Protocells Featuring Membrane-Bound and Dynamic Membraneless Organelles.

Living cells, especially eukaryotic ones, use multicompartmentalization to regulate intra- and extracellular activities, featuring membrane-bound and membraneless organelles. These structures govern numerous biological and chemical processes spatially and temporally. Synthetic cell models, primarily utilizing lipidic and polymeric vesicles, have been developed to carry out cascade reactions within their compartments. However, these reconstructions often segregate membrane-bound and membraneless organelles, neglecting their collaborative role in cellular regulation. To address this, we propose a structural design incorporating microfluidic-produced liposomes housing synthetic membrane-bound organelles made from self-assembled poly(ethylene glycol)-block-poly(trimethylene carbonate) nanovesicles and synthetic membraneless organelles formed via temperature-sensitive elastin-like polypeptide phase separation. This architecture mirrors natural cellular organization, facilitating a detailed examination of the interactions for a comprehensive understanding of cellular dynamics.

Controlling polymersome size through microfluidic-assisted self-assembly: Enabling 'ready to use' formulations for biological applications.

The self-assembly of poly(ethylene glycol)-block-poly(trimethylene carbonate) PEG-b-PTMC copolymers into vesicles, also referred as polymersomes, was evaluated by solvent displacement using microfluidic systems. Two microfluidic chips with different flow regimes (micromixer and Herringbone) were used and the impact of process conditions on vesicle formation was evaluated. As polymersomes are sensitive to osmotic variations, their preparation under conditions allowing their direct use in biological medium is of major importance. We therefore developed a solvent exchange approach from DMSO (Dimethylsulfoxide) to aqueous media with an osmolarity of 300 mOsm L-1, allowing their direct use for biological evaluation. We evidenced that the organic/aqueous solvent ratio does not impact vesicle size, but the total flow rate and copolymer concentration have been observed to influence the size of polymersomes. Finally, nanoparticles with diameters ranging from 76 nm to 224 nm were confirmed to be vesicles through the use of multi-angle light scattering in combination with cryo-TEM (Cryo-Transmission Electron Microscopy) characterization.