Shear Stress and Its Effect on the Interaction of Myoblast Cells with Nanosized Drug Delivery Vehicles

An important aspect to ensure progress in biomedicine is the fundamental understanding of the interaction of cells and tissue with (bio)­materials. The consideration of shear stress in drug delivery and/or tissue engineering remains largely unexplored. To illustrate the fundamental relevance, we employ a microfluidic setup to evaluate the myoblast cell response to two prominent drug carrier systems, namely, liposomes and nanoparticles, in the presence of low shear stress. We show that positively charged carriers have an enhanced interaction with myoblast cells in the presence of shear stress. This effect can be translated into improved therapeutic response in terms of reduction in cell viability when delivering a cytotoxic compound or into a better translocation efficiency when using lipoplexes. Taken together, our fundamental findings open up new possibilities in tissue engineering and drug delivery by considering an additional parameter when delivering beneficial compounds

Confined Multiple Enzymatic (Cascade) Reactions within Poly(dopamine)-based Capsosomes

The design of compartmentalized carriers as artificial cells is envisioned to be an efficient tool with potential applications in the biomedical field. The advent of this area has witnessed the assembly of functional, bioinspired systems attempting to tackle challenges in cell mimicry by encapsulating multiple compartments and performing controlled encapsulated enzymatic catalysis. Although capsosomes, which consist of liposomes embedded within a polymeric carrier capsule, are among the most advanced systems, they are still amazingly simple in their functionality and cumbersome in their assembly. We report on capsosomes by embedding liposomes within a poly­(dopamine) (PDA) carrier shell created in a solution-based single-step procedure. We demonstrate for the first time the potential of PDA-based capsosomes to act as artificial cell mimics by performing a two-enzyme coupled reaction in parallel with a single-enzyme conversion by encapsulating three different enzymes into separated liposomal compartments. In the former case, the enzyme uricase converts uric acid into hydrogen peroxide, CO₂ and allantoin, followed by the reaction of hydrogen peroxide with the reagent Amplex Ultra Red in the presence of the enzyme horseradish peroxidase to generate the fluorescent product resorufin. The parallel enzymatic catalysis employs the enzyme ascorbate oxidase to convert ascorbic acid into 2-<i>L</i>-dehydroascorbic acid

Engineering Advanced Capsosomes: Maximizing the Number of Subcompartments, Cargo Retention, and Temperature-Triggered Reaction

Advanced mimics of cells require a large yet controllable number of subcompartments encapsulated within a scaffold, equipped with a trigger to initiate, terminate, and potentially restart an enzymatic reaction. Recently introduced capsosomes, polymer capsules containing thousands of liposomes, are a promising platform for the creation of artificial cells. Capsosomes are formed by sequentially layering liposomes and polymers onto particle templates, followed by removal of the template cores. Herein, we engineer advanced capsosomes and demonstrate the ability to control the number of subcompartments and hence the degree of cargo loading. To achieve this, we employ a range of polymer separation layers and liposomes to form functional capsosomes comprising multiple layers of enzyme-loaded liposomes. Differences in conversion rates of an enzymatic assay are used to verify that multilayers of intact enzyme-loaded liposomes are assembled within a polymer hydrogel capsule. The size-dependent retention of the cargo encapsulated within the liposomal subcompartments during capsosome assembly and its dependence on environmental pH changes are also examined. We further show that temperature can be used to trigger an enzymatic reaction at the phase transition temperature of the liposomal subcompartments, and that the encapsulated enzymes can be utilized repeatedly in several subsequent conversions. These engineered capsosomes with tailored properties present new opportunities <i>en route</i> to the development of functional artificial cells

One-Pot Green Synthesis of Biocompatible Graphene Quantum Dots and Their Cell Uptake Studies

Graphene-based quantum dots (GQDs) are attractive fluorophores due to their excellent photoluminescence properties, water solubility, low cost, and low toxicity. However, the lack of simple, efficient, and environmental-friendly synthesis methods often limits their biological applications. Herein, we explore a novel, one-pot, green synthesis approach for the fabrication of fluorescent GQDs without involving any harsh reagents. Graphene oxide is used as a precursor, and a 2 h hydrothermal synthesis is carried out with assistance of hydrogen peroxide; no further post purification steps are required. The effects of reaction conditions on the characteristics of GQDs are comprehensively investigated. The as-synthesized GQDs show a high photostability and excellent biocompatibility as revealed by cell viability assays for three different cell lines, namely, macrophages, endothelial cells, and a model cancer cell line. The detailed studies of cellular uptake mechanisms suggest that for all of the three cell lines the major internalization route for GQDs is caveolae-mediated endocytosis followed by clathrin-mediated endocytosis at a less extent. Our results demonstrate the great potential of the as-synthesized GQDs as fluorescent nanoprobes. The study also provides unique insight into the cell–GQDs interactions, which is highly valuable for bioimaging and other related applications such as diagnostics and drug delivery

Liposomes as Drug Deposits in Multilayered Polymer Films

The ex vivo growth of implantable hepatic or cardiac tissue remains a challenge and novel approaches are highly sought after. We report an approach to use liposomes embedded within multilayered films as drug deposits to deliver active cargo to adherent cells. We verify and characterize the assembly of poly­(l-lysine) (PLL)/alginate, PLL/poly­(l-glutamic acid), PLL/poly­(methacrylic acid) (PMA), and PLL/cholesterol-modified PMA (PMA_c) films, and assess the myoblast and hepatocyte adhesion to these coatings using different numbers of polyelectrolyte layers. The assembly of liposome-containing multilayered coatings is monitored by QCM-D, and the films are visualized using microscopy. The myoblast and hepatocyte adhesion to these films using PLL/PMA_c or poly­(styrenesulfonate) (PSS)/poly­(allyl amine hydrochloride) (PAH) as capping layers is evaluated. Finally, the uptake of fluorescent lipids from the surface by these cells is demonstrated and compared. The activity of this liposome-containing coating is confirmed for both cell lines by trapping the small cytotoxic compound thiocoraline within the liposomes. It is shown that the biological response depends on the number of capping layers, and is different for the two cell lines when the compound is delivered from the surface, while it is similar when administered from solution. Taken together, we demonstrate the potential of liposomes as drug deposits in multilayered films for surface-mediated drug delivery

Assembly of Poly(dopamine)/Poly(<i>N</i>‑isopropylacrylamide) Mixed Films and Their Temperature-Dependent Interaction with Proteins, Liposomes, and Cells

Many biomedical applications benefit from responsive polymer coatings. The properties of poly­(dopamine) (PDA) films can be affected by codepositing dopamine (DA) with the temperature-responsive polymer poly­(<i>N</i>-isopropylacrylamide) (pNiPAAm). We characterize the film assembly at 24 and 39 °C using DA and aminated or carboxylated pNiPAAm by a quartz crystal microbalance with dissipation monitoring (QCM-D), X-ray photoelectron spectroscopy, UV–vis, ellipsometry, and atomic force microscopy. It was found that pNiPAAm with both types of end groups are incorporated into the films. We then identified a temperature-dependent adsorption behavior of proteins and liposomes to these PDA and pNiPAAm containing coatings by QCM-D and optical microscopy. Finally, a difference in myoblast cell response was found when these cells were allowed to adhere to these coatings. Taken together, these fundamental findings considerably broaden the potential biomedical applications of PDA films due to the added temperature responsiveness