Liposomes Embedded in Layer by Layer Constructs as Simplistic Exosome Transfer Model

Abstract Background. Exosomes are extracellular vesicles originating from the exfoliation of the cellular membrane. They are involved in cell-to-cell and cell-to-matrix signaling, exchange of bioactive molecules, tumorigenesis and metastasis, among others. To mitigate the limited understanding of exosome transfer phenomena, we developed a simplistic model that mimics exosomes and their interactions with cells and the extracellular matrix. The proposed model is a layer-by-layer (LbL) film built from the polycationic poly-L-lysine (PLL) and the glycosaminoglycan hyaluronic acid (HA) to provide ECM mimicry. Positively charged 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and N1,N1,N14,N14-tetramethyl-N1,N14-ditetradecyltetradecane-1,14-diaminium dibromide (GS14) liposomes were embedded in this construct to act as exosome analogs.Results. To simulate exosomes carrying substances, Nile Red was loaded as a model of lipophilic cargo molecules. The integration of each component was followed by quartz-crystal microbalance measurements, which confirmed the immobilization of intact liposomes on the underlying (PLL/HA)3 soft film. The release of Nile Red from liposomes either embedded in the LbL construct or exposed at its surface revealed a fast first order release. This system was validated as a model for exosome/cell interactions by incubation with breast cancer cells MDA-MB-231. We observed higher internalization for embedded liposomes when compared with surface-exposed ones, showcasing that the ECM mimic layers do not constitute a barrier to liposome/cell interactions but favor them.Conclusions. Our findings indicate that the developed model enhances the structural stability of liposomes and induces endocytosis from breast cancer cells. We envisage that the internalization can be tuned by exploring different levels of embedment to achieve a cellular uptake modulated in a spatiotemporal dependent manner. The versatility provided by the LbL technique will allow incorporating additional specialized biomaterials to better mimic the structure and composition of exosomes and their role in cell-to-cell communications.