Biosilk 521 is a 3D platform combining the biological instructive cues from laminins with mechanical and topological support from silk allowing engineered, controllable and tunable organoids

Background & Aim Tissues are 3D formations of cells integrated in an extracellular matrix (ECM) with specific sites for cell anchorage offering positional and instructive information regulating cell behavior. A vast number of publications in the last decades have proven the reciprocal interaction between the ECM, cytoskeleton and nuclear matrix, showing that these structures exert a physical and chemical influence over gene expression. Thus, to culture authentic cells from which safe and therapeutically relevant cells can be generated or real biological questions can be answered, environmental context is pivotal. Laminins comprise a family of 16 unique heterotrimeric glycoproteins naturally present in the body. Laminins are tissue specific, supply a natural environment for all cell types, and influence cell adhesion, differentiation, migration, phenotypic stability, and cell functionality. As such, recombinant laminins are biologically relevant ECM protein substrates that can be used to mimic the in vivo cell niche in vitro. The laminin 521 isoform is a critical factor supporting pluripotent stem cells in culture, and given the right context, this isoform also provides support for other differentiated cell types. Methods, Results & Conclusion Biosilk 521 is a biomaterial made of recombinant silk, which is functionalized with human recombinant laminin 521 protein (Biolaminin™ 521). This material supports integration and proliferation of human pluripotent stem cells in vitro and serves as a viable base for the development of subsequent lineage-specific differentiation in a 3D format. Human PSCs (2 hESC and 2 hiPSC lines) seeded in the Biosilk 521 scaffold integrate, migrate, and form small colonies at day 1 post seeding and retain pluripotency thereafter. Human PSCs in the Biosilk 521 scaffold could be cultured using different types of medium, express pluripotent markers, generate all three tissues germ layers upon spontaneous differentiation, and directly to different neural fates. Furthermore, the Biosilk 521 can be used for organoid culture, without the need of Matrigel. Forebrain, midbrain and glia organoids were started in a controlled manner by using the Biosilk 521 and correct regional specific markers, cell morphology and layers were present. In sum, Biosilk 521 is a xenofree and defined, biorelevant 3D system that is biodegradable and non-immunogenic, and thus ideal for cell culturing and many other biomedical applications.