Modular Click Assembly of Degradable Capsules Using Polyrotaxanes

A modular approach for the formation of degradable capsules using polyrotaxanes (PRXs) is described. The PRXs consist of α-cyclodextrin (αCD) and poly(ethylene glycol) (PEG), which are both biologically benign and the main degradation products of the capsules. The PRXs were equipped with three alkyne groups at their ends and could be successfully grafted to azide-functionalized silica particles (2.76 μm diameter) using azide–alkyne click chemistry. The assembled PRXs were then cross-linked using a degradable linker. The cross-linked structure was sufficiently robust to allow the formation of capsules after dissolving the template silica particles. The formation of capsules of <i>ca</i>. 2 μm diameter was verified by optical microscopy, TEM, and AFM imaging. The capsules were loaded with the chemotherapy drug doxorubicin (DOX) by conjugating it to the threaded αCDs <i>via</i> their free OH groups, while maintaining degradability of the capsules. Alkyne moieties at the surface of the cross-linked PRX architecture were available for further functionalization of the capsules, as is demonstrated by clicking on fluorescent PEG moieties. The DOX-loaded capsules were degraded within 90 min at 37 °C upon exposure to a 5 mM solution of glutathione in water

Formation and Degradation of Layer-by-Layer-Assembled Polyelectrolyte Polyrotaxane Capsules

We report the preparation of degradable capsules via layer-by-layer assembly using polyelectrolyte (PE) polyrotaxanes (PRXs). The PRX capsules were prepared by the sequential deposition of PRXs onto silica particles followed by the dissolution of the silica cores. The colloidal stability of the PRX capsules that are formed depends on the salt/buffer solution used in the assembly process. Various salt/buffer combinations were examined to avoid aggregation of the core–shell particles during PRX assembly and core dissolution. Using appropriate assembly conditions, we prepared colloidally stable, robust capsules. PRX capsules consisting of eight layers of PE PRXs had a wall thickness of ∼15 nm. The degradation of the PRX capsules was demonstrated through the disassembly of the PE PRXs using glutathione, which cleaves the disulfide bonds linking the end-capping groups of the PE PRXs. Given the supramolecular noncovalent structure of PRXs and their adjustable properties, it is expected that PRXs will be used as building blocks for assembling advanced capsules with unique and tailored properties

Probing the Dynamic Nature of DNA Multilayer Films Using Förster Resonance Energy Transfer

DNA films are of interest for use in a number of areas, including sensing, diagnostics, and as drug/gene delivery carriers. The specific base pairing of DNA materials can be used to manipulate their architecture and degradability. The programmable nature of these materials leads to complex and unexpected structures that can be formed from solution assembly. Herein, we investigate the structure of DNA multilayer films using Förster resonance energy transfer (FRET). The DNA films are assembled on silica particles by depositing alternating layers of homopolymeric diblocks (polyA₁₅G₁₅ and polyT₁₅C₁₅) with fluorophore (polyA₁₅G₁₅-TAMRA) and quencher (polyT₁₅C₁₅-BHQ2) layers incorporated at predesigned locations throughout the films. Our results show that DNA films are dynamic structures that undergo rearrangement. This occurs when the multilayer films are perturbed during new layer formation through hybridization but can also take place spontaneously when left over time. These films are anticipated to be useful in drug delivery applications and sensing applications

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

Confinement of Acoustic Cavitation for the Synthesis of Protein-Shelled Nanobubbles for Diagnostics and Nucleic Acid Delivery

We report a novel flow-through sonication technique for synthesizing stable and monodispersed nano- and micrometer-sized bubbles that have potential applications in diagnostics and gene therapy. The size and size distribution of the bubbles are controlled by the active cavitation zone generated by ultrasound. These bubbles are shown to possess echogenic properties and can be used for loading oligonucleotides

Tuning the Mechanical Properties of Nanoporous Hydrogel Particles via Polymer Cross-Linking

Soft hydrogel particles with tunable mechanical properties are promising for next-generation therapeutic applications. This is due to the increasingly proven role that physicochemical properties play in particulate-based delivery vectors, both <i>in vitro</i> and <i>in vivo</i>. The ability to understand and quantify the mechanical properties of such systems is therefore essential to optimize function and performance. We report control over the mechanical properties of poly­(methacrylic acid) (PMA) hydrogel particles based on a mesoporous silica templating method. The mechanical properties of the obtained particles can be finely tuned through variation of the cross-linker concentration, which is hereby quantified using a cross-linking polymer with a fluorescent tag. We demonstrate that the mechanical properties of the particles can be elucidated using an atomic force microscopy (AFM) force spectroscopy method, which additionally allows for the study of hydrogel material properties at the nanoscale through high-resolution force mapping. Young’s modulus and stiffness of the particles were tuned between 0.04 and 2.53 MPa and between 1.6 and 28.4 mN m^–1, respectively, through control over the cross-linker concentration. The relationship between the concentration of the cross-linker added and the amount of adsorbed polymer was observed to follow a Langmuir isotherm, and this relationship was found to correlate linearly with the particle mechanical properties

Microfluidic Examination of the “Hard” Biomolecular Corona Formed on Engineered Particles in Different Biological Milieu

The formation of a biomolecular corona around engineered particles determines, in large part, their biological behavior in vitro and in vivo. To gain a fundamental understanding of how particle design and the biological milieu influence the formation of the “hard” biomolecular corona, we conduct a series of in vitro studies using microfluidics. This setup allows the generation of a dynamic incubation environment with precise control over the applied flow rate, stream orientation, and channel dimensions, thus allowing accurate control of the fluid flow and the shear applied to the proteins and particles. We used mesoporous silica particles, poly­(2-methacryl­oyloxyethyl­phosphoryl­choline) (PMPC)-coated silica hybrid particles, and PMPC replica particles (obtained by removal of the silica particle templates), representing high-, intermediate-, and low-fouling particle systems, respectively. The protein source used in the experiments was either human serum or human full blood. The effects of flow, particle surface properties, incubation medium, and incubation time on the formation of the biomolecular corona formation are examined. Our data show that protein adhesion on particles is enhanced after incubation in human blood compared to human serum and that dynamic incubation leads to a more complex corona. By varying the incubation time from 2 s to 15 min, we demonstrate that the “hard” biomolecular corona is kinetically subdivided into two phases comprising a tightly bound layer of proteins interacting directly with the particle surface and a loosely associated protein layer. Understanding the influence of particle design parameters and biological factors on the corona composition, as well as its dynamic assembly, may facilitate more accurate prediction of corona formation and therefore assist in the design of advanced drug delivery vehicles

Dynamic Flow Impacts Cell–Particle Interactions: Sedimentation and Particle Shape Effects

The interaction of engineered particles with biological systems determines their performance in biomedical applications. Although standard static cell cultures remain the norm for in vitro studies, modern models mimicking aspects of the dynamic in vivo environment have been developed. Herein, we investigate fundamental cell–particle interactions under dynamic flow conditions using a simple and self-contained device together with standard multiwell cell culture plates. We engineer two particle systems and evaluate their cell interactions under dynamic flow, and we compare the results to standard static cell cultures. We find substantial differences between static and dynamic flow conditions and attribute these to particle shape and sedimentation effects. These results demonstrate how standard static assays can be complemented by dynamic flow assays for a more comprehensive understanding of fundamental cell–particle interactions

Emerging Techniques in Proteomics for Probing Nano–Bio Interactions

Nanoengineered particles that can facilitate drug formulation and improve specificity of delivery afford exciting opportunities for improved lesion-specific therapy. Understanding and controlling the nano–bio interactions of these materials is central to future developments in this area. Mass-spectrometry-based proteomics techniques, in conjunction with other emerging technologies, are enabling novel insights into the modulation of particle surfaces by biological fluids (formation of the protein corona) and subsequent particle-induced cellular responses. In this Perspective, we summarize important recent developments using proteomics-based techniques to understand nano–bio interactions and discuss the impact of such knowledge on improving particle design

Improved Auditory Nerve Survival with Nanoengineered Supraparticles for Neurotrophin Delivery into the Deafened Cochlea - Fig 4

<p>(A) Average of the SGN density measured in each cochlear region after one month of treatment with BDNF-SP (black) and the Control-SPs (Grey). There was a significantly greater density of SGNs in cochleae treated with BDNF-SPs compared to the Control-SPs (two way ANOVA, p = 0.009) with post hoc analysis indicated (Holm-Sidak, **p<0.005, *p<0.05). Error bars ± 1 SEM. (B) Analysis of the cochlear tissue response measured in the scala tympani (ST) in cochlear Regions A and B showed a tissue response for Region A (near the site of the cochleostomy) that was significantly larger than the tissue response in Region B (Post Hoc Holm-Sidak; **p<0.001). The tissue response measured in Region A in the BDNF-SP treated cochleae was greater than that in the Control-SP treated cochlea (Post Hoc Holm-Sidak; *p = 0.003).</p