Search

Physiologically relevant human skin models that include key skin cell types can be used for in vitro drug testing, skin pathology studies, or clinical applications such as skin grafts. However, there is still no golden standard for such a model. We investigated the potential of a recombinant functionalized spider silk protein, FN-silk, for the construction of a dermal, an epidermal, and a bilayered skin equivalent (BSE). Specifically, two formats of FN-silk (i.e. 3D network and nanomembrane) were evaluated. The 3D network was used as an elastic ECM-like support for the dermis, and the thin, permeable nanomembrane was used as a basement membrane to support the epidermal epithelium. Immunofluorescence microscopy and spatially resolved transcriptomics analysis demonstrated the secretion of key ECM components and the formation of microvascular-like structures. Furthermore, the epidermal layer exhibited clear stratification and the formation of a cornified layer, resulting in a tight physiologic epithelial barrier. Our findings indicate that the presented FN-silk-based skin models can be proposed as physiologically relevant standalone epidermal or dermal models, as well as a combined BSE., (Creative Commons Attribution license.)

Replicating tissue barriers is critical for generating relevant in vitro models for evaluating novel therapeutics. Today, this is commonly done using tissue culture inserts with a plastic membrane, which generates an apical and a basal side. Besides providing support for the cells, these membranes come far from emulating their native counterpart, the basement membrane, which is a nanofibrillar, protein-based matrix. In this work, we show a simple way to considerably improve the biological relevance of the tissue culture inserts by replacing the plastic membrane with one made from a pure recombinant functionalized spider silk protein. The silk membrane forms through self-assembly and will spontaneously adhere to a membrane-free tissue culture insert, where it can provide support for cells. Custom-designed tissue culture inserts can be printed using a standard 3D printer, following the instructions provided in the protocol, or commercial ones can be purchased and used instead. This protocol shows how the culture system with silk membranes in inserts is set up and, subsequently, how the same cell culturing techniques that are used with traditional, commercially available inserts can be implemented.

Traditional cancer models rely on 2D cell cultures or 3D spheroids, which fail to recapitulate cell-extracellular matrix (ECM) interactions, a key element of tumor development. Existing hydrogel-based 3D alternatives lack mechanical support for cell growth and often suffer from low reproducibility. Here we report a novel strategy to make 3D models of breast cancer using a tissue-like, well-defined network environment based on recombinant spider silk, functionalized with a cell adhesion motif from fibronectin (FN-silk). With this approach, the canonical cancer cells SK-BR-3, MCF-7, and MDA-MB-231, maintain their characteristic expression of markers (i.e., ERα, HER2, and PGR) while developing distinct morphology. Transcriptomic analyses demonstrate how culture in the FN-silk networks modulates the biological processes of cell adhesion and migration while affecting physiological events involved in malignancy, such as inflammation, remodeling of the ECM, and resistance to anticancer drugs. Finally, we show that integration in FN-silk networks promotes the viability of cells obtained from the superficial scraping of patients' breast tumors., Competing Interests: My Hedhammar has shares in Spiber Technologies AB, a company that aims to commercialize recombinant spider silk., (© 2023 The Authors. Bioengineering & Translational Medicine published by Wiley Periodicals LLC on behalf of American Institute of Chemical Engineers.)

This work describes the design and validation of a novel device, the High-Throughput Degradation Monitoring Device (HDD), for monitoring the degradation of 24 soft tissue samples over incubation periods of several days inside a cell culture incubator. The device quantifies sample degradation by monitoring its deformation induced by a static gravity load. Initial instrument design and experimental protocol development focused on quantifying cartilage degeneration. Characterization of measurement errors, caused mainly by thermal transients and by translating the instrument sensor, demonstrated that HDD can quantify sample degradation with <6 μm precision and <10 μm temperature-induced errors. HDD capabilities were evaluated in a pilot study that monitored the degradation of fresh ex vivo human cartilage samples by collagenase solutions over three days. HDD could robustly resolve the effects of collagenase concentration as small as 0.5 mg/ml. Careful sample preparation resulted in measurements that did not suffer from donor-to-donor variation (coefficient of variance <70%). Due to its unique combination of sample throughput, measurement precision, temporal sampling and experimental versality, HDD provides a novel biomechanics-based experimental platform for quantifying the effects of proteins (cytokines, growth factors, enzymes, antibodies) or small molecules on the degradation of soft tissues or tissue engineering constructs. Thereby, HDD can complement established tools and in vitro models in important applications including drug screening and biomaterial development., (Copyright © 2018 Elsevier Ltd. All rights reserved.)