Pancreatic Islet Survival and Engraftment Is Promoted by Culture on Functionalized Spider Silk Matrices

Abstract

Transplantation of pancreatic islets is one approach for treatment of diabetes, however, hampered by the low availability of viable islets. Islet isolation leads to disruption of the environment surrounding the endocrine cells, which contributes to eventual cell death. The reestablishment of this environment is vital, why we herein investigated the possibility of using recombinant spider silk to support islets in vitro after isolation. The spider silk protein 4RepCT was formulated into three different formats; 2D-film, fiber mesh and 3D-foam, in order to provide a matrix that can give the islets physical support in vitro. Moreover, cell-binding motifs from laminin were incorporated into the silk protein in order to create matrices that mimic the natural cell environment. Pancreatic mouse islets were thoroughly analyzed for adherence, necrosis and function after in vitro maintenance on the silk matrices. To investigate their suitability for transplantation, we utilized an eye model which allows in vivo imaging of engraftment. Interestingly, islets that had been maintained on silk foam during in vitro culture showed improved revascularization. This coincided with the observation of preserved islet architecture with endothelial cells present after in vitro culture on silk foam. Selected matrices were further evaluated for long-term preservation of human islets. Matrices with the cell-binding motif RGD improved human islet maintenance (from 36% to 79%) with preserved islets architecture and function for over 3 months in vitro. The islets established cell-matrix contacts and formed vessel-like structures along the silk. Moreover, RGD matrices promoted formation of new, insulin-positive islet-like clusters that were connected to the original islets via endothelial cells. On silk matrices with islets from younger donors (<35 year), the amount of newly formed islet-like clusters found after 1 month in culture were almost double compared to the initial number of islets added.

Survival of encapsulated islets: More than a membrane story

“…Some groups have added RGD or IKVAV (another integrin binding epitope) to the intracapsular environment and demonstrated improved viability and functionality under culture Barkai U et al . Survival of encapsulated islets conditions (for examples, see [209,210] ) and in animal models [211] . However, ECM molecules may be necessary for additional processes contributing to prevention of anoikis and prolonging survival of islet cells as they are anchoring sites for many essential growth factors.…”

Improving gelation efficiency and cytocompatibility of visible light polymerized thiol-norbornene hydrogels via addition of soluble tyrosine

“…In this experiment, we utilized non-degradable 8-arm PEGNB (PEG8aNB, 20 kDa) as the base macromer to afford hydrolytically stable hydrogels throughout the course of study ( Figure S4). In addition, we synthesized a laminin-derived peptide flanking with terminal cysteine residues (i.e., KCYIGSRCK, terminal lysine residues added to increase peptide solubility) as the hydrogel crosslinker to provide biomimicry to the otherwise inert PEG-based hydrogels [56][57][58]. Since the addition of soluble tyrosine during thiol-norbornene photopolymerization increased stiffness of the resulting gels, which could be a confounding factor when evaluating cellular fate, we formulated the cellladen hydrogels such that the elastic moduli of all hydrogels were similar ( Figure S4, ~2 kPa).…”

Biomedical Applications of Recombinant Silk‐Based Materials

“…The islets were evaluated for cell viability and attachment, as well as insulin production and the amount of intracellular calcium. The best performance was on RGD functionalized 4RepCT assembled into foams, and the details of these results are therefore described in the foams section of this review …”