Silk fibroin, derived from Bombyx mori cocoons, is a widely used and studied protein polymer for biomaterial applications. Silk fibroin has remarkable mechanical properties when formed into different materials, demonstrates biocompatibility, has controllable degradation rates from hours to years, and it can be chemically modified to alter surface properties or to immobilize growth factors. A variety of aqueous or organic solvent processing methods can be used to generate silk biomaterials for a range of applications. In this protocol we include methods to extract silk from B. mori cocoons in order to fabricate hydrogels, tubes, sponges, composites, fibers, microspheres and thin films. These materials can be used directly as biomaterials for implants, as scaffolding in tissue engineering and in vitro disease models, and for drug delivery.

Tissue engineering is currently limited by the inability to adequately vascularize tissues in vitro or in vivo. Issues of nutrient perfusion and mass transport limitations, especially oxygen diffusion, restrict construct development to smaller than clinically relevant dimensions and limit the ability for in vivo integration. There is much interest in the field as researchers have undertaken a variety of approaches to vascularization, including material functionalization, scaffold design, microfabrication, bioreactor development, endothelial cell seeding, modular assembly, and in vivo systems. Efforts to model and measure oxygen diffusion and consumption within these engineered tissues have sought to quantitatively assess and improve these design strategies. This review assesses the current state of the field by outlining the prevailing approaches taken toward producing vascularized tissues and highlighting their strengths and weaknesses.

Silk presents a rare combination of desirable properties for sustained drug delivery, including aqueous-based purification and processing options without chemical cross-linkers, compatibility with common sterilization methods, controllable and surface-mediated biodegradation into non-inflammatory by-products, biocompatibility, utility in drug stabilization, and robust mechanical properties. A versatile silk-based toolkit is currently available for sustained drug delivery formulations of small molecule through macromolecular drugs, with a promise to mitigate several drawbacks associated with other degradable sustained delivery technologies in the market. Silk-based formulations utilize silk’s well-defined nano- through microscale structural hierarchy, stimuli-responsive self-assembly pathways and crystal polymorphism, as well as sequence and genetic modification options towards targeted pharmaceutical outcomes. Furthermore, by manipulating the interactions between silk and drug molecules, near-zero order sustained release may be achieved through diffusion- and degradation-based release mechanisms. Because of these desirable properties, there has been increasing industrial interest in silk-based drug delivery systems currently at various stages of the developmental pipeline from pre-clinical to FDA-approved products. Here, we discuss the unique aspects of silk technology as a sustained drug delivery platform and highlight the current state of the art in silk-based drug delivery. We also offer a potential early development pathway for silk-based sustained delivery products.

Currently available synthetic grafts demonstrate moderate success at the macrovascular level, but fail at the microvascular scale (<6mm inner diameter). We report on the development of silk fibroin microtubes for blood vessel repair with several advantages over existing scaffold materials/designs. These microtubes were prepared by dipping straight lengths of stainless steel wire into aqueous silk fibroin, where the addition of poly(ethylene oxide) (PEO) enabled control of microtube porosity. The microtube properties were characterized in terms of pore size, burst strength, protein permeability, enzymatic degradation, and cell migration. Low porosity microtubes demonstrated superior mechanical properties in terms of higher burst pressures, but displayed poor protein permeability; whereas higher porosity tubes had lower burst strengths but increased permeability and enhanced protein transport. The microtubes also exhibited cellular barrier functions as low porosity tubes prevented outward migration of GFP-transduced HUVECs, while the high porosity microtubes allowed a few cells per tube to migrate outward during perfusion. When combined with the biocompatible and suturability features of silk fibroin, these results suggest that silk microtubes, either implanted directly or preseeded with cells, are an attractive biomaterial for microvascular grafts.

Vascular surgeries such as coronary artery bypass require small diameter vascular grafts with properties that are not available at this time. Approaches using synthetic biomaterials have been not completely successful in producing non-thrombogenic grafts with inner diameters less than 6 mm, and there is a need for new biomaterials and graft designs. We propose silk fibroin as a microvascular graft material and describe tubular silk scaffolds that demonstrate improved properties over existing vascular graft materials. Silk tubes produced using an aqueous gel spinning technique were first assessed in vitro in terms of thrombogenicity (thrombin and fibrinogen adsorption, platelet adhesion) and vascular cell responses (endothelial and smooth muscle cell attachment and proliferation) in comparison with polytetrafluoroethylene (PTFE), a synthetic material most frequently used for vascular grafts. Silk tubes were then implanted into the abdominal aortas of Sprague-Dawley rats. At time points of 2 weeks and 4 weeks post implantation, tissue outcomes were assessed through gross observation (acute thrombosis, patency) and histological staining (H&E, Factor VIII, smooth muscle actin). Over the 4-week time period, we observed graft patency and endothelial cell lining of the lumen surfaces. These results demonstrate the feasibility of using silk fibroin as a vascular graft material and some advantages of silk tubes over the currently used synthetic grafts.