Phospholipid-Block Copolymer Hybrid Vesicles with Lysosomal Escape Ability.

The success of nanoparticulate formulations in drug delivery depends on various aspects including their toxicity, internalization, and intracellular location. Vesicular assemblies consisting of phospholipids and amphiphilic block copolymers are an emerging platform, which combines the benefits from liposomes and polymersomes while overcoming their challenges. We report the synthesis of poly(cholesteryl methacrylate)- block-poly(2-(dimethylamino) ethyl methacrylate) (pCMA- b-pDMAEMA) block copolymers and their assembly with phospholipids into hybrid vesicles. Their geometry, their ζ-potential, and their ability to adsorb onto polymer-coated surfaces were assessed. Giant unilamellar vesicles were employed to confirm the presence of both the phospholipids and the block copolymer in the same membrane. Furthermore, the cytotoxicity of selected hybrid vesicles was determined in RAW 264.7 mouse macrophages, primary rat Kupffer cells, and human macrophages. The internalization and lysosomal escape ability of the hybrid vesicles were confirmed using RAW 264.7 mouse macrophages. Taken together, our findings illustrate that the reported hybrid vesicles are a promising complementary drug delivery platform for existing liposomes and polymersomes.

Platinum Nanoparticle-Based Microreactors as Support for Neuroblastoma Cells.

Excitotoxicity is a common phenomenon in several neurological diseases, associated with an impaired clearance of synaptically released glutamate, which leads to an overactivation of postsynaptic glutamate receptors. This will, in turn, start an intracellular cascade of neurotoxic events, which include exacerbated production of reactive oxygen species and ammonia toxicity. We report the assembly of microreactors equipped with platinum nanoparticles as artificial enzymes and polymer terminating layers including poly(dopamine). The biological response to these microreactors is assessed in human neuroblastoma cell culture. The microreactors' function to deplete hydrogen peroxide (H2O2) and ammonia is confirmed. While the proliferation of the cells depends on the number of microreactors present, no inherent toxicity is found. Furthermore, the microreactors are able to ameliorate the effects of excitotoxicity in cell culture by scavenging H2O2 and ammonia, thus having the potential to provide a therapeutic approach for several neurological diseases in which excitotoxicity is observed.

Mucopenetrating micelles with a PEG corona.

Crossing the intestinal mucus layer is a long-standing challenge for orally delivered nanoparticles carrying therapeutic cargo. We report the assembly of mucopenetrating cargo-loaded micelles using block copolymers consisting of either linear poly(ethylene glycol) (PEG) or bottle-brush poly(oligo(ethylene glycol)methacrylate) (PEGb) as the hydrophilic block and poly(caprolactone) (PCL) or poly(cholesteryl methacrylate) (PCMA) as the hydrophobic extension. The micelles were shown to preserve their stability and retain ∼50% of their cargo in simulated gastric fluid. The ability of micelles to diffuse through reconstituted porcine mucus was assessed in a microfluidic set-up. Finally, the delivery of Nile Red as a hydrophobic model cargo across a mucus layer produced by epithelial cells was demonstrated. These engineered mucopenetrating micelles have potential to be developed into efficient absorption enhancers, contributing a nanotechnology solution to oral drug delivery.

Enzymes as key features in therapeutic cell mimicry.

Cell mimicry is a nature inspired concept that aims to substitute for missing or lost (sub)cellular function. This review focuses on the latest advancements in the use of enzymes in cell mimicry for encapsulated catalysis and artificial motility in synthetic bottom-up assemblies with emphasis on the biological response in cell culture or more rarely in animal models. Entities across the length scale from nano-sized enzyme mimics, sub-micron sized artificial organelles and self-propelled particles (swimmers) to micron-sized artificial cells are discussed. Although the field remains in its infancy, the primary aim of this review is to illustrate the advent of nature-mimicking artificial molecules and assemblies on their way to become a complementary alternative to their role models for diverse biomedical purposes.

A Polymer Chemistry Point of View on Mucoadhesion and Mucopenetration.

Although oral is the preferred route of administration of pharmaceutical formulations, the long-standing challenge for medically active compounds to efficiently cross the mucus layer barrier limits its wider applicability. Efforts in nanomedicine to overcome this hurdle consider mucoadhesive and mucopenetrating drug carriers by selectively designing (macromolecular) building blocks. This review highlights and critically discusses recent strategies developed in this context including poly(ethylene glycol)-based modifications, cationic and thiolated polymers, as well as particles with high charge density, zeta-potential shifting ability, or mucolytic properties. The latest advances in ex vivo test platforms are also reviewed.

Double-Fueled Janus Swimmers with Magnetotactic Behavior.

Self-propelled particles attract a great deal of attention due to the auspicious range of applications for which nanobots can be used. In a biomedical context, self-propelled swimmers hold promise to autonomously navigate to a desired location in an attempt to counteract cell/tissue defects either by releasing drugs or by performing surgical tasks. The vast majority of prior reports deal with single engine assemblies, often utilizing fuel molecules which are considered to be highly cytotoxic. Herein, we introduce two engines: (1) a motor which couples enzymes (i.e., glucose oxidase) and inorganic nanoparticles (i.e., platinum nanoparticles) to gain power and (2) a peptide-fueled trypsin motor. We demonstrate that both engines can induce enhanced diffusion properties of (Janus) particles using bioavailable and completely harmless fuel molecules. By combining both engines on the same carrier, we show self-propelled particles employing two independent engines, using two different fuels. A collaborative enhancement of the swimmer's diffusion properties upon powering-up both engines simultaneously is observed. Additionally, the incorporation of magnetic nanoparticles allows for the swimmer to move in a magnetic gradient upon applying an external magnetic field, yielding in directional motion of the double-fueled particles. These multiple-fueled biocompatible swimmers are a significant contribution to make them applicable in a biomedical context.

Planar and Cell Aggregate-Like Assemblies Consisting of Microreactors and HepG2 Cells.

The assembly of microreactors has made considerable progress toward the fabrication of artificial cells. However, their characterization remains largely limited to buffer solution-based assays in the absence of their natural role model-the biological cells. Herein, the combination of microreactors with HepG2 cells either in planar cell cultures or in the form of cell aggregates is reported. Alginate (Alg)-based microreactors loaded with catalase are assembled by droplet microfluidics, and their activity is confirmed. The acceptance of polymer-coated ∼40 µm Alg particles by proliferating HepG2 cells is depending on the terminating polymer layer. When these functional microreactors are cocultured with HepG2 cells, they can be employed for detoxification, that is, hydrogen peroxide removal, and by doing so, they assist the cells to survive. This report is among the first successful combination of microreactors with biological cells, that is, HepG2 cells, contributing to the fundamental understanding of integrating synthetic and biological partners toward the maturation of this semisynthetic concept for biomedical applications.

Subcompartmentalized Nanoreactors as Artificial Organelle with Intracellular Activity.

Cell mimicry is an approach which aims at substituting missing or lost activity. In this context, the goal of artificial organelles is to provide intracellularly active nanoreactors to affect the cellular performance. So far, only a handful of reports discuss concepts addressing this challenge based on single-component reactors. Here, the assembly of nanoreactors equipped with glucose oxidase (GOx)-loaded liposomal subunits coated with a poly(dopamine) polymer layer and RGD targeting units is reported. When comparing different surface modifications, the uptake of the nanoreactors by endothelial cells and macrophages with applied shear stress is confirmed without inherent cytotoxicity. Furthermore, the encapsulation and preserved activity of GOx within the nanoreactors is shown. The intracellular activity is demonstrated by exposing macrophages with internalized nanoreactors to glucose and assessment of the cell viability after 6 and 24 h. The macrophage viability is found to be reduced due to the intracellularly produced hydrogen peroxide by GOx. This report on the first intracellular active subcompartmentalized nanoreactors is a considerable step in therapeutic cell mimicry.

Recent developments in poly(dopamine)-based coatings for biomedical applications.

The success of polymer coatings for biomedical applications is undeniable. Among the very successful examples are poly(dopamine) (PDA) films due to their simplicity in deposition and beneficial interaction with biomolecules and cells. The aim of this review is to highlight the findings and achievement of PDA in nanomedicine since 2011. We discuss the progress that has been made to elucidate the structure of PDA and novel aspects considering the assembly of PDA-based films on diverse substrates. We highlight the newest results considering the biological evaluation PDA-based coatings to control cell behavior and the use of PDA in biosensing. The popularity of PDA remains unchanged, but the research efforts start to be consolidated toward more specific aims and clinical applications.

Janus subcompartmentalized microreactors.

We report on Janus subcompartmentalized assemblies with enzyme loaded liposomes entrapped within a polymer carrier capsule - Janus subcompartmentalized microreactors. The concept is based on the use of Pickering emulsions and the subsequent deposition of interacting liposomes and polymer layers. We demonstrate the adjustment of the size of the Janus domains and the control over the amount of trapped liposomes using multiple liposome deposition steps. The obtained Janus capsosomes feature a distinct liposome domain within a closed polymeric hydrogel shell. The assembly of functional Janus microreactors using trypsin as cargo within the liposomal subcompartments is shown by performing locally confined enzymatic encapsulated catalysis. The presented assemblies with spatial control over the position of their liposomal subunits are a fascinating first step towards artificial cells with polarity.

Recent progress of liposomes in nanomedicine.

Liposomes, spherical vesicles consisting of one or more lipid bilayer membrane(s) encapsulating an aqueous medium, are among the prominent players in the field of nanomedicine. Herein, we highlight the newest, atypical applications of liposomes towards their use in biomedicine. In particular, we put special emphasis on innovative chemical modification of liposomes, the interactions of liposomes with cells under the influence of shear stress, and the utilisation of liposomes as drug deposits in polymeric films and as components in synthetic cell mimics.

Multi-responsive copolymers: using thermo-, light- and redox stimuli as three independent inputs towards polymeric information processing.

We report on triple responsive polymers, exhibiting a distinct and reversible lower critical solution temperature in water that can be altered by light and redox stimuli, and we suggest their evaluation for molecular information processing.

Nanotube Friendly Poly(N-isopropylacrylamide).

Poly(N-ispropylacrylamide) [PNIPAM] is a widely studied polymer for use in biological applications due to its lower critical solution temperature (LCST) being so close to the human body temperature. Unfortunately, attempts to combine carbon nanotubes (CNTs) with PNIPAM have been unsuccessful due to poor interactions between these two materials. In this work, a PNIPAM copolymer with 1 mol-% pyrene side group [p-PNIPAM] was used to produce a thermoresponsive polymer capable of stabilizing both single and multi-walled carbon nanotubes (MWNTs) in water. The presence of pyrene in the polymer chain lowers the LCST less than 4 °C and the interaction with nanotubes does not show any influence on LCST. Moreover, p-PNIPAM stabilized nanotubes show a temperature-dependent dispersion in water that allows the level of nanotube exfoliation/bundling to be controlled. Cryo-TEM images, turbidity, and viscosity of these suspensions were used to characterize these thermoresponsive changes. This ability to manipulate the dispersion state of CNTs in water with p-PNIPAM will likely benefit many biological applications, such as drug delivery, optical sensors, and hydrogels.

Covalently bonded layer-by-layer assembly of multifunctional thin films based on activated esters.

We demonstrate that chemically stable, multifunctional polymer thin films can be obtained using the layer-by-layer (LbL) deposition based on covalent bonds between adsorbing chains. Poly(pentafluorophenyl-4-vinylbenzoate) (P1) or poly(pentafluorophenylacrylate) (P2) polymers were assembled with poly(allyl amine) (PAAm) to yield LbL multilayer films through amide bond formation by the reaction between activated esters of P1 or P2 and amine groups in PAAm, which was quantitatively monitored by Fourier transform infrared spectroscopy (FT-IR). It was found that the difference in the solubility of P1 and P2 against ethanol, which was used as the solvent for PAAm, during the LbL deposition yields different reaction conversion for the activated esters in either P1 or P2: the reaction conversion of P2 is higher than the conversion with P1. In addition, free (or unreacted) activated esters and amine groups remaining in the PAAm/P1 LbL film were further utilized for the incorporation of multiple functional materials (5-((2-aminoethyl)amino)naphthalene-1-sulfonic acid (EDANS) and Rhodamine B dyes in the present case) by post-treatments in order to further tailor the film properties. It was also demonstrated that the surface functional groups (activated esters) in the LbL films can also be utilized for surface patterning with one functional material, followed by functionalization with a second functional material during the post-treatment throughout the whole film. Finally, the PAAm/P1 and PAAm/P2 LbL films were shown to be quite stable in the extreme pH range, and free-standing films can easily be obtained by the treatment of the films with mild acidic conditions. The versatility of incorporating multiple functional materials into a single multilayer film as well as the excellent physicochemical stability of the covalently bonded multilayer free-standing films proves to be quite useful to design flexible and multifunctional thin film structures for many chemical and biological applications.