Development of a microfluidic device to produce asymmetric biomimetic liposomes

Abstract

The plasma membrane is a complex structure that plays a key role in the function of the cell and some proteins and also in the recognition of “allogenic” molecules. In plants, this structure, and more particularly its lipid fraction, is thought to be involved in the perception of elicitors such as surfactin, a molecule representing a potential alternative to conventional pesticides. However, the development of such alternatives requires understanding the biological mechanisms at the molecular level. As the plant plasma membrane is a very intricate structure with a huge molecular diversity, simplified biomimetic models such as liposomes have been developed to decipher the molecular mechanisms at stake. Still, to be correlated with the biological phenomena, these simplified biomimetic models have to take into account some important biological facts such as the asymmetric lipid composition between the two lipid leaflets composing the plasma membrane. The production of such asymmetric models represents a current challenge and has been hardly investigated yet. In this context, this master thesis aimed to use the promising microfluidic approach to conceive a device able to produce asymmetric biomimetic liposomes. To perform the liposome assembly, a microfluidic chip with two successive channel junctions was designed to form the two leaflets of the liposome one after the other. The channels following the second junction were then render hydrophilic with PVA coating. This modification is required to produce liposome and is the most critical step to build a functional microfluidic device. Two approaches were then considered to produce liposomes with the developed device: the spontaneous transfer across a laminar interface and the w/o/w double emulsion. With the experimental conditions chosen, only the second approach led to the production of liposomes with a size ranging from 92 to 111 μm that still contains some residual octanol. In this approach, W1 droplets formed at the first channel junction, where the inner aqueous phase containing lipids (W1) is sheared by the octanol phase, gave rise to the formation of liposomes at the second channel junction with a maximal success rate of 59%. This success rate is affected when varying the flow rate ratio between the W1 and the octanol phases, which impacts the W1 droplets size. The non-confinement of W1 droplet also impacted negatively the liposome formation. In addition, it was observed that the correct timing for the W1 droplet arrival at the second channel junction is a condition for a successful liposome formation. Even though the developed device still need to be improved and the liposome asymmetry has not been assessed yet, this work has put strong basis for the conception of a microfluidic device able to generate biomimetic liposomes.