Chemoselective liposome fusion for cell-surface and tissue engineering applications

Abstract

Proper cell-cell communication through physical contact is crucial for a range of fundamental biological processes including, cell proliferation, migration, differentiation, and apoptosis and for the correct function of organs and other multi-cellular tissues. The spatial and temporal arrangements of these cellular interactions in vivo are dynamic and lead to higher-order function that is extremely difficult to recapitulate in vitro. The development of 3-dimensional (3D), in vitro model systems to investigate these complex, in vivo interconnectivities would generate novel methods to study the biochemical signaling of these processes, as well as provide platforms for tissue engineering technologies. Herein, we develop and employ a strategy to induce specific and stable cell-cell contacts in 3D through chemoselective cell-surface engineering based on liposome delivery and fusion to display bio-orthogonal functional groups from cell membranes. This strategy uses liposome fusion for the delivery of ketone or oxyamine groups to different populations of cells for subsequent cell assembly via oxime ligation. We demonstrate how this method can be used for several applications including, the delivery of reagents to cells for fluorescent labeling, the formation of small, 3D spheroid cell assemblies, and the generation of large and dense, 3D multi-layered tissue-like structures. We were also able to create dynamic and switchable cell tissue assemblies through chemoselective conjugation and release chemistry. Cell membranes are decorated with a range of molecules that can be released in vitro for subsequent rounds of molecular conjugation and release. Each step to modify the cell surface: activation, conjugation, release, and regeneration can be monitored and modulated by non-invasive, label-free analytical techniques. Additionally, we also develop and demonstrate a novel liposome fusion based delivery strategy to incorporate a unique bio-orthogonal lipid that has the dual ability to serve as a receptor for chemoselective cell surface tailoring and as a reporter to track cell behavior