Keratin-Alginate Sponges Support Healing of Partial-Thickness Burns.

Deep partial-thickness burns damage most of the dermis and can cause severe pain, scarring, and mortality if left untreated. This study serves to evaluate the effectiveness of crosslinked keratin-alginate composite sponges as dermal substitutes for deep partial-thickness burns. Crosslinked keratin-alginate sponges were tested for the ability to support human dermal fibroblasts in vitro and to support the closure and healing of partial-thickness burn wounds in Sus scrofa pigs. Keratin-alginate composite sponges supported the enhanced proliferation of human dermal fibroblasts compared to alginate-only sponges and exhibited decreased contraction in vitro when compared to keratin only sponges. As dermal substitutes in vivo, the sponges supported the expression of keratin 14, alpha-smooth muscle actin, and collagen IV within wound sites, comparable to collagen sponges. Keratin-alginate composite sponges supported the regeneration of basement membranes in the wounds more than in collagen-treated wounds and non-grafted controls, suggesting the subsequent development of pathological scar tissues may be minimized. Results from this study indicate that crosslinked keratin-alginate sponges are suitable alternative dermal substitutes for clinical applications in wound healing and skin regeneration.

Anisotropic hair keratin-dopamine composite scaffolds exhibit strain-stiffening properties.

Human hair keratin (HHK) has been successfully explored as raw materials for three-dimensional scaffolds for soft tissue regeneration due to its excellent biocompatibility and bioactivity. However, none of the reported HHK based scaffolds is able to replicate the strain-stiffening capacity of living tissues when responding to large deformations. In the present study, strain-stiffening property was achieved in scaffolds fabricated from HHK via a synergistic effect of well-defined, aligned microstructure and chemical crosslinking. Directed ice-templating method was used to fabricate HHK-based scaffolds with highly aligned (anisotropic) microstructure while oxidized dopamine (ODA) was used to crosslink covalently to HHKs. The resultant HHK-ODA scaffolds exhibited strain-stiffening behavior characterized by the increased gradient of the stress-strain curve after the yield point. Both ultimate tensile strength and the elongation at break were enhanced significantly (~700 kPa, ~170%) in comparison to that of HHK scaffolds lacking of aligned microstructure or ODA crosslinking. In vitro cell culture studies indicated that HHK-ODA scaffolds successfully supported human dermal fibroblasts (HDFs) adhesion, spreading and proliferation. Moreover, anisotropic HHK-ODA scaffolds guided cell growth in alignment with the defined microstructure as shown by the highly organized cytoskeletal networks and nuclei distribution. The findings suggest that HHK-ODA scaffolds, with strain-stiffening properties, biocompatibility and bioactivity, have the potential to be applied as biomimetic matrices for soft tissue regeneration.

Characterization of Anisotropic Human Hair Keratin Scaffolds Fabricated via Directed Ice Templating.

Human hair keratin (HHK) is successfully exploited as raw materials for 3D scaffolds for soft tissue regeneration owing to its excellent biocompatibility and bioactivity. However, most HHK scaffolds are not able to achieve the anisotropic mechanical properties of soft tissues such as tendons and ligaments due to lack of tunable, well-defined microstructures. In this study, directed ice templating method is used to fabricate anisotropic HHK scaffolds that are characterized by aligned pores (channels) in between keratin layers in the longitudinal plane. In contrast, pores in the transverse plane maintain a homogenous rounded morphology. Channel widths throughout the scaffolds range from ≈5 to ≈15 µm and are tunable by varying the freezing temperature. In comparison with HHK scaffolds with random, isotropic pore structures, the tensile strength of anisotropic HHK scaffolds is enhanced significantly by up to fourfolds (≈200 to ≈800 kPa) when the tensile load is applied in the direction parallel to the aligned pores. In vitro results demonstrate that the anisotropic HHK scaffolds are able to support human dermal fibroblast adhesion, spreading, and proliferation. The findings suggest that HHK scaffolds with well-defined, aligned microstructure hold promise as templates for soft tissues regeneration by mimicking their anisotropic properties.

Composite Hydrogels in Three-Dimensional in vitro Models.

3-dimensional (3D) in vitro models were developed in order to mimic the complexity of real organ/tissue in a dish. They offer new possibilities to model biological processes in more physiologically relevant ways which can be applied to a myriad of applications including drug development, toxicity screening and regenerative medicine. Hydrogels are the most relevant tissue-like matrices to support the development of 3D in vitro models since they are in many ways akin to the native extracellular matrix (ECM). For the purpose of further improving matrix relevance or to impart specific functionalities, composite hydrogels have attracted increasing attention. These could incorporate drugs to control cell fates, additional ECM elements to improve mechanical properties, biomolecules to improve biological activities or any combinations of the above. In this Review, recent developments in using composite hydrogels laden with cells as biomimetic tissue- or organ-like constructs, and as matrices for multi-cell type organoid cultures are highlighted. The latest composite hydrogel systems that contain nanomaterials, biological factors, and combinations of biopolymers (e.g., proteins and polysaccharide), such as Interpenetrating Networks (IPNs) and Soft Network Composites (SNCs) are also presented. While promising, challenges remain. These will be discussed in light of future perspectives toward encompassing diverse composite hydrogel platforms for an improved organ environment in vitro.

Enhanced performance of chitosan/keratin membranes with potential application in peripheral nerve repair.

Although surgical management of peripheral nerve injuries (PNIs) has improved over time, autografts are still the current "gold standard" treatment for PNIs, which presents numerous limitations. In an attempt to improve natural biomaterial-based nerve guidance conduits (NGCs), chitosan (CHT), a derivative of the naturally occurring biopolymer chitin, has been explored for peripheral nerve regeneration (PNR). In addition to CHT, keratin has gained enormous attention as a biomaterial and tissue engineering scaffolding. In this study, biomimetic CHT/keratin membranes were produced using a solvent casting technique. These membranes were broadly characterized in terms of their surface topography and physicochemical properties, with techniques such as Fourier Transform Infrared Spectroscopy (FTIR), Differential Scanning Calorimetry (DSC), contact angle, weight loss and water uptake measurements, Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM). Biological in vitro assays were also performed, where a preliminary cytotoxicity screening with the L929 fibroblast cell line revealed that the membranes and respective materials are suitable for cell culture. In addition, Schwann cells, fibroblasts and endothelial cells were directly seeded in the membranes. Quantitative and qualitative assays revealed that the addition of keratin enhanced cell viablity and adhesion. Based on the encouraging in vitro results, the in vivo angiogenic/antiangiogenic potential of CHT and CHT/keratin membranes was assessed, using an optimized chick embryo chorioallantoic membrane assay, where higher angiogenic responses were seen in keratin-enriched materials. Overall, the obtained results indicate the higher potential of CHT/keratin membranes for guided tissue regeneration applications in the field of PNR.