While ligand-functionalized polymer gels of adjustable stiffness have been widely used to study cellular mechanosensory in an extracellular matrix-like environment, less is known about cellular mechanosensing in multicellular systems, considered to be important in processes, such as wound healing and tumor-to-metastatic transition of cancer cells. This shortcoming can be attributed, at least in part, to the lack of suitable cell surface mimetics of adjustable viscoelasticity. Polymer-tethered lipid membranes comprised of phospholipids and lipopolymers were previously introduced as potentially promising cell surface mimetics that allow spreading and migration of adherend cells. However, their substrate mechanical properties remained ill-defined, thus making systematic studies of cellular mechanosensitivity challenging. To overcome this limitation, here we describe the assembly and characterization of a polymer gel-tethered lipid bilayer (PGTB), in which a lipid bilayer containing about 5 mol% of SH-lipids (DPTE) is added to the surface of a maleimide-functionalized polyacrylamide (PAA) gel of well-defined stiffness. Specifically, two assembly strategies are described. In the first case, a glass slide with a polymer gel-tethered lipid monolayer is brought in contact with another glass slide containing a lipid monolayer, resulting in the formation of the PGTB sandwiched between two glass slides. In the second case, the lipid bilayer is attached to the surface of the functionalized polymer gel via fusion of giant unilamellar vesicles (GUVs). In both cases, formation of the lipid bilayer on the polymer gel surface was confirmed using epifluorescence microscopy, fluorescence recovery after photobleaching (FRAP), and fluorescence correlation spectroscopy (FCS).

Assembly and characterization of polymer gel-tethered lipid bilayer (PGTB)

ACS Spring 2022

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