Manipulation of encapsulated artificial phospholipid membranes using sub-micellar lysolipid concentrations.

Droplet Interface Bilayers (DIBs) constitute a commonly used model of artificial membranes for synthetic biology research applications. However, their practical use is often limited by their requirement to be surrounded by oil. Here we demonstrate in-situ bilayer manipulation of submillimeter, hydrogel-encapsulated droplet interface bilayers (eDIBs). Monolithic, Cyclic Olefin Copolymer/Nylon 3D-printed microfluidic devices facilitated the eDIB formation through high-order emulsification. By exposing the eDIB capsules to varying lysophosphatidylcholine (LPC) concentrations, we investigated the interaction of lysolipids with three-dimensional DIB networks. Micellar LPC concentrations triggered the bursting of encapsulated droplet networks, while at lower concentrations the droplet network endured structural changes, precisely affecting the membrane dimensions. This chemically-mediated manipulation of enclosed, 3D-orchestrated membrane mimics, facilitates the exploration of readily accessible compartmentalized artificial cellular machinery. Collectively, the droplet-based construct can pose as a chemically responsive soft material for studying membrane mechanics, and drug delivery, by controlling the cargo release from artificial cell chassis.

Building programmable multicompartment artificial cells incorporating remotely activated protein channels using microfluidics and acoustic levitation.

Intracellular compartments are functional units that support the metabolism within living cells, through spatiotemporal regulation of chemical reactions and biological processes. Consequently, as a step forward in the bottom-up creation of artificial cells, building analogous intracellular architectures is essential for the expansion of cell-mimicking functionality. Herein, we report the development of a droplet laboratory platform to engineer complex emulsion-based, multicompartment artificial cells, using microfluidics and acoustic levitation. Such levitated models provide free-standing, dynamic, definable droplet networks for the compartmentalisation of chemical species. Equally, they can be remotely operated with pneumatic, heating, and magnetic elements for post-processing, including the incorporation of membrane proteins; alpha-hemolysin; and mechanosensitive channel of large-conductance. The assembly of droplet networks is three-dimensionally patterned with fluidic input configurations determining droplet contents and connectivity, whilst acoustic manipulation can be harnessed to reconfigure the droplet network in situ. The mechanosensitive channel can be repeatedly activated and deactivated in the levitated artificial cell by the application of acoustic and magnetic fields to modulate membrane tension on demand. This offers possibilities beyond one-time chemically mediated activation to provide repeated, non-contact, control of membrane protein function. Collectively, this expands our growing capability to program and operate increasingly sophisticated artificial cells as life-like materials.

Droplet Microfluidics for Tumor Drug-Related Studies and Programmable Artificial Cells.

Anticancer drug development is a crucial step toward cancer treatment, that requires realistic predictions of malignant tissue development and sophisticated drug delivery. Tumors often acquire drug resistance and drug efficacy, hence cannot be accurately predicted in 2D tumor cell cultures. On the other hand, 3D cultures, including multicellular tumor spheroids (MCTSs), mimic the in vivo cellular arrangement and provide robust platforms for drug testing when grown in hydrogels with characteristics similar to the living body. Microparticles and liposomes are considered smart drug delivery vehicles, are able to target cancerous tissue, and can release entrapped drugs on demand. Microfluidics serve as a high-throughput tool for reproducible, flexible, and automated production of droplet-based microscale constructs, tailored to the desired final application. In this review, it is described how natural hydrogels in combination with droplet microfluidics can generate MCTSs, and the use of microfluidics to produce tumor targeting microparticles and liposomes. One of the highlights of the review documents the use of the bottom-up construction methodologies of synthetic biology for the formation of artificial cellular assemblies, which may additionally incorporate both target cancer cells and prospective drug candidates, as an integrated "droplet incubator" drug assay platform.