Keratin hydrogel carrier system for simultaneous delivery of exogenous growth factors and muscle progenitor cells

Tomblyn, Seth; Kneller, Elizabeth L. Pettit; Walker, Stephen J.; Ellenburg, Mary D.; Kowalczewski, Christine; Dyke, Mark Van; Burnett, Luke; Saul, Justin M.

doi:10.1002/jbm.b.33438

Cited by 64 publications

(81 citation statements)

References 62 publications

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“…Ideally, cells would have been incorporated directly into the keratin hydrogels for these studies. However, kerateine does not gel in the presence of the salts present in cell culture media, so we were unable to incorporate cells directly into the keratin (though we note that we have recently shown cell viability in keratose gels [46]).…”

Section: Resultsmentioning

confidence: 99%

Alkylation of human hair keratin for tunable hydrogel erosion and drug delivery in tissue engineering applications

et al. 2015

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Polymeric biomaterials that provide a matrix for cell attachment and proliferation while achieving delivery of therapeutic agents are an important component of tissue engineering and regenerative medicine strategies. Keratins are a class of proteins that have received attention for numerous tissue engineering applications because, like other natural polymers, they promote favorable cell interactions and have non-toxic degradation products. Keratins can be extracted from various sources including human hair, and they are characterized by a high percentage of cysteine residues. Thiol groups on reductively extracted keratin (kerateine) form disulfide bonds, providing a more stable cross-linked hydrogel network than oxidatively extracted keratin (keratose) that cannot form disulfide crosslinks. We hypothesized that an iodoacetamide alkylation (or “capping”) of cysteine thiol groups on the kerateine form of keratin could be used as a simple method to modulate the levels of disulfide crosslinking in keratin hydrogels, providing tunable rates of gel erosion and therapeutic agent release. After alkylation, the alkylated kerateines still formed hydrogels and the alkylation led to changes in the mechanical and visco-elastic properties of the materials consistent with loss of disulfide crosslinking. The alkylated kerateines did not lead to toxicity in MC3T3-E1 pre-osteoblasts. These cells adhered to keratin at levels comparable to fibronectin and greater than collagen. Alkylated kerateine gels eroded more rapidly than non-alkylated kerateine and this control over erosion led to tunable rates of delivery of rhBMP-2, rhIGF-1, and ciprofloxacin. These results demonstrate that alkylation of kerateine cysteine residues provides a cell-compatible approach to tune rates of hydrogel erosion and therapeutic agent release within the context of a naturally-derived polymeric system.

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Section: Resultsmentioning

confidence: 99%

Alkylation of human hair keratin for tunable hydrogel erosion and drug delivery in tissue engineering applications

et al. 2015

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“…This surprising result suggests that these materials could be used to deliver viable cells in vivo directly from within injectable keratin hydrogels. 24 Given the flowable nature of these materials within tunable erosion timeframes noted above, this has clear advantages for tissue repair strategies. More in-depth studies on cell viability and function will be required to further assess the suitability of using these materials for cell delivery.…”

Section: Discussionmentioning

confidence: 99%

Tunable Keratin Hydrogels for Controlled Erosion and Growth Factor Delivery

et al. 2015

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Tunable erosion of polymeric materials is an important aspect of tissue engineering for reasons that include cell infiltration, controlled release of therapeutic agents, and ultimately to tissue healing. In general, the biological response to proteinaceous polymeric hydrogels is favorable (e.g., minimal inflammatory response). However, unlike synthetic polymers, achieving tunable erosion with natural materials is a challenge. Keratins are a class of intermediate filament proteins that can be obtained from several sources including human hair and have gained increasing levels of use in tissue engineering applications. An important characteristic of keratin proteins is the presence of a large number of cysteine residues. Two classes of keratins with different chemical properties can be obtained by varying the extraction techniques: (1) keratose by oxidative extraction and (2) kerateine by reductive extraction. Cysteine residues of keratose are “capped” by sulfonic acid and are unable to form covalent crosslinks upon hydration, whereas cysteine residues of kerateine remain as sulfhydryl groups and spontaneously form covalent disulfide crosslinks. Here, we describe a straightforward approach to fabricate keratin hydrogels with tunable rates of erosion by mixing keratose and kerateine. SEM imaging and mechanical testing of freeze-dried materials showed similar pore diameters and compressive moduli, respectively, for each keratose-kerateine mixture formulation (~1200 kPa for freeze-dried materials and ~1.5 kPa for hydrogels). However, the elastic modulus (G’) determined by rheology varied in proportion with the keratose-kerateine ratios, as did the rate of hydrogel erosion and the release rate of thiol from the hydrogels. The variation in keratose-kerateine ratios also led to tunable control over release rates of recombinant human insulin-like growth factor 1.

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“…Keratins are a family of structural proteins that can be found either as a major cytoskeletal component of keratinocyte cells in the epidermis layers of skin (“soft” keratin) or as a fibrous extracellular protein in hair, wool, quills and horns (“hard” keratin) [19–21]. Keratin, particularly “hard” keratin, has been used in scaffolds and drug delivery carriers for skin [22], muscle [23], and nerve tissue engineering [23–29]. Furthermore, extracellular “hard” keratin has been successfully used in the treatment of dermal burn wounds on animal models (split-thickness burns in mice, rats [30], or pigs [31]) and on clinical patients (split-thickness burns <10% of total body surface area [22]).…”

Section: Introductionmentioning

confidence: 99%

Development of keratin-based membranes for potential use in skin repair

et al. 2019

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The layers in skin determine its protective and hemostasis functions. This layered microstructure cannot be naturally regenerated after severe burns; we aim to reconstruct it using guided tissue regeneration (GTR). In GTR, a membrane is used to regulate tissue growth by stopping fast-proliferating cells and allowing slower cells to migrate and reconstruct specialized microstructures. Here, we proposed the use of keratin membranes crosslinked via dityrosine bonding. Variables from the crosslinking process were grouped within an energy density (ED) parameter to manufacture and evaluate the membranes. Sol fraction, spectrographs, and thermograms were used to quantify the non-linear relation between ED and the resulting crosslinking degree (CD). Mechanical and swelling properties increased until an ED threshold was reached; at higher ED, the CD and properties of the membranes remained invariable indicating that all possible dityrosine bonds were formed. Transport assays showed that the membranes allow molecular diffusion; low ED membranes retain solutes within their structure while the high ED samples allow higher transport rates indicating that uncrosslinked proteins can be responsible of reducing transport. This was confirmed with lower transport of adipogenic growth factors to stem cells when using low ED membranes; high ED samples resulted in increased production of intracellular lipids. Overall, we can engineer keratin membranes with specific CD, a valuable tool to tune microstructural and transport properties.

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Keratin hydrogel carrier system for simultaneous delivery of exogenous growth factors and muscle progenitor cells

Cited by 64 publications

References 62 publications

Alkylation of human hair keratin for tunable hydrogel erosion and drug delivery in tissue engineering applications

Alkylation of human hair keratin for tunable hydrogel erosion and drug delivery in tissue engineering applications

Tunable Keratin Hydrogels for Controlled Erosion and Growth Factor Delivery

Development of keratin-based membranes for potential use in skin repair

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