Mussel-inspired cross-linking mechanisms enhance gelation and adhesion of multifunctional mucin-derived hydrogels

Abstract

Mucus supports human health by hydrating, lubricating, and preventing infection of wet epithelial surfaces. The beneficial material properties and bioactivity of mucus stem from glycoproteins called mucins, motivating the development of mucin-derived hydrogels for wound dressings and antifouling coatings. However, these applications require robust gelation and adhesion to a wide range of substrates. Inspired by the chemical cross-linking and water-tolerant adhesion of marine mussel adhesive structures, we use catechol-thiol bonding to drive gelation of native mucin proteins and synthetic mucin-inspired polymers, forming soft, adhesive hydrogels that can be coated onto diverse surfaces. The gelation dynamics and adhesive properties can be systematically tuned by varying the hydrogel composition, polymer architecture, and thiol availability, with gelation timescales adjustable from seconds to hours, and values of elastic modulus, failure stress, and debonding work spanning orders of magnitude. We demonstrate the functionality of these gels in two applications: as tissue adhesives, using porcine skin as a proxy for human skin, and as bioactive surface coatings to prevent bacterial colonization. The results highlight the potential of catechol-thiol cross-linking as a versatile platform for engineering multifunctional glycoprotein hydrogels with applications in wound repair and antimicrobial surface engineering.

Keywords: antifouling; biomaterials; catechol-thiol bond; mucus; tissue adhesive.

Fig. 1.

Schematic overview of the polymers investigated in this work. (A) Thiol-functionalized polymers and (B) catechol-functionalized cross-linkers. The structures shown are ideal; the average functionalization of thiol or catechol groups per molecule is stated in the image labels.

Fig. 2.

Gelation dynamics of catechol- and thiol-functionalized polymers. Evolution of the storage modulus G′ vs time t at frequency ω = 5 rad/s for (A) 1% PEG-8C, 0 to 1% PEG-4SH, 0.16 mg/mL tyrosinase and (B) 4% hPG-C, 0 to 2% PEG-4SH, 0.64 mg/mL tyrosinase. The dashed lines show the minimum measurable moduli for different parallel plate test fixtures (diameter d = 8 or 25 mm). The gelation timescale t_gel, fastest mutation timescale λ*, storage modulus G_λ=3h, and final storage modulus G_f are indicated. (C) Average values of t_gel, λ*, and G_λ=3h as a function of the thiol:catechol ratio (N ≥ 3); error bars show the SD.

Fig. 3.

Adhesion of hydrogels composed of catechol- and thiol-functionalized polymers measured after the gelation shown in Fig. 2. (A) Evolution of the tensile engineering stress σ vs engineering strain ε for hydrogels composed of PEG-4SH cross-linked by (A) PEG-8C and (B) hPG-C, where negative stresses correspond to tension. The largest tensile engineering stress is denoted the failure stress σ_f, and the integrated area under the stress–strain curve gives the debonding work W_d. (C) Average values of σ_f and W_d as a function of the catechol:thiol ratio (N ≥ 3); error bars show the SD.

Fig. 4.

Gelation and adhesion of 1.4% or 2.8% (2x) MIP or mucin protein (MUC2) cross-linked by 1% PEG-8C and 0.16 mg/mL tyrosinase. Subscripts indicate reduction “r” or alkylation “a” of mucin. (A) Evolution of the storage modulus G′ vs time t at frequency ω = 5 rad/s. Dashed line shows the minimum measurable modulus for the parallel plate test fixture (d = 8 mm). (B) Evolution of the tensile engineering stress σ vs engineering strain ε measured after gelation. (C) Average values of the final modulus G_f, failure stress σ_f, and debonding work W_d (N ≥ 2); error bars show the SD for N ≥ 3. (D) Sketches of the proposed cross-linking of different preparations of mucin proteins by PEG-8C in the presence of tyrosinase. 8-spoked asterisks represent PEG-8C. Yellow triangles represent thiol groups, merged yellow triangles represent disulfide bonds, and purple triangles represent alkylated thiol groups. Cross-linking between catechol groups and either amine, imidazole, or other catechol groups is proposed to contribute to gelation regardless of mucin preparation, whereas catechol–thiol cross-linking requires reduced thiol groups, and is prevented by disulfide bonding or thiol alkylation.

Fig. 5.

Applications of mucin-inspired and mucin-derived hydrogels as tissue adhesives and antifouling coatings. (A) Adhesion measurements of hydrogels composed of MIP, PEG-8C, and tyrosinase (blue) after gelation between moistened pigskin surfaces. Control measurements of PEG-4SH gels (gray/black) cross-linked by disulfide bonds are shown for comparison. Three replicates for each gel are shown, indicated by different shadings. The Upper Inset shows the evolution of the storage modulus G′ vs time t at frequency ω = 5 rad/s during gelation of each replicate; the lower inset shows the correlation between the failure stress σ_f and the final storage modulus G_f of the replicates. (B) Box plot representation of biofilm volume on bare or coated glass surfaces after incubation in Pseudomonas aeruginosa, for coatings composed of in-lab purified mucin (MUC2, MUC5AC), commercial BSM, and MIP, each cross-linked by PEG-8C and tyrosinase. Open squares show the mean of each dataset. Representative fluorescence images are shown on Right. (Scale bars, 50 μm.)