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Designing biomimetic scaffolds with in vivo–like microenvironments using biomaterials is an essential component of successful tissue engineering approaches. The intestinal smooth muscle layers exhibit a complex tubular structure consisting of two concentric muscle layers in which the inner circular layer is orthogonally oriented to the outer longitudinal layer. Here, a 3D bi‐layered tubular scaffold is presented based on flexible, mechanically robust, and well aligned silk protein microfibers to mimic the native human intestinal smooth muscle structure. The scaffolds are seeded with primary human intestinal smooth muscle cells to replicate intestinal muscle tissues in vitro. Characterization of the tissue constructs reveals good biocompatibility and support for cell alignment and elongation in the different scaffold layers to enhance cell differentiation and functions. Furthermore, the engineered smooth muscle constructs support oriented neurite outgrowth, a requisite step to achieve functional innervation. These results suggest these microfiber scaffolds as functional templates for in vitro regeneration of human intestinal smooth muscle systems. The scaffolding provides a crucial step toward engineering functional human intestinal tissue in vitro, as well as engineering other types of smooth muscles in terms of their similar phenotypes. Such utility may lead to a better understanding of smooth muscle associated diseases and treatments. [ABSTRACT FROM AUTHOR]

There is great need for soft biomaterials that match the stiffness of human tissues for tissue engineering and regeneration. Hydrogels are frequently employed for extracellular matrix functionalization and to provide appropriate mechanical cues. It is challenging, however, to achieve structural integrity and retain bioactive molecules in hydrogels for complex tissue formation that may take months to develop. This work aims to investigate mechanical and biochemical characteristics of silk hydrogels for soft tissue engineering, specifically for the nervous system. The stiffness of 1 to 8% silk hydrogels, measured by atomic force microscopy, is 4 to 33 kPa. The structural integrity of silk gels is maintained throughout embryonic chick dorsal root ganglion (cDRG) explant culture over 4 days whereas fibrin and collagen gels decrease in mass over time. Neurite extension of cDRGs cultured on 2 and 4% silk hydrogels exhibit greater growth than softer or stiffer gels. Silk hydrogels release <5% of neurotrophin-3 (NT-3) over 2 weeks and 11-day old gels show maintenance of growth factor bioactivity. Finally, fibronectin- and NT-3-functionalized silk gels elicit increased axonal bundling suggesting their use in bridging nerve injuries. These results support silk hydrogels as soft and sustainable biomaterials for neural tissue engineering. [ABSTRACT FROM AUTHOR]

In the field of regenerative medicine there is a need for scaffolds that support large, critically-sized tissue formation. Major limitations in reaching this goal are the delivery of oxygen and nutrients throughout the bulk of the engineered tissue as well as host tissue integration and vascularization upon implantation. To address these limitations, the development of a porous scaffold platform made from biodegradable silk protein that contains an array of vascular-like structures that extend through the bulk of the scaffold was previously reported. Here, the hollow channels play a pivotal role in enhancing cell infiltration, delivering oxygen and nutrients to the scaffold bulk, and promoting in vivo host tissue integration and vascularization. The unique features of this protein biomaterial system, including the vascular structures and tunable material properties, render this scaffold a robust and versatile tool for implementation in a variety of tissue engineering, regenerative medicine, and disease modeling applications. [ABSTRACT FROM AUTHOR]

The goals of the present study are to establish an in vitro co-culture model of osteoblast and osteoclast function and to quantify the resulting bone remodeling. The bone is tissue engineered using well-defined silk protein biomaterials in 2D and 3D formats in combination with human cells. Parathyroid hormone (PTH) and glucose-dependent insulinotropic peptide (GIP) are selected because of their roles in bone remodeling for expression in tethered format on human mesenchymal stem cells (hMSCs). The cell-modified biomaterial surfaces are reconstructed from scanning electron microscopy images into 3D models for quantitative measurement of surface characteristics. Increased calcium deposition and surface roughness are found in 3D surface models of silk protein films remodeled by co-cultures containing tethered PTH, and decreased surface roughness is found for the films remodeled by tethered GIP co-cultures. Increased surface roughness is not found in monocultures of hMSCs expressing tethered PTH, suggesting that osteoclast-osteoblast interactions in the presence of PTH signaling are responsible for the increased mineralization. These data point towards the design of in vitro bone models in which osteoblast-osteoclast interactions are mimicked for a better understanding of bone remodeling. [ABSTRACT FROM AUTHOR]

Tunable protein composites are important for constructing extracellular matrix mimics of human nerve tissues with control of charge, structural, and mechanical properties. Molecular interaction mechanisms between silk fibroin protein and recombinant human tropoelastin, based on charge, are utilized to generate a new group of multifunctional protein alloys with different net charges. These new biomaterials are then utilized as a biomaterial platform to control neuron cell response. With a +38 net charge in water, tropoelastin molecules provide extraordinary elasticity and selective interactions with cell surface integrins. In contrast, negatively charged silk fibroin protein (net charge −36) provides remarkable toughness and stiffness with morphologic stability in material formats via autoclaving-induced beta-sheet crystal physical crosslinks. The combination of these properties in alloy format extends the versatility of both structural proteins, providing a new biocompatible, biodegradable, and charge-tunable biomaterial platform for neural repair. The data point to these protein alloys as an alternative to commonly used charged synthetic polymers, particularly with regard to the versatility of material formats (e.g., gels, sponges, films, fibers). The results also provide a practical example of physically designed protein materials with control of net charge to direct biological outcomes, in this case for neuronal tissue engineering. [ABSTRACT FROM AUTHOR]

There is critical clinical demand for tissue-engineered (TE), 3D constructs for tissue repair and organ replacements. Current efforts toward this goal are prone to necrosis at the core of larger constructs because of limited oxygen and nutrient diffusion. Therefore, critically sized 3D TE constructs demand an immediate vascular system for sustained tissue function upon implantation. To address this challenge the goal of this project was to develop a strategy to incorporate microchannels into a porous silk TE scaffold that could be fabricated reproducibly using microfabrication and soft lithography. Silk is a suitable biopolymer material for this application because it is mechanically robust, biocompatible, slowly degrades in vivo, and has been used in a variety of TE constructs. Here, the fabrication of a silk-based TE scaffold that contains an embedded network of porous microchannels is reported. Enclosed porous microchannels support endothelial lumen formation, a critical step toward development of the vascular niche, while the porous scaffold surrounding the microchannels supports tissue formation, demonstrated using human mesenchymal stem cells. This approach for fabricating vascularized TE constructs is advantageous compared to previous systems, which lack porosity and biodegradability or degrade too rapidly to sustain tissue structure and function. The broader impact of this research will enable the systemic study and development of complex, critically-sized engineered tissues, from regenerative medicine to in vitro tissue models of disease states. [ABSTRACT FROM AUTHOR]

Development and maturation of vascular and neuronal tissues occurs simultaneously in utero, and are regulated by significant crosstalk. Here, the development of a 3D tissue system to model neurogenesis and recapitulate developmental signaling conditions is reported. Human umbilical vein endothelial cells (HUVECs) are seeded inside channels within collagen gels to represent nascent vascular networks. Axons extending from chicken dorsal root ganglia grow significantly longer and preferentially toward the HUVEC seeded channels with respect to unloaded channels. To replicate these findings without the vascular component, channels are loaded with brain‐derived neurotrophic factor (BDNF), the principle signaling molecule in HUVEC‐stimulated axonal growth, and axons likewise are significantly longer and grow preferentially toward the BDNF‐loaded channels with respect to controls. This 3D tissue system is then used as an in vitro replicate for peripheral nerve injury, with neural repair observed within 2 weeks. These results demonstrate that the 3D tissue system can model neural network formation, can repair after laceration injuries, and can be utilized to further study how these networks form and interact with other tissues, such as skin or skeletal muscle. A 3D tissue system is developed to model neurogenesis and understand the development, maturation, and repair of neural networks. Neural networks are patterned on collagen gels and align along channel dimensions from neurotrophic cues embedded within the gel. A laceration injury model is described to study mechanisms involved in neural repair and regeneration. [ABSTRACT FROM AUTHOR]