Design of Bio-Conjugated Hydrogels for Regenerative Medicine Applications: From Polymer Scaffold to Biomolecule Choice

Abstract

Bio-conjugated hydrogels merge the functionality of a synthetic network with the activity of a biomolecule, becoming thus an interesting class of materials for a variety of biomedical applications. This combination allows the fine tuning of their functionality and activity, whilst retaining biocompatibility, responsivity and displaying tunable chemical and mechanical properties. A complex scenario of molecular factors and conditions have to be taken into account to ensure the correct functionality of the bio-hydrogel as a scaffold or a delivery system, including the polymer backbone and biomolecule choice, polymerization conditions, architecture and biocompatibility. In this review, we present these key factors and conditions that have to match together to ensure the correct functionality of the bio-conjugated hydrogel. We then present recent examples of bio-conjugated hydrogel systems paving the way for regenerative medicine applications.

Keywords: bio-conjugated hydrogels; biomolecule; polymer scaffold; regenerative medicine.

Scheme 1

Schematic presentation of bio-conjugated hydrogels based on the combination of biomolecules (DNA, proteins, and peptides) and different polymers for regenerative medicine.

Figure 1

Scanning electron microscope (SEM) micrographs of dried (a) PVA hydrogel and (b–d) macroporous PVA hydrogels treated with (b) 2 wt% (c) 4 wt% and (d) 6 wt% of agarose as pore formation agent [93].

Figure 2

Schematic representation of the polymerization procedure for synthesis of a biocompatible and biodegradable EPC copolymer [119].

Figure 3

Schematics presenting several functional groups involved in hydrogel formation [128] and attachment of biomolecules via covalent bonding.

Scheme 2

Schematic representation of the most common biomolecule immobilization approaches: (a) non-covalent interaction; (b) covalent attachment, and (c) physical entrapment.

Figure 4

(A) Designing microgels for loading and release of the model protein, cytochrome c (cyt-c), by changing the distribution of ionizable groups into the microgel structure: N-pure PNIPAM microgel; V-pure PVCL microgel; V⁻-anionic PVCL microgel; V^±-PVCL polyampholyte microgel with a random distribution of ionizable groups; V⁻N⁺-core−shell polyampholyte microgel consisting of an anionic PVCL core (black network) and a cationic PNIPAM shell (grey network); (B) Schematic representation of the protein loading and release into/from the polyampholyte microgels by reversibly changing the hydrogel charges from positive to negative using the pH switch from acidic to basic; (C) Effect of changing the pH value from 8 to 6, and respectively 3, on the amount of bound cyt-c for the polyampholyte microgels comprising of V10⁻N10⁺ (a) and V10⁻5N10⁺ (b); Variation of the amount of bound cyt-c vs. time at pH 3 for V10⁻N10⁺ and V10⁻5N10⁺ polyampholyte core-shell microgels in comparison to the V20⁻ polyelectrolyte microgels (c) [205].

Scheme 3

Schematics of DNA-functionalized hybrid hydrogels: (left) single-stranded DNA covalently attached to the polymer scaffold, and (right) plasmids or long DNA sequences conjugated by electrostatic interactions.

Figure 5

Design of hydrogel networks with mechanical properties similar to naturally occurring tissues: (a) The hydrogel network is formed from two mechanical responsive elements: cross-linkers (CLs) and load-bearing modules (LBMs). The binding/unbinding kinetics of the CL and the folding/unfolding of the LBM are regulated by the applied mechanical force; (b) Structure of the protein domains (LBMs) used in the hydrogel design to form the TIP-1:Kir complex: GB1, HP67, and SH3 [178]. The figure is reproduced from an open access article published by Nature under a Creative Commons Attribution 4.0 International License (https://www.nature.com/articles/s41467-018-02917-6#rightslink).

Figure 6

The bulk mechanical properties of the designed hydrogels: (a) General procedure for preparation of protein hydrogel rings for mechanical testing. All protein gels were cross-linked using TIP-1:Kir complexes. (GB1)8, (GB1-HP67)4 and SH3 were used as the LBMs and the corresponding hydrogels were denoted as Gel-1, Gel-2 and Gel-3, respectively; (b) Setup for the mechanical tests of ring-shaped hydrogel. Stress-strain curves for the Gel-1, Gel-2 and Gel-3 until their breakage (c–e). Representative stretching-relaxation curves for the Gel-1, Gel-2 and Gel-3 (f–h). The curves are horizontally offset for clarity. The final strains are shown on the curves. Insets show the superposition of the stretching-relaxation curves at different strains [178]. The figure is reproduced from an open access article published by Nature under a Creative Commons Attribution 4.0 International License (https://www.nature.com/articles/s41467-018-02917-6#rightslink).

Figure 7

Histology of a self-assembled peptide hydrogel with mMA encoded within after 3 days (A,C) and after 7 days (B,D). (C) and (D) show close to no cellular growth within the gel matrix [257].

Figure 8

(A) Multimethacrylated HA with m + n (m + n ≈ 10,000) repeat units (Mr ≈ 400 Da). In this study, ~1 in 5 disaccharide units was functionalized with a methacrylate group. (B) Dimethacrylated PEG chains with n repeat units (Mr ≈ 44, n ≈ 105) were end-capped with methacrylate groups. (C) Hydrogel networks formed by radical initiated chain copolymerization of HA with varying amounts of PEG. Primary radicals were produced by the dissociation of the photoinitiator, 1-[4-(2-hydroxyethoxy)-phenyl]-2-hydroxy-2-methyl-1-propanone, in the presence of UV light (l = 365 nm) [180].

Scheme 4

Schematic representation of peptide-functionalized hybrid hydrogels.

Figure 9

(A) Fluorescence image of hMSC cells stained with Calcein, entrapped within the non-stained gel matrix. (B) Cryo-SEM image of hMSC entrapped within the gel pores. (C) Scheme of hydrogel formation via the reaction between an 8-arm maleimide functional PEG and a collagen triple helical peptide [182].