Dynamic peptide-folding mediated biofunctionalization and modulation of hydrogels for 4D bioprinting

Abstract

Hydrogels are used in a wide range of biomedical applications, including three-dimensional (3D) cell culture, cell therapy and bioprinting. To enable processing using advanced additive fabrication techniques and to mimic the dynamic nature of the extracellular matrix (ECM), the properties of the hydrogels must be possible to tailor and change over time with high precision. The design of hydrogels that are both structurally and functionally dynamic, while providing necessary mechanical support is challenging using conventional synthesis techniques. Here, we show a modular and 3D printable hydrogel system that combines a robust but tunable covalent bioorthogonal cross-linking strategy with specific peptide-folding mediated interactions for dynamic modulation of cross-linking and functionalization. The hyaluronan-based hydrogels were covalently cross-linked by strain-promoted alkyne-azide cycloaddition using multi-arm poly(ethylene glycol). In addition, a de novo designed helix-loop-helix peptide was conjugated to the hyaluronan backbone to enable specific peptide-folding modulation of cross-linking density and kinetics, and hydrogel functionality. An array of complementary peptides with different functionalities was developed and used as a toolbox for supramolecular tuning of cell-hydrogel interactions and for controlling enzyme-mediated biomineralization processes. The modular peptide system enabled dynamic modifications of the properties of 3D printed structures, demonstrating a novel route for design of more sophisticated bioinks for four-dimensional bioprinting.

Bioorthogonally Cross-Linked Hyaluronan-Laminin Hydrogels for 3D Neuronal Cell Culture and Biofabrication

“…Laminin labeling and formation of hydrogel: Hyaluronan-poly(ethylene glycol) (HA:PEG) hybrid hydrogels were prepared by combining bicyclo [6.1.0]nonyne (BCN) modified HA (~100 kDa) and an 8-arm PEG with terminal azides ((PEG-Az)8) as previously described. [50,51] LN was modified with azide (Az) moieties using linkers of different lengths (LN-Az and LNp-AZ) and was conjugated to HA-BCN, after which (PEG-Az)8 was added to form the final hydrogel at 37°C. The hydrogels were analyzed by rheology and scanning electron microscopy (SEM).…”

3D Bioprinting and Translation of Beta Cell Replacement Therapies for Type 1 Diabetes

“…Aspects of 3D bioprinting, which will need to be addressed, include the need for incorporating vascular beds for perfusion of large constructs to ensure nutrient delivery and cell viability, faster printing capability at high resolution, islet/beta cell-compatible bioinks, and defined universal media to ensure functional potency of heterogenous constructs, among various other recommendations. 135,167 Looking ahead, exploring new frontiers such as 4D bioprinting, [168][169][170] where stimuli-responsive biomaterials and/or cells can be used to include conformational or biochemical changes in printed structures over time in a predetermined manner, presents the possibility of constructing dynamic, complex structures.…”

Natural Origin Biomaterials for 4D Bioprinting Tissue‐Like Constructs

“…[16] This strategy can also be applied to achieve other desirable properties, [16] such as solubility in water, [54] control over the degradation rate, [55] and enhanced mechanical properties. [56] Gathering on this, some of the most explored natural polymers for 4D bioprinting including alginate, [33,57] collagen, [58] gelatin, [34,59,60] hyaluronic acid, [18,33] and chitosan [61] are depicted in Figure 1. Within 3D bioprinting applications, alginate hydrogel bioinks are undoubtedly one of the most broadly researched natural biomaterials.…”

“…[2] Moreover, cellular spatial distribution, either in single or multicellular 3D aggregates, has shown to considerably influence numerous intra-/intercellular processes occurring in constructs during in vitro maturation or upon in vivo implantation, and must also be considered in the design stages. [2,18] Fortunately, both 3D and 4D bioprinting techniques allow a suitable control over this parameter, thus granting the possibility of producing heterogeneous scaffolds, comprised of several biomaterials and cell types, with a more controlled spatial arrangement for each final application when compared to other available technologies [19] (e.g., electrospinning, solution casting, particulate leaching, and micromolding, in which cell distribution is generally random [20,21] ). Additionally, it is extremely important for future engineered biomimetic scaffolds to be able to grasp the intrinsic dynamism of the supporting extracellular matrix (ECM), moving away from the traditional 3D paradigm of bioprinted constructs as inanimate structures, and toward a timespanned 4D approach.…”