Extracellular Matrix Mimetic Hydrogels Research Articles

Extracellular Matrix-Mimetic Hydrogels for Treating Neural Tissue Injury: A Focus on Fibrin, Hyaluronic Acid, and Elastin-Like Polypeptide Hydrogels.

Neurological and functional recovery is limited following central nervous system injury and severe injury to the peripheral nervous system. Extracellular matrix (ECM)-mimetic hydrogels are of particular interest as regenerative scaffolds for the injured nervous system as they provide 3D bioactive interfaces that modulate cellular response to the injury environment and provide naturally degradable scaffolding for effective tissue remodeling. In this review, three unique ECM-mimetic hydrogels used in models of neural injury are reviewed: fibrin hydrogels, which rely on a naturally occurring enzymatic gelation, hyaluronic acid hydrogels, which require chemical modification prior to chemical crosslinking, and elastin-like polypeptide (ELP) hydrogels, which exhibit a temperature-sensitive gelation. The hydrogels are reviewed by summarizing their unique biological properties, their use as drug depots, and their combination with other biomaterials, such as electrospun fibers and nanoparticles. This review is the first to focus on these three ECM-mimetic hydrogels for their use in neural tissue engineering. Additionally, this is the first review to summarize the use of ELP hydrogels for nervous system applications. ECM-mimetic hydrogels have shown great promise in preclinical models of neural injury and future advancements in their design and use can likely lead to viable treatments for patients with neural injury.

An Optogenetic Platform to Dynamically Control the Stiffness of Collagen Hydrogels.

The extracellular matrix (ECM) comprises a meshwork of biomacromolecules whose composition, architecture, and macroscopic properties, such as mechanics, instruct cell fate decisions during development and disease progression. Current methods implemented in mechanotransduction studies either fail to capture real-time mechanical dynamics or utilize synthetic polymers that lack the fibrillar nature of their natural counterparts. Here we present an optogenetic-inspired tool to construct light-responsive ECM mimetic hydrogels comprised exclusively of natural ECM proteins. Optogenetic tools offer seconds temporal resolution and submicron spatial resolution, permitting researchers to probe cell signaling dynamics with unprecedented precision. Here we demonstrated our approach of using SNAP-tag and its thiol-targeted substrate, benzylguanine-maleimide, to covalently attach blue-light-responsive proteins to collagen hydrogels. The resulting material (OptoGel), in addition to encompassing the native biological activity of collagen, stiffens upon exposure to blue light and softens in the dark. Optogels have immediate use in dissecting the cellular response to acute mechanical inputs and may also have applications in next-generation biointerfacing prosthetics.

Fabrication of modular hyaluronan-PEG hydrogels to support 3D cultures of hepatocytes in a perfused liver-on-a-chip device

Liver cell culture models are attractive in both tissue engineering and for development of assays for drug toxicology research. To retain liver specific cell functions, the use of adequate cell types and culture conditions, such as a 3D orientation of the cells and a proper supply of nutrients and oxygen, are critical. In this article, we show how extracellular matrix mimetic hydrogels can support hepatocyte viability and functionality in a perfused liver-on-a-chip device. A modular hydrogel system based on hyaluronan and poly(ethylene glycol) (HA-PEG), modified with cyclooctyne moieties for bioorthogonal strain-promoted alkyne-azide 1, 3-dipolar cycloaddition (SPAAC), was developed, characterized, and compared for cell compatibility to hydrogels based on agarose and alginate. Hepatoma cells (HepG2) formed spheroids with viable cells in all hydrogels with the highest expression of albumin and urea in alginate hydrogels. By including an excess of cyclooctyne in the HA backbone, azide-modified cell adhesion motifs (linear and cyclic RGD peptides) could be introduced in order to enhance viability and functionality of human induced pluripotent stem cell derived hepatocytes (hiPS-HEPs). In the HA-PEG hydrogels modified with cyclic RGD peptides hiPS-HEPs migrated and grew in 3D and showed an increased viability and higher albumin production compared to when cultured in the other hydrogels. This flexible SPAAC crosslinked hydrogel system enabled fabrication of perfused 3D cell culture of hiPS-HEPs and is a promising material for further development and optimization of liver-on-a-chip devices.

Mechanical Properties, Cytocompatibility and Manufacturability of Chitosan:PEGDA Hybrid-Gel Scaffolds by Stereolithography.

Extracellular matrix mimetic hydrogels which hybridize synthetic and natural polymers offer molecularly-tailored, bioactive properties and tunable mechanical strength. In addition, 3D bioprinting by stereolithography allows fabrication of internal pores and defined macroscopic shapes. In this study, we formulated a hybrid biocompatible resin using natural and synthetic polymers (chitosan and polyethylene glycol diacrylate (PEGDA), respectively) by controlling molecular weight of chitosan, feed-ratios, and photo-initiator concentration. Ear-shaped, hybrid scaffolds were fabricated by a stereolithographic method using a 405nm laser. Hybrid hydrogel scaffolds of chitosan (50-190kDa) and PEGDA (575Da) were mixed at varying feed-ratios. Some of the cationic, amino groups of chitosan were neutralized by dialysis in acidic solution containing chitosan in excess of sodium acetate solution to inhibit quenching of newly formed photoradicals. A feed-ratio of 1:7.5 was found to be the most appropriate of the formulations considered in this study in terms of mechanical properties, cell adhesion, and printability. The biofabricated hybrid scaffold showed interconnected, homogeneous pores with a nominal pore size of 50µm and an elastic modulus of~400kPa. Moreover, long-term cell viability and cell spreading was observed via actin filament staining. Printability of the biocompatible resin was confirmed by printing thresholded MR images of an ear and the feed ratio of 1:7.5 provided the most faithful reproduction of the shape. To the best of our knowledge, this is the first report of stereolithographic printing hybridizing cell-adhesive properties of chitosan with mechanical robustness of PEG in scaffolds suitable for repair of complex tissue geometries, such as those of the human ear.

Tuning the Microenvironment: Click‐Crosslinked Hyaluronic Acid‐Based Hydrogels Provide a Platform for Studying Breast Cancer Cell Invasion

A big challenge in cell culture is the non‐natural environment in which cells are routinely screened, making in vivo phenomena, such as cell invasion, difficult to understand and predict. To study cancer cell invasion, extracellular matrix (ECM) analogs with decoupled mechanical and chemical properties are required. Hyaluronic acid (HA)‐based hydrogels crosslinked with matrix‐metalloproteinase (MMP)‐cleavable peptides are developed to study MDA‐MB‐231 breast cancer cell invasion. Hydrogels are synthesized by reacting furan‐modified HA with bismaleimide peptide crosslinkers in a Diels–Alder click reaction. This new hydrogel takes advantage of the biomimetic properties of HA, which is overexpressed in breast cancer, and eliminates the use of nonadhesive crosslinkers, such as poly(ethylene glycol) (PEG). The crosslink (mechanical) and ligand (chemical) densities are varied independently to evaluate the effects of each parameter on cell migration. Increased crosslink density correlates with decreased MDA‐MB‐231 cell invasion whereas incorporation of MMP‐cleavable sequences within the peptide crosslinker enhances invasion. Increasing the ligand density of pendant GRGDS groups induces cell proliferation, but has no significant impact on invasion. By independently tuning the mechanical and chemical environment of ECM mimetic hydrogels, a platform is provided that recapitulates variable tissue properties and elucidates the role of the microenvironment in cancer cell invasion.

Towards constructing extracellular matrix-mimetic hydrogels: An elastic hydrogel constructed from tandem modular proteins containing tenascin FnIII domains

Protein-based hydrogels have been developed for various biomedical applications where they provide artificial extracellular microenvironments that mimic the physical and biochemical characteristics of natural extracellular matrices (ECMs). In natural ECMs, a large number of proteins are tandem modular proteins consisting of many individually folded functional domains that confer structural and biological functionalities. Such tandem modular proteins are promising building blocks for constructing ECM-mimetic biomaterials. However, their use for such purposes has not been explored extensively. Tenascin-C (TNC) is an ECM tandem modular protein and plays an important role in mechanotransduction by regulating important cell–matrix interactions. The third FnIII domain of TNC (TNfn3) contains an RGD sequence and is known to bind integrins. Here we use the TNfn3 domain and resilin sequence-based tandem modular protein FRF4RF4R (F represents the TNfn3 domain and R represents the resilin sequence, respectively) as a building block to construct protein-based ECM-mimetic hydrogels. The tandem modular protein-based building block FRF4RF4R closely mimics the architecture of the naturally occurring tandem modular ECM protein TNC and incorporates intact RGD-containing FnIII domains. Our results demonstrate that tandem modular proteins containing TNfn3 can be readily photochemically crosslinked into elastic hydrogels, whose Young’s modulus can be tuned by the concentration of the tandem modular protein solution. In vitro studies demonstrate that none of the photochemical crosslinking reaction components are cytotoxic at the level tested, and the hydrogel supports the spread of human lung fibroblast cells. Our results demonstrate that FRF4RF4R-based hydrogel is a novel ECM-mimetic hydrogel.

Tandem Modular Protein-Based Hydrogels Constructed Using a Novel Two-Component Approach

Leucine zipper sequences have been widely used to engineer protein-based hydrogels for biomedical applications. Previously, we have used this method to engineer tandem modular protein-based hydrogels as a step toward developing extracellular matrix-mimetic hydrogels. However, the spontaneous self-association of leucine zipper sequences in solution has made it challenging to express and purify tandem modular proteins carrying leucine zipper under native conditions. To obviate this problem, here we report a novel two-component approach to engineer tandem modular protein-based hydrogels. This methodology makes use of two complementary leucine zipper sequences (CCE and CCK), which do not self-associate but self-assemble into heterodimeric coiled-coils at neutral pH, as functional groups to drive the self-assembly of protein hydrogels. The two protein components are bifunctional and trifunctional tandem modular proteins carrying the leucine zipper functional groups. We found that the two proteins carrying CCE or CCK can be expressed and purified under native conditions with high yield. Upon mixing, the aqueous solution of the two proteins readily forms a transparent hydrogel. The resultant hydrogel can undergo reversible sol-gel transitions as a function of temperature, and shows much improved erosion properties. This method provides a new approach to tune the topology and physical properties of the protein hydrogels via genetic engineering, and opens the possibility to systematically explore the use of large native extracellular proteins to engineer extracellular matrix-mimetic hydrogels.