submicromolar 50% inhibitory concentration (IC50) values over eight cell lines of eight different cancer types, and high efficacy with total growth inhibition (TGI) and 50% lethal concentration (LC50) values less than 4.22 µM over three colon, ovarian, and prostate cancer cell lines. It showed superior potency and efficacy to Sorafenib and Imatinib over most of the tested cell lines.

  19. In vivo testing of a 3D bifurcating microchannel scaffold inducing separation of regenerating axon bundles in peripheral nerves

    NASA Astrophysics Data System (ADS)

    Stoyanova, Irina I.; van Wezel, Richard J. A.; Rutten, Wim L. C.

    2013-12-01

    Artificial nerve guidance channels enhance the regenerative effectiveness in an injured peripheral nerve but the existing design so far has been limited to basic straight tubes simply guiding the growth to bridge the gap. Hence, one of the goals in development of more effective neuroprostheses is to create bidirectional highly selective neuro-electronic interface between a prosthetic device and the severed nerve. A step towards improving selectivity for both recording and stimulation have been made with some recent in vitro studies which showed that three-dimensional (3D) bifurcating microchannels can separate neurites growing on a planar surface and bring them into contact with individual electrodes. Since the growing axons in vivo have the innate tendency to group in bundles surrounded by connective tissue, one of the big challenges in neuro-prosthetic interface design is how to overcome it. Therefore, we performed experiments with 3D bifurcating guidance scaffolds implanted in the sciatic nerve of rats to test if this new channel architecture could trigger separation pattern of ingrowth also in vivo. Our results showed that this new method enabled the re-growth of neurites into channels with gradually diminished width (80, 40 and 20 µm) and facilitated the separation of the axonal bundles with 91% success. It seems that the 3D bifurcating scaffold might contribute towards conveying detailed neural control and sensory feedback to users of prosthetic devices, and thus could improve the quality of their daily life.

  20. Integration of 3D Printed and Micropatterned Polycaprolactone Scaffolds for Guidance of Oriented Collagenous Tissue Formation In vivo

    PubMed Central

    Pilipchuk, Sophia P; Monje, Alberto; Jiao, Yizu; Hao, Jie; Kruger, Laura; Flanagan, Colleen L; Hollister, Scott J

    2016-01-01

    Scaffold design incorporating multi-scale cues for clinically-relevant, aligned tissue regeneration has potential to improve structural and functional integrity of multi-tissue interfaces. The objective of this pre-clinical study was to develop poly(ε-caprolactone) (PCL) scaffolds with mesoscale and microscale architectural cues specific to human ligament progenitor cells and assess their ability to form aligned bone-ligament-cementum complexes in vivo. PCL scaffolds were designed to integrate a 3D printed bone region with a micropatterned PCL thin film consisting of grooved pillars. The patterned film region was seeded with human ligament cells, fibroblasts transduced with BMP-7 genes seeded within the bone region, and a tooth dentin segment positioned on the ligament region prior to subcutaneous implantation into a murine model. Results indicated increased tissue alignment in vivo using micropatterned PCL films, compared to random-porous PCL. At 6 weeks, 30um groove depth significantly enhanced oriented collagen fiber thickness, overall cell alignment, and nuclear elongation relative to 10um groove depth. This study demonstrates for the first time that scaffolds with combined hierarchical mesoscale and microscale features can align cells in vivo for oral tissue repair with potential for improving the regenerative response of other bone-ligament complexes. PMID:26820240

  1. Systematical Evaluation of Mechanically Strong 3D Printed Diluted magnesium Doping Wollastonite Scaffolds on Osteogenic Capacity in Rabbit Calvarial Defects

    PubMed Central

    Sun, Miao; Liu, An; Shao, Huifeng; Yang, Xianyan; Ma, Chiyuan; Yan, Shigui; Liu, Yanming; He, Yong; Gou, Zhongru

    2016-01-01

    Wollastonite (CaSiO3; CSi) ceramic is a promising bioactive material for bone defect repair due to slightly fast degradation of its porous constructs in vivo. In our previous strategy some key features of CSi ceramic have been significantly improved by dilute magnesium doping for regulating mechanical properties and biodegradation. Here we demonstrate that 6 ~ 14% of Ca substituted by Mg in CSi (CSi-Mgx, x = 6, 10, 14) can enhance the mechanical strength (>40 MPa) but not compromise biological performances of the 3D printed porous scaffolds with open porosity of 60‒63%. The in vitro cell culture tests in vitro indicated that the dilute Mg doping into CSi was beneficial for ALP activity and high expression of osteogenic marker genes of MC3T3-E1 cells in the scaffolds. A good bone tissue regeneration response and elastoplastic response in mechanical strength in vivo were determined after implantation in rabbit calvarial defects for 6‒12 weeks. Particularly, the CSi-Mg10 and CSi-Mg14 scaffolds could enhance new bone regeneration with a significant increase of newly formed bone tissue (18 ~ 22%) compared to the pure CSi (~14%) at 12 weeks post-implantation. It is reasonable to consider that, therefore, such CSi-Mgx scaffolds possessing excellent strength and reasonable degradability are promising for bone reconstruction in thin-wall bone defects. PMID:27658481

  2. Integration of 3D Printed and Micropatterned Polycaprolactone Scaffolds for Guidance of Oriented Collagenous Tissue Formation In Vivo.

    PubMed

    Pilipchuk, Sophia P; Monje, Alberto; Jiao, Yizu; Hao, Jie; Kruger, Laura; Flanagan, Colleen L; Hollister, Scott J; Giannobile, William V

    2016-03-01

    Scaffold design incorporating multiscale cues for clinically relevant, aligned tissue regeneration has potential to improve structural and functional integrity of multitissue interfaces. The objective of this preclinical study is to develop poly(ε-caprolactone) (PCL) scaffolds with mesoscale and microscale architectural cues specific to human ligament progenitor cells and assess their ability to form aligned bone-ligament-cementum complexes in vivo. PCL scaffolds are designed to integrate a 3D printed bone region with a micropatterned PCL thin film consisting of grooved pillars. The patterned film region is seeded with human ligament cells, fibroblasts transduced with bone morphogenetic protein-7 genes seeded within the bone region, and a tooth dentin segment positioned on the ligament region prior to subcutaneous implantation into a murine model. Results indicate increased tissue alignment in vivo using micropatterned PCL films, compared to random-porous PCL. At week 6, 30 μm groove depth significantly enhances oriented collagen fiber thickness, overall cell alignment, and nuclear elongation relative to 10 μm groove depth. This study demonstrates for the first time that scaffolds with combined hierarchical mesoscale and microscale features can align cells in vivo for oral tissue repair with potential for improving the regenerative response of other bone-ligament complexes.

  3. Recombinant human bone morphogenetic protein 2 (rhBMP-2) immobilized on laser-fabricated 3D scaffolds enhance osteogenesis.

    PubMed

    Chatzinikolaidou, Maria; Pontikoglou, Charalampos; Terzaki, Konstantina; Kaliva, Maria; Kalyva, Athanasia; Papadaki, Eleni; Vamvakaki, Maria; Farsari, Maria

    2017-01-01

    The regeneration of bone via a tissue engineering approach involves components from the macroscopic to the nanoscopic level, including appropriate 3D scaffolds, cells and growth factors. In this study, hexagonal scaffolds of different diagonals were fabricated by Direct Laser Writing using a photopolymerizable hybrid material. The proliferation of bone marrow (BM) mesenchymal stem cells (MSCs) cultured on structures with various diagonals, 50, 100, 150 and 200μm increased significantly after 10days in culture, however without significant differences among them. Next, recombinant human bone morphogenetic protein 2 (rhBMP-2) was immobilized onto the hybrid material both via covalent binding and physical adsorption. Both immobilization types exhibited similar high releaseate bioactivity profiles and a sustained delivery of rhBMP-2. The collagen and calcium levels produced in the extracellular matrix (ECM) were significantly elevated for the samples functionalized with BMP-2 compared to those in the osteogenic medium. Furthermore, significant upregulation of gene expression in both types of BMP-2 immobilized scaffolds was observed for alkaline phosphatase (ALPL) and osteocalcin (BGLAP) at days 7, 14, and 21, for RUNX2 at day 21, and for osteonectin (SPARC) at days 7 and 14. The results suggest that the release of bioactive rhBMP-2 from the hybrid scaffolds enhance the control over the osteogenic differentiation during cell culture.

  4. Synthesis of highly interconnected 3D scaffold from Arothron stellatus skin collagen for tissue engineering application.

    PubMed

    Ramanathan, Giriprasath; Singaravelu, Sivakumar; Raja, M D; Sivagnanam, Uma Tiruchirapalli

    2015-11-01

    The substrate which is avidly used for tissue engineering applications should have good mechanical and biocompatible properties, and all these parameters are often considered as essential for dermal reformation. Highly interconnected three dimensional (3D) wound dressing material with enhanced structural integrity was synthesized from Arothron stellatus fish skin (AsFS) collagen for tissue engineering applications. The synthesized 3D collagen sponge (COL-SPG) was further characterized by different physicochemical methods. The scanning electron microscopy analysis of the material demonstrated that well interconnected pores with homogeneous microstructure on the surface aids higher swelling index and that the material also possessed good mechanical properties with a Young's modulus of 0.89±0.2 MPa. Biocompatibility of the 3D COL-SPG showed 92% growth for both NIH 3T3 fibroblasts and keratinocytes. Overall, the study revealed that synthesized 3D COL-SPG from fish skin will act as a promising wound dressing in skin tissue engineering.

  5. Characterization of Silk Fibroin/Chitosan 3D Porous Scaffold and In Vitro Cytology

    PubMed Central

    Zeng, Shuguang; Liu, Lei; Shi, Yong; Qiu, Junqi; Fang, Wei; Rong, Mingdeng; Guo, Zehong; Gao, Wenfeng

    2015-01-01

    Bone tissue engineering is a powerful tool to treat bone defects caused by trauma, infection, tumors and other factors. Both silk fibroin (SF) and chitosan (CS) are non-toxic and have good biocompatibility, but are poor biological scaffolds when used alone. In this study, the microscopic structure and related properties of SF/CS composite scaffolds with different component ratios were examined. The scaffold material most suitable for osteoblast growth was determined, and these results offer an experimental basis for the future reconstruction of bone defects. First, via freeze-drying and chemical crosslinking methods, SF/CS composites with different component ratios were prepared and their structure was characterized. Changes in the internal structure of the SF and CS mixture were observed, confirming that the mutual modification between the two components was complete and stable. The internal structure of the composite material was porous and three-dimensional with a porosity above 90%. We next studied the pore size, swelling ratio, water absorption ratio, degradation and in vitro cell proliferation. For the 40% SF-60% CS group, the pore size of the scaffold was suitable for the growth of osteoblasts, and the rate of degradation was steady. This favors the early adhesion, growth and proliferation of MG-63 cells. In addition to good biocompatibility and satisfactory cell affinity, this material promotes the secretion of extracellular matrix materials by osteoblasts. Thus, 40% SF-60% CS is a good material for bone tissue engineering. PMID:26083846

  6. Micro-computed tomography image-based evaluation of 3D anisotropy degree of polymer scaffolds.

    PubMed

    Pérez-Ramírez, Ursula; López-Orive, Jesús Javier; Arana, Estanislao; Salmerón-Sánchez, Manuel; Moratal, David

    2015-01-01

    Anisotropy is one of the most meaningful determinants of biomechanical behaviour. This study employs micro-computed tomography (μCT) and image techniques for analysing the anisotropy of regenerative medicine polymer scaffolds. For this purpose, three three-dimensional anisotropy evaluation image methods were used: ellipsoid of inertia (EI), mean intercept length (MIL) and tensor scale (t-scale). These were applied to three patterns (a sphere, a cube and a right prism) and to two polymer scaffold topologies (cylindrical orthogonal pore mesh and spherical pores). For the patterns, the three methods provided good results. Regarding the scaffolds, EI mistook both topologies (0.0158, [-0.5683; 0.6001]; mean difference and 95% confidence interval), and MIL showed no significant differences (0.3509, [0.0656; 0.6362]). T-scale is the preferable method because it gave the best capability (0.3441, [0.1779; 0.5102]) to differentiate both topologies. This methodology results in the development of non-destructive tools to engineer biomimetic scaffolds, incorporating anisotropy as a fundamental property to be mimicked from the original tissue and permitting its assessment by means of μCT image analysis.

  7. 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 (Ca3SiO5 and C3S) can be self-set in water to obtain high mechanical strength under mild conditions. Another advantage of using C3S to prepare 3D scaffolds is the possibility of simultaneous drug loading. Herein, we, for the first time, demonstrated successful preparation of uniform 3D-printed C3S 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 C3S bone cement scaffolds with nanoneedle-structured surfaces significantly improved bone regeneration, as compared to pure C3S bone cement scaffolds, suggesting that 3D-printed C3S bone cement scaffolds with controllable nanotopography surface are bioactive implantable biomaterials for bone repair.

  8. Migration and Proliferative Activity of Mesenchymal Stem Cells in 3D Polylactide Scaffolds Depends on Cell Seeding Technique and Collagen Modification.

    PubMed

    Rodina, A V; Tenchurin, T Kh; Saprykin, V P; Shepelev, A D; Mamagulashvili, V G; Grigor'ev, T E; Lukanina, K I; Orekhov, A S; Moskaleva, E Yu; Chvalun, S N

    2016-11-01

    We analyzed viability of mesenchymal stem cells seeded by static and dynamic methods to highly porous fibrous 3D poly-L-lactide scaffolds with similar physical and chemical properties, but different spatial organization modified with collagen. Standard collagen coating promoted protein adsorption on the scaffold surface and improved adhesive properties of 100 μ-thick scaffolds. Modification of 600-μ scaffolds with collagen under pressure increased proliferative activity of mesenchymal stem cells seeded under static and dynamic (delivery of 100,000 cells in 10 ml medium in a perfusion system at a rate of 1 ml/min) conditions by 47 and 648%, respectively (measured after 120-h culturing by MTT test). Dynamic conditions provide more uniform distribution of collagen on scaffold fibers and promote cell penetration into 3D poly-L-lactide scaffolds with thickness >600 μ.

  9. Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part 6: 3D hydrogels with positive and negative surface charges and polyelectrolyte complexes in spinal cord injury repair.

    PubMed

    Hejcl, A; Lesný, P; Prádný, M; Sedý, J; Zámecník, J; Jendelová, P; Michálek, J; Syková, E

    2009-07-01

    Macroporous hydrogels are artificial biomaterials commonly used in tissue engineering, including central nervous system (CNS) repair. Their physical properties may be modified to improve their adhesion properties and promote tissue regeneration. We implanted four types of hydrogels based on 2-hydroxyethyl methacrylate (HEMA) with different surface charges inside a spinal cord hemisection cavity at the Th8 level in rats. The spinal cords were processed 1 and 6 months after implantation and histologically evaluated. Connective tissue deposition was most abundant in the hydrogels with positively-charged functional groups. Axonal regeneration was promoted in hydrogels carrying charged functional groups; hydrogels with positively charged functional groups showed increased axonal ingrowth into the central parts of the implant. Few astrocytes grew into the hydrogels. Our study shows that HEMA-based hydrogels carrying charged functional groups improve axonal ingrowth inside the implants compared to implants without any charge. Further, positively charged functional groups promote connective tissue infiltration and extended axonal regeneration inside a hydrogel bridge.

  10. Osteogenic Differentiation of Human Mesenchymal Stem Cells in 3-D Zr-Si Organic-Inorganic Scaffolds Produced by Two-Photon Polymerization Technique

    PubMed Central

    Koroleva, Anastasia; Deiwick, Andrea; Nguyen, Alexander; Schlie-Wolter, Sabrina; Narayan, Roger; Timashev, Peter; Popov, Vladimir; Bagratashvili, Viktor; Chichkov, Boris

    2015-01-01

    Two-photon polymerization (2PP) is applied for the fabrication of 3-D Zr-Si scaffolds for bone tissue engineering. Zr-Si scaffolds with 150, 200, and 250 μm pore sizes are seeded with human bone marrow stem cells (hBMSCs) and human adipose tissue derived stem cells (hASCs) and cultured in osteoinductive and control media for three weeks. Osteogenic differentiation of hASCs and hBMSCs and formation of bone matrix is comparatively analyzed via alkaline phosphatase activity (ALP), calcium quantification, osteocalcin staining and scanning electron microscopy (SEM). It is observed that the 150 μm pore size Zr-Si scaffolds support the strongest matrix mineralization, as confirmed by calcium deposition. Analysis of ALP activity, osteocalcin staining and SEM observations of matrix mineralization reveal that mesenchymal stem cells cultured on 3-D scaffolds without osteogenic stimulation spontaneously differentiate towards osteogenic lineage. Nanoindentation measurements show that aging of the 2PP-produced Zr-Si scaffolds in aqueous or alcohol media results in an increase in the scaffold Young’s modulus and hardness. Moreover, accelerated formation of bone matrix by hASCs is noted, when cultured on the scaffolds with lower Young’s moduli and hardness values (non aged scaffolds) compared to the cells cultured on scaffolds with higher Young’s modulus and hardness values (aged scaffolds). Presented results support the potential application of Zr-Si scaffolds for autologous bone tissue engineering. PMID:25706270

  11. The influence of plasma technology coupled to chemical grafting on the cell growth compliance of 3D hydroxyapatite scaffolds.

    PubMed

    Russo, Laura; Zanini, Stefano; Giannoni, Paolo; Landi, Elena; Villa, Anna; Sandri, Monica; Riccardi, Claudia; Quarto, Rodolfo; Doglia, Silvia M; Nicotra, Francesco; Cipolla, Laura

    2012-11-01

    The development of advanced materials with biomimetic features in order to elicit desired biological responses and to guarantee tissue biocompatibility is recently gaining attention for tissue engineering applications. Bioceramics, such as hydroxyapatite-based biomaterials are now used in a number of different applications throughout the body, covering all areas of the skeleton, due to their biological and chemical similarity to the inorganic phases of bones. When bioactive sintered hydroxyapatite (HA) is desired, biomolecular modification of these materials is needed. In the present work, we investigated the influence of plasma surface modification coupled to chemical grafting on the cell growth compliance of HA 3D scaffolds.

  12. Differentiation capacity and maintenance of differentiated phenotypes of human mesenchymal stromal cells cultured on two distinct types of 3D polymeric scaffolds.

    PubMed

    Leferink, A M; Santos, D; Karperien, M; Truckenmüller, R K; van Blitterswijk, C A; Moroni, L

    2015-12-01

    Many studies have shown the influence of soluble factors and material properties on the differentiation capacity of mesenchymal stromal cells (MSCs) cultured as monolayers. These types of two-dimensional (2D) studies can be used as simplified models to understand cell processes related to stem cell sensing and mechano-transduction in a three-dimensional (3D) context. For several other mechanisms such as cell-cell signaling, cell proliferation and cell morphology, it is well-known that cells behave differently on a planar surface compared to cells in 3D environments. In classical tissue engineering approaches, a combination of cells, 3D scaffolds and soluble factors are considered as the key ingredients for the generation of mechanically stable 3D tissue constructs. However, when MSCs are used for tissue engineering strategies, little is known about the maintenance of their differentiation potential in 3D scaffolds after the removal of differentiation soluble factors. In this study, the differentiation potential of human MSCs (hMSCs) into the chondrogenic and osteogenic lineages on two distinct 3D scaffolds, additive manufactured electrospun scaffolds, was assessed and compared to conventional 2D culture. Human MSCs cultured in the presence of soluble factors in 3D showed to differentiate to the same extent as hMSCs cultured as 2D monolayers or as scaffold-free pellets, indicating that the two scaffolds do not play a consistent role in the differentiation process. In the case of phenotypic changes, the achieved differentiated phenotype was not maintained after the removal of soluble factors, suggesting that the plasticity of hMSCs is retained in 3D cell culture systems. This finding can have implications for future tissue engineering approaches in which the validation of hMSC differentiation on 3D scaffolds will not be sufficient to ensure the maintenance of the functionality of the cells in the absence of appropriate differentiation signals.

  13. Composite lithium metal anode by melt infusion of lithium into a 3D conducting scaffold with lithiophilic coating

    PubMed Central

    Liang, Zheng; Lin, Dingchang; Zhao, Jie; Lu, Zhenda; Liu, Yayuan; Liu, Chong; Lu, Yingying; Wang, Haotian; Yan, Kai; Tao, Xinyong; Cui, Yi

    2016-01-01

    Lithium metal-based battery is considered one of the best energy storage systems due to its high theoretical capacity and lowest anode potential of all. However, dendritic growth and virtually relative infinity volume change during long-term cycling often lead to severe safety hazards and catastrophic failure. Here, a stable lithium–scaffold composite electrode is developed by lithium melt infusion into a 3D porous carbon matrix with “lithiophilic” coating. Lithium is uniformly entrapped on the matrix surface and in the 3D structure. The resulting composite electrode possesses a high conductive surface area and excellent structural stability upon galvanostatic cycling. We showed stable cycling of this composite electrode with small Li plating/stripping overpotential (<90 mV) at a high current density of 3 mA/cm2 over 80 cycles. PMID:26929378

  14. Laser Speckle Rheology for evaluating the viscoelastic properties of hydrogel scaffolds

    NASA Astrophysics Data System (ADS)

    Hajjarian, Zeinab; Nia, Hadi Tavakoli; Ahn, Shawn; Grodzinsky, Alan J.; Jain, Rakesh K.; Nadkarni, Seemantini K.

    2016-12-01

    Natural and synthetic hydrogel scaffolds exhibit distinct viscoelastic properties at various length scales and deformation rates. Laser Speckle Rheology (LSR) offers a novel, non-contact optical approach for evaluating the frequency-dependent viscoelastic properties of hydrogels. In LSR, a coherent laser beam illuminates the specimen and a high-speed camera acquires the time-varying speckle images. Cross-correlation analysis of frames returns the speckle intensity autocorrelation function, g2(t), from which the frequency-dependent viscoelastic modulus, G*(ω), is deduced. Here, we establish the capability of LSR for evaluating the viscoelastic properties of hydrogels over a large range of moduli, using conventional mechanical rheometry and atomic force microscopy (AFM)-based indentation as reference-standards. Results demonstrate a strong correlation between |G*(ω)| values measured by LSR and mechanical rheometry (r = 0.95, p < 10‑9), and z-test analysis reports that moduli values measured by the two methods are identical (p > 0.08) over a large range (47 Pa – 36 kPa). In addition, |G*(ω)| values measured by LSR correlate well with indentation moduli, E, reported by AFM (r = 0.92, p < 10‑7). Further, spatially-resolved moduli measurements in micro-patterned substrates demonstrate that LSR combines the strengths of conventional rheology and micro-indentation in assessing hydrogel viscoelastic properties at multiple frequencies and small length-scales.

  15. The self-crosslinking smart hyaluronic acid hydrogels as injectable three-dimensional scaffolds for cells culture.

    PubMed

    Bian, Shaoquan; He, Mengmeng; Sui, Junhui; Cai, Hanxu; Sun, Yong; Liang, Jie; Fan, Yujiang; Zhang, Xingdong

    2016-04-01

    Although the disulfide bond crosslinked hyaluronic acid hydrogels have been reported by many research groups, the major researches were focused on effectively forming hydrogels. However, few researchers paid attention to the potential significance of controlling the hydrogel formation and degradation, improving biocompatibility, reducing the toxicity of exogenous and providing convenience to the clinical operations later on. In this research, the novel controllable self-crosslinking smart hydrogels with in-situ gelation property was prepared by a single component, the thiolated hyaluronic acid derivative (HA-SH), and applied as a three-dimensional scaffold to mimic native extracellular matrix (ECM) for the culture of fibroblasts cells (L929) and chondrocytes. A series of HA-SH hydrogels were prepared depending on different degrees of thiol substitution (ranging from 10 to 60%) and molecule weights of HA (0.1, 0.3 and 1.0 MDa). The gelation time, swelling property and smart degradation behavior of HA-SH hydrogel were evaluated. The results showed that the gelation and degradation time of hydrogels could be controlled by adjusting the component of HA-SH polymers. The storage modulus of HA-SH hydrogels obtained by dynamic modulus analysis (DMA) could be up to 44.6 kPa. In addition, HA-SH hydrogels were investigated as a three-dimensional scaffold for the culture of fibroblasts cells (L929) and chondrocytes cells in vitro and as an injectable hydrogel for delivering chondrocytes cells in vivo. These results illustrated that HA-SH hydrogels with controllable gelation process, intelligent degradation behavior, excellent biocompatibility and convenient operational characteristics supplied potential clinical application capacity for tissue engineering and regenerative medicine.

  16. Bioengineered 3D brain tumor model to elucidate the effects of matrix stiffness on glioblastoma cell behavior using PEG-based hydrogels.

    PubMed

    Wang, Christine; Tong, Xinming; Yang, Fan

    2014-07-07

    Glioblastoma (GBM) is the most common and aggressive form of primary brain tumor with a median survival of 12-15 months, and the mechanisms underlying GBM tumor progression remain largely elusive. Given the importance of tumor niche signaling in driving GBM progression, there is a strong need to develop in vitro models to facilitate analysis of brain tumor cell-niche interactions in a physiologically relevant and controllable manner. Here we report the development of a bioengineered 3D brain tumor model to help elucidate the effects of matrix stiffness on GBM cell fate using poly(ethylene-glycol) (PEG)-based hydrogels with brain-mimicking biochemical and mechanical properties. We have chosen PEG given its bioinert nature and tunable physical property, and the resulting hydrogels allow tunable matrix stiffness without changing the biochemical contents. To facilitate cell proliferation and migration, CRGDS and a MMP-cleavable peptide were chemically incorporated. Hyaluronic acid (HA) was also incorporated to mimic the concentration in the brain extracellular matrix. Using U87 cells as a model GBM cell line, we demonstrate that such biomimetic hydrogels support U87 cell growth, spreading, and migration in 3D over the course of 3 weeks in culture. Gene expression analyses showed U87 cells actively deposited extracellular matrix and continued to upregulate matrix remodeling genes. To examine the effects of matrix stiffness on GBM cell fate in 3D, we encapsulated U87 cells in soft (1 kPa) or stiff (26 kPa) hydrogels, which respectively mimics the matrix stiffness of normal brain or GBM tumor tissues. Our results suggest that changes in matrix stiffness induce differential GBM cell proliferation, morphology, and migration modes in 3D. Increasing matrix stiffness led to delayed U87 cell proliferation inside hydrogels, but cells formed denser spheroids with extended cell protrusions. Cells cultured in stiff hydrogels also showed upregulation of HA synthase 1 and matrix

  17. 3D cell culture and osteogenic differentiation of human bone marrow stromal cells plated onto jet-sprayed or electrospun micro-fiber scaffolds.

    PubMed

    Brennan, Meadhbh Á; Renaud, Audrey; Gamblin, Anne-Laure; D'Arros, Cyril; Nedellec, Steven; Trichet, Valerie; Layrolle, Pierre

    2015-08-04

    A major limitation of the 2D culture systems is that they fail to recapitulate the in vivo 3D cellular microenvironment whereby cell-cell and cell-extracellular matrix (ECM) interactions occur. In this paper, a biomaterial scaffold that mimics the structure of collagen fibers was produced by jet-spraying. This micro-fiber polycaprolactone (PCL) scaffold was evaluated for 3D culture of human bone marrow mesenchymal stromal cells (MSCs) in comparison with a commercially available electrospun scaffold. The jet-sprayed scaffolds had larger pore diameters, greater porosity, smaller diameter fibers, and more heterogeneous fiber diameter size distribution compared to the electrospun scaffolds. Cells on jet-sprayed constructs exhibited spread morphology with abundant cytoskeleton staining, whereas MSCs on electrospun scaffolds appeared less extended with fewer actin filaments. MSC proliferation and cell infiltration occurred at a faster rate on jet-sprayed compared to electrospun scaffolds. Osteogenic differentiation of MSCs and ECM production as measured by ALP, collagen and calcium deposition was superior on jet-sprayed compared to electrospun scaffolds. The jet-sprayed scaffold which mimics the native ECM and permits homogeneous cell infiltration is important for 3D in vitro applications such as bone cellular interaction studies or drug testing, as well as bone tissue engineering strategies.

  18. 3D Scaffold of Electrosprayed Fibers with Large Pore Size for Tissue Regeneration

    PubMed Central

    Hong, Jong Kyu; Madihally, Sundararajan V.

    2010-01-01

    Regeneration of tissues using biodegradable porous scaffolds has been an intensely investigated area. Since electrospinning can produce scaffolds mimicking nanofibrous architecture found in the body, it recently has gained widespread attention. However, a major problem is the lack of pore size necessary for infiltration of cells into the layers below the surface, restricting cell colonization to the surfaces only. This study describes a novel twist to the traditional electrospinning technology. In particular, collector plates are designed which allows forming very thin layers with pore sizes suitable for cell infiltration. Thin samples can be handled without mechanically damaging the structure and can be transferred into cell culture. These thin layers were stacked by layer-by-layer assembly to develop thick structures. Thirty day cultures of fibroblasts show attachment and spreading of cells in every layer. This concept is useful in regenerating thick tissues with uniformly distributed cells and others in vitro cell culture. PMID:20620245

  19. Design and Printing Strategies in 3D Bioprinting of Cell-Hydrogels: A Review.

    PubMed

    Lee, Jia Min; Yeong, Wai Yee

    2016-11-01

    Bioprinting is an emerging technology that allows the assembling of both living and non-living biological materials into an ideal complex layout for further tissue maturation. Bioprinting aims to produce engineered tissue or organ in a mechanized, organized, and optimized manner. Various biomaterials and techniques have been utilized to bioprint biological constructs in different shapes, sizes and resolutions. There is a need to systematically discuss and analyze the reported strategies employed to fabricate these constructs. We identified and discussed important design factors in bioprinting, namely shape and resolution, material heterogeneity, and cellular-material remodeling dynamism. Each design factors are represented by the corresponding process capabilities and printing parameters. The process-design map will inspire future biomaterials research in these aspects. Design considerations such as data processing, bio-ink formulation and process selection are discussed. Various printing and crosslinking strategies, with relevant applications, are also systematically reviewed. We categorized them into 5 general bioprinting strategies, including direct bioprinting, in-process crosslinking, post-process crosslinking, indirect bioprinting and hybrid bioprinting. The opportunities and outlook in 3D bioprinting are highlighted. This review article will serve as a framework to advance computer-aided design in bioprinting technologies.

  20. 3D printing enables separation of orthogonal functions within a hydrogel particle.

    PubMed

    Raman, Ritu; Clay, Nicholas E; Sen, Sanjeet; Melhem, Molly; Qin, Ellen; Kong, Hyunjoon; Bashir, Rashid

    2016-06-01

    Multifunctional particles with distinct physiochemical phases are required by a variety of applications in biomedical engineering, such as diagnostic imaging and targeted drug delivery. This motivates the development of a repeatable, efficient, and customizable approach to manufacturing particles with spatially segregated bioactive moieties. This study demonstrates a stereolithographic 3D printing approach for designing and fabricating large arrays of biphasic poly (ethylene glycol) diacrylate (PEGDA) gel particles. The fabrication parameters governing the physical and biochemical properties of multi-layered particles are thoroughly investigated, yielding a readily tunable approach to manufacturing customizable arrays of multifunctional particles. The advantage in spatially organizing functional epitopes is examined by loading superparamagnetic iron oxide nanoparticles (SPIONs) and bovine serum albumin (BSA) in separate layers of biphasic PEGDA gel particles and examining SPION-induced magnetic resonance (MR) contrast and BSA-release kinetics. Particles with spatial segregation of functional moieties have demonstrably higher MR contrast and BSA release. Overall, this study will contribute significant knowledge to the preparation of multifunctional particles for use as biomedical tools.

  1. Compensation of spherical aberration influences for two-photon polymerization patterning of large 3D scaffolds

    NASA Astrophysics Data System (ADS)

    Stichel, T.; Hecht, B.; Houbertz, R.; Sextl, G.

    2015-10-01

    Two-photon polymerization using femtosecond laser pulses at a wavelength of 515 nm is used for three-dimensional patterning of photosensitive, biocompatible inorganic-organic hybrid polymers (ORMOCER®s). In order to fabricate millimeter-sized biomedical scaffold structures with interconnected pores, medium numerical aperture air objectives with long working distances are applied which allow voxel lengths of several micrometers and thus the solidification of large scaffolds in an adequate time. It is demonstrated that during processing the refraction of the focused laser beam at the air/material interface leads to strong spherical aberration which decreases the peak intensity of the focal point spread function along with shifting and severely extending the focal region in the direction of the beam propagation. These effects clearly decrease the structure integrity, homogeneity and the structure details and therefore are minimized by applying a positioning and laser power adaptation throughout the fabrication process. The results will be discussed with respect to the resulting structural homogeneity and its application as biomedical scaffold.

  2. Fabrication and characterization of graphene hydrogel via hydrothermal approach as a scaffold for preliminary study of cell growth

    PubMed Central

    Lim, HN; Huang, NM; Lim, SS; Harrison, I; Chia, CH

    2011-01-01

    Background Three-dimensional assembly of graphene hydrogel is rapidly attracting the interest of researchers because of its wide range of applications in energy storage, electronics, electrochemistry, and waste water treatment. Information on the use of graphene hydrogel for biological purposes is lacking, so we conducted a preliminary study to determine the suitability of graphene hydrogel as a substrate for cell growth, which could potentially be used as building blocks for biomolecules and tissue engineering applications. Methods A three-dimensional structure of graphene hydrogel was prepared via a simple hydrothermal method using two-dimensional large-area graphene oxide nanosheets as a precursor. Results The concentration and lateral size of the graphene oxide nanosheets influenced the structure of the hydrogel. With larger-area graphene oxide nanosheets, the graphene hydrogel could be formed at a lower concentration. X-ray diffraction patterns revealed that the oxide functional groups on the graphene oxide nanosheets were reduced after hydrothermal treatment. The three-dimensional graphene hydrogel matrix was used as a scaffold for proliferation of a MG63 cell line. Conclusion Guided filopodia protrusions of MG63 on the hydrogel were observed on the third day of cell culture, demonstrating compatibility of the graphene hydrogel structure for bioapplications. PMID:21931479

  3. Decellularized Wharton’s Jelly from human umbilical cord as a novel 3D scaffolding material for tissue engineering applications

    PubMed Central

    Jadalannagari, Sushma; Converse, Gabriel; McFall, Christopher; Buse, Eric; Filla, Michael; Villar, Maria T.; Artigues, Antonio; Mellot, Adam J.; Wang, Jinxi; Detamore, Michael S.; Hopkins, Richard A.; Aljitawi, Omar S.

    2017-01-01

    In tissue engineering, an ideal scaffold attracts and supports cells thus providing them with the necessary mechanical support and architecture as they reconstruct new tissue in vitro and in vivo. This manuscript details a novel matrix derived from decellularized Wharton’s jelly (WJ) obtained from human umbilical cord for use as a scaffold for tissue engineering application. This decellularized Wharton’s jelly matrix (DWJM) contained 0.66 ± 0.12 μg/mg sulfated glycosaminoglycans (GAGs), and was abundant in hyaluronic acid, and completely devoid of cells. Mass spectroscopy revealed the presence of collagen types II, VI and XII, fibronectin-I, and lumican I. When seeded onto DWJM, WJ mesenchymal stem cells (WJMSCs), successfully attached to, and penetrated the porous matrix resulting in a slower rate of cell proliferation. Gene expression analysis of WJ and bone marrow (BM) MSCs cultured on DWJM demonstrated decreased expression of proliferation genes with no clear pattern of differentiation. When this matrix was implanted into a murine calvarial defect model with, green fluorescent protein (GFP) labeled osteocytes, the osteocytes were observed to migrate into the matrix as early as 24 hours. They were also identified in the matrix up to 14 days after transplantation. Together with these findings, we conclude that DWJM can be used as a 3D porous, bioactive and biocompatible scaffold for tissue engineering and regenerative medicine applications. PMID:28222169

  4. Pore structure and dielectric behaviour of the 3D collagen-DAC scaffolds designed for nerve tissue repair.

    PubMed

    Pietrucha, Krystyna; Marzec, Ewa; Kudzin, Marcin

    2016-11-01

    The design and selection of a suitable scaffold with well-defined pores size distribution and dielectric properties are critical features for neural tissue engineering. In this study we use mercury porosimetry and the dielectric spectroscopy in the alpha-dispersion region of the electric field to determine the microarchitecture and activation energy of collagen (Col) modified by 2,3 dialdehyde cellulose (DAC). The scaffold was synthesized in three steps: (i) preparation of DAC by oxidation of cellulose, (ii) construction of a 3D Col sponge-shape or film, (iii) cross-linkage of the Col samples using DAC. The activation energy needed to break the bonds formed by water in the Col-DAC composite is approximately 2 times lower than that in the unmodified Col. In addition, the magnitude of conductivity for modified Col at 70°C is approximately 40% lower than that recorded for the unmodified Col. The largest fraction, of which at least 70% of the total pore volume comprises the sponge, is occupied by pores ranging from 20 to 100μm in size. The knowledge on the dielectric behaviour and microstructure of the Col-DAC scaffold may prove relevant to neural tissue engineering focused on the regeneration of the nervous system.

  5. A transferrin variant as the targeting ligand for polymeric nanoparticles incorporated in 3-D PLGA porous scaffolds.

    PubMed

    Lopes, André M; Chen, Kevin Y; Kamei, Daniel T

    2017-04-01

    We have developed doxorubicin (DOX)-loaded poly(lactide-co-glycolide) (PLGA) nanoparticles (DP) conjugated with polyethylene glycol (PEG) and transferrin (Tf) to form Tf-PEG-DPs (TPDPs), and incorporated these TPDPs into three-dimensional (3-D) PLGA porous scaffolds to form a controlled delivery system. To our knowledge, this represents the first use of a Tf variant (oxalate Tf) to improve the targeted delivery of drug-encapsulated nanoparticles (NPs) in PLGA scaffolds to PC3 prostate cancer cells. The PLGA scaffolds with TPDPs incorporated have been shown to release drugs for sustained delivery and provided a continuous release of DOX. The MTS assay was also performed to determine the potency of native and oxalate TPDPs, and a 3.0-fold decrease in IC50 values were observed between the native and oxalate TPDPs. The lower IC50 value for the oxalate version signifies greater potency compared to the native version, since a lower concentration of drug was required to achieve the same therapeutic effect. These results suggest that this technology has potential to become a new implantable polymeric device to improve the controlled and targeted drug delivery of Tf-conjugated NPs for cancer therapy.

  6. Changes in morphology of actin filaments and expression of alkaline phosphatase at 3D cultivation of MG-63 osteoblast-like cells on mineralized fibroin scaffolds.

    PubMed

    Goncharenko, A V; Malyuchenko, N V; Moisenovich, A M; Kotlyarova, M S; Arkhipova, A Yu; Kon'kov, A S; Agapov, I I; Molochkov, A V; Moisenovich, M M; Kirpichnikov, M P

    2016-09-01

    3D cultivation of MG-63 osteoblast-like cells on mineralized fibroin scaffolds leads to an increase in the expression of alkaline phosphatase, an early marker of bone formation. Increased expression is associated with the actin cytoskeleton reorganization under the influence of 3D cultivation and osteogenic calcium phosphate component of the microcarrier.

  7. Biomimetic poly(ethylene glycol)-based hydrogels as scaffolds for inducing endothelial adhesion and capillary-like network formation.

    PubMed

    Zhu, Junmin; He, Ping; Lin, Lin; Jones, Derek R; Marchant, Roger E

    2012-03-12

    The extracellular matrix (ECM) is an attractive model for designing synthetic scaffolds with a desirable environment for tissue engineering. Here, we report on the synthesis of ECM-mimetic poly(ethylene glycol) (PEG) hydrogels for inducing endothelial cell (EC) adhesion and capillary-like network formation. A collagen type I-derived peptide GPQGIAGQ (GIA)-containing PEGDA (GIA-PEGDA) was synthesized with the collagenase-sensitive GIA sequence attached in the middle of the PEGDA chain, which was then copolymerized with RGD capped-PEG monoacrylate (RGD-PEGMA) to form biomimetic hydrogels. The hydrogels degraded in vitro with the rate dependent on the concentration of collagenase and also supported the adhesion of human umbilical vein ECs (HUVECs). Biomimetic RGD/GIA-PEGDA hydrogels with incorporation of 1% RGD-PEGDA into GIA-PEGDA hydrogels induced capillary-like organization when HUVECs were seeded on the hydrogel surface, while RGD/PEGDA and GIA-PEGDA hydrogels did not. These results indicate that both cell adhesion and biodegradability of scaffolds play important roles in the formation of capillary-like networks.

  8. 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.

  9. Subcellular stretch-induced cytoskeletal response of single fibroblasts within 3D designer scaffolds.

    PubMed

    Scheiwe, Andrea C; Frank, Stephanie C; Autenrieth, Tatjana J; Bastmeyer, Martin; Wegener, Martin

    2015-03-01

    In vivo, cells are exposed to mechanical forces in many different ways. These forces can strongly influence cell functions or may even lead to diseases. Through their sensing machinery, cells are able to perceive the physical information of the extracellular matrix and translate it into biochemical signals resulting in cellular responses. Here, by virtue of two-component polymer scaffolds made via direct laser writing, we precisely control the cell matrix adhesions regarding their spatial arrangement and size. This leads to highly controlled and uniform cell morphologies, thereby allowing for averaging over the results obtained from several different individual cells, enabling quantitative analysis. We transiently deform these elastic structures by a micromanipulator, which exerts controlled stretching forces on primary fibroblasts grown in these scaffolds on a subcellular level. We find stretch-induced remodeling of both actin cytoskeleton and cell matrix adhesions. The responses to static and periodic stretching are significantly different. The amount of paxillin and phosphorylated focal adhesion kinase increases in cell matrix adhesions at the manipulated pillar after static stretching whereas it decreases after periodic stretching.

  10. 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

  11. Mitigating Scarring and Inflammation during Corneal Wound Healing using Nanofiber-Hydrogel Scaffolds

    NASA Astrophysics Data System (ADS)

    Fu, Amy

    Due to the universal lack of donor tissue, there has been emerging interest in engineering materials to stimulate living cells to restore the features and functions of injured organs. We are particularly interested in developing materials for corneal use, where the necessity to maintain the tissue's transparency presents an additional challenge. Every year, there are 1.5 -- 2 million new cases of monocular blindness due to irregular healing of corneal injuries, dwarfing the approximately 150,000 corneal transplants performed. The large gap between the need and availability of cornea transplantation motivates us to develop a wound-healing scaffold that can prevent corneal blindness. To develop such a scaffold, it is necessary to regulate the cells responsible for repairing the damaged cornea, namely myofibroblasts, which are responsible for the disordered and non-refractive index matched scar that leads to corneal blindness. Using in vitro assays, we identified that protein nanofibers of certain orientation can promote cell migration and modulate the myofibroblast phenotype. The nanofibers are also transparent, easy to handle and non-cytotoxic. To adhere the nanofibers to a wound bed, we examined the use of two different in situ forming hydrogels: an artificial extracellular matrix protein (aECM)-based gel and a photo-crosslinkable heparin-based gel. Both hydrogels can be formed within minutes, are transparent upon gelation and are easily tunable. Using an in vivo mouse model for epithelial defects, we show that our corneal scaffolds (nanofibers together with hydrogel) are well-tolerated (no inflammatory response or turbidity) and support epithelium regrowth. We developed an ex vivo corneal tissue culture model where corneas that are wounded and treated with our scaffold can be cultured while retaining their ability to repair wounds for up to 21 days. Using this technique, we found that the aECM-based treatment induced a more favorable wound response than the

  12. Structure, Properties, and In Vitro Behavior of Heat-Treated Calcium Sulfate Scaffolds Fabricated by 3D Printing

    PubMed Central

    Asadi-Eydivand, Mitra; Solati-Hashjin, Mehran; Shafiei, Seyedeh Sara; Mohammadi, Sepideh; Hafezi, Masoud; Abu Osman, Noor Azuan

    2016-01-01

    The ability of inkjet-based 3D printing (3DP) to fabricate biocompatible ceramics has made it one of the most favorable techniques to generate bone tissue engineering (BTE) scaffolds. Calcium sulfates exhibit various beneficial characteristics, and they can be used as a promising biomaterial in BTE. However, low mechanical performance caused by the brittle character of ceramic materials is the main weakness of 3DP calcium sulfate scaffolds. Moreover, the presence of certain organic matters in the starting powder and binder solution causes products to have high toxicity levels. A post-processing treatment is usually employed to improve the physical, chemical, and biological behaviors of the printed scaffolds. In this study, the effects of heat treatment on the structural, mechanical, and physical characteristics of 3DP calcium sulfate prototypes were investigated. Different microscopy and spectroscopy methods were employed to characterize the printed prototypes. The in vitro cytotoxicity of the specimens was also evaluated before and after heat treatment. Results showed that the as-printed scaffolds and specimens heat treated at 300°C exhibited severe toxicity in vitro but had almost adequate strength. By contrast, the specimens heat treated in the 500°C–1000°C temperature range, although non-toxic, had insufficient mechanical strength, which was mainly attributed to the exit of the organic binder before 500°C and the absence of sufficient densification below 1000°C. The sintering process was accelerated at temperatures higher than 1000°C, resulting in higher compressive strength and less cytotoxicity. An anhydrous form of calcium sulfate was the only crystalline phase existing in the samples heated at 500°C–1150°C. The formation of calcium oxide caused by partial decomposition of calcium sulfate was observed in the specimens heat treated at temperatures higher than 1200°C. Although considerable improvements in cell viability of heat-treated scaffolds

  13. Structure, Properties, and In Vitro Behavior of Heat-Treated Calcium Sulfate Scaffolds Fabricated by 3D Printing.

    PubMed

    Asadi-Eydivand, Mitra; Solati-Hashjin, Mehran; Shafiei, Seyedeh Sara; Mohammadi, Sepideh; Hafezi, Masoud; Abu Osman, Noor Azuan

    2016-01-01

    The ability of inkjet-based 3D printing (3DP) to fabricate biocompatible ceramics has made it one of the most favorable techniques to generate bone tissue engineering (BTE) scaffolds. Calcium sulfates exhibit various beneficial characteristics, and they can be used as a promising biomaterial in BTE. However, low mechanical performance caused by the brittle character of ceramic materials is the main weakness of 3DP calcium sulfate scaffolds. Moreover, the presence of certain organic matters in the starting powder and binder solution causes products to have high toxicity levels. A post-processing treatment is usually employed to improve the physical, chemical, and biological behaviors of the printed scaffolds. In this study, the effects of heat treatment on the structural, mechanical, and physical characteristics of 3DP calcium sulfate prototypes were investigated. Different microscopy and spectroscopy methods were employed to characterize the printed prototypes. The in vitro cytotoxicity of the specimens was also evaluated before and after heat treatment. Results showed that the as-printed scaffolds and specimens heat treated at 300°C exhibited severe toxicity in vitro but had almost adequate strength. By contrast, the specimens heat treated in the 500°C-1000°C temperature range, although non-toxic, had insufficient mechanical strength, which was mainly attributed to the exit of the organic binder before 500°C and the absence of sufficient densification below 1000°C. The sintering process was accelerated at temperatures higher than 1000°C, resulting in higher compressive strength and less cytotoxicity. An anhydrous form of calcium sulfate was the only crystalline phase existing in the samples heated at 500°C-1150°C. The formation of calcium oxide caused by partial decomposition of calcium sulfate was observed in the specimens heat treated at temperatures higher than 1200°C. Although considerable improvements in cell viability of heat-treated scaffolds were

  14. Design, fabrication and in vitro evaluation of a novel polymer-hydrogel hybrid scaffold for bone tissue engineering.

    PubMed

    Igwe, John C; Mikael, Paiyz E; Nukavarapu, Syam P

    2014-02-01

    The development of a bone mechanically-compatible and osteoinductive scaffold is important for bone tissue engineering applications, particularly for the repair and regeneration of large area critically-sized bone defects. Although previous studies with weight-bearing scaffolds have shown promising results, there is a clear need to develop better osteoinductive strategies for effective scaffold-based bone regeneration. In this study, we designed and fabricated a novel polymer-hydrogel hybrid scaffold system in which a load-bearing polymer matrix and a peptide hydrogel allowed for the synergistic combination of mechanical strength and great potential for osteoinductivity in a single scaffold. The hybrid scaffold system promoted increased pre-osteoblastic cell proliferation. Further, we biotinylated human recombinant bone morphogenetic protein 2 (rhBMP2), and characterized the biotin addition and its effect on rhBMP2 biological activity. The biotinylated rhBMP2 was tethered to the hybrid scaffold using biotin-streptavidin complexation. Controlled release studies demonstrated increased rhBMP2 retention with the tethered rhBMP2 hybrid scaffold group. In vitro evaluation of the hybrid scaffold was performed with rat bone marrow stromal cells and mouse pre-osteoblast cell line MC3T3-E1 cells. Gene expression of alkaline phosphatase (ALP), collagen I (Col I), osteopontin (OPN), bone sialoprotein (BSP), Runx-2 and osteocalcin (OC) increased in MC3T3-E1 cells seeded on the rhBMP2 tethered hybrid scaffolds over the untethered counterparts, demonstrating osteoinductive potential of the hybrid graft. These findings suggest the possibility of developing a novel polymer-hydrogel hybrid system that is weight bearing and osteoinductive for effective bone tissue engineering.

  15. 3D-Cultivation of bone marrow stromal cells on hydroxyapatite scaffolds fabricated by dispense-plotting and negative mould technique.

    PubMed

    Detsch, R; Uhl, F; Deisinger, U; Ziegler, G

    2008-04-01

    The main principle of a bone tissue engineering (BTE) strategy is to cultivate osteogenic cells in an osteoconductive porous scaffold. Ceramic implants for osteogenesis are based mainly on hydroxyapatite (HA), since this is the inorganic component of bone. Rapid Prototyping (RP) is a new technology in research for producing ceramic scaffolds. This technology is particularly suitable for the fabrication of individually and specially tailored single implants. For tissue engineering these scaffolds are seeded with osteoblast or osteoblast precursor cells. To supply the cultured osteoblastic cells efficiently with nutrition in these 3D-geometries a bioreactor system can be used. The aim of this study was to analyse the influence of differently fabricated HA-scaffolds on bone marrow stromal cells. For this, two RP-techniques, dispense-plotting and a negative mould method, were used to produce porous ceramics. The manufactured HA-scaffolds were then cultivated in a dynamic system (bioreactor) with an osteoblastic precursor cell line. In our study, the applied RP-techniques give the opportunity to design and process HA-scaffolds with defined porosity, interconnectivity and 3D pore distribution. A higher differentiation of bone marrow stromal cells could be detected on the negative mould fabricated scaffolds, while cell proliferation was higher on the dispense-plotted scaffolds. Nevertheless, both scaffold types can be used in tissue engineering applications.

  16. Laser-based three-dimensional multiscale micropatterning of biocompatible hydrogels for customized tissue engineering scaffolds.

    PubMed

    Applegate, Matthew B; Coburn, Jeannine; Partlow, Benjamin P; Moreau, Jodie E; Mondia, Jessica P; Marelli, Benedetto; Kaplan, David L; Omenetto, Fiorenzo G

    2015-09-29

    Light-induced material phase transitions enable the formation of shapes and patterns from the nano- to the macroscale. From lithographic techniques that enable high-density silicon circuit integration, to laser cutting and welding, light-matter interactions are pervasive in everyday materials fabrication and transformation. These noncontact patterning techniques are ideally suited to reshape soft materials of biological relevance. We present here the use of relatively low-energy (< 2 nJ) ultrafast laser pulses to generate 2D and 3D multiscale patterns in soft silk protein hydrogels without exogenous or chemical cross-linkers. We find that high-resolution features can be generated within bulk hydrogels through nearly 1 cm of material, which is 1.5 orders of magnitude deeper than other biocompatible materials. Examples illustrating the materials, results, and the performance of the machined geometries in vitro and in vivo are presented to demonstrate the versatility of the approach.

  17. Laser-based three-dimensional multiscale micropatterning of biocompatible hydrogels for customized tissue engineering scaffolds

    PubMed Central

    Applegate, Matthew B.; Coburn, Jeannine; Partlow, Benjamin P.; Moreau, Jodie E.; Mondia, Jessica P.; Marelli, Benedetto; Kaplan, David L.; Omenetto, Fiorenzo G.

    2015-01-01

    Light-induced material phase transitions enable the formation of shapes and patterns from the nano- to the macroscale. From lithographic techniques that enable high-density silicon circuit integration, to laser cutting and welding, light–matter interactions are pervasive in everyday materials fabrication and transformation. These noncontact patterning techniques are ideally suited to reshape soft materials of biological relevance. We present here the use of relatively low-energy (< 2 nJ) ultrafast laser pulses to generate 2D and 3D multiscale patterns in soft silk protein hydrogels without exogenous or chemical cross-linkers. We find that high-resolution features can be generated within bulk hydrogels through nearly 1 cm of material, which is 1.5 orders of magnitude deeper than other biocompatible materials. Examples illustrating the materials, results, and the performance of the machined geometries in vitro and in vivo are presented to demonstrate the versatility of the approach. PMID:26374842

  18. A Hydrogel-Mineral Composite Scaffold for Osteochondral Interface Tissue Engineering

    PubMed Central

    Khanarian, Nora T.; Jiang, Jie; Wan, Leo Q.; Mow, Van C.

    2012-01-01

    Osteoarthritis is the leading cause of physical disability among Americans, and tissue engineered cartilage grafts have emerged as a promising treatment option for this debilitating condition. Currently, the formation of a stable interface between the cartilage graft and subchondral bone remains a significant challenge. This study evaluates the potential of a hybrid scaffold of hydroxyapatite (HA) and alginate hydrogel for the regeneration of the osteochondral interface. Specifically, the effects of HA on the response of chondrocytes were determined, focusing on changes in matrix production and mineralization, as well as scaffold mechanical properties over time. Additionally, the optimal chondrocyte population for interface tissue engineering was evaluated. It was observed that the HA phase of the composite scaffold promoted the formation of a proteoglycan- and type II collagen–rich matrix when seeded with deep zone chondrocytes. More importantly, the elevated biosynthesis translated into significant increases in both compressive and shear moduli relative to the mineral-free control. Presence of HA also promoted chondrocyte hypertrophy and type X collagen deposition. These results demonstrate that the hydrogel–calcium phosphate composite supported the formation of a calcified cartilage-like matrix and is a promising scaffold design for osteochondral interface tissue engineering. PMID:21919797

  19. Hyaluronan (HA) Interacting Proteins RHAMM and Hyaluronidase Impact Prostate Cancer Cell Behavior and Invadopodia Formation in 3D HA-Based Hydrogels

    PubMed Central

    Gurski, Lisa A.; Nguyen, Ngoc T.; Xiao, Longxi; van Golen, Kenneth L.; Jia, Xinqiao; Farach-Carson, Mary C.

    2012-01-01

    To study the individual functions of hyaluronan interacting proteins in prostate cancer (PCa) motility through connective tissues, we developed a novel three-dimensional (3D) hyaluronic acid (HA) hydrogel assay that provides a flexible, quantifiable, and physiologically relevant alternative to current methods. Invasion in this system reflects the prevalence of HA in connective tissues and its role in the promotion of cancer cell motility and tissue invasion, making the system ideal to study invasion through bone marrow or other HA-rich connective tissues. The bio-compatible cross-linking process we used allows for direct encapsulation of cancer cells within the gel where they adopt a distinct, cluster-like morphology. Metastatic PCa cells in these hydrogels develop fingerlike structures, “invadopodia”, consistent with their invasive properties. The number of invadopodia, as well as cluster size, shape, and convergence, can provide a quantifiable measure of invasive potential. Among candidate hyaluronan interacting proteins that could be responsible for the behavior we observed, we found that culture in the HA hydrogel triggers invasive PCa cells to differentially express and localize receptor for hyaluronan mediated motility (RHAMM)/CD168 which, in the absence of CD44, appears to contribute to PCa motility and invasion by interacting with the HA hydrogel components. PCa cell invasion through the HA hydrogel also was found to depend on the activity of hyaluronidases. Studies shown here reveal that while hyaluronidase activity is necessary for invadopodia and inter-connecting cluster formation, activity alone is not sufficient for acquisition of invasiveness to occur. We therefore suggest that development of invasive behavior in 3D HA-based systems requires development of additional cellular features, such as activation of motility associated pathways that regulate formation of invadopodia. Thus, we report development of a 3D system amenable to dissection of

  20. High-resolution direct 3D printed PLGA scaffolds: print and shrink.

    PubMed

    Chia, Helena N; Wu, Benjamin M

    2014-12-17

    Direct three-dimensional printing (3DP) produces the final part composed of the powder and binder used in fabrication. An advantage of direct 3DP is control over both the microarchitecture and macroarchitecture. Prints which use porogen incorporated in the powder result in high pore interconnectivity, uniform porosity, and defined pore size after leaching. The main limitations of direct 3DP for synthetic polymers are the use of organic solvents which can dissolve polymers used in most printheads and limited resolution due to unavoidable spreading of the binder droplet after contact with the powder. This study describes a materials processing strategy to eliminate the use of organic solvent during the printing process and to improve 3DP resolution by shrinking with a non-solvent plasticizer. Briefly, poly(lactic-co-glycolic acid) (PLGA) powder was prepared by emulsion solvent evaporation to form polymer microparticles. The printing powder was composed of polymer microparticles dry mixed with sucrose particles. After printing with a water-based liquid binder, the polymer microparticles were fused together to form a network by solvent vapor in an enclosed vessel. The sucrose is removed by leaching and the resulting scaffold is placed in a solution of methanol. The methanol acts as a non-solvent plasticizer and allows for polymer chain rearrangement and efficient packing of polymer chains. The resulting volumetric shrinkage is ∼80% at 90% methanol. A complex shape (honey-comb) was designed, printed, and shrunken to demonstrate isotropic shrinking with the ability to reach a final resolution of ∼400 μm. The effect of type of alcohol (i.e. methanol or ethanol), concentration of alcohol, and temperature on volumetric shrinking was studied. This study presents a novel materials processing strategy to overcome the main limitations of direct 3DP to produce high resolution PLGA scaffolds.

  1. BMP2 induced osteogenic differentiation of human umbilical cord stem cells in a peptide-based hydrogel scaffold

    NASA Astrophysics Data System (ADS)

    Lakshmana, Shruthi M.

    Craniofacial tissue loss due to traumatic injuries and congenital defects is a major clinical problem around the world. Cleft palate is the second most common congenital malformation in the United States occurring with an incidence of 1 in 700. Some of the problems associated with this defect are feeding difficulties, speech abnormalities and dentofacial anomalies. Current treatment protocol offers repeated surgeries with extended healing time. Our long-term goal is to regenerate bone in the palatal region using tissue-engineering approaches. Bone tissue engineering utilizes osteogenic cells, osteoconductive scaffolds and osteoinductive signals. Mesenchymal stem cells derived from human umbilical cord (HUMSCs) are highly proliferative with the ability to differentiate into osteogenic precursor cells. The primary objective of the study was to characterize HUMSCs and culture them in a 3D hydrogel scaffold and investigate their osteogenic potential. PuraMatrix(TM) is an injectable 3D nanofiber scaffold capable of self-assembly when exposed to physiologic conditions. Our second objective was to investigate the effect of Bone Morphogenic Protein 2 (BMP2) in enhancing the osteogenic differentiation of HUMSCs encapsulated in PuraMatrix(TM). We isolated cells isolated from Wharton's Jelly region of the umbilical cord obtained from NDRI (New York, NY). Isolated cells satisfied the minimal criteria for mesenchymal stem cells (MSCs) as defined by International Society of Cell Therapy in terms of plastic adherence, fibroblastic phenotype, surface marker expression and osteogenic differentiation. Flow Cytometry analysis showed that cells were positive for CD73, CD90 and CD105 while negative for hematopoietic marker CD34. Alkaline phosphatase activity (ALP) of HUMSCs showed peak activity at 2 weeks (p<0.05). Cells were encapsulated in 0.2% PuraMatrix(TM) at cell densities of 10x104, 20x104, 40x10 4 and 80x104. Cell viability with WST and proliferation with Live-Dead cell assays

  2. Tetronic(®)-based composite hydrogel scaffolds seeded with rat bladder smooth muscle cells for urinary bladder tissue engineering applications.

    PubMed

    Sivaraman, Srikanth; Ostendorff, Rachel; Fleishman, Benjamin; Nagatomi, Jiro

    2015-01-01

    Natural hydrogels such as collagen offer desirable properties for tissue engineering, including cell adhesion sites, but their low mechanical strength is not suitable for bladder tissue regeneration. In contrast, synthetic hydrogels such as poly (ethylene glycol) allow tuning of mechanical properties, but do not elicit protein adsorption or cell adhesion. For this reason, we explored the use of composite hydrogel blends composed of Tetronic (BASF) 1107-acrylate (T1107A) in combination with extracellular matrix moieties collagen and hyaluronic acid seeded with bladder smooth muscle cells (BSMC). This composite hydrogel supported BSMC growth and distribution throughout the construct. When compared to the control (acellular) hydrogels, mechanical properties (peak stress, peak strain, and elastic modulus) of the cellular hydrogels were significantly greater. When compared to the 7-day time point after BSMC seeding, results of mechanical testing at the 14-day time point indicated a significant increase in both ultimate tensile stress (4.1-11.6 kPa) and elastic modulus (11.8-42.7 kPa) in cellular hydrogels. The time-dependent improvement in stiffness and strength of the cellular constructs can be attributed to the continuous collagen deposition and reconstruction by BSMC seeded in the matrix. The composite hydrogel provided a biocompatible scaffold for BSMC to thrive and strengthen the matrix; further, this trend could lead to strengthening the construct to match the mechanical properties of the bladder.

  3. Repair of peripheral nerve defects in rabbits using keratin hydrogel scaffolds.

    PubMed

    Hill, Paulina S; Apel, Peter J; Barnwell, Jonathan; Smith, Tom; Koman, L Andrew; Atala, Anthony; Van Dyke, Mark

    2011-06-01

    Entubulation of transected nerves using bioabsorbable conduits is a promising alternative to sural nerve autografting, but full functional recovery is rarely achieved. Numerous studies have suggested that scaffold-based conduit fillers may promote axon regeneration, but no neuroinductive biomaterial filler has been identified. We previously showed that a nerve guide filled with keratin hydrogel actively stimulates regeneration in a mouse model, and results in functional outcomes superior to empty conduits at early time points. The goal of the present study was to develop a peripheral nerve defect model in a rabbit and assess the effectiveness of a keratin hydrogel filler. Although repairs with keratin-filled conduits were not as consistently successful as autograft overall, the use of keratin resulted in a significant improvement in conduction delay compared to both empty conduits and autograft, as well as a significant improvement in amplitude recovery compared to empty conduits when measurable regeneration did occur. Taking into account all study animals (i.e., regenerated and nonregenerated), histological assessment showed that keratin-treated nerves had significantly greater myelin thickness than empty conduits. These data support the findings of our earlier study and suggest that keratin hydrogel fillers have the potential to be used clinically to improve conduit repair.

  4. Mechanical, permeability, and degradation properties of 3D designed poly(1,8 octanediol-co-citrate) scaffolds for soft tissue engineering.

    PubMed

    Jeong, Claire G; Hollister, Scott J

    2010-04-01

    Poly(1,8-octanediol-co-citric acid) (POC) is a synthetic biodegradable elastomer that can be processed into three-dimensional (3D) scaffolds for tissue engineering. We investigated the effect of designed porosity on the mechanical properties, permeability, and degradation profiles of the POC scaffolds. For mechanical properties, scaffold compressive data were fitted to a one-dimensional (1D) nonlinear elastic model, and solid tensile data were fitted to a Neohookean incompressible nonlinear elastic model. Chondrocytes were seeded on scaffolds to assess the biocompatibility of POC. Increased porosity was associated with increased degradation rate, increased permeability, and decreased mechanical stiffness, which also became less nonlinear. Scaffold characterization in this article will provide design guidance for POC scaffolds to meet the mechanical and biological parameters needed for engineering soft tissues such as cartilage.

  5. Mechanical, Permeability, and Degradation Properties of 3D Designed Poly(1,8 Octanediol-co-Citrate)(POC) Scaffolds for Soft Tissue Engineering

    PubMed Central

    Jeong, Claire G.; Hollister, Scott J.

    2015-01-01

    Poly(1,8-octanediol-co-citric acid) (POC) is a synthetic biodegradable elastomer that can be processed into 3D scaffolds for tissue engineering. We investigated the effect of designed porosity on the mechanical properties, permeability and degradation profiles of the POC scaffolds. For mechanical properties, scaffold compressive data was fit to a 1D nonlinear elastic model and solid tensile data was fit to a Neohookean incompressible nonlinear elastic model. Chondrocytes were seeded on scaffolds to assess the biocompatibility of POC. Increased porosity was associated with increased degradation rate, increased permeability, and decreased mechanical stiffness which also became less nonlinear. Scaffold characterization in this paper will provide design guidance for POC scaffolds to meet the mechanical and biological parameters needed for engineering soft tissues such as cartilage. PMID:20091910

  6. Effects of permeability and living space on cell fate and neo-tissue development in hydrogel-based scaffolds: a study with cartilaginous model.

    PubMed

    Fan, Changjiang; Wang, Dong-An

    2015-04-01

    One bottleneck in tissue regeneration with hydrogel scaffolds is the limited understanding of the crucial factors for controlling hydrogel's physical microenvironments to regulate cell fate. Here, the effects of permeability and living space of hydrogels on encapsulated cells' behavior were evaluated, respectively. Three model hydrogel-based constructs are fabricated by using photo-crosslinkable hyaluronic acid as precursor and chondrocytes as model cell type. The better permeable hydrogels facilitate better cell viability and rapid proliferation, which lead to increased production of extracellular matrix (ECM), e.g. collagen, glycosaminoglycan. By prolonged culture, nano-sized hydrogel networks inhibit neo-tissue development, and the presence of macro-porous living spaces significantly enhance ECM deposition via forming larger cell clusters and eventually induce formation of scaffold-free neo-tissue islets. The results of this work demonstrate that the manipulation and optimization of hydrogel microenvironments, namely permeability and living space, are crucial to direct cell fate and neo-tissue formation.

  7. Using a decellularized splenic matrix as a 3D scaffold for hepatocyte cultivation in vitro: a preliminary trial.

    PubMed

    Zheng, Xing-Long; Xiang, Jun-Xi; Wu, Wan-Quan; Wang, Bo; Liu, Wen-Yan; Gao, Rui; Dong, Ding-Hui; Lv, Yi

    2015-08-18

    Using a decellularized liver matrix (DLM) to reengineer liver tissue is a promising therapy for end-stage liver disease. However, the limited supply of donor organs still hampers its potential clinical application, while a xenogenic decellularized matrix may bring a risk of zoonosis and immunological rejection. Therefore, an appropriate alternative scaffold is needed. In this research, we established a decellularized splenic matrix (DSM) in a rodent model, which preserved the 3D ultrastructure, the components of the extracellular matrix (ECM) and the native vascular network. The DSM and DLM had similar components of ECM, and similar mechanical properties. Hepatocytes were seeded to the DSM and DLM for dynamic culturing up to 6 d, and distributed both in decellularized sinusoidal spaces and around the vessels. The TUNEL-positive cell percentage in a dynamic culturing decellularized splenic matrix (dDSM) was 10.7%  ±  3.6% at 3d and 25.8%  ±  5.6% at 5d, although 14.2%  ±  4.5% and 24.8%  ±  2.9%, respectively, in a dynamic culturing decellularized liver matrix (dDLM) at the same time point (p  >  0.05). Primary hepatocytes in the dDSM and dDLM expressed albumin, G6pc and Ugt1a1. The gene expression of Cyp2b1, Cyp1a2 and HNF1α in the gene transcription level revealed hepatocytes had lower gene expression levels in the dDSM compared with the dDLM at 3d, but better than those in a sandwich culture. The cumulative albumin production at 6 d of culture was 80.7   ±   9.6 μg per million cells in the dDSM and 89.6   ±   4.6 μg per million cells in the dDLM (p  >  0.05). In summary, the DSM is a promising 3D scaffold for hepatocyte cultivation in vitro.

  8. Layer-by-layer assembly of antibacterial coating on interbonded 3D fibrous scaffolds and its cytocompatibility assessment.

    PubMed

    Tang, Yanwei; Zhao, Yan; Wang, Hongxia; Gao, Yuan; Liu, Xin; Wang, Xungai; Lin, Tong

    2012-08-01

    Bonded fibrous matrices have shown great potential in tissue engineering because of their unique 3D structures and pore characteristics. For some applications, bacterial infections must be taken into account, and antibacterial function is highly desired. In this study, an antibacterial polymer, polyhexamethylene biguanide (PHMB), was applied onto the fiber surface of a bonded poly(ε-caprolactone) (PCL) fibrous matrix with the objective to achieve both strong antibacterial effect and good cell compatibility. The coatings were prepared by using an electrostatic layer-by-layer (LbL) assembly technique, which allowed the control of PHMB loading and coating uniformity on the fiber surface. The PHMB coating provided antibacterial activities, but had no toxicity on mammalian cells. This bonded PCL fibrous matrix with electrostatically self-assembled PHMB may provide a new antiinfective tissue scaffold for various biomedical applications.

  9. Microarray analyses to quantify advantages of 2D and 3D hydrogel culture systems in maintaining the native valvular interstitial cell phenotype.

    PubMed

    Mabry, Kelly M; Payne, Samuel Z; Anseth, Kristi S

    2016-01-01

    Valvular interstitial cells (VICs) actively maintain and repair heart valve tissue; however, persistent activation of VICs to a myofibroblast phenotype can lead to aortic stenosis. To better understand and quantify how microenvironmental cues influence VIC phenotype and myofibroblast activation, we compared expression profiles of VICs cultured on poly(ethylene glycol) (PEG) gels to those cultured on tissue culture polystyrene (TCPS), as well as fresh isolates. In general, VICs cultured in hydrogel matrices had lower levels of activation (<10%), similar to levels seen in healthy valve tissue, while VICs cultured on TCPS were ∼75% activated myofibroblasts. VICs cultured on TCPS also exhibited a higher magnitude of perturbations in gene expression than soft hydrogel cultures when compared to the native phenotype. Using peptide-modified PEG gels, VICs were seeded on (2D), as well as encapsulated in (3D), matrices of the same composition and modulus. Despite similar levels of activation, VICs cultured in 2D had distinct variations in transcriptional profiles compared to those in 3D hydrogels. Genes related to cell structure and motility were particularly affected by the dimensionality of the culture platform, with higher expression levels in 2D than in 3D. These results indicate that dimensionality may play a significant role in dictating cell phenotype (e.g., through differences in polarity, diffusion of soluble signals), and emphasize the importance of using multiple metrics when characterizing cell phenotype.

  10. Scaffolds fabricated by 3D two-photon photopolymerization for live cell studies

    NASA Astrophysics Data System (ADS)

    Teplicky, T.; Cunderlikova, B.; Mateasik, A.; Vincze, A.; Chorvat, D.; Marcek Chorvatova, A.

    2016-12-01

    Design and fabrication of appropriate biocompatible microstructures that ensure fixation and control of experimental conditions for live cell and bacteria observations is an important prerequisite for number of real time experiments. Our approach is to design engineered microfabricated 3D structures for growth of cells in culture without significant modification of their metabolic state. Presented approach is aimed at evaluation of the potential applicability of biocompatible constructs in the biomedical field and thus live cell monitoring in controlled conditions. Design and evaluation of properties of materials and structures with mesoscopic arrangement and their interaction with biological objects is a prerequisite for establishment of physiologically relevant in vitro models of pathologies as well as for development of a new generation of nano / micro / bio-sensors.

  11. 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.

  12. Synthesis, Structural and Micromechanical Properties of 3D Hyaluronic Acid-Based Cryogel Scaffolds.

    PubMed

    Oelschlaeger, C; Bossler, F; Willenbacher, N

    2016-02-08

    In this study, macroporous, elastic, three-dimensional scaffolds formed of hyaluronic acid mixed with ethylene glycol diglycidyl ether as a chemical cross-linker have been prepared by cryogelation for application in tissue engineering. These cryogels are characterized by large interconnected pores of size ∼50-300 μm and pore wall thickness of ∼5-30 μm as determined from confocal microscopy images. Variation of pH, freezing temperature, and polymerization time allows for control of pore size and shape as well as matrix thickness. These structural properties then determine mechanical strength as well as swelling capacity. Furthermore, increasing hyaluronic acid concentration decreases cryogel pore size, reduces swelling properties, and reinforces mechanical properties. On the other hand, decreasing cross-linker concentration, at a constant hyaluronic acid concentration, increases pore size and swelling capacity but provides less rigidity. Additionally, for the first time, local elastic properties of the polymer matrix and viscous properties of the pores have been characterized using multiple particle tracking microrheology. Local matrix elasticity, relaxation time of hyaluronic acid chains, and the degree of heterogeneity are discussed in detail. These latter properties are crucial for the development of new tissue engineering constructs and will help to understand how local matrix viscoelasticity affects cell cultivation. Finally, elastic moduli obtained in bulk rheology are much higher than corresponding values deduced from microrheology. This discrepancy might be explained by the formation of very highly cross-linked cores in the network where no tracer particle can penetrate.

  13. Computer-aided multiple-head 3D printing system for printing of heterogeneous organ/tissue constructs

    PubMed Central

    Jung, Jin Woo; Lee, Jung-Seob; Cho, Dong-Woo

    2016-01-01

    Recently, much attention has focused on replacement or/and enhancement of biological tissues via the use of cell-laden hydrogel scaffolds with an architecture that mimics the tissue matrix, and with the desired three-dimensional (3D) external geometry. However, mimicking the heterogeneous tissues that most organs and tissues are formed of is challenging. Although multiple-head 3D printing systems have been proposed for fabricating heterogeneous cell-laden hydrogel scaffolds, to date only the simple exterior form has been realized. Here we describe a computer-aided design and manufacturing (CAD/CAM) system for this application. We aim to develop an algorithm to enable easy, intuitive design and fabrication of a heterogeneous cell-laden hydrogel scaffolds with a free-form 3D geometry. The printing paths of the scaffold are automatically generated from the 3D CAD model, and the scaffold is then printed by dispensing four materials; i.e., a frame, two kinds of cell-laden hydrogel and a support. We demonstrated printing of heterogeneous tissue models formed of hydrogel scaffolds using this approach, including the outer ear, kidney and tooth tissue. These results indicate that this approach is particularly promising for tissue engineering and 3D printing applications to regenerate heterogeneous organs and tissues with tailored geometries to treat specific defects or injuries. PMID:26899876

  14. Computer-aided multiple-head 3D printing system for printing of heterogeneous organ/tissue constructs

    NASA Astrophysics Data System (ADS)

    Jung, Jin Woo; Lee, Jung-Seob; Cho, Dong-Woo

    2016-02-01

    Recently, much attention has focused on replacement or/and enhancement of biological tissues via the use of cell-laden hydrogel scaffolds with an architecture that mimics the tissue matrix, and with the desired three-dimensional (3D) external geometry. However, mimicking the heterogeneous tissues that most organs and tissues are formed of is challenging. Although multiple-head 3D printing systems have been proposed for fabricating heterogeneous cell-laden hydrogel scaffolds, to date only the simple exterior form has been realized. Here we describe a computer-aided design and manufacturing (CAD/CAM) system for this application. We aim to develop an algorithm to enable easy, intuitive design and fabrication of a heterogeneous cell-laden hydrogel scaffolds with a free-form 3D geometry. The printing paths of the scaffold are automatically generated from the 3D CAD model, and the scaffold is then printed by dispensing four materials; i.e., a frame, two kinds of cell-laden hydrogel and a support. We demonstrated printing of heterogeneous tissue models formed of hydrogel scaffolds using this approach, including the outer ear, kidney and tooth tissue. These results indicate that this approach is particularly promising for tissue engineering and 3D printing applications to regenerate heterogeneous organs and tissues with tailored geometries to treat specific defects or injuries.

  15. Computer-aided multiple-head 3D printing system for printing of heterogeneous organ/tissue constructs.

    PubMed

    Jung, Jin Woo; Lee, Jung-Seob; Cho, Dong-Woo

    2016-02-22

    Recently, much attention has focused on replacement or/and enhancement of biological tissues via the use of cell-laden hydrogel scaffolds with an architecture that mimics the tissue matrix, and with the desired three-dimensional (3D) external geometry. However, mimicking the heterogeneous tissues that most organs and tissues are formed of is challenging. Although multiple-head 3D printing systems have been proposed for fabricating heterogeneous cell-laden hydrogel scaffolds, to date only the simple exterior form has been realized. Here we describe a computer-aided design and manufacturing (CAD/CAM) system for this application. We aim to develop an algorithm to enable easy, intuitive design and fabrication of a heterogeneous cell-laden hydrogel scaffolds with a free-form 3D geometry. The printing paths of the scaffold are automatically generated from the 3D CAD model, and the scaffold is then printed by dispensing four materials; i.e., a frame, two kinds of cell-laden hydrogel and a support. We demonstrated printing of heterogeneous tissue models formed of hydrogel scaffolds using this approach, including the outer ear, kidney and tooth tissue. These results indicate that this approach is particularly promising for tissue engineering and 3D printing applications to regenerate heterogeneous organs and tissues with tailored geometries to treat specific defects or injuries.

  16. Collagen/heparin sulfate scaffolds fabricated by a 3D bioprinter improved mechanical properties and neurological function after spinal cord injury in rats.

    PubMed

    Chen, Chong; Zhao, Ming-Liang; Zhang, Ren-Kun; Lu, Gang; Zhao, Chang-Yu; Fu, Feng; Sun, Hong-Tao; Zhang, Sai; Tu, Yue; Li, Xiao-Hong

    2017-01-25

    Effective treatments promoting axonal regeneration and functional recovery for spinal cord injury (SCI) are still in the early stages of development. Most approaches have been focused on providing supportive substrates for guiding neurons and overcoming the physical and chemical barriers to healing that arise after SCI. Although collagen has become a promising natural substrate with good compatibility, its low mechanical properties restrict its potential applications. The mechanical properties mainly rely on the composition and pore structure of scaffolds. For the composition of a scaffold, we used heparin sulfate to react with collagen by crosslinking. For the structure, we adopted a three-dimensional (3D) printing technology to fabricate a scaffold with a uniform pore distributions. We observed that the internal structure of the scaffold printed with a 3D bioprinter was regular and porous. We also found that both the compression modulus and strengths of the scaffold were significantly enhanced by the collagen/heparin sulfate composition compared to a collagen scaffold. Meanwhile, the collagen/heparin sulfate scaffold presented good biocompatibility when it was co-cultured with neural stem cells in vitro. We also demonstrated that heparin sulfate modification significantly improved bFGF immobilization and absorption to the collagen by examining the release kinetics of bFGF from scaffolds. Two months after implantating the scaffold into transection lesions in T10 of the spinal cord in rats, the collagen/heparin sulfate group demonstrated significant recovery of locomotor function and according to electrophysiological examinations. Parallel to functional recovery, collagen/heparin sulfate treatment further ameliorated the pathological process and markedly increased the number of neurofilament (NF) positive cells compared to collagen treatment alone. These data suggested that a collagen/heparin sulfate scaffold fabricated by a 3D bioprinter could enhance the

  17. Laser Speckle Rheology for evaluating the viscoelastic properties of hydrogel scaffolds

    PubMed Central

    Hajjarian, Zeinab; Nia, Hadi Tavakoli; Ahn, Shawn; Grodzinsky, Alan J.; Jain, Rakesh K.; Nadkarni, Seemantini K.

    2016-01-01

    Natural and synthetic hydrogel scaffolds exhibit distinct viscoelastic properties at various length scales and deformation rates. Laser Speckle Rheology (LSR) offers a novel, non-contact optical approach for evaluating the frequency-dependent viscoelastic properties of hydrogels. In LSR, a coherent laser beam illuminates the specimen and a high-speed camera acquires the time-varying speckle images. Cross-correlation analysis of frames returns the speckle intensity autocorrelation function, g2(t), from which the frequency-dependent viscoelastic modulus, G*(ω), is deduced. Here, we establish the capability of LSR for evaluating the viscoelastic properties of hydrogels over a large range of moduli, using conventional mechanical rheometry and atomic force microscopy (AFM)-based indentation as reference-standards. Results demonstrate a strong correlation between |G*(ω)| values measured by LSR and mechanical rheometry (r = 0.95, p < 10−9), and z-test analysis reports that moduli values measured by the two methods are identical (p > 0.08) over a large range (47 Pa – 36 kPa). In addition, |G*(ω)| values measured by LSR correlate well with indentation moduli, E, reported by AFM (r = 0.92, p < 10−7). Further, spatially-resolved moduli measurements in micro-patterned substrates demonstrate that LSR combines the strengths of conventional rheology and micro-indentation in assessing hydrogel viscoelastic properties at multiple frequencies and small length-scales. PMID:27905494

  18. Alginate/nanohydroxyapatite scaffolds with designed core/shell structures fabricated by 3D plotting and in situ mineralization for bone tissue engineering.

    PubMed

    Luo, Yongxiang; Lode, Anja; Wu, Chengtie; Chang, Jiang; Gelinsky, Michael

    2015-04-01

    Composite scaffolds, especially polymer/hydroxyapatite (HAP) composite scaffolds with predesigned structures, are promising materials for bone tissue engineering. Various methods including direct mixing of HAP powder with polymers or incubating polymer scaffolds in simulated body fluid for preparing polymer/HAP composite scaffolds are either uncontrolled or require long times of incubation. In this work, alginate/nano-HAP composite scaffolds with designed pore parameters and core/shell structures were fabricated using 3D plotting technique and in situ mineralization under mild conditions (at room temperature and without the use of any organic solvents). Light microscopy, scanning electron microscopy, microcomputer tomography, X-ray diffraction, and Fourier transform infrared spectroscopy were applied to characterize the fabricated scaffolds. Mechanical properties and protein delivery of the scaffolds were evaluated, as well as the cell response to the scaffolds by culturing human bone-marrow-derived mesenchymal stem cells (hBMSC). The obtained data indicate that this method is suitable to fabricate alginate/nano-HAP composite scaffolds with a layer of nano-HAP, coating the surface of the alginate strands homogeneously and completely. The surface mineralization enhanced the mechanical properties and improved the cell attachment and spreading, as well as supported sustaining protein release, compared to pure alginate scaffolds without nano-HAP shell layer. The results demonstrated that the method provides an interesting option for bone tissue engineering application.

  19. Balancing mechanical strength with bioactivity in chitosan-calcium phosphate 3D microsphere scaffolds for bone tissue engineering: air- vs. freeze-drying processes.

    PubMed

    Nguyen, D T; McCanless, J D; Mecwan, M M; Noblett, A P; Haggard, W O; Smith, R A; Bumgardner, J D

    2013-01-01

    The objective of this study was to evaluate the potential benefit of 3D composite scaffolds composed of chitosan and calcium phosphate for bone tissue engineering. Additionally, incorporation of mechanically weak lyophilized microspheres within those air-dried (AD) was considered for enhanced bioactivity. AD microsphere, alone, and air- and freeze-dried microsphere (FDAD) 3D scaffolds were evaluated in vitro using a 28-day osteogenic culture model with the Saos-2 cell line. Mechanical testing, quantitative microscopy, and lysozyme-driven enzymatic degradation of the scaffolds were also studied. FDAD scaffold showed a higher concentration (p < 0.01) in cells per scaffold mass vs. AD constructs. Collagen was ∼31% greater (p < 0.01) on FDAD compared to AD scaffolds not evident in microscopy of microsphere surfaces. Alternatively, AD scaffolds demonstrated a superior threefold increase in compressive strength over FDAD (12 vs. 4 MPa) with minimal degradation. Inclusion of FD spheres within the FDAD scaffolds allowed increased cellular activity through improved seeding, proliferation, and extracellular matrix production (as collagen), although mechanical strength was sacrificed through introduction of the less stiff, porous FD spheres.

  20. Biofunctionalization of electrospun PCL-based scaffolds with perlecan domain IV peptide to create a 3-D pharmacokinetic cancer model

    PubMed Central

    Hartman, Olga; Zhang, Chu; Adams, Elizabeth L.; Farach-Carson, Mary C.; Petrelli, Nicholas J.; Chase, Bruce D.; Rabolt, John F.

    2010-01-01

    Because prostate cancer cells metastasize to bone and exhibit osteoblastic features (osteomimicry), the interrelationships between bone-specific microenvironment and prostate cancer cells at sites of bone metastasis are critical to disease progression. In this work the bone marrow microenvironment in vitro was recreated both by tailoring scaffolds physical properties and by functionalizing electrospun polymer fibers with a bioactive peptide derived from domain IV of perlecan heparan sulfate proteoglycan. Electrospun poly (ε-caprolactone) (PCL) fibers and PCL/gelatin composite scaffolds were modified covalently with perlecan domain IV (PlnDIV) peptide. The expression of tight junction protein (E-cadherin) and focal adhesion kinase (FAK) phosphorylation on tyrosine 397 also were investigated. The described bioactive motif significantly enhanced adherence and infiltration of the metastatic prostate cancer cells on all modified electrospun substrates by day 5 post-seeding. Cells cultured on PlnDIV-modified matrices organized stress fibers and increased proliferation at statistically significant rates. Additional findings suggest that presence of PlnDIV peptide in the matrix reduced expression of tight junction protein and binding to PlnDIV peptide was accompanied by increased focal adhesion kinase (FAK) phosphorylation on tyrosine 397. We conclude that PlnDIV peptide supports key signaling events leading to proliferation, survival, and migration of C4-2B cancer cells; hence its incorporation into electrospun matrix is a key improvement to create a successful three-dimensional (3-D) pharmacokinetic cancer model. PMID:20417554

  1. Image-based analysis of the internal microstructure of bone replacement scaffolds fabricated by 3D printing

    NASA Astrophysics Data System (ADS)

    Irsen, Stephan H.; Leukers, Barbara; Bruckschen, Björn; Tille, Carsten; Seitz, Hermann; Beckmann, Felix; Müller, Bert

    2006-08-01

    Rapid Prototyping and especially the 3D printing, allows generating complex porous ceramic scaffolds directly from powders. Furthermore, these technologies allow manufacturing patient-specific implants of centimeter size with an internal pore network to mimic bony structures including vascularization. Besides the biocompatibility properties of the base material, a high degree of open, interconnected porosity is crucial for the success of the synthetic bone graft. Pores with diameters between 100 and 500 μm are the prerequisite for vascularization to supply the cells with nutrients and oxygen, because simple diffusion transport is ineffective. The quantification of porosity on the macro-, micro-, and nanometer scale using well-established techniques such as Hg-porosimetry and electron microscopy is restricted. Alternatively, we have applied synchrotron-radiation-based micro computed tomography (SRμCT) to determine the porosity with high precision and to validate the macroscopic internal structure of the scaffold. We report on the difficulties in intensity-based segmentation for nanoporous materials but we also elucidate the power of SRμCT in the quantitative analysis of the pores at the different length scales.

  2. Novel 3D scaffold with enhanced physical and cell response properties for bone tissue regeneration, fabricated by patterned electrospinning/electrospraying.

    PubMed

    Hejazi, Fatemeh; Mirzadeh, Hamid

    2016-09-01

    Developing three dimensional scaffolds mimicking the nanoscale structure of native extracellular matrix is a key parameter in tissue regeneration. In this study, we aimed to introduce a novel 3D structures composed of nanofibers (NF) and micro particles (MP) and compare their efficiency with 2D nanofibrous scaffold. The conventional nanofibrous PCL scaffolds are 2D mats fabricated by the electrospinning technique, whereas the NF/MP and patterned NF/MP PCL scaffolds are three dimensional structures fabricated by a modified electrospinning/electrospraying technique. The mentioned method was carried out by varying the electrospinning solution parameters and use of a metal mesh as the collector. Detailed fabrication process and morphological properties of the fabricated structures is discussed and porosity, pore size and PBS solution absorption value of the prepared structures are reported. Compared with the 2D structure, 3D scaffolds possessed enhanced porosity and pore size which led to the significant increase in their water uptake capacity. In vitro cell experiments were carried out on the prepared structures by the use of MG-63 osteosarcoma cell line. The fabricated 3D structures offered significantly increased cell attachment, spread and diffusion which were confirmed by SEM analysis. In vitro cytocompatibility assessed by MTT colorimetric assay indicated a continuous cell proliferation over 21 days on the innovative 3D structure, while on 2D mat cell proliferation stopped at early time points. Enhanced osteogenic differentiation of the seeded MG-63 cells on 3D scaffold was confirmed by the remarkable ALP activity together with increased and accelerated calcium deposition on this structure compared to 2D mat. Massive and well distributed bone minerals formed on patterned 3D structure were shown by EDX analysis. In comparison between NF/MP quasi-3D and Patterned NF/MP 3D scaffolds, patterned structures proceeded in all of the above properties. As such, the

  3. Comparing different methods to fix and to dehydrate cells on alginate hydrogel scaffolds using scanning electron microscopy.

    PubMed

    Santana, Bianca Palma; Nedel, Fernanda; Perelló Ferrúa, Camila; Marques e Silva, Ricardo; da Silva, Adriana Fernandes; Demarco, Flávio Fernando; Lenin Villarreal Carreño, Neftali

    2015-07-01

    Scanning electron microscopy (SEM) is commonly used in the analysis of scaffolds morphology, as well as cell attachment, morphology and spreading on to the scaffolds. However, so far a specific methodology to prepare the alginate hydrogel (AH) scaffolds for SEM analysis has not been evaluated. This study compared different methods to fix/dehydrate cells in AH scaffolds for SEM analysis. AH scaffolds were prepared and seeded with NIH/3T3 cell line; fixed with glutaraldehyde, osmium tetroxide, or the freeze drying method and analyzed by SEM. Results demonstrated that the freeze dried method interferes less with cell morphology and density, and preserves the scaffolds structure. The fixation with glutaraldehyde did not affect cells morphology and density; however, the scaffolds morphology was affected in some level. The fixation with osmium tetroxide interfered in the natural structure of cells and scaffold. In conclusion the freeze drying and glutaraldehyde are suitable methods for cell fixation in AH scaffold for SEM, although scaffolds structure seems to be affected by glutaraldehyde.

  4. First step in developing a 3D biodegradable fibrin scaffold for an artificial ovary

    PubMed Central

    2013-01-01

    Background Although transplantation of cryopreserved ovarian tissue is a promising approach to restore fertility in cancer patients, it is not advisable for women at risk of ovarian involvement due to the threat of reintroducing malignant cells. The aim of this study was therefore to find an alternative for these patients by development of an artificial ovary. Methods For construction of the artificial ovary matrix, we used a central composite design to investigate nine combinations of fibrinogen (mg/ml) and thrombin (IU/mL) (F/T): F1/T4, F12.5/T1, F12.5/T20, F25/T0.1, F25/T4, F25/T500, F50/T1, F50/T20 and F100/T4. From the first qualitative analyses (handling and matrix size), five combinations (F12.5/T1, F25/T4, F50/T20, F50/T1 and F100/T4) yielded positive results. They were further evaluated in order to assess fibrin matrix degradation and homogeneous cell encapsulation (density), survival and proliferation (Ki67), and atresia (TUNEL) before and after 7 days of in vitro culture. To determine the best compromise between maximizing the dynamic density (Y1) and minimizing the apoptosis rate (Y2), we used the desirability function approach. Results Two combinations (F12.5/T1 and F25/T4) showed greater distribution of cells before in vitro culture, reproducible degradation of the fibrin network and adequate support for isolated human ovarian stromal cells, with a high proportion of Ki67-positive cells. SEM analysis revealed a network of fibers with regular pores and healthy stromal cells after in vitro culture with both F/T combinations. Conclusion This study reports two optimal F/T combinations that allow survival and proliferation of isolated human ovarian cells. Further studies are required to determine if such a scaffold will also be a suitable environment for isolated ovarian follicles. PMID:24274108

  5. Association of electrospinning with electrospraying: a strategy to produce 3D scaffolds with incorporated stem cells for use in tissue engineering

    PubMed Central

    Braghirolli, Daikelly Iglesias; Zamboni, Fernanda; Acasigua, Gerson AX; Pranke, Patricia

    2015-01-01

    In tissue engineering, a uniform cell occupation of scaffolds is crucial to ensure the success of tissue regeneration. However, this point remains an unsolved problem in 3D scaffolds. In this study, a direct method to integrate cells into fiber scaffolds was investigated by combining the methods of electrospinning of fibers and bioelectrospraying of cells. With the associating of these methods, the cells were incorporated into the 3D scaffolds while the fibers were being produced. The scaffolds containing cells (SCCs) were produced using 20% poly(lactide-co-glycolide) solution for electrospinning and mesenchymal stem cells from deciduous teeth as a suspension for bioelectrospraying. After their production, the SCCs were cultivated for 15 days at 37°C with an atmosphere of 5% CO2. The 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide test demonstrated that the cells remained viable and were able to grow between the fibers. Scanning electron microscopy showed the presence of a high number of cells in the structure of the scaffolds and confocal images demonstrated that the cells were able to adapt and spread between the fibers. Histological analysis of the SCCs after 1 day of cultivation showed that the cells were uniformly distributed throughout the thickness of the scaffolds. Some physicochemical properties of the scaffolds were also investigated. SCCs exhibited good mechanical properties, compatible with their handling and further implantation. The results obtained in the present study suggest that the association of electrospinning and bioelectrospraying provides an interesting tool for forming 3D cell-integrated scaffolds, making it a viable alternative for use in tissue engineering. PMID:26316747

  6. 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.

  7. Prostate cancer xenografts engineered from 3D precision-porous poly(2-hydroxyethyl methacrylate) hydrogels as models for tumorigenesis and dormancy escape

    PubMed Central

    Long, Thomas J.; Sprenger, Cynthia C.; Plymate, Stephen R.; Ratner, Buddy D.

    2014-01-01

    Synthetic biomaterial scaffolds show promise for in vitro and in vivo 3D cancer models. Tumors engineered in biomaterial scaffolds have shown evidence of being more physiologically relevant than some traditional preclinical model systems, and synthetic biomaterials provide the added benefit of defined and consistent microenvironmental control. Here, we examine sphere-templated poly(2-hydroxyethyl methacrylate) (pHEMA) scaffolds as the basis for engineering xenografts from multiple human prostate cancer cell lines. pHEMA scaffolds seeded and pre-cultured with tumorigenic M12 cells prior to implantation generated tumors in athymic nude mice, demonstrating the ability of the scaffolds to be used as a synthetic vehicle for xenograft generation. pHEMA scaffolds seeded with LNCaP C4-2 cells, which require Matrigel or stromal cell support for tumor formation, were poorly tumorigenic up to twelve weeks after implantation even when Matrigel was infused into the scaffold, demonstrating a lack of necessary pro-tumorigenic signaling within the scaffolds. Finally, M12mac25 cells, which are ordinarily rendered non-tumorigenic through the expression of the tumor suppressor insulin-like growth factor binding protein 7 (IGFBP7), displayed a tumorigenic response when implanted within porous pHEMA scaffolds. These M12mac25 tumors showed a significantly higher macrophage infiltration within the scaffolds driven by the foreign body response to the materials. These findings show the potential for this biomaterials-based model system to be used in the study of prostate cancer tumorigenesis and dormancy escape. PMID:24942815

  8. Printing Thermoresponsive Reverse Molds for the Creation of Patterned Two-component Hydrogels for 3D Cell Culture

    PubMed Central

    Müller, Michael; Becher, Jana; Schnabelrauch, Matthias; Zenobi-Wong, Marcy

    2013-01-01

    Bioprinting is an emerging technology that has its origins in the rapid prototyping industry. The different printing processes can be divided into contact bioprinting1-4 (extrusion, dip pen and soft lithography), contactless bioprinting5-7 (laser forward transfer, ink-jet deposition) and laser based techniques such as two photon photopolymerization8. It can be used for many applications such as tissue engineering9-13, biosensor microfabrication14-16 and as a tool to answer basic biological questions such as influences of co-culturing of different cell types17. Unlike common photolithographic or soft-lithographic methods, extrusion bioprinting has the advantage that it does not require a separate mask or stamp. Using CAD software, the design of the structure can quickly be changed and adjusted according to the requirements of the operator. This makes bioprinting more flexible than lithography-based approaches. Here we demonstrate the printing of a sacrificial mold to create a multi-material 3D structure using an array of pillars within a hydrogel as an example. These pillars could represent hollow structures for a vascular network or the tubes within a nerve guide conduit. The material chosen for the sacrificial mold was poloxamer 407, a thermoresponsive polymer with excellent printing properties which is liquid at 4 °C and a solid above its gelation temperature ~20 °C for 24.5% w/v solutions18. This property allows the poloxamer-based sacrificial mold to be eluted on demand and has advantages over the slow dissolution of a solid material especially for narrow geometries. Poloxamer was printed on microscope glass slides to create the sacrificial mold. Agarose was pipetted into the mold and cooled until gelation. After elution of the poloxamer in ice cold water, the voids in the agarose mold were filled with alginate methacrylate spiked with FITC labeled fibrinogen. The filled voids were then cross-linked with UV and the construct was imaged with an epi

  9. Printing thermoresponsive reverse molds for the creation of patterned two-component hydrogels for 3D cell culture.

    PubMed

    Müller, Michael; Becher, Jana; Schnabelrauch, Matthias; Zenobi-Wong, Marcy

    2013-07-10

    Bioprinting is an emerging technology that has its origins in the rapid prototyping industry. The different printing processes can be divided into contact bioprinting(1-4) (extrusion, dip pen and soft lithography), contactless bioprinting(5-7) (laser forward transfer, ink-jet deposition) and laser based techniques such as two photon photopolymerization(8). It can be used for many applications such as tissue engineering(9-13), biosensor microfabrication(14-16) and as a tool to answer basic biological questions such as influences of co-culturing of different cell types(17). Unlike common photolithographic or soft-lithographic methods, extrusion bioprinting has the advantage that it does not require a separate mask or stamp. Using CAD software, the design of the structure can quickly be changed and adjusted according to the requirements of the operator. This makes bioprinting more flexible than lithography-based approaches. Here we demonstrate the printing of a sacrificial mold to create a multi-material 3D structure using an array of pillars within a hydrogel as an example. These pillars could represent hollow structures for a vascular network or the tubes within a nerve guide conduit. The material chosen for the sacrificial mold was poloxamer 407, a thermoresponsive polymer with excellent printing properties which is liquid at 4 °C and a solid above its gelation temperature ~20 °C for 24.5% w/v solutions(18). This property allows the poloxamer-based sacrificial mold to be eluted on demand and has advantages over the slow dissolution of a solid material especially for narrow geometries. Poloxamer was printed on microscope glass slides to create the sacrificial mold. Agarose was pipetted into the mold and cooled until gelation. After elution of the poloxamer in ice cold water, the voids in the agarose mold were filled with alginate methacrylate spiked with FITC labeled fibrinogen. The filled voids were then cross-linked with UV and the construct was imaged with an

  10. Antibacterial and cell-adhesive polypeptide and poly(ethylene glycol) hydrogel as a potential scaffold for wound healing.

    PubMed

    Song, Airong; Rane, Aboli A; Christman, Karen L

    2012-01-01

    The ideal wound-healing scaffold should provide the appropriate physical and mechanical properties to prevent secondary infection, as well as an excellent physiological environment to facilitate cell adhesion, proliferation and/or differentiation. Therefore, we developed a synthetic cell-adhesive polypeptide hydrogel with inherent antibacterial activity. A series of polypeptides, poly(Lys)(x)(Ala)(y) (x+y=100), with varied hydrophobicity via metal-free ring-opening polymerization of NCA-Lys(Boc) and NCA-Ala monomers (NCA=N-carboxylic anhydride) mediated by hexamethyldisilazane (HMDS) were synthesized. These polypeptides were cross-linked with 6-arm polyethylene glycol (PEG)-amide succinimidyl glutarate (ASG) (M(w)=10K) to form hydrogels with a gelation time of five minutes and a storage modulus (G') of 1400-3000 Pa as characterized by rheometry. The hydrogel formed by cross-linking of poly(Lys)(60)(Ala)(40) (5 wt.%) and 6-arm PEG-ASG (16 wt.%) (Gel-III) exhibited cell adhesion and cell proliferation activities superior to other polypeptide hydrogels. In addition, Gel-III displays significant antibacterial activity against Escherichia coli JM109 and Staphylococcus aureus ATCC25923. Thus, we have developed a novel, cell-adhesive hydrogel with inherent antibacterial activity as a potential scaffold for cutaneous wound healing.

  11. Antibacterial and Cell-adhesive Polypeptide and Poly(ethylene glycol) Hydrogel as a Potential Scaffold for Wound Healing

    PubMed Central

    Song, Airong; Rane, Aboli A.; Christman, Karen L.

    2011-01-01

    The ideal wound healing scaffold should provide the appropriate physical and mechanical properties to prevent secondary infection, as well as an excellent physiological environment to facilitate cell adhesion, proliferation and/or differentiation. Therefore, we developed a synthetic cell-adhesive polypeptide hydrogel with inherent antibacterial activity. A series of polypeptides, poly(Lys)x(Ala)y (x+y=100) with varied hydrophobicity via metal-free ring-opening polymerization of NCA-Lys(Boc) and NCA-Ala monomers (NCA = N-carboxylic anhydride) mediated by hexamethyldisilazane (HMDS) were synthesized. These polypeptides were cross-linked with 6-arm PEG-amide succinimidyl glutarate (ASG) (Mw = 10K) to form hydrogels with a gelation time of five minutes and a storage modulus (G') of 1400–3000 Pa as characterized by rheometry. The hydrogel formed by cross-linking of poly(Lys)60(Ala)40 (5 wt%) and 6-arm PEG-ASG (16 wt%) (Gel-III) exhibited cell adhesion and cell proliferation activities superior to other polypeptide hydrogels. In addition, Gel-III displays significant antibacterial activity against E. coli JM109 and S. aureus ATCC25923. Thus, we have developed a novel, cell-adhesive hydrogel with inherent antibacterial activity as a potential scaffold for cutaneous wound healing. PMID:22023748

  12. Alginate hydrogel enriched with enamel matrix derivative to target osteogenic cell differentiation in TiO2 scaffolds.

    PubMed

    Pullisaar, Helen; Verket, Anders; Szoke, Krisztina; Tiainen, Hanna; Haugen, Håvard J; Brinchmann, Jan E; Reseland, Janne E; Østrup, Esben

    2015-01-01

    The purpose of bone tissue engineering is to employ scaffolds, cells, and growth factors to facilitate healing of bone defects. The aim of this study was to assess the viability and osteogenic differentiation of primary human osteoblasts and adipose tissue-derived mesenchymal stem cells from various donors on titanium dioxide (TiO2) scaffolds coated with an alginate hydrogel enriched with enamel matrix derivative. Cells were harvested for quantitative reverse transcription polymerase chain reaction on days 14 and 21, and medium was collected on days 2, 14, and 21 for protein analyses. Neither coating with alginate hydrogel nor alginate hydrogel enriched with enamel matrix derivative induced a cytotoxic response. Enamel matrix derivative-enriched alginate hydrogel significantly increased the expression of osteoblast markers COL1A1, TNFRSF11B, and BGLAP and secretion of osteopontin in human osteoblasts, whereas osteogenic differentiation of human adipose tissue-derived mesenchymal stem cells seemed unaffected by enamel matrix derivative. The alginate hydrogel coating procedure may have potential for local delivery of enamel matrix derivative and other stimulatory factors for use in bone tissue engineering.

  13. Alginate hydrogel enriched with enamel matrix derivative to target osteogenic cell differentiation in TiO2 scaffolds

    PubMed Central

    Pullisaar, Helen; Verket, Anders; Szoke, Krisztina; Tiainen, Hanna; Haugen, Håvard J; Brinchmann, Jan E; Reseland, Janne E

    2015-01-01

    The purpose of bone tissue engineering is to employ scaffolds, cells, and growth factors to facilitate healing of bone defects. The aim of this study was to assess the viability and osteogenic differentiation of primary human osteoblasts and adipose tissue–derived mesenchymal stem cells from various donors on titanium dioxide (TiO2) scaffolds coated with an alginate hydrogel enriched with enamel matrix derivative. Cells were harvested for quantitative reverse transcription polymera