Silk fibroin (SF) and mucin are extensively recognized as promising biomaterials for wound dressings due to their outstanding biocompatibility, biodegradability, and ability to support cell growth and tissue regeneration. In this study, we developed a hybrid SF/mucin wound dressing (HYB) using tetrazine and norbornene click chemistry to enhance its structural and functional properties. The robust assembly resulted in a dual-phase material with a dense SF membrane and a porous mucin hydrogel (MH). Scanning electron microscopy confirmed the successful integration and tight adhesion between these polymers. The hybrid material exhibited a controlled release of bioactive agents, with epidermal growth factor (EGF) showing a sustained release of up to 48% over 48 h. The optimized 25 mg/mL mucin hydrogel showed efficient EGF release and performance comparable to higher concentrations. It was selected for papain loading to reduce material usage without compromising efficacy. HYB showed a higher papain release rate of 36% compared to the bare SF membrane. Additionally, the hybrid material exhibited enhanced mechanical strength, optimized water vapor permeability comparable to commercial wound dressings, and improved cell proliferation relative to its individual components. Cytotoxicity assays demonstrated that the papain-loaded hybrid material is a viable candidate for wound dressing applications. These results suggest that the click-chemistry-functionalized SF/mucin hybrid material holds significant potential as an advanced wound dressing, capable of promoting tissue regeneration while maintaining a moist environment conducive to healing.

Biofilms constitute one of the most common forms of living matter, playing an increasingly important role in technology, health, and ecology. While it is well established that biofilm growth and morphology are highly dependent on the external flow environment, the precise role of fluid friction has remained elusive. We grew Bacillus subtilis biofilms on flat surfaces of a channel in a laminar flow at wall shear stresses spanning one order of magnitude (τw = 0.068 Pa to τw = 0.67 Pa). By monitoring the three-dimensional distribution of biofilm over seven days, we found that the biofilms consist of smaller microcolonies, shaped like leaning pillars, many of which feature a streamer in the form of a thin filament that originates near the tip of the pillar. While the shape, size, and distribution of these microcolonies depend on the imposed shear stress, the same structural features appear consistently for all shear stress values. The formation of streamers occurs after the development of a base structure, suggesting that the latter induces a secondary flow that triggers streamer formation. Moreover, we observed that the biofilm volume grows approximately linearly over seven days for all shear stress values, with a growth rate inversely proportional to the wall shear stress. We develop a scaling model, providing insight into the mechanisms by which friction limits biofilm growth.

Saliva plays a crucial role in lubricating the oral cavity and maintaining a normal hydrated mouthfeel, with the main macromolecule in saliva, MUC5B, playing a significant role in its lubricating properties. Mucins form a hydrated biogel that covers the oral epithelium and forms part of the mucosal pellicle, maintaining proper hydration. This study investigates the relationship between ionic concentration, saliva lubrication, and the perception of oral dryness. At baseline, sodium was found to be ten times more concentrated in the mucosal pellicle compared to bulk saliva, in contrast to potassium which was only 1.5 times more concentrated in the pellicle. Increasing concentrations of mouthwashes containing sodium or potassium indicated a preferential sequestering of sodium in the mucosal pellicle with sodium chloride rinses being associated with an increased thirst perception. We therefore propose that sodium binds to MUC5B, reducing the electrostatic repulsions between its negatively charged moieties. This interaction subsequently leads to a compaction of the mucin structure, which decreases the hydration of MUC5B and reduces its lubricating properties. We determined that increased sodium concentrations compact the mucin structure as indicated by transmission electron microscopy. However, we found that increased sodium concentrations were associated with improved the lubrication of salivary and MUC5B films in an oral tribology mimic, decreasing the coefficient of friction from 0.044 to 0.018, and 0.416 to 0.100 with the addition of 200 mM sodium chloride for saliva and MUC5B, respectively. MUC5B was also demonstrated to become more hydrated with increasing ionic concentrations, as indicated by Quartz Crystal Microbalance with Dissipation. We therefore propose that increased hydration shells surrounding the sodium counterions are responsible for this improved lubrication via the hydration lubrication mechanism. Lastly, it was observed that the spinnbarkeit of saliva decreased with increasing ionic concentrations, indicating a reduced stretchiness in its rheological properties. However, the concentration of MUC5B alone did not appear to affect spinnbarkeit, highlighting that many factors influence the biophysical properties of saliva.

Histoplasmosis is a respiratory disease caused by Histoplasma capsulatum, a dimorphic fungus, with high mortality and morbidity rates, especially in immunocompromised patients. Considering the small existing therapeutic arsenal, new treatment approaches are still required. Chitosan, a linear polysaccharide obtained from partial chitin deacetylation, has anti-inflammatory, antimicrobial, biocompatibility, biodegradability, and non-toxicity properties. Chitosan with different deacetylation degrees and molecular weights has been explored as a potential agent against fungal pathogens. In this study, the chitosan antifungal activity against H. capsulatum was evaluated using the broth microdilution assay, obtaining minimum inhibitory concentrations (MIC) ranging from 32 to 128 µg/mL in the filamentous phase and 8 to 64 µg/mL in the yeast phase. Chitosan combined with classical antifungal drugs showed a synergic effect, reducing chitosan’s MICs by 32 times, demonstrating that there were no antagonistic interactions relating to any of the strains tested. A synergism between chitosan and amphotericin B or itraconazole was detected in the yeast-like form for all strains tested. For H. capsulatum biofilms, chitosan reduced biomass and metabolic activity by about 40% at 512 µg/mL. In conclusion, studying chitosan as a therapeutic strategy against Histoplasma capsulatum is promising, mainly considering its numerous possible applications, including its combination with other compounds.

Biomaterials made of self-assembling protein building blocks are widely explored for biomedical applications, for example, as drug carriers, tissue engineering scaffolds, and functionalized coatings. It has previously been shown that a recombinant spider silk protein functionalized with a cell binding motif from fibronectin, FN-4RepCT (FN-silk), self-assembles into fibrillar structures at interfaces, i.e., membranes, fibers, or foams at liquid/air interfaces, and fibrillar coatings at liquid/solid interfaces. Recently, we observed that FN-silk also assembles into microspheres in the bulk of a physiological buffer (PBS) solution. Herein, we investigate the self-assembly process of FN-silk into microspheres in the bulk and how its progression is affected by the presence of hyaluronic acid (HA), both in solution and in a cross-linked HA hydrogel. Moreover, we characterize the size, morphology, mesostructure, and protein secondary structure of the FN-silk microspheres prepared in PBS and HA. Finally, we examine how the FN-silk microspheres can be used to mediate cell adhesion and spreading of human mesenchymal stem cells (hMSCs) during cell culture. These investigations contribute to our fundamental understanding of the self-assembly of silk protein into materials and demonstrate the use of silk microspheres as additives for cell culture applications.

Mucus reduces friction between epithelial surfaces by providing lubrication in the boundary and mixed regime. Mucins, the main macromolecule, are heavily glycosylated proteins that polymerise and retain water molecules, resulting in a hydrated biogel. It is assumed that positively charged ions can influence mucin film structure by screening the electrostatic repulsions between the negatively charged glycans on mucin moieties and draw in water molecules via hydration shells. The ionic concentration can vary significantly in different mucus systems and here we show that increasing the ionic concentration in mucin films leads to an increase in lubrication between two polydimethylsiloxane surfaces at sliding contact in a compliant oral mimic. Mucins were found to bind sodium ions in a concentration-dependent manner and increased ionic concentration appears to cause mucin films to swell when assessed by Quartz Crystal hiMicrobalance with Dissipation (QCM-D) analysis. Furthermore, we determined that the removal of negatively charged sialic acid moieties by sialidase digestion resulted in reduced adsorption to hydrophilic surfaces but did not affect the swelling of mucin films with increasing ionic concentrations. Moreover, the coefficient of friction was increased with sialic acid removal, but lubrication was still increased with increasing ionic concentrations. Taken together this suggests that sialic acids are important for lubrication and may exert this through the sacrificial layer mechanism. Ionic concentration appears to influence mucin films and their lubrication, and sialic acids, at least partly, may be important for ion binding.

Hydrogels of cross-linked mucin glycoproteins (Muc-gel) have shown strong immune-modulating properties toward macrophages in vitro, which are translated in vivo by the dampening of the foreign body response to implantation in mice. Beyond mucin hydrogels, other biomaterials such as sensors, electrodes, and other long-term implants would also benefit from such immune-modulating properties. In this work, we aimed to transfer the bioactivity observed for three-dimensional Muc-gels to the surface of two model materials by immobilizing mucin into thin films (Muc-film) using covalent layer-by-layer assembly. We tested how the surface immobilization of mucins affects macrophage responses compared to Muc-gels. We showed that Muc-films on soft polyacrylamide gels mimic Muc-gel in their modulation of macrophage responses with activated gene expression of inflammatory cytokines on day 1 and then dampening them on day 3. Also, the markers of polarized macrophages, M1 and M2, were expressed at the same level for macrophages on Muc-film-coated soft polyacrylamide gels and Muc-gel. In contrast, Muc-film-coated hard polystyrene led to a different macrophage response compared to Muc-gel, having no activated expression of inflammatory cytokines and a different M1 marker expression. This suggested that the substrate mechanical properties and mucin molecular configuration determined by substrate-mucin interactions affect mucin immune-modulating properties. We conclude that mucin immune-modulating properties can be transferred to materials by mucin surface immobilization but will be dependent on the substrate chemical and mechanical properties. 

Biocatalytical upgrading of side streams from agricultural biomass into multifunctional materials constitutes a very attractive option to increase the circularity of food and material systems. We propose the design of radical scavenging hydrogels with mechanical integrity and protective effects against reactive oxygen species by enzymatic crosslinking of arabinoxylans (AX) with high ferulic acid content extracted from corn fibre using subcritical water. We have compared the influence of two enzymatic systems, laccase/O2 and peroxidase/H2O2, on the biochemical structure, multiscale assembly, physicochemical properties, and radical scavenging activity of the polysaccharide hydrogels. Peroxidase crosslinking results in instant hydrogel formation, whereas laccase shows slower crosslinking kinetics, resulting in a more elastic gel network. Characterization by size exclusion chromatography, small angle X-ray scattering, and microscopy revealed structural differences in the network organization of the hydrogels produced by the two enzymes. Laccase crosslinking leads to smaller polymeric aggregates, promoting their progressive organization in network clusters that impact the overall ultrastructure. Conversely, the fast crosslinking induced by peroxidase results in higher porosity and forms larger and potentially more heterogeneous aggregates, which seem to hinder their subsequent association in clusters. Both AX hydrogels exhibit adequate biocompatibility and protective effects against in vitro cellular oxidative stress compared to an alginate reference. This constitutes a proof of concept of the potential application of radical scavenging hydrogels from agricultural side streams for biomedical and nutritional applications in wound healing, cellular repair and targeted delivery.