3D printed gelatin/decellularized bone composite scaffolds for bone tissue engineering: Fabrication, characterization and cytocompatibility study.
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SUBMITTER: Kara A
PROVIDER: S-EPMC9213825 | biostudies-literature |
REPOSITORIES: biostudies-literature
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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: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.
| S-EPMC9640992 | biostudies-literature
Project description:Biofabrication has been adapted in engineering patient-specific biosynthetic grafts for bone regeneration. Herein, we developed a three-dimensional (3D) high-resolution, room-temperature printing approach to fabricate osteoconductive scaffolds using calcium phosphate cement (CPC). The non-aqueous CPC bioinks were composed of tetracalcium phosphate, dicalcium phosphate anhydrous, and Polyvinyl butyral (PVB) dissolved in either ethanol (EtOH) or tetrahydrofuran (THF). They were printed in an aqueous sodium phosphate bath, which performs as a hardening accelerator for hydroxyapatite formation and as a retainer for 3D microstructure. The PVB solvents, EtOH or THF, affected differently the slurry rheological properties, scaffold microstructure, mechanical properties, and osteoconductivity. Our proposed approach overcomes limitations of conventional fabrication methods, which require high-temperature (>50 °C), low-resolution (>400 μm) printing with an inadequate amount of large ceramic particles (>35 μm). This proof-of-concept study opens venues in engineering high-resolution, implantable, and osteoconductive scaffolds with predetermined properties for bone regeneration.
| S-EPMC8164647 | biostudies-literature
Project description:The use of porous three-dimensional (3D) composite scaffolds has attracted great attention in bone tissue engineering applications because they closely simulate the major features of the natural extracellular matrix (ECM) of bone. This study aimed to prepare biomimetic composite scaffolds via a simple 3D printing of gelatin/hyaluronic acid (HA)/hydroxyapatite (HAp) and subsequent biomineralization for improved bone tissue regeneration. The resulting scaffolds exhibited uniform structure and homogeneous pore distribution. In addition, the microstructures of the composite scaffolds showed an ECM-mimetic structure with a wrinkled internal surface and a porous hierarchical architecture. The results of bioactivity assays proved that the morphological characteristics and biomineralization of the composite scaffolds influenced cell proliferation and osteogenic differentiation. In particular, the biomineralized gelatin/HA/HAp composite scaffolds with double-layer staggered orthogonal (GEHA20-ZZS) and double-layer alternative structure (GEHA20-45S) showed higher bioactivity than other scaffolds. According to these results, biomineralization has a great influence on the biological activity of cells. Hence, the biomineralized composite scaffolds can be used as new bone scaffolds in bone regeneration.
| S-EPMC8267715 | biostudies-literature
Project description:Recent developments in 3D printing (3DP) research have led to a variety of scaffold designs and techniques for osteochondral tissue engineering; however, the simultaneous incorporation of multiple types of gradients within the same construct remains a challenge. Herein, we describe the fabrication and mechanical characterization of porous poly(?-caprolactone) (PCL) and PCL-hydroxyapatite (HA) scaffolds with incorporated vertical porosity and ceramic content gradients via a multimaterial extrusion 3DP system. Scaffolds of 0?wt% HA (PCL), 15?wt% HA (HA15), or 30?wt% HA (HA30) were fabricated with uniform composition and porosity (using 0.2?mm, 0.5?mm, or 0.9?mm on-center fiber spacing), uniform composition and gradient porosity, and gradient composition (PCL-HA15-HA30) and porosity. Micro-CT imaging and porosity analysis demonstrated the ability to incorporate both vertical porosity and pore size gradients and a ceramic gradient, which collectively recapitulate gradients found in native osteochondral tissues. Uniaxial compression testing demonstrated an inverse relationship between porosity, ?, and compressive modulus, E, and yield stress, ?y, for uniform porosity scaffolds, however, no differences were observed as a result of ceramic incorporation. All scaffolds demonstrated compressive moduli within the appropriate range for trabecular bone, with average moduli between 86?±?14-220?±?26?MPa. Uniform porosity and pore size scaffolds for all ceramic levels had compressive moduli between 205?±?37-220?±?26?MPa, 112?±?13-118?±?23?MPa, and 86?±?14-97?±?8?MPa respectively for porosities ranging between 14?±?4-20?±?6%, 36?±?3-43?±?4%, and 54?±?2-57?±?2%, with the moduli and yield stresses of low porosity scaffolds being significantly greater (p?<?0.05) than those of all other groups. Single (porosity) gradient and dual (composition/porosity) gradient scaffolds demonstrated compressive properties similar (p?>?0.05) to those of the highest porosity uniform scaffolds (porosity gradient scaffolds 98?±?23-107?±?6?MPa, and 102?±?7?MPa for dual composition/porosity gradient scaffolds), indicating that these properties are more heavily influenced by the weakest section of the gradient. The compression data for uniform scaffolds were also readily modeled, yielding scaling laws of the form E???(1?-??)1.27 and ?y???(1?-??)1.37, which demonstrated that the compressive properties evaluated in this study were well-aligned with expectations from previous literature and were readily modeled with good fidelity independent of polymer scaffold geometry and ceramic content. All uniform scaffolds were similarly deformed and recovered despite different porosities, while the large-pore sections of porosity gradient scaffolds were significantly more deformed than all other groups, indicating that porosity may not be an independent factor in determining strain recovery. Moving forward, the technique described here will serve as the template for more complex multimaterial constructs with bioactive cues that better match native tissue physiology and promote tissue regeneration. STATEMENT OF SIGNIFICANCE: This manuscript describes the fabrication and mechanical characterization of "dual" porosity/ceramic content gradient scaffolds produced via a multimaterial extrusion 3D printing system for osteochondral tissue engineering. Such scaffolds are designed to better address the simultaneous gradients in architecture and mineralization found in native osteochondral tissue. The results of this study demonstrate that this technique may serve as a template for future advances in 3D printing technology that may better address the inherent complexity in such heterogeneous tissues.
| S-EPMC6744258 | biostudies-literature
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.
| S-EPMC8588077 | biostudies-literature
Project description:Functional reconstruction of craniomaxillofacial defects is challenging, especially for the patients who suffer from traumatic injury, cranioplasty, and oncologic surgery. Three-dimensional (3D) printing/bioprinting technologies provide a promising tool to fabricate bone tissue engineering constructs with complex architectures and bioactive components. In this study, we implemented multi-material 3D printing to fabricate 3D printed PCL/hydrogel composite scaffolds loaded with dual bioactive small molecules (i.e. resveratrol and strontium ranelate). The incorporated small molecules are expected to target several types of bone cells. We systematically studied the scaffold morphologies and small molecule release profiles. We then investigated the effects of the released small molecules from the drug loaded scaffolds on the behavior and differentiation of mesenchymal stem cells (MSCs), monocyte-derived osteoclasts, and endothelial cells. The 3D printed scaffolds, with and without small molecules, were further implanted into a rat model with a critical-sized mandibular bone defect. We found that the bone scaffolds containing the dual small molecules had combinational advantages in enhancing angiogenesis and inhibiting osteoclast activities, and they synergistically promoted MSC osteogenic differentiation. The dual drug loaded scaffolds also significantly promoted in vivo mandibular bone formation after 8 week implantation. This work presents a 3D printing strategy to fabricate engineered bone constructs, which can likely be used as off-the-shelf products to promote craniomaxillofacial regeneration.
| S-EPMC8059098 | biostudies-literature
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:In the field of bone tissue, maintaining adequate mechanical strength and tissue volume is an important part. Recently, biphasic calcium phosphate (BCP) was fabricated to solve the shortcomings of hydroxyapatite (HA) and beta-tricalcium phosphate (β-TCP), and it is widely studied in the field of bone-tissue engineering. In this study, a composite hydrogel was fabricated by applying BCP to gelatin methacrylate (GelMA). It was tested by using a mechanical tester, to characterize the mechanical properties of the prepared composite hydrogel. The fabricated BCP was analyzed through FTIR and XRD. As a result, a different characteristic pattern from hydroxyapatite (HA) and beta-tricalcium phosphate (β-TCP) was observed, and it was confirmed that it was successfully bound to the hydrogel. Then, the proliferation and differentiation of preosteoblasts were checked to evaluate cell viability. The analysis results showed high cell viability and relatively high bone differentiation ability in the composite hydrogel to which BCP was applied. These features have been shown to be beneficial for bone regeneration by maintaining the volume and shape of the hydrogel. In addition, hydrogels can be advantageous for clinical use, as they can shape the structure of the material for custom applications.
| S-EPMC7999599 | biostudies-literature
Project description:It is common knowledge that pure alginate hydrogel is more likely to have weak mechanical strength, a lack of cell recognition sites, extensive swelling and uncontrolled degradation, and thus be unable to satisfy the demands of the ideal scaffold. To address these problems, we attempted to fabricate alginate/bacterial cellulose nanocrystals-chitosan-gelatin (Alg/BCNs-CS-GT) composite scaffolds using the combined method involving the incorporation of BCNs in the alginate matrix, internal gelation through the hydroxyapatite-d-glucono-δ-lactone (HAP-GDL) complex, and layer-by-layer (LBL) electrostatic assembly of polyelectrolytes. Meanwhile, the effect of various contents of BCNs on the scaffold morphology, porosity, mechanical properties, and swelling and degradation behavior was investigated. The experimental results showed that the fabricated Alg/BCNs-CS-GT composite scaffolds exhibited regular 3D morphologies and well-developed pore structures. With the increase in BCNs content, the pore size of Alg/BCNs-CS-GT composite scaffolds was gradually reduced from 200 μm to 70 μm. Furthermore, BCNs were fully embedded in the alginate matrix through the intermolecular hydrogen bond with alginate. Moreover, the addition of BCNs could effectively control the swelling and biodegradation of the Alg/BCNs-CS-GT composite scaffolds. Furthermore, the in vitro cytotoxicity studies indicated that the porous fiber network of BCNs could fully mimic the extracellular matrix structure, which promoted the adhesion and spreading of MG63 cells and MC3T3-E1 cells on the Alg/BCNs-CS-GT composite scaffolds. In addition, these cells could grow in the 3D-porous structure of composite scaffolds, which exhibited good proliferative viability. Based on the effect of BCNs on the cytocompatibility of composite scaffolds, the optimum BCNs content for the Alg/BCNs-CS-GT composite scaffolds was 0.2% (w/v). On the basis of good merits, such as regular 3D morphology, well-developed pore structure, controlled swelling and biodegradation behavior, and good cytocompatibility, the Alg/BCNs-CS-GT composite scaffolds may exhibit great potential as the ideal scaffold in the bone tissue engineering field.
| S-EPMC8400783 | biostudies-literature