Micropatterned composite membranes of polymerized and fluid lipid bilayers

Special Division for Human Life Technology, National Institute of Advanced Industrial Science and Technology (AIST), Ikeda 563-8577, Japan.
Langmuir (Impact Factor: 4.38). 09/2004; 20(18):7729-35. DOI: 10.1021/la049340e
Source: PubMed

ABSTRACT Micropatterned composite membranes of polymerized and fluid lipid bilayers were constructed on solid substrates. Lithographic photopolymerization of a diacetylene-containing phospholipid, 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine (DiynePC), and subsequent removal of nonreacted monomers by a detergent solution (0.1 M sodium dodecyl sulfate (SDS)) yielded a patterned polymeric bilayer matrix on the substrate. Fluid lipid bilayers of phosphatidylcholine from egg yolk (egg-PC) were incorporated into the lipid-free wells surrounded by the polymeric bilayers through the process of fusion and reorganization of suspended small unilamellar vesicles. Spatial distribution of the fluid bilayers in the patterned bilayer depended on the degree of photopolymerization that in turn could be modulated by varying the applied UV irradiation dose. The polymeric bilayer domains blocked lateral diffusion of the fluid lipid bilayers and confined them in the defined areas (corrals), if the polymerization was conducted with a sufficiently large UV dose. On the other hand, lipid molecules of the fluid bilayers penetrated into the polymeric bilayer domains, if the UV dose was relatively small. A direct correlation was observed between the applied UV dose and the lateral diffusion coefficient of fluorescent marker molecules in the fluid bilayers embedded within the polymeric bilayer domains. Artificial control of lateral diffusion by polymeric bilayers may lead to the creation of complex and versatile biomimetic model membrane arrays.

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    ABSTRACT: Supported lipid bilayer (SLB) platforms have been developed to transport and separate membrane-embedded species in the species' native bilayer environment. In this study, we used the phase segregation phenomenon of lipid mixtures containing a polymerizable diacetylene phospholipid, 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine (DiynePC), and a nonpolymerizable phospholipid, 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), to create filter barrier structures in SLBs. Upon exposing the phase segregated samples to UV light, the DiynePC-rich domains could become crosslinked and remain fixed on the surface of the support, while the DOPC-rich regions, where no crosslinking could happen, could be removed later by detergent washing, and thus became the void regions in the filter. During the filter fabrication process, we used the laminar flow configuration in a microfluidic channel to control the spatial locations of the feed region and filter region in the SLB. The flow in a microfluidic channel was also used to apply a strong hydrodynamic shear stress to the SLB to transport the membrane-embedded species from the feed region to the filter region. We varied the DiynePC/DOPC molar ratio from 60/40 to 80/20 to adjust the cutoff size of the filter barriers and used two model membrane-embedded species of different sizes to examine the filtering capability. One of the model species, Texas Red 1,2-dihexa-decanoyl-sn-glycero-3-phosphoethanolamine triethylammonium salt (Texas Red DHPE), had a single-lipid size, and the other species, cholera toxin subunit B-GM1 complex, had a multilipid size. When the DiynePC/DOPC molar ratio was 60/40, both species had high penetration ratios in the filter region. However, when the ratio was increased to 70/30, only the Texas Red DHPE, which was the smaller of the two model species, could penetrate the filter to a considerable extent. When the ratio was increased to 80/20, neither of the model species could penetrate the filter region. The results showed the possibility of using phase segregation of a mixture containing a polymerizable lipid and a nonpolymerizable lipid to fabricate filter barrier structures with tunable cutoff sizes in SLBs.
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    ABSTRACT: The localization of lipids and proteins in microdomains (lipid rafts) is believed to play important functional roles in the biological membrane. Herein, we report on a micropatterned model membrane that mimics lipid rafts by quantitatively controlling the spatial distribution of lipid phases. We generated a composite membrane of polymeric and fluid lipid bilayers by lithographic polymerization of diacetylene phospholipid(1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine: DiynePC). The composite membrane comprised polymer free-region (R0), partially polymerized region (R1), and fully polymerized region (R2). As a ternary mixture of saturated lipid, unsaturated lipid, and cholesterol was introduced into the voids between polymeric bilayers, liquid-ordered (Lo) and liquid-disordered (Ld) lipid phases were accumulated in R0 and R1, respectively. Local enrichment of Ld phase in R1 (and Lo phase in R0) was enhanced with a heightened coverage of polymeric bilayer in R1, supporting the premise that polymeric bilayer domains are inducing the phase separation. The pattern geometry (the area fractions of R0 and R1) also affected the enrichment due to the balance of gross Lo/Ld area fractions. Therefore, we could control the local Lo/Ld ratios by modulating the pattern geometry and polymer coverage in R1. Micropatterned model membrane with quantitatively controlled distribution of Lo/Ld phases offers a new tool to study the functional roles of lipid rafts by enabling to separate membrane-bound molecules according to their affinities to Lo and Ld phases.
    RSC Advances 01/2015; 5(2):1507-1513. DOI:10.1039/C4RA09981H · 3.71 Impact Factor