Click chemistry reactions have become an important tool for synthesizing user-defined hydrogels consisting of poly(ethylene glycol) (PEG) and bioactive peptides for tissue engineering. However, because click crosslinking proceeds via a step-growth mechanism, multi-arm telechelic precursors are required, which has some disadvantages. Here, we report for the first time that this requirement can be circumvented to create PEG–peptide hydrogels solely from linear precursors through the use of two orthogonal click reactions, the thiol–maleimide Michael addition and thiol–norbornene click reaction. The rapid kinetics of both click reactions allowed for quick formation of norbornene-functionalized PEG–peptide block copolymers via Michael addition, which were subsequently photocrosslinked into hydrogels with a dithiol linker. Characterization and in vitro testing demonstrated that the hydrogels have highly tunable physicochemical properties and excellent cytocompatibility. In addition, stoichiometric control over the crosslinking reaction can be leveraged to leave unreacted norbornene groups in the hydrogel for subsequent hydrogel functionalization via bioorthogonal inverse-electron demand Diels–Alder click reactions with s -tetrazines. After selectively capping norbornene groups in a user-defined region with cysteine, this feature was leveraged for protein patterning. Collectively, these results demonstrate that our novel chemical strategy is a simple and versatile approach to the development of hydrogels for tissue engineering that could be useful for a variety of applications.
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Time-dependent covalent network formation in extrudable hydrogels
The fabrication of hydrogel materials has gained increased attention for a broad range of biomedical and biotechnological applications. However, one longstanding challenge in the field is to develop hydrogels that can be easily processed into the desired form factor, while achieving the necessary final physical and biochemical properties. Herein, we report a shear-thinning hydrogel ink that can be photo-cured to create a stretchable, suturable hydrogel whose polymer network is formed via the combination of thiol-Michael addition and radical polymerization. A shear-thinning hydrogel based on bis-methacrylated Pluronic® F-127 was modified with varying equivalents of 2,2′-(ethylenedioxy)diethanethiol (EDT) as an additive. We observed that aging the hydrogel over time prior to extrusion allowed the relatively slow thiol-Michael addition to occur (between thiol and methacrylate) prior to UV initiated photopolymerization of the methacrylates. The viscoelastic properties of these hydrogels could be tuned based on the amount of EDT added, and the aging time of the hydrogel formulation. The changes to the physical properties of the hydrogels were attributed to the increased chain length between network junctions that resulted from the thiol-Michael addition reactions. The optimized hydrogel composition was then extruded from a coaxial nozzle to produce hydrogel tubes that, after curing, were resistant to tearing and were suturable. These extrudable synthetic hydrogels with tunable viscoelastic properties are promising for tissue engineering applications and as surgical training models for human vasculature.
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- Award ID(s):
- 1752972
- NSF-PAR ID:
- 10223082
- Date Published:
- Journal Name:
- Polymer Chemistry
- Volume:
- 11
- Issue:
- 43
- ISSN:
- 1759-9954
- Page Range / eLocation ID:
- 6910 to 6918
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Abstract 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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Abstract 3D‐printing is emerging as a technology to introduce microchannels into hydrogels, for the perfusion of engineered constructs. Although numerous techniques have been developed, new techniques are still needed to obtain the complex geometries of blood vessels and with materials that permit desired cellular responses. Here, a printing process where a shear‐thinning and self‐healing hydrogel “ink” is injected directly into a “support” hydrogel with similar properties is reported. The support hydrogel is further engineered to undergo stabilization through a thiol‐ene reaction, permitting (i) the washing of the ink to produce microchannels and (ii) tunable properties depending on the crosslinker design. When adhesive peptides are included in the support hydrogel, endothelial cells form confluent monolayers within the channels, across a range of printed configurations (e.g., straight, stenosis, spiral). When protease‐degradable crosslinkers are used for the support hydrogel and gradients of angiogenic factors are introduced, endothelial cells sprout into the support hydrogel in the direction of the gradient. This printing approach is used to investigate the influence of channel curvature on angiogenic sprouting and increased sprouting is observed at curved locations. Ultimately, this technique can be used for a range of biomedical applications, from engineering vascularized tissue constructs to modeling in vitro cultures.
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null (Ed.)Fibrotic disorders account for over one third of mortalities worldwide. Despite great efforts to study the cellular and molecular processes underlying fibrosis, there are currently few effective therapies. Dual-stage polymerization reactions are an innovative tool for recreating heterogeneous increases in extracellular matrix (ECM) modulus, a hallmark of fibrotic diseases in vivo . Here, we present a clickable decellularized ECM (dECM) crosslinker incorporated into a dynamically responsive poly(ethylene glycol)-α-methacrylate (PEGαMA) hybrid-hydrogel to recreate ECM remodeling in vitro . An off-stoichiometry thiol–ene Michael addition between PEGαMA (8-arm, 10 kg mol −1 ) and the clickable dECM resulted in hydrogels with an elastic modulus of E = 3.6 ± 0.24 kPa, approximating healthy lung tissue (1–5 kPa). Next, residual αMA groups were reacted via a photo-initiated homopolymerization to increase modulus values to fibrotic levels ( E = 13.4 ± 0.82 kPa) in situ . Hydrogels with increased elastic moduli, mimicking fibrotic ECM, induced a significant increase in the expression of myofibroblast transgenes. The proportion of primary fibroblasts from dual-reporter mouse lungs expressing collagen 1a1 and alpha-smooth muscle actin increased by approximately 60% when cultured on stiff and dynamically stiffened hybrid-hydrogels compared to soft. Likewise, fibroblasts expressed significantly increased levels of the collagen 1a1 transgene on stiff regions of spatially patterned hybrid-hydrogels compared to the soft areas. Collectively, these results indicate that hybrid-hydrogels are a new tool that can be implemented to spatiotemporally induce a phenotypic transition in primary murine fibroblasts in vitro .more » « less
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Abstract 3D printing involves the development of inks that exhibit the requisite properties for both printing and the intended application. In bioprinting, these inks are often hydrogels with controlled rheological properties that can be stabilized after deposition. Here, an alternate approach is developed where the ink is composed exclusively of jammed microgels, which are designed to incorporate a range of properties through microgel design (e.g., composition, size) and through the mixing of microgels. The jammed microgel inks are shear‐thinning to permit flow and rapidly recover upon deposition, including on surfaces or when deposited in 3D within hydrogel supports, and can be further stabilized with secondary cross‐linking. This platform allows the use of microgels engineered from various materials (e.g., thiol‐ene cross‐linked hyaluronic acid (HA), photo‐cross‐linked poly(ethylene glycol), thermo‐sensitive agarose) and that incorporate cells, where the jamming process and printing do not decrease cell viability. The versatility of this particle‐based approach opens up numerous potential biomedical applications through the printing of a more diverse set of inks.
- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1039/D0PY01129K
