Flow Simulation and Gradient Printing of Fluorapatite- and Cell-Loaded Recombinant Spider Silk Hydrogels.

Hierarchical structures are abundant in almost all tissues of the human body. Therefore, it is highly important for tissue engineering approaches to mimic such structures if a gain of function of the new tissue is intended. Here, the hierarchical structures of the so-called enthesis, a gradient tissue located between tendon and bone, were in focus. Bridging the mechanical properties from soft to hard secures a perfect force transmission from the muscle to the skeleton upon locomotion. This study aimed at a novel method of bioprinting to generate gradient biomaterial constructs with a focus on the evaluation of the gradient printing process. First, a numerical approach was used to simulate gradient formation by computational flow as a prerequisite for experimental bioprinting of gradients. Then, hydrogels were printed in a single cartridge printing set-up to transfer the findings to biomedically relevant materials. First, composites of recombinant spider silk hydrogels with fluorapatite rods were used to generate mineralized gradients. Then, fibroblasts were encapsulated in the recombinant spider silk-fluorapatite hydrogels and gradually printed using unloaded spider silk hydrogels as the second component. Thereby, adjustable gradient features were achieved, and multimaterial constructs were generated. The process is suitable for the generation of gradient materials, e.g., for tissue engineering applications such as at the tendon/bone interface.

Recombinant Spider Silk Gels Derived from Aqueous-Organic Solvents as Depots for Drugs.

Hydrogels are widely used in various biomedical applications, as they cannot only serve as materials for biofabrication but also as depots for the administration of drugs. However, the possibilities of formulation of water-insoluble drugs in hydrogels are rather limited. Herein, we assembled recombinant spider silk gels using a new processing route with aqueous-organic co-solvents, and the properties of these gels could be controlled by the choice of the co-solvent. The presence of the organic co-solvent further enabled the incorporation of hydrophobic drugs as exemplarily shown for 6-mercaptopurine. The developed gels showed shear-thinning behaviour and could be easily injected to serve, for example, as drug depots, and they could even be 3D printed to serve as scaffolds for biofabrication. With this new processing route, the formulation of water-insoluble drugs in spider silk-based depots is possible, circumventing common pharmaceutical solubility issues.

Silk-Based Materials for Hard Tissue Engineering.

Hard tissues, e.g., bone, are mechanically stiff and, most typically, mineralized. To design scaffolds for hard tissue regeneration, mechanical, physico-chemical and biological cues must align with those found in the natural tissue. Combining these aspects poses challenges for material and construct design. Silk-based materials are promising for bone tissue regeneration as they fulfill several of such necessary requirements, and they are non-toxic and biodegradable. They can be processed into a variety of morphologies such as hydrogels, particles and fibers and can be mineralized. Therefore, silk-based materials are versatile candidates for biomedical applications in the field of hard tissue engineering. This review summarizes silk-based approaches for mineralized tissue replacements, and how to find the balance between sufficient material stiffness upon mineralization and cell survival upon attachment as well as nutrient supply.

Spider Silk Fusion Proteins for Controlled Collagen Binding and Biomineralization.

The development of biomaterials for the interface between tendon and bone is important for realizing functional tendon replacements. Toward the development of new materials for such applications, engineered recombinant spider silk proteins were modified with peptide tag sequences derived from noncollagenous proteins in bone, so-called SIBLING proteins, such as osteopontin and sialoprotein, which are known to interact with collagen and to initiate mineralization. Materials made of these spider silk-SIBLING hybrids were analyzed concerning mineralization and interaction with cells. They showed enhanced calcium phosphate formation upon incubation in mineralization agents. In gradient films, MC3T3-E1 mouse preosteoblasts adhered preferentially along the gradient toward the variant with a collagen binding motif.

Free-standing spider silk webs of the thomisid Saccodomus formivorus are made of composites comprising micro- and submicron fibers.

Our understanding of the extraordinary mechanical and physico-chemical properties of spider silk is largely confined to the fibers produced by orb-weaving spiders, despite the diversity of foraging webs that occur across numerous spider families. Crab spiders (Thomisidae) are described as ambush predators that do not build webs, but nevertheless use silk for draglines, egg cases and assembling leaf-nests. A little-known exception is the Australian thomisid Saccodomus formivorus, which constructs a basket-like silk web of extraordinary dimensional stability and structural integrity that facilitates the capture of its ant prey. We examined the physical and chemical properties of this unusual web and revealed that the web threads comprise microfibers that are embedded within a biopolymeric matrix containing additionally longitudinally-oriented submicron fibers. We showed that the micro- and submicron fibers differ in their chemical composition and that the web threads show a remarkable lateral resilience compared with that of the major ampullate silk of a well-investigated orb weaver. Our novel analyses of these unusual web and silk characteristics highlight how investigations of non-model species can broaden our understanding of silks and the evolution of foraging webs.