14759
2005, 109, 14759-14763 Published on Web 07/20/2005
Controlled Hydrogel Formation in the Internal Compartment of Giant Unilamellar Vesicles Aldo Jesorka,* Martin Markstro1 m, Mattias Karlsson, and Owe Orwar Department of Chemistry and Bioscience, Chalmers UniVersity of Technology, SE-41296 Go¨teborg, Sweden ReceiVed: April 26, 2005; In Final Form: June 23, 2005
The introduction of poly(ethylene dioxythiophene) (PEDOT)/poly(styrene sulfonate) (PSS) polyelectrolyte into giant unilamellar phospholipid vesicles (GUVs) and cross-linking with Ca2+ ions to generate a hydrogel within the internal compartment are reported. The aqueous colloidal suspension of PEDOT with excess PSS was microinjected into the internal compartment of liposomes as well as networks of GUVs and lipid nanotubes. The subsequent introduction of calcium ions as cross-linking agent in order to induce hydrogel formation was achieved by three different methods: vesicle fusion, electroporation, and direct microinjection. Gel formation was probed by coinjection of fluorescent nanoparticles and tracking of Brownian motion. Particle mobility was shown to be distinctly reduced in the gel-filled vesicles. Diffusion constants for the particles were calculated from the projected movement of the particles and compared to particles in reference gels and solutions.
Giant unilamellar vesicles are flexible, spherical microcompartments made from surfactant materials. They are mostly composed of phospholipid membranes and can harbor volumes smaller than 10-12 L in an aqueous environment. The fluid character of the membrane gives the vesicles dynamic properties, meaning that introduction of liquid through the membrane can be easily achieved, for example, by electroinjection or vesicle fusion. The nature of the liquid content of each vesicle can therefore be freely chosen, provided that the filling solution does not disrupt, destabilize, or dissolve the membrane.1 Micro- and nanofluidic devices based on phospholipid membrane vesicles and interconnecting lipid nanotubes have been developed quite recently, and a number of systematic studies have been conducted (e.g., the controlled generation of the micrometer scale networks, their structure and topology as well as modification with biomacromolecules within the compartments, membranes, and nanotubes2). Previously, it has been shown that, with these vesicles and their artificially created networks of interconnecting nanotubes with a typical diameter of 100 nm, a variety of functional aspects can be studied (e.g., reaction kinetics in confined volumes3). Applications as a model for cellular functions4 and the interaction with functional membrane proteins5 were demonstrated. The biomimetic properties of the membrane also make the vesicles attractive as vessels for model systems in cellular biology (e.g., to study membrane transport phenomena6 or exo/endocytosis7). In comparison to a living cell, such a model system is, because of its lesser complexity and low sensitivity to external stress, far easier to create, to maintain, and to manipulate during an experiment. However, the lack of equivalents to cytoplasm and cytoskeleton within their primitive structures renders giant vesicles as model systems for chemical reaction kinetics, migration, or * Corresponding author. E-mail:
[email protected]. Fax: +4631-7722785. Telephone: +46-31-7723069.
10.1021/jp052162t CCC: $30.25
diffusion studies in the context of biological cells only valuable with limitations. Fundamental differences in kinetics in cellular environments are attributed to macromolecular crowding, resulting from hindered diffusion, size exclusion, and limited molecular motion in such systems.8,9 The effect of macromolecular crowding cannot be studied in such simple vesicular microreactors containing only buffer solution. A cell-sized membrane compartment featuring macromolecular crowding conditions would represent a novel, useful model for quantitative studies of diffusion-controlled kinetics in comparison to macroscopic chemical reactors. Modification of GUVs and vesicle/nanotube networks with a highly viscous or soft solid medium, which can be used to accurately mimic intracellular environments, is desirable. Presented here is a basic feasibility study with the focus on the combination of functional polymeric materials with fluid membrane devices, adding internal complexity to a structurally simple vesicular system. The goals of the study were the formation of a polymer-based, dense, and biocompatible hydrogel in single giant unilamellar vesicles and networks of such vesicles with nanotubes and the trapping of coinjected nanoparticles in the gel. One important rationale for the study is the development of methods for the stabilization of GUVs, aiming at a well-defined internal structure of significantly higher density than that of the buffer solution, practically the most basic feature of a biological cell. A second major point of interest in such a system is the possibility of controlled immobilization of chemical and biological material in the context of artificial cells.10,11 Both aspects are important prerequisites for the study of cellular kinetics in artificially created microreactors under simulated macromolecular crowding conditions. An attractive material for sol-gel transition studies is the aqueous suspension of the electrically conducting “synthetic metal” poly(ethylene dioxythiophene) (PEDOT) in a solution © 2005 American Chemical Society
14760 J. Phys. Chem. B, Vol. 109, No. 31, 2005
Letters
Figure 1. GUV after injection of PEDOT/PSS suspension (A) and after subsequent injection of CaCl2 solution (B). A deformed, gel-filled vesicle in (B) is suspended between the injection needle (left) and the carbon fiber counter electrode (right).
of excess poly(styrene sulfonate) (PSS). This polyelectrolyte has been reported to form hydrogels with high ion conductivity and an electron-conducting polymer backbone12 and found its first applications in the biosensor area.13 We established an experimental protocol to directly introduce PEDOT/PSS into giant unilamellar vesicles and to cross-link its main constituent, water-soluble PSS, to form a hydrogel in the internal compartment. Figure 1A shows a 50 µm vesicle microinjected with PEDOT suspension in PSS solution, containing up to 6 wt % of PEDOT with a ratio of PEDOT to PSS of 40:60 and a PEDOT particle size of
