Polymer Hydrogel Capsules Research Articles

Shape-Dependent Cellular Processing of Polyelectrolyte Capsules

Particle shape is emerging as a key design parameter for tailoring the interactions between particles and cells. Herein, we report the preparation of rod-shaped layer-by-layer (LbL)-assembled polymer hydrogel capsules with tunable aspect ratios (ARs). By templating spherical and rodlike silica particles, disulfide-stabilized poly(methacrylic acid) hydrogel capsules (PMA HCs) with different ARs (from 1 to 4) are generated. The influence of capsule AR on cellular internalization and intracellular fate was quantitatively investigated by flow cytometry, imaging flow cytometry, and fluorescence deconvolution microscopy. These experiments reveal that the cellular internalization kinetics of PMA HCs are dependent on the AR, with spherical capsules being internalized more rapidly and to a greater extent compared with rod-shaped capsules. In contrast, the capsules with different ARs are colocalized with the lysosomal marker LAMP1, suggesting that the lysosomal compartmentalization is independent of shape for these soft polymer capsules.

Biodistribution of polymer hydrogel capsules for the delivery of therapeutics

A key phase in the development of intelligently designed nanoparticle delivery vehicles for new therapeutic agents is to gain an understanding of their interaction with tissues and cells. We report a series of in vitro and in vivo experiments aimed at tracking a potential delivery vehicle for therapeutic agents, including vaccine peptides and drugs derived from poly(methacrylic acid) hydrogel capsules in certain organs and cell types. For the in vitro studies, two immortal liver-derived cell lines (Huh7 and Hepa1-6) and primary cultures of mouse hepatocytes were incubated with Alexa 647 labelled fluorescent capsules to track their internalization and intracellular distribution by confocal microscopy. Capsules, 500nm in diameter, were taken up into the cells in a time-dependent manner in all three cell lines. Capsules were observed in plasma membrane-derived vesicles within the cells. After 24h a significant proportion of the capsules was observed in lysosomes. To understand the behaviour of the capsules in vivo, Alexa 488 labelled fluorescent capsules were intravenously injected into Sprague–Dawley rats and after 24h the fate of the capsules in a number of organs was determined by flow cytometry and confocal microscopy. By flow cytometry, the majority of the capsules were detected in the spleen whilst similar numbers were found in the lung and liver. By confocal microscopy, the majority of the capsules were found in the liver and spleen with significantly less capsules in the lung, heart and kidney. Colocalization of capsules with cell-type specific markers indicated that in lung, heart and kidney, the majority of the capsules were located in endothelial cells. In the spleen ∼50% of the capsules were found in CD163-positive cells, whereas in the liver, almost all capsules were located in CD163-positive cells, indicating uptake by Kupffer cells. Electron microscopy confirmed the presence of capsules within Kupffer cells.

Macromolecule Functionalization of Disulfide-Bonded Polymer Hydrogel Capsules and Cancer Cell Targeting

We present a generic and versatile method for functionalization of disulfide-stabilized PMA hydrogel capsules (HCs) with macromolecules, including a number of specific antibodies to cancer cells. Functionalization was achieved by reversible addition-fragmentation chain transfer (RAFT) polymerization of poly(N-vinyl pyrrolidone) (PVPON), which introduced biorelevant heterotelechelic end groups (thiol and amine) to the polymer chain. The PVPON with heterotelechelic end groups was conjugated to the outermost layer of PMA HCs through the thiol groups and reacted with biotin via the amine groups to generate PMA/PVPON(biotin) HCs. On the basis of the high specific interaction and high affinity between biotin and avidin, and its derivates, such as NeutrAvidin (NAv), we functionalized the PMA HCs with biotinylated antibodies. We demonstrate significantly enhanced cellular binding and internalization of the antibody (Ab)-functionalized capsules compared with control human immunoglobulin (IgG)-functionalized capsules, suggesting these capsules can specifically interact with cells through antibody/antigen recognition. We anticipate that the versatility of the functionalization approach reported in this study will assist in targeted therapeutic delivery applications.

Tuning the Permeability of Polymer Hydrogel Capsules: An Investigation of Cross-Linking Density, Membrane Thickness, and Cross-Linkers

Nanoengineered poly(methacrylic acid) hydrogel capsules (PMA HCs) are promising candidate carriers for biomedical applications, especially in the areas of drug delivery, encapsulated catalysis, and cell mimicry. The assembly, stability, and degradation of these carriers, as well as their use for the encapsulation of therapeutics, have received considerable attention. However, tailoring the permeability properties of PMA HCs to various types of cargo remains largely unexplored. Herein, we investigate fundamental parameters that govern the structural integrity and the capability of PMA HCs to encapsulate macromolecular cargo. The thiol content of the constituent polymers and the number of deposited polymer layers are shown to be key factors in controlling cargo retention within the PMA HCs. We further introduce a new strategy to achieve disulfide cross-linking for PMA HCs via a thiol-disulfide exchange in order to obtain capsules with superior cargo retention characteristics. Finally, we provide evidence for the semipermeable nature of PMA HCs based on the charge of the solutes and demonstrate that rational design of these systems can yield capsules with specific cargo retention properties. This work contributes toward the development of multilayered polymer capsules and PMA HCs and associated applications in biomedicine.

Degradation of liposomal subcompartments in PEGylated capsosomes

Capsosomes, polymer hydrogel capsules containing intact liposomal subcompartments, represent a promising platform toward the assembly of cell-like systems. Compartmentalized assembly plays a key role in the creation of therapeutic artificial cells, which focuses on enzymatic activities to degrade waste products or to support the synthesis of biomolecules. A fundamental aspect that governs the success of continuous enzymatic reactions in capsosomes is the long-term stability of the liposomal subunits within the polymer hydrogel capsules at physiological conditions, as they are subject to degradation in the presence of lipases. In this study, the outer membrane of the capsosomes was PEGylated by immobilizing graft copolymers of poly(methacrylic acid) (PMA) and poly(ethylene glycol) (PEG), and the fouling characteristics of this carrier system were investigated by incubation with bovine serum albumin. The influence of PEGylation on the stability of the liposomal subcompartments in the presence of phospholipases in different media was assessed. Diffusion of lipases across the multilayered membrane of the carrier capsules was hindered when the capsules were coated with PEG molecules. Furthermore, the rate of degradation of the liposomal subunits in PEGylated capsosomes was found to be three times higher when the subcompartments were “free-floating” (i.e., not membrane associated) in the capsules. The outlined surface modification of capsosomes contributes to the development of these systems toward functional therapeutic artificial cells.

Noncovalent Liposome Linkage and Miniaturization of Capsosomes for Drug Delivery

We report the synthesis of poly(methacrylic acid)-co-(oleyl methacrylate) with three different amounts of oleyl methacrylate and compare the ability of these polymers with that of poly(methacrylic acid)-co-(cholesteryl methacrylate) (PMA(c)) to noncovalently anchor liposomes to polymer layers. We subsequently assembled ∼1 μm diameter PMA(c)-based capsosomes, polymer hydrogel capsules that contain up to ∼2000 liposomal subcompartments, and investigate the potential of these carriers to deliver water-insoluble drugs by encapsulating two different antitumor compounds, thiocoraline or paclitaxel, into the liposomes. The viability of lung cancer cells is used to substantiate the cargo concentration-dependent activity of the capsosomes. These findings cover several crucial aspects for the application of capsosomes as potential drug delivery vehicles.

Cytotoxicity and Internalization of Polymer Hydrogel Capsules by Mammalian Cells

Polymer hydrogel capsules comprised of poly(methacrylic acid) chains and cross-linked via disulfide linkages were investigated for their cytotoxicity and mechanism of internalization in a variety of mammalian cells. The capsules were internalized by all the tested cell lines which differed in their morphology and function and over short to medium term (24 h) revealed no reduction in viability and metabolic activity of cells. The mechanism of capsule uptake was analyzed using inhibitors of various cellular entry pathways. Of these, blocking the clathrin-mediated endocytotic pathway resulted in a statistically significant reduction in capsule uptake, suggesting this was the predominant pathway of capsule entry in these cell lines. The uptake of solid particles with similar surface chemistry was not significantly decreased by the inhibitor of the clathrin-mediated pathway, which suggested that softness and concomitant flexibility of the hydrogel capsules were factors governing the entry mechanism. This work represents the first systematic study of the interaction of polymer hydrogel capsules with mammalian cells and provides essential information for the application of these capsules in biomedicine.

Subcompartmentalized Polymer Hydrogel Capsules with Selectively Degradable Carriers and Subunits

Subcompartmentalized hydrogel capsules (SHCs) with selectively degradable carriers and subunits are designed for potential applications in drug delivery and microencapsulated biocatalysis. Thiolated poly(methacrylic acid) and poly(N-vinyl pyrrolidone) are used to assemble 3-microm-diameter carrier capsules and 300-nm-diameter subunits, independently stabilized by a diverse range of covalent linkages. This paper presents examples of SHCs with tens of subcompartments and their successful drug loading, as well as selective degradation of the SHC carrier and/or subunits in response to multiple chemical stimuli.

Uptake and Intracellular Fate of Disulfide-Bonded Polymer Hydrogel Capsules for Doxorubicin Delivery to Colorectal Cancer Cells

Understanding the interactions between drug carriers and cells is of importance to enhance the delivery of therapeutics. The release of therapeutics into different intracellular environments, such as the lysosomes or the cell cytoplasm, will impact their pharmacological activity. Herein, we investigate the intracellular fate of layer-by-layer (LbL)-assembled, submicrometer-sized polymer hydrogel capsules in a human colon cancer derived cell line, LIM1899. The cellular uptake of the disulfide-stabilized poly(methacrylic acid) (PMA(SH)) capsules by colon cancer cells is a time-dependent process. Confocal laser scanning microscopy and transmission electron microscopy reveal that the internalized capsules are deformed in membrane-enclosed compartments, which further mature to late endosomes or lysosomes. We further demonstrate the utility of these redox-responsive PMA(SH) capsules for the delivery of doxorubicin (DOX) to colon cancer cells. The DOX-loaded PMA(SH) capsules demonstrate a 5000-fold enhanced cytotoxicity in cell viability studies compared to free DOX.

Tuning the Formation and Degradation of Layer-by-Layer Assembled Polymer Hydrogel Microcapsules

Engineered polymer capsules are finding widespread importance in the delivery of encapsulated toxic or fragile drugs. The effectiveness of polymer capsules as therapeutic delivery vehicles is often dependent on the degradation behavior of the capsules because it is often necessary to release the encapsulated drugs at specific times and in certain locations. Herein we investigate the parameters that govern the formation and degradation of a recently introduced new class of polymer hydrogel capsules based on disulfide cross-linked poly(methacrylic acid). We report a new and efficient method for the synthesis of thiol-functionalized poly(methacrylic acid) (PMA(SH)), the main component of the capsules. Polymeric capsules were synthesized by the layer-by-layer deposition of PMA(SH) and poly(vinylpyrrolidone) (PVPON) on silica particle templates, followed by cross-linking the PMA(SH) layers and removing PVPON and the template particles. The disulfide cross-links provided a redox-active trigger for degradation that was initiated by a cellular concentration of glutathione. We demonstrate that increasing the degree of PMA(SH) thiol modification affords direct control over the thickness of the polymer film and the degradation rate of the polymer capsules. Furthermore, the degradation rate of the PMA(SH) capsules was independent of film thickness, suggesting a bulk erosion process.

Polymer hydrogel capsules: en route toward synthetic cellular systems

Engineered synthetic cellular systems are expected to become a powerful biomedical platform for the development of next-generation therapeutic carrier vehicles. In this mini-review, we discuss the potential of polymer capsules derived by the layer-by-layer assembly as a platform system for the construction of artificial cells and organelles. We outline the characteristics of polymer capsules that make them unique for these applications, and we describe several successful examples of microencapsulated catalysis, including biologically relevant enzymatic reactions. We also provide examples of subcompartmentalized polymer capsules, which represent a major step toward the creation of synthetic cells.