Extrusion-Based 3D Bioprinting of Gradients of Stiffness, Cell Density, and Immobilized Peptide Using Thermogelling Hydrogels.
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ABSTRACT:
SUBMITTER: Kuzucu M
PROVIDER: S-EPMC8207502 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Gelatin-methacryloyl (GelMA) is a semi-synthetic hydrogel which consists of gelatin derivatized with methacrylamide and methacrylate groups. These hydrogels provide cells with an optimal biological environment (e.g., RGD motifs for adhesion) and can be quickly photo-crosslinked, which provides shape fidelity and stability at physiological temperature. In the present work, we demonstrated how GelMA hydrogels can be synthesized with a specific degree of functionalization (DoF) and adjusted to the intended application as a three-dimensional (3D) cell culture platform. The focus of this work lays on producing hydrogel scaffolds which provide a cell promoting microenvironment for human adipose tissue-derived mesenchymal stem cells (hAD-MSCs) and are conductive to their adhesion, spreading, and proliferation. The control of mechanical GelMA properties by variation of concentration, DoF, and ultraviolet (UV) polymerization conditions is described. Moreover, hAD-MSC cell viability and morphology in GelMA of different stiffness was evaluated and compared. Polymerized hydrogels with and without cells could be digested in order to release encapsulated cells without loss of viability. We also demonstrated how hydrogel viscosity can be increased by the use of biocompatible additives, in order to enable the extrusion bioprinting of these materials. Taken together, we demonstrated how GelMA hydrogels can be used as a versatile tool for 3D cell cultivation.
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Project description: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.
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Project description:Cellular models are needed to study human development and disease in vitro, and to screen drugs for toxicity and efficacy. Current approaches are limited in the engineering of functional tissue models with requisite cell densities and heterogeneity to appropriately model cell and tissue behaviors. Here, we develop a bioprinting approach to transfer spheroids into self-healing support hydrogels at high resolution, which enables their patterning and fusion into high-cell density microtissues of prescribed spatial organization. As an example application, we bioprint induced pluripotent stem cell-derived cardiac microtissue models with spatially controlled cardiomyocyte and fibroblast cell ratios to replicate the structural and functional features of scarred cardiac tissue that arise following myocardial infarction, including reduced contractility and irregular electrical activity. The bioprinted in vitro model is combined with functional readouts to probe how various pro-regenerative microRNA treatment regimes influence tissue regeneration and recovery of function as a result of cardiomyocyte proliferation. This method is useful for a range of biomedical applications, including the development of precision models to mimic diseases and the screening of drugs, particularly where high cell densities and heterogeneity are important.
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Project description:Breast cancer cell invasion is influenced by growth factor concentration gradients in the tumor microenvironment. However, studying the influence of growth factor gradients on breast cancer cell invasion is challenging due to both the complexities of in vivo models and the difficulties in recapitulating the tumor microenvironment with defined gradients using in vitro models. A defined hyaluronic acid (HA)-based hydrogel crosslinked with matrix metalloproteinase (MMP) cleavable peptides and modified with multiphoton labile nitrodibenzofuran (NDBF) was synthesized to photochemically immobilize epidermal growth factor (EGF) gradients. We demonstrate that EGF gradients can differentially influence breast cancer cell invasion and drug response in cell lines with different EGF receptor (EGFR) expression levels. Photopatterned EGF gradients increase the invasion of moderate EGFR expressing MDA-MB-231?cells, reduce invasion of high EGFR expressing MDA-MB-468?cells, and have no effect on invasion of low EGFR-expressing MCF-7?cells. We evaluate MDA-MB-231 and MDA-MB-468?cell response to the clinically tested EGFR inhibitor, cetuximab. Interestingly, the cellular response to cetuximab is completely different on the EGF gradient hydrogels: cetuximab decreases MDA-MB-231?cell invasion but increases MDA-MB-468?cell invasion and cell number, thus demonstrating the importance of including cell-microenvironment interactions when evaluating drug targets.
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Project description:The fabrication of three-dimensional (3D) scaffolds is indispensable to tissue engineering and 3D printing is emerging as an important approach towards this. Hydrogels are often used as inks in extrusion-based 3D printing, including with encapsulated cells; however, numerous challenging requirements exist, including appropriate viscosity, the ability to stabilize after extrusion, and cytocompatibility. Here, we present a shear-thinning and self-healing hydrogel crosslinked through dynamic covalent chemistry for 3D bioprinting. Specifically, hyaluronic acid was modified with either hydrazide or aldehyde groups and mixed to form hydrogels containing a dynamic hydrazone bond. Due to their shear-thinning and self-healing properties, the hydrogels could be extruded for 3D printing of structures with high shape fidelity, stability to relaxation, and cytocompatibility with encapsulated fibroblasts (>80% viability). Forces for extrusion and filament sizes were dependent on parameters such as material concentration and needle gauge. To increase scaffold functionality, a second photocrosslinkable interpenetrating network was included that was used for orthogonal photostiffening and photopatterning through a thiol-ene reaction. Photostiffening increased the scaffold's modulus (?300%) while significantly decreasing erosion (?70%), whereas photopatterning allowed for spatial modification of scaffolds with dyes. Overall, this work introduces a simple approach to both fabricate and modify 3D printed scaffolds. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 865-875, 2018.
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Project description:Purpose: Bioprinting is becoming an increasingly popular platform technology for engineering a variety of tissue types. Our aim was to identify biomaterials that have been found to be suitable for extrusion 3D bioprinting, outline their biomechanical properties and biocompatibility towards their application for bioprinting specific tissue types. This systematic review provides an in-depth overview of current biomaterials suitable for extrusion to aid bioink selection for specific research purposes and facilitate design of novel tailored bioinks. Methods: A systematic search was performed on EMBASE, PubMed, Scopus and Web of Science databases according to the PRISMA guidelines. References of relevant articles, between December 2006 to January 2018, on candidate bioinks used in extrusion 3D bioprinting were reviewed by two independent investigators against standardised inclusion and exclusion criteria. Data was extracted on bioprinter brand and model, printing technique and specifications (speed and resolution), bioink material and class of mechanical assessment, cell type, viability, and target tissue. Also noted were authors, study design (in vitro/in vivo), study duration and year of publication. Results: A total of 9,720 studies were identified, 123 of which met inclusion criteria, consisting of a total of 58 reports using natural biomaterials, 26 using synthetic biomaterials and 39 using a combination of biomaterials as bioinks. Alginate (n = 50) and PCL (n = 33) were the most commonly used bioinks, followed by gelatin (n = 18) and methacrylated gelatin (GelMA) (n = 16). Pneumatic extrusion bioprinting techniques were the most common (n = 78), followed by piston (n = 28). The majority of studies focus on the target tissue, most commonly bone and cartilage, and investigate only one bioink rather than assessing a range to identify those with the most promising printability and biocompatibility characteristics. The Bioscaffolder (GeSiM, Germany), 3D Discovery (regenHU, Switzerland), and Bioplotter (EnvisionTEC, Germany) were the most commonly used commercial bioprinters (n = 35 in total), but groups most often opted to create their own in-house devices (n = 20). Many studies also failed to specify whether the mechanical data reflected pre-, during or post-printing, pre- or post-crosslinking and with or without cells. Conclusions: Despite the continued increase in the variety of biocompatible synthetic materials available, there has been a shift change towards using natural rather than synthetic bioinks for extrusion bioprinting, dominated by alginate either alone or in combination with other biomaterials. On qualitative analysis, no link was demonstrated between the type of bioink or extrusion technique and the target tissue, indicating that bioprinting research is in its infancy with no established tissue specific bioinks or bioprinting techniques. Further research is needed on side-by-side characterisation of bioinks with standardisation of the type and timing of biomechanical assessment.
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Project description:Three-dimensional (3D) bioprinting has become a promising strategy for bone manufacturing, with excellent control over geometry and microarchitectures of the scaffolds. The bioprinting ink for bone and cartilage engineering has thus become the key to developing 3D constructs for bone and cartilage defect repair. Maintaining the balance of cellular viability, drugs or cytokines' function, and mechanical integrity is critical for constructing 3D bone and/or cartilage scaffolds. Photo-crosslinkable hydrogel is one of the most promising materials in tissue engineering; it can respond to light and induce structural or morphological transition. The biocompatibility, easy fabrication, as well as controllable mechanical and degradation properties of photo-crosslinkable hydrogel can meet various requirements of the bone and cartilage scaffolds, which enable it to serve as an effective bio-ink for 3D bioprinting. Here, in this review, we first introduce commonly used photo-crosslinkable hydrogel materials and additives (such as nanomaterials, functional cells, and drugs/cytokine), and then discuss the applications of the 3D bioprinted photo-crosslinkable hydrogel scaffolds for bone and cartilage engineering. Finally, we conclude the review with future perspectives about the development of 3D bioprinting photo-crosslinkable hydrogels in bone and cartilage engineering.
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Project description:Bioinks with shear-thinning/rapid solidification properties and strong mechanics are usually needed for the bioprinting of three-dimensional (3D) cell-laden constructs. As such, it remains challenging to generate soft constructs from bioinks at low concentrations that are favorable for cellular activities. Herein, we report a strategy to fabricate cell-laden constructs with tunable 3D microenvironments achieved by bioprinting of gelatin methacryloyl (GelMA)/alginate core/sheath microfibers, where the alginate sheath serves as a template to support and confine the GelMA pre-hydrogel in the core during the extrusion process, allowing for subsequent UV crosslinking. This novel strategy minimizes the bioprinting requirements for the core bioink, and facilitates the fabrication of cell-laden GelMA constructs at low concentrations. We first showed the capability of generating various alginate hollow microfibrous constructs using a coaxial nozzle setup, and verified the diffusibility and perfusability of the bioprinted hollow structures that are important for the tissue engineering applications. More importantly, the hollow alginate microfibers were then used as templates for generating cell-laden GelMA constructs with soft microenvironments, by using GelMA pre-hydrogel as the bioink for the core phase during bioprinting. As such, GelMA constructs at extremely low concentrations (<2.0%) could be extruded to effectively support cellular activities including proliferation and spreading for various cell types. We believe that our strategy is likely to provide broad opportunities in bioprinting of 3D constructs with cell-favorable microenvironments for applications in tissue engineering and pharmaceutical screening.
| S-EPMC5837947 | biostudies-literature
Project description: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, while VIC expressed elevated vimentin. These results demonstrate that anatomically complex, heterogeneously encapsulated aortic valve hydrogel conduits can be fabricated with 3D bioprinting.
| S-EPMC3694360 | biostudies-literature