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Energetic electrons have recently evolved as powerful tool for crosslinking bio-derived hydrogels without the need for adding potentially hazardous reagents. Application of this approach allows for synthesis of biomimetic collagen-derived networks of highly tunable properties and functionalization. Yet, the underlying reaction kinetics it still not sufficiently established at this point. While hydroxyl radicals are generated by energetic electron-induced hydrolysis of water and play a key role in introducing covalent bonds between network fibers, a detailed mechanistic understanding would significantly increase applicability. We present a comprehensive analysis of central aspects of the reactivity between the hydroxyl radical (•OH) and collagen, elastin, glycine (Gly) and L-lysine (Lys). Pulse radiolysis (PR), solid state nuclear magnetic resonance (NMR), ultraviolet-visible absorption spectroscopy (UV/VIS) and electron spray ionization mass spectrometry (ESI-MS) shine light on distinct features of the crosslinking process. These highlight retained protein backbone integrity in collagen and elastin whilst Lys's ability to form several imine bonded Lys-Lys species suggests striking similarities to crosslinking via lysyloxidase catalysis in vivo. Thus, energetic electron based crosslinking opens the venue for customized hybrid gels of outstanding biomimicry and –compatibility. Statement of significance Energetic electron beam treatment constitutes a highly attractive approach to establish chemical bonds between (bio) molecules. Although a convincing number of publications showed the versatility regarding crosslinking of bioderived hydrogels, insights into the underlying chemistry are still unestablished at this point. The present work unravels the mechanistics of energetic electron induced processes in collagen and elastin hydrogels as well as several abundant amino acids in aqueous solution. As key finding we demonstrate, that i) the connection between polymer chains is dominated by amino acid side chain interaction and ii) two single L-lysine molecules form an imine bond between the terminal amino group of one molecule and the delta carbon of the second molecule. We also consider the formation of H-bonds as a second crosslinking pathway. These findings open up for advanced, optionally spatially resolved biomaterials design.

Iron oxide nanoparticles with a mean size of approximately 5 nm were synthesized by irradiating micro-emulsions containing iron salts with energetic electrons. The properties of the nanoparticles were investigated using scanning electron microscopy, high-resolution transmission electron microscopy, selective area diffraction and vibrating sample magnetometry. It was found that formation of superparamagnetic nanoparticles begins at a dose of 50 kGy, though these particles show low crystallinity, and a higher portion is amorphous. With increasing doses, an increasing crystallinity and yield could be observed, which is reflected in an increasing saturation magnetization. The blocking temperature and effective anisotropy constant were determined via zero-field cooling and field cooling measurements. The particles tend to form clusters with a size of 34 nm to 73 nm. Magnetite/maghemite nanoparticles could be identified via selective area electron diffraction patterns. Additionally, goethite nanowires could be observed.

Biopolymer blends are advantageous materials with novel properties that may show performances way beyond their individual constituents. Collagen elastin hybrid gels are a new representative of such materials as they employ elastin’s thermo switching behavior in the physiological temperature regime. Although recent studies highlight the potential applications of such systems, little is known about the interaction of collagen and elastin fibers during polymerization. In fact, the final network structure is predetermined in the early and mostly arbitrary association of the fibers. We investigated type I collagen polymerized with bovine neck ligament elastin with up to 33.3 weight percent elastin and showed, by using a plate reader, zeta potential and laser scanning microscopy (LSM) experiments, that elastin fibers bind in a lateral manner to collagen fibers. Our plate reader experiments revealed an elastin concentration-dependent increase in the polymerization rate, although the rate increase was greatest at intermediate elastin concentrations. As elastin does not significantly change the structural metrics pore size, fiber thickness or 2D anisotropy of the final gel, we are confident to conclude that elastin is incorporated homogeneously into the collagen fibers.

Networks of crosslinked keratin filaments are abundant in epithelial cells and tissues, providing resilience against mechanical forces and ensuring cellular integrity. Although studies of in vitro models of reconstituted keratin networks have revealed important mechanical aspects, the mechanical properties of crosslinked keratin structures remain poorly understood. Here, we exploited the power of electron beam irradiation (EBI) to crosslink in vitro networks of soft epithelial keratins 8 and 18 (k8–k18) filaments with different irradiation doses (30 kGy, 50 kGy, 80 kGy, 100 kGy, and 150 kGy). We combined bulk shear rheology with confocal microscopy to investigate the impact of crosslinking on the mechanical and structural properties of the resultant keratin gels. We found that irradiated keratin gels display higher linear elastic modulus than the unirradiated, entangled networks at all doses tested. However, at the high doses (80 kGy, 100 kGy, and 150 kGy), we observed a remarkable drop in the elastic modulus compared to 50 kGy. Intriguingly, the irradiation drastically changed the behavior for large, nonlinear deformations. While untreated keratin networks displayed a strong strain stiffening, increasing irradiation doses shifted the system to a strain softening behavior. In agreement with the rheological behavior in the linear regime, the confocal microscopy images revealed fully isotropic networks with high percolation in 30 kGy and 50 kGy-treated keratin samples, while irradiation with 100 kGy induced the formation of thick bundles and clusters. Our results demonstrate the impact of permanent crosslinking on k8–k18 mechanics and provide new insights into the potential contribution of intracellular covalent crosslinking to the loss of mechanical resilience in some human keratin diseases. These insights will also provide inspiration for the synthesis of new keratin-based biomaterials.

As emerging responsive materials, ferrogels have become highly attractive for biomedical and technical applications in terms of soft actuation, tissue engineering or controlled drug release. In the present study, bioderived ferrogels were fabricated and successfully deformed within moderate, heterogeneous magnetic fields. Synthesis was realized by arresting iron oxide nanoparticles in porcine gelatin by introduction of covalent crosslinks via treatment with energetic electrons for mesh refinement. This approach also allows for tuning thermal and mechanical stability of the gelatin matrix. Operating the bioferrogel in compression, magnetic forces on the nanoparticles are counterbalanced by the stiffness of the hydrogel matrix that is governed by a shift in thermodynamic equilibrium of swelling, as derived in the framework of osmosis. As gelatin and iron oxide nanoparticles are established as biocompatible constituents, these findings promise potential for in vivo use as contactless mechanical transducers.

Smart materials such as stimuli responsive polymeric hydrogels offer unique possibilities for tissue engineering and regenerative medicine. As, however, most synthetic polymer systems and their degradation products lack complete biocompatibility and biodegradability, this study aims to synthesize a highly magnetic responsive hydrogel, based on the abundant natural biopolymer collagen. As the main component of vertebratal extracellular matrix, it reveals excellent biocompatibility. In combination with incorporated magnetic iron oxide nanoparticles, a novel smart nano-bio-ferrogel can be designed. While retaining its basic biophysical properties and interaction with living cells, this collagen-nanoparticle hydrogel can be compressed to 38% of its original size and recovers to 95% in suitable magnetic fields. Besides the phenomenology of this scenario, the underlying physical scenarios are also discussed within the framework of network models. The observed reversible peak strains as large as 150% open up possibilities for the fields of biomedical actuation, soft robotics and beyond.

The motility of cells plays an important role for many processes such as wound healing and malignant progression of cancer. The efficiency of cell motility is affected by the microenvironment. The connection between the cell and its microenvironment is facilitated by cell-matrix adhesion receptors and upon their activation focal adhesion proteins such as integrin-linked kinase (ILK) are recruited to sites of focal adhesion formation. In particular, ILK connects cell-matrix receptors to the actomyosin cytoskeleton. However, ILK's role in cell mechanics regulating cellular motility in 3D collagen matrices is still not well understood. We suggest that ILK facilitates 3D motility by regulating cellular mechanical properties such as stiffness and force transmission. Thus, ILK wild-type and knock-out cells are analyzed for their ability to migrate on 2D substrates serving as control and in dense 3D extracellular matrices. Indeed, ILK wild-type cells migrated faster on 2D substrates and migrated more numerous and deeper in 3D matrices. Hence, we analyzed cellular deformability, Young's modulus (stiffness) and adhesion forces. We found that ILK wild-type cells are less deformable (stiffer) and produce higher cell-matrix adhesion forces compared to ILK knock-out cells. Finally, ILK is essential for providing cellular mechanical stiffness regulating 3D motility.