Lithographically Defined Macroscale Modulation of Lateral Fluidity and Phase Separation Realized via Patterned Nanoporous Silica-Supported Phospholipid Bilayers

Abstract

Using lithographically defined surfaces consisting of hydrophilic patterns of nanoporous and nonporous (bulk) amorphous silica, we show that fusion of small, unilamellar lipid vesicles produces a single, contiguous, fluid bilayer phase experiencing a predetermined pattern of interfacial interactions. Although long-range lateral fluidity of the bilayer, characterized by fluorescence recovery after photobleaching, indicates a nominally single average diffusion constant, fluorescence microscopy-based measurements of temperature-dependent onset of fluidity reveals a locally enhanced fluidity for bilayer regions supported on nanoporous silica in the vicinity of the fluid-gel transition temperature. Furthermore, thermally quenching lipid bilayers composed of a binary lipid mixture below its apparent miscibility transition temperature induces qualitatively different lateral phase separation in each region of the supported bilayer: The nanoporous substrate produces large, microscopic domains (and domain-aggregates), whereas surface texture characterized by much smaller domains and devoid of any domain-aggregates appears on bulk glass-supported regions of the single-lipid bilayer. Interestingly, lateral distribution of the constituent molecules also reveals an enrichment of gel-phase lipids over nanoporous regions, presumably as a consequence of differential mobilities of constituent lipids across the topographic bulk/nanoporous boundary. Together, these results reveal that subtle local variations in constraints imposed at the bilayer interface, such as by spatial variations in roughness and substrate adhesion, can give rise to significant differences in macroscale biophysical properties of phospholipid bilayers even within a single, contiguous phase.

Nanoporous Silica-Based Protocells at Multiple Scales for Designs of Life and Nanomedicine

“…In particular, protocells were found to bind to hepatocellular carcinoma cells (HCCs) with low targeting-peptide densities while liposomes failed to do so, because the enhanced fluidity of lipid bilayers on the protocell facilitated multivalent peptide binding to the cell surface [ 58 ]. This unique advantage of the protocells is critical to enhance specific affinity of the targeting peptide and guide receptor-mediated endocytosis of protocells.…”

Micropatterned model membrane with quantitatively controlled separation of lipid phases

“…It is in contrast with other approaches which generally utilize the interactions of the membrane with the substrate surface for patterning L o and L d phases. [11][12][13][14][15][16] The fact that the present approach does not rely on the interaction with the substrate should allow us to construct a model membrane on a wider variety of substrates. In the future, it may be possible to detach the membrane from the substrate with a hydrophilic polymer cushion and suspend it in a similar fashion as black lipid membranes.…”

“…11 Some other studies have also exploited the different bending energies of L o and L d phases to realize patterned accumulation. [12][13][14] Alternatively, some studies utilized kinetic effects to realize a patterned phase separation by using photolithography and micro-uidics. 15,16 Patterned L o /L d phases with controlled size and spatial distribution would provide a model membrane for systematic in vitro parallel assays of the lipid-ra-related functions.…”

“…4 In the case of SPBs, micropatterning techniques have been applied to generate arrayed patches of L o and L d phases in the model membranes. [11][12][13][14][15][16] For example, Yoon et al accumulated L o and L d phases on a silicon substrate by locally modulating the surface curvatures. L o and L d phases were enriched on the at and corrugated surfaces, respectively, due to the difference in bending energy.…”

Bilayer membrane interactions with nanofabricated scaffolds

“…This issue is being addressed by many groups by investigating lateral diffusion in lipid bilayers supported on high-curvature micro/nanoporous silica substrates. 81,82 Spherically supported bilayer lipid (SSBL) membranes consist of silica beads ranging in diameter from about 50 nm to about 80 m, which support a fluid lipid bilayer membrane in the same manner as traditional supported lipid bilayer membranes on flat hydrophilic substrates, but on supports with radii of curvature in the size range from tens of nanometers, corresponding to small unilamellar vesicles (SUVs), to tens of micrometers, corresponding to cells and giant unilamellar vesicles (GUVs). 83 In addition to interactions with micro/nanofabricated scaffolds that form corrals in the membrane, demixing involving phase separation of binary and ternary mixtures of lipids in the membrane itself can also induce lateral organization with the formation of microdomains.…”