Silk nanocoatings of mammalian cells for cytoprotection against mechanical stress

Mammalian cells are widely used in biotechnology and regenerative medicine applications, including recombinant protein production, cell therapy, 3D bioprinting, and ex vivo engineering of tissues and organs. Unlike unicellular organisms, fungi, or plants, animal cells lack a protective cell wall, and therefore are more sensitive to processing conditions, particularly mechanical forces. Hydrodynamic forces during capillary flow can damage the plasma membrane and impact cell viability and functions, limiting the yields of protein production or the success of injection-based cell delivery and 3D bioprinting of artificial tissues and organs. Here, we present nanocoating individual murine fibroblasts as a model organism with silk-based artificial cell walls to protect against mechanical stress. Cells were coated with three bilayers of silk polyelectrolytes through layer-by-layer electrostatic deposition and subjected to mechanical stress by extrusion through needles with small inner diameters or shearing using a rheometer. The silk nanocoatings preserved membrane integrity, cell survival, and proliferation after exposure to stress in viscous polyethylene glycol solution, providing a useful strategy for cytoprotection during cell delivery and 3D bioprinting applications. Impact statement Use of mammalian cells is a necessity in the industrial production of biopharmaceuticals because of proper protein folding through chaperone systems and post-translational modifications such as glycosylation being essential for functional products. Moreover, transplantation of human cells holds great promise in regeneration of serious injuries in the bone, brain, or spinal cord as well as debilitating diseases such as stroke, diabetes, arterial dysfunctions, and cancers. Cell delivery by direct injection or within 3D-printed scaffolds with well-defined, patient-specific geometry and composition has potential in filling the gap of insufficient organ transplantation. Both the delivery of mammalian cells into a bioreactor through transfer pipes or cell extrusion through fine needles or nozzles for injection-based delivery or bioprinting, however, may expose the cells to extremes of mechanical stress that impair cell viability and reduce the yield of production or overall success of the transplantation therapy. The silk nanocoatings on individual cells demonstrated in this study acted as artificial cell walls for mammalian cells and provided protection against mechanical stress during capillary flow or exposure to a constant shear force and significantly limited the loss of membrane integrity and necrotic or apoptotic cell death. The safety and simplicity of the approach involving protein-based, biocompatible silk fibroin processed through organic solvent-free, nature-friendly modification pathways allows for rapid translation in biotechnology and regenerative medicine to improve cell delivery applications.

Automated summary

Mammalian cells lack a protective cell wall and are more sensitive to mechanical forces, making them vulnerable during processing. Nanocoating individual cells with silk-based artificial cell walls can protect against mechanical stress, preserving membrane integrity, cell survival, and proliferation during cell delivery and 3D bioprinting applications.