Enhancing the Hydrophilicity and Cell Attachment of 3D Printed PCL/Graphene Scaffolds for Bone Tissue Engineering.
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ABSTRACT:
SUBMITTER: Wang W
PROVIDER: S-EPMC5456956 | biostudies-other | 2016 Dec
REPOSITORIES: biostudies-other
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Project description:Multifunctional and resistant 3D structures represent a great promise and a great challenge in bone tissue engineering. This study addresses this problem by employing polycaprolactone (PCL)-based scaffolds added with hydroxyapatite (HAp) and superparamagnetic iron oxide nanoparticles (SPION), able to drive on demand the necessary cells and other bioagents for a high healing efficiency. PCL-HAp-SPION scaffolds with different concentrations of the superparamagnetic component were developed through the 3D-printing technology and the specific topographical features were detected by Atomic Force and Magnetic Force Microscopy (AFM-MFM). AFM-MFM measurements confirmed a homogenous distribution of HAp and SPION throughout the surface. The magnetically assisted seeding of cells in the scaffold resulted most efficient for the 1% SPION concentration, providing good cell entrapment and adhesion rates. Mesenchymal Stromal Cells (MSCs) seeded onto PCL-HAp-1% SPION showed a good cell proliferation and intrinsic osteogenic potential, indicating no toxic effects of the employed scaffold materials. The performed characterizations and the collected set of data point on the inherent osteogenic potential of the newly developed PCL-HAp-1% SPION scaffolds, endorsing them towards next steps of in vitro and in vivo studies and validations.
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Project description:This work aims at providing guidance through systematic experimental characterization, for the design of 3D printed scaffolds for potential orthopaedic applications, focusing on fused deposition modeling (FDM) with a composite of clinically available polycaprolactone (PCL) and ?-tricalcium phosphate (?-TCP). First, we studied the effect of the chemical composition (0% to 60% ?-TCP/PCL) on the scaffold's properties. We showed that surface roughness and contact angle were respectively proportional and inversely proportional to the amount of ?-TCP, and that degradation rate increased with the amount of ceramic. Biologically, the addition of ?-TCP enhanced proliferation and osteogenic differentiation of C3H10. Secondly, we systematically investigated the effect of the composition and the porosity on the 3D printed scaffold mechanical properties. Both an increasing amount of ?-TCP and a decreasing porosity augmented the apparent Young's modulus of the 3D printed scaffolds. Third, as a proof-of-concept, a novel multi-material biomimetic implant was designed and fabricated for potential disk replacement.
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Project description:The regeneration of bone remains one of the main challenges in the biomedical field, with the need to provide more personalized and multifunctional solutions. The other persistent challenge is related to the local prevention of infections after implantation surgery. To fulfill the first one and provide customized scaffolds with complex geometries, 3D printing is being investigated, with polylactic acid (PLA) as the biomaterial mostly used, given its thermoplastic properties. The 3D printing of PLA in combination with hydroxyapatite (HA) is also under research, to mimic the native mechanical and biological properties, providing more functional scaffolds. Finally, to fulfill the second one, antibacterial drugs locally incorporated into biodegradable scaffolds are also under investigation. This work aims to develop vancomycin-loaded 3D-printed PLA-HA scaffolds offering a dual functionality: local prevention of infections and personalized biodegradable scaffolds with osseointegrative properties. For this, the antibacterial drug vancomycin was incorporated into 3D-printed PLA-HA scaffolds using three loading methodologies: (1) dip coating, (2) drop coating, and (3) direct incorporation in the 3D printing with PLA and HA. A systematic characterization was performed, including release kinetics, Staphylococcus aureus antibacterial/antibiofilm activities and cytocompatibility. The results demonstrated the feasibility of the vancomycin-loaded 3D-printed PLA-HA scaffolds as drug-releasing vehicles with significant antibacterial effects for the three methodologies. In relation to the drug release kinetics, the (1) dip- and (2) drop-coating methodologies achieved burst release (first 60 min) of around 80-90% of the loaded vancomycin, followed by a slower release of the remaining drug for up to 48 h, while the (3) 3D printing presented an extended release beyond 7 days as the polymer degraded. The cytocompatibility of the vancomycin-loaded scaffolds was also confirmed.
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Project description:Recent challenges in post-surgical bone tumor management have elucidated the need for a multifunctional scaffold, which can be used for residual tumor-cell suppression, defect repair, and simultaneous bone regeneration. In this perspective, 3D printing allows to create a wide variety of patient-specific implant with complex porous architecture and compatible mechanical strength to that of cancellous bone. Here, a multifunctional bone graft substitute is designed by incorporating the three primary soy isoflavones: genistein, daidzein, and glycitein onto a 3D printed (3DP) tricalcium phosphate (TCP) scaffolds with designed pores, endowing them with in vitro chemopreventive, bone-cell proliferating and immune-modulatory potential. The interconnected porosity and biodegradability of 3DP TCP ceramics have allowed controlled release kinetics of genistein, daidzein and glycitein in acidic and physiological buffer medium for 16 days, which is fitted with Korsmeyer-Peppas model. Presence of genistein, a well-known natural biomolecule shows a 90% reduction in vitro osteosarcoma (MG-63) cell viability and proliferation after 11 days. Meanwhile, daidzein, the other primary isoflavone, promotes in vitro cellular attachment and enhances viability and proliferation of human fetal osteoblast cell (hFOB). Furthermore, controlled release of genistein, daidzein, and glycitein from 3DP TCP scaffold demonstrates improved hFOB cell proliferation, viability, and differentiation in a dynamic flow-perfusion bioreactor, which is utilized to better simulate the clinical microenvironment. Finally, in vivo H&E staining confirms controlled co-delivery of genistein-daidzein-glycitein from 3DP scaffold carefully modulated neutrophil recruitment to the surgery site after 24 h of implantation in a rat distal femur model. These results advance our understanding towards multipronged therapeutic approaches utilizing synthetic bone graft substitutes as a drug delivery vehicle, and more importantly, demonstrate the feasibility of localized tumor cell suppression and bone cell proliferation for post-surgical defect repair application. STATEMENT OF SIGNIFICANCE: Designed multimodal porosity of 3D printed TCP scaffold allows a controlled and sustained release of soy isoflavones, genistein, daidzein and glycitein in both physiological and acidic pH. Presence of genistein shows 90% reduction in vitro bone cancer cell viability and proliferation. Meanwhile, controlled release of genistein, daidzein, and glycitein from 3DP TCP scaffolds demonstrate improved osteoblast cell proliferation, viability, and differentiation in static and dynamic flow-perfusion bioreactor. Furthermore, H&E staining at 24 h post-surgical specimens from rat distal femur model shows neutrophil recruitment at the surgery site is significantly decreased, suggesting the anti-inflammatory property of soy isoflavones. This work provides deeper understanding on the design of a multifunctional 3D printed patient-specific scaffold with enhanced in vitro chemopreventive, osteogenic and in vivo anti-inflammatory ability.
| S-EPMC8009492 | biostudies-literature
Project description:Bioengineering scaffolds have been improved to achieve efficient regeneration of various damaged tissues. In this study, we attempted to fabricate mechanically and biologically activated 3D printed scaffold in which porous gelatin/hydroxyapatite (G/H) as a matrix material provided outstanding mechanical properties with recoverable behavior, and human placental extracts (hPE) embedded in the scaffold were used as bioactive components. Methods: Various cell types (human adipose-derived stem cells; hASCs, pre-osteoblast; MC3T3-E1, human endothelial cell line; EA.hy926, and human dermal fibroblast; hDFs) were used to assess the effect of the hPE on cellular responses. High weight fraction (~ 70 wt%) of hydroxyapatite (HA) in a gelatin solution supplemented with glycerol was used for the G/H scaffold fabrication, and the scaffolds were immersed in hPE for the embedding (G/H/hPE scaffold). The osteogenic abilities of the scaffolds were investigated in cultured cells (hASCs) assaying for ALP activity and expression of osteogenic genes. For the in vivo test, the G/H and G/H/hPE scaffolds were implanted in the rat mastoid obliteration model. Results: The G/H/hPE scaffold presented unique elastic recoverable properties, which are important for efficient usage of implantable scaffolds. The effects of G/H and G/H/hPE scaffold on various in vitro cell-activities including non-toxicity, biocompatibility, and cell proliferation were investigated. The in vitro results indicated that proliferation (G/H = 351.1 ± 13.3%, G/H/hPE = 430.9 ± 8.7% at day 14) and expression of osteogenic markers (ALP: 3.4-fold, Runx2: 3.9-fold, BMP2: 1.7-fold, OPN: 2.4-fold, and OCN: 4.8-fold at day 21) of hASCs grown in the G/H/hPE scaffold were significantly enhanced compared with that in cells grown in the G/H scaffold. In addition, bone formation was also observed in an in vivo model using rat mastoid obliteration. Conclusion: In vitro and in vivo results suggested that the G/H/hPE scaffold is a potential candidate for use in bone tissue engineering.
| S-EPMC9169369 | biostudies-literature
Project description:Three-dimensional (3D) printing technology enables the design of personalized scaffolds with tunable pore size and composition. Combining decellularization and 3D printing techniques provides the opportunity to fabricate scaffolds with high potential to mimic native tissue. The aim of this study is to produce novel decellularized bone extracellular matrix (dbECM)-reinforced composite-scaffold that can be used as a biomaterial for bone tissue engineering. Decellularized bone particles (dbPTs, ∼100 μm diameter) were obtained from rabbit femur and used as a reinforcement agent by mixing with gelatin (GEL) in different concentrations. 3D scaffolds were fabricated by using an extrusion-based bioprinter and crosslinking with microbial transglutaminase (mTG) enzyme, followed by freeze-drying to obtain porous structures. Fabricated 3D scaffolds were characterized morphologically, mechanically, and chemically. Furthermore, MC3T3-E1 mouse pre-osteoblast cells were seeded on the dbPTs reinforced GEL scaffolds (GEL/dbPTs) and cultured for 21 days to assess cytocompatibility and cell attachment. We demonstrate the 3D-printability of dbPTs-reinforced GEL hydrogels and the achievement of homogenous distribution of the dbPTs in the whole scaffold structure, as well as bioactivity and cytocompatibility of GEL/dbPTs scaffolds. It was shown that Young's modulus and degradation rate of scaffolds were enhanced with increasing dbPTs content. Multiphoton microscopy imaging displayed the interaction of cells with dbPTs, indicating attachment and proliferation of cells around the particles as well as into the GEL-particle hydrogels. Our results demonstrate that GEL/dbPTs hydrogel formulations have potential for bone tissue engineering.
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Project description:Knee osteoarthritis is a chronic disease caused by the deterioration of the knee joint due to various factors such as aging, trauma, and obesity, and the nonrenewable nature of the injured cartilage makes the treatment of osteoarthritis challenging. Here, we present a three-dimensional (3D) printed porous multilayer scaffold based on cold-water fish skin gelatin for osteoarticular cartilage regeneration. To make the scaffold, cold-water fish skin gelatin was combined with sodium alginate to increase viscosity, printability, and mechanical strength, and the hybrid hydrogel was printed according to a pre-designed specific structure using 3D printing technology. Then, the printed scaffolds underwent a double-crosslinking process to enhance their mechanical strength even further. These scaffolds mimic the structure of the original cartilage network in a way that allows chondrocytes to adhere, proliferate, and communicate with each other, transport nutrients, and prevent further damage to the joint. More importantly, we found that cold-water fish gelatin scaffolds were nonimmunogenic, nontoxic, and biodegradable. We also implanted the scaffold into defective rat cartilage for 12 weeks and achieved satisfactory repair results in this animal model. Thus, cold-water fish skin gelatin scaffolds may have broad application potential in regenerative medicine.
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Project description:The functionality or survival of tissue engineering constructs depends on the adequate vascularization through oxygen transport and metabolic waste removal at the core. This study reports the presence of magnesium and silicon in direct three dimensional printed (3DP) tricalcium phosphate (TCP) scaffolds promotes in vivo osteogenesis and angiogenesis when tested in rat distal femoral defect model. Scaffolds with three different interconnected macro pore sizes were fabricated using direct three dimensional printing. In vitro ion release in phosphate buffer for 30 days showed sustained Mg2+ and Si4+ release from these scaffolds. Histolomorphology and histomorphometric analysis from the histology tissue sections revealed a significantly higher bone formation, between 14 and 20% for 4-16 weeks, and blood vessel formation, between 3 and 6% for 4-12 weeks, due to the presence of magnesium and silicon in TCP scaffolds compared to bare TCP scaffolds. The presence of magnesium in these 3DP TCP scaffolds also caused delayed TRAP activity. These results show that magnesium and silicon incorporated 3DP TCP scaffolds with multiscale porosity have huge potential for bone tissue repair and regeneration.
| S-EPMC5149117 | biostudies-literature
Project description:Background/Objective: Tissue engineering involves scaffolds, cells and growth factors, among which growth factors have limited applications due to potential safety risks and high costs. Therefore, an alternative approach to exogenously induce osteogenesis is desirable. Considering that osteogenesis and angiogenesis are coupled, a system of human umbilical vein endothelial cells (HUVECs) and human bone mesenchymal stem cells (hBMSCs) coculture is more biologically adapted to the microenvironment in vivo and can mediate osteogenesis and angiogenesis via paracrine signalling. Hence, in this study, a HUVECs/hBMSCs coculture system with appropriate cell and medium proportions was established. The substrate for the coculture system was a 3D-printed composite bioceramic scaffold (β-TCP/CaSiO3) based on a previous study. The aim of this study was to explore the potential of this system for bone tissue engineering.MethodsBioactive ceramic scaffolds for tissue engineering were fabricated via a 3D Bioplotter™ system. The coculture system for in vitro and in vivo studies consisted of direct contact between HUVECs and hBMSCs cultured on the 3D-printed scaffolds.ResultsThe proportions of HUVECs/hBMSCs and medium components were determined by cell viability, and the coculture system showed negligible cytotoxicity. CD31 secreted by HUVECs formed strings, and cells tended to aggregate in island chain-like arrays. Real-time cell tracking showed that HUVECs were recruited by hBMSCs, and the integrin expression by HUVECs was upregulated. Ultimately, osteogenic and angiogenic marker gene expression and protein secretion were upregulated. Moreover, the obtained bone tissue engineering scaffolds could induce early osteogenic protein secretion and capillary tube formation in nude rats.ConclusionThese bone tissue engineering scaffolds without exogenous growth factors exhibited the ability to promote osteogenesis/angiogenesis.Translational potential of this articleThe fabricated 3D-printed bioactive ceramic scaffolds could provide mechanical, biodegradable and bioadaptive support for personalized bone regeneration. In addition, the bone tissue engineering scaffolds exhibited the ability to promote osteogenesis/angiogenesis without the addition of exogenous growth factors, thus mitigating safety risks. Although application of the HUVECs/hBMSCs coculture system might be a time-consuming process, further development of cord blood storage could be beneficial for multicell coculture.
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Project description:In this work, we describe a new 3D printing methodology for the fabrication of multimaterial scaffolds involving the combination of thermoplastic extrusion and low temperature extrusion of bioinks. A fiber engraving technique was used to create a groove on the surface of a thermoplastic printed fiber using a commercial 3D printer and a low viscosity bioink was deposited into this groove. In contrast to traditional extrusion bioinks that rely on increased viscosity to prevent lateral spreading, this groove creates a defined space for bioink deposition. By physically constraining bioink spreading, a broader range of viscosities can be used. As proof-of-concept, we fabricated and characterized a multimaterial scaffold containing poly(ε-caprolactone) (PCL) as the thermoplastic polymer and a gelatin-based bioink. A 7.5 w/v% gelatin methacryloyl (GelMA) bioink loaded with either 5 w/v% poly(lactic-co-glycolic acid) (PLGA) microparticles containing fluorescent albumin or mouse fibroblasts (1 × 106 cell/mL) was printed at 24 °C. The structure of the composite scaffolds had no significant decrease in porosity or mechanical properties as compared to the PCL control scaffolds, demonstrating the engraving technique did not significantly compromise the mechanical or structural integrity of the scaffold. The encapsulated PLGA microparticles were homogeneously distributed in the GelMA and remained in the scaffolds after incubation in PBS for 24 h at 37 °C. In addition, the viability of the fibroblasts encapsulated in the GelMA bioink and printed in the grooves of the PCL scaffolds was confirmed after 24 h of incubation. Overall, this work provides a new methodology for the preparation of 3D printed scaffolds containing a robust thermoplastic structure in combination with low viscosity bioinks.
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