Endosomal Escape of Polymer-Coated Silica Nanoparticles in Endothelial Cells.

Current investigations into hazardous nanoparticles (i.e., nanotoxicology) aim to understand the working mechanisms that drive toxicity. This understanding has been used to predict the biological impact of the nanocarriers as a function of their synthesis, material composition, and physicochemical characteristics. It is particularly critical to characterize the events that immediately follow cell stress resulting from nanoparticle internalization. While reactive oxygen species and activation of autophagy are universally recognized as mechanisms of nanotoxicity, the progression of these phenomena during cell recovery has yet to be comprehensively evaluated. Herein, primary human endothelial cells are exposed to controlled concentrations of polymer-functionalized silica nanoparticles to induce lysosomal damage and achieve cytosolic delivery. In this model, the recovery of cell functions lost following endosomal escape is primarily represented by changes in cell distribution and the subsequent partitioning of particles into dividing cells. Furthermore, multilamellar bodies are found to accumulate around the particles, demonstrating progressive endosomal escape. This work provides a set of biological parameters that can be used to assess cell stress related to nanoparticle exposure and the subsequent recovery of cell processes as a function of endosomal escape.

Infusion of iodine-based contrast agents into poly(p-dioxanone) as a radiopaque resorbable IVC filter.

To determine the feasibility of infusing resorbable inferior vena cava (IVC) filter with iodine-based contrast agents to produce a radiopaque, computed tomography (CT)-visible IVC filter. Infused poly(p-dioxanone) (PPDO) was obtained by incubating PPDO in different concentrations of 4-iodobenzoyl chloride (IBC) and 2,3,5-triiodobenzoic acid (TIBA). Characterizations of infused and nascent PPDO were done using elemental analysis, micro-CT, tensile strength analysis, scanning electron microscopy, and differential scanning calorimetry. Elemental analysis showed percentage loading of 1.07 ± 0.08 for IBC and 0.73 ± 0.01 for TIBA. The iodine loading remained the same within 2 weeks for TIBA but decreased to about 80 % with IBC when subjected to physiological conditions. Micro-CT images showed increased attenuation of the infused PPDO compared with the nascent PPDO. The Hounsfield unit values for infused and nascent sutures were 110 ± 40 and 153 ± 53 for PPDO infused with 2 mg/mL IBC and TIBA, respectively, but only 11.35 ± 2 for nascent PPDO. In contrast the HU for bone was 116 ± 37. Tensile strength analysis showed maximum loads of 1.01 ± 0.43 kg and 10.02 ± 0.54 kg for IBC and TIBA, respectively, and 10.10 ± 0.64 kg for nascent PPDO. Scanning electron microscopy showed that the morphology of the PPDO surface did not change after coating and preliminary cytotoxicity assay showed no killing effect on Hela cells. PPDO infused with a contrast agent is significantly more radiopaque than nascent PPDO on micro-CT imaging. This radiopacity could allow the position and integrity of infused resorbable IVC filter to be monitored while it is in place, thus increasing its safety and efficacy as a medical device.

Multiscale patterning of a biomimetic scaffold integrated with composite microspheres.

The ideal scaffold for regenerative medicine should concurrently mimic the structure of the original tissue from the nano- up to the macroscale and recapitulate the biochemical composition of the extracellular matrix (ECM) in space and time. In this study, a multiscale approach is followed to selectively integrate different types of nanostructured composite microspheres loaded with reporter proteins, in a multi-compartment collagen scaffold. Through the preservation of the structural cues of the functionalized collagen scaffold at the nano- and microscale, its macroscopic features (pore size, porosity, and swelling) are not altered. Additionally, the spatial confinement of the microspheres allows the release of the reporter proteins in each of the layers of the scaffold. Finally, the staged and zero-order release kinetics enables the temporal biochemical patterning of the scaffold. The versatile manufacturing of each component of the scaffold results in the ability to customize it to better mimic the architecture and composition of the tissues and biological systems.

Addition of platelet-rich plasma supports immune modulation and improved mechanical integrity in Alloderm mesh for ventral hernia repair in a rat model.

The recurrence of ventral hernias continues to be a problem faced by surgeons, in spite of efforts toward implementing novel repair techniques and utilizing different materials to promote healing. Cadaveric acellular dermal matrices (Alloderm) have shown some promise in numerous surgical subspecialties, but these meshes still suffer from subsequent failure and necessitation of re-intervention. Here, it is demonstrated that the addition of platelet rich plasma to Alloderm meshes temporally modulates both the innate and cytotoxic inflammatory responses to the implanted material. This results in decreased inflammatory cytokine production at early time points, decreased matrix metalloproteinase expression, and decreased CD8+ T cell infiltration. Collectively, these immune effects result in a healing phenotype that is free from mesh thinning and characterized by increased material stiffness.

Biomimetic cellular vectors for enhancing drug delivery to the lungs.

Despite recent advances in drug delivery, the targeted treatment of unhealthy cells or tissues continues to remain a priority. In cancer (much like other pathologies), delivery vectors are designed to exploit physical and biological features of unhealthy tissues that are not always homogenous across the disease. In some cases, shifting the target from unhealthy tissues to the whole organ can represent an advantage. Specifically, the natural organ-specific retention of nanotherapeutics following intravenous administration as seen in the lung, liver, and spleen can be strategically exploited to enhance drug delivery. Herein, we outline the development of a cell-based delivery system using macrophages as a delivery vehicle. When loaded with a chemotherapeutic payload (i.e., doxorubicin), these cellular vectors (CELVEC) were shown to provide continued release within the lung. This study provides proof-of-concept evidence of an alternative class of biomimetic delivery vectors that capitalize on cell size to provide therapeutic advantages for pulmonary treatments.

A New Class of Phantom Materials for Poroelastography Imaging Techniques.

Poroelastography is an elastographic technique used to image the temporal mechanical behavior of tissues. One of the major challenges in determining experimental potentials and limitations of this technique has been the lack of complex and realistic controlled phantoms that could be used to corroborate the limited number of theoretical and simulation studies available in the literature as well as to predict its performance in complex experimental situations and in a variety of conditions. In the study described here, we propose and analyze a new class of phantom materials for temporal elastography imaging. The results indicate that, by using polyacrylamide, we can generate inhomogeneous elastographic phantoms with controlled fluid content and fluid flow properties, while maintaining mechanical and ultrasonic properties similar to those of soft tissues.

One-pot synthesis of pH-responsive hybrid nanogel particles for the intracellular delivery of small interfering RNA.

This report describes a novel, one-pot synthesis of hybrid nanoparticles formed by a nanostructured inorganic silica core and an organic pH-responsive hydrogel shell. This easy-to-perform, oil-in-water emulsion process synthesizes fluorescently-doped silica nanoparticles wrapped within a tunable coating of cationic poly(2-diethylaminoethyl methacrylate) hydrogel in one step. Transmission electron microscopy and dynamic light scattering analysis demonstrated that the hydrogel-coated nanoparticles are uniformly dispersed in the aqueous phase. The formation of covalent chemical bonds between the silica and the polymer increases the stability of the organic phase around the inorganic core as demonstrated by thermogravimetric analysis. The cationic nature of the hydrogel is responsible for the pH buffering properties of the nanostructured system and was evaluated by titration experiments. Zeta-potential analysis demonstrated that the charge of the system was reversed when transitioned from acidic to basic pH and vice versa. Consequently, small interfering RNA (siRNA) can be loaded and released in an acidic pH environment thereby enabling the hybrid particles and their payload to avoid endosomal sequestration and enzymatic degradation. These nanoparticles, loaded with specific siRNA molecules directed towards the transcript of the membrane receptor CXCR4, significantly decreased the expression of this protein in a human breast cancer cell line (i.e., MDA-MB-231). Moreover, intravenous administration of siRNA-loaded nanoparticles demonstrated a preferential accumulation at the tumor site that resulted in a reduction of CXCR4 expression.

Cell source determines the immunological impact of biomimetic nanoparticles.

Recently, engineering the surface of nanotherapeutics with biologics to provide them with superior biocompatibility and targeting towards pathological tissues has gained significant popularity. Although the functionalization of drug delivery vectors with cellular materials has been shown to provide synthetic particles with unique biological properties, these approaches may have undesirable immunological repercussions upon systemic administration. Herein, we comparatively analyzed unmodified multistage nanovectors and particles functionalized with murine and human leukocyte cellular membrane, dubbed Leukolike Vectors (LLV), and the immunological effects that may arise in vitro and in vivo. Previously, LLV demonstrated an avoidance of opsonization and phagocytosis, in addition to superior targeting of inflammation and prolonged circulation. In this work, we performed a comprehensive evaluation of the importance of the source of cellular membrane in increasing their systemic tolerance and minimizing an inflammatory response. Time-lapse microscopy revealed LLV developed using a cellular coating derived from a murine (i.e., syngeneic) source resulted in an active avoidance of uptake by macrophage cells. Additionally, LLV composed of a murine membrane were found to have decreased uptake in the liver with no significant effect on hepatic function. As biomimicry continues to develop, this work demonstrates the necessity to consider the source of biological material in the development of future drug delivery carriers.

PLGA-Mesoporous Silicon Microspheres for the in Vivo Controlled Temporospatial Delivery of Proteins.

In regenerative medicine, the temporospatially controlled delivery of growth factors (GFs) is crucial to trigger the desired healing mechanisms in the target tissues. The uncontrolled release of GFs has been demonstrated to cause severe side effects in the surrounding tissues. The aim of this study was to optimize a translational approach for the fine temporal and spatial control over the release of proteins, in vivo. Hence, we proposed a newly developed multiscale composite microsphere based on a core consisting of the nanostructured silicon multistage vector (MSV) and a poly(dl-lactide-co-glycolide) acid (PLGA) outer shell. Both of the two components of the resulting composite microspheres (PLGA-MSV) can be independently tailored to achieve multiple release kinetics contributing to the control of the release profile of a reporter protein in vitro. The influence of MSV shape (hemispherical or discoidal) and size (1, 3, or 7 µm) on PLGA-MSV's morphology and size distribution was investigated. Second, the copolymer ratio of the PLGA used to fabricate the outer shell of PLGA-MSV was varied. The composites were fully characterized by optical microscopy, scanning electron microscopy, ζ potential, Fourier transform infrared spectroscopy, and thermogravimetric analysis-differential scanning calorimetry, and their release kinetics over 30 days. PLGA-MSV's biocompatibility was assessed in vitro with J774 macrophages. Finally, the formulation of PLGA-MSV was selected, which concurrently provided the most consistent microsphere size and allowed for a zero-order release kinetic. The selected PLGA-MSVs were injected in a subcutaneous model in mice, and the in vivo release of the reporter protein was followed over 2 weeks by intravital microscopy, to assess if the zero-order release was preserved. PLGA-MSV was able to retain the payload over 2 weeks, avoiding the initial burst release typical of most drug delivery systems. Finally, histological evaluation assessed the biocompatibility of the platform in vivo.

Potential avoidance of adverse analgesic effects using a biologically "smart" hydrogel capable of controlled bupivacaine release.

Acute pain remains a tremendous clinical and economic burden, as its prevalence and common narcotic-based treatments are associated with poorer outcomes and higher costs. Multimodal analgesia portends great therapeutic promise, but rarely allows opioid sparing, and new alternatives are necessary. Microparticles (MPs) composed of biodegradable polymers [e.g., poly(lactic-co-glycolic acid) or PLGA] have been applied for controlled drug release and acute pain treatment research. However, foreign particles' presence within inflamed tissue may affect the drug release or targeting, and/or cause a secondary inflammatory reaction. We examined how small alterations in the particulate nature of MPs affect both their uptake into and subsequent activation of macrophages. MPs composed of PLGA and chitosan (PLGA-Chi) loaded with bupivacaine (BP) were engineered at different sizes and their opsonization by J774 macrophages was assessed. Uptake of PLGA-Chi by macrophages was found to be size dependent, but they were not cytotoxic or proinflammatory in effect. Moreover, encapsulation of MPs in a thermoresponsive loading gel (pluronic F-127) effectively prevented opsonization. Finally, MPs displayed sustained, tunable release of BP up to 7 days. These results demonstrate our ability to develop a drug delivery system capable of controlled release of local anesthetics to treat acute/subacute pain while concurrently avoiding enhanced inflammation.

Synthetic nanoparticles functionalized with biomimetic leukocyte membranes possess cell-like functions.

The therapeutic efficacy of systemic drug-delivery vehicles depends on their ability to evade the immune system, cross the biological barriers of the body and localize at target tissues. White blood cells of the immune system--known as leukocytes--possess all of these properties and exert their targeting ability through cellular membrane interactions. Here, we show that nanoporous silicon particles can successfully perform all these actions when they are coated with cellular membranes purified from leukocytes. These hybrid particles, called leukolike vectors, can avoid being cleared by the immune system. Furthermore, they can communicate with endothelial cells through receptor-ligand interactions, and transport and release a payload across an inflamed reconstructed endothelium. Moreover, leukolike vectors retained their functions when injected in vivo, showing enhanced circulation time and improved accumulation in a tumour.

Silicon micro- and nanofabrication for medicine.

This manuscript constitutes a review of several innovative biomedical technologies fabricated using the precision and accuracy of silicon micro- and nanofabrication. The technologies to be reviewed are subcutaneous nanochannel drug delivery implants for the continuous tunable zero-order release of therapeutics, multi-stage logic embedded vectors for the targeted systemic distribution of both therapeutic and imaging contrast agents, silicon and porous silicon nanowires for investigating cellular interactions and processes as well as for molecular and drug delivery applications, porous silicon (pSi) as inclusions into biocomposites for tissue engineering, especially as it applies to bone repair and regrowth, and porous silica chips for proteomic profiling. In the case of the biocomposites, the specifically designed pSi inclusions not only add to the structural robustness, but can also promote tissue and bone regrowth, fight infection, and reduce pain by releasing stimulating factors and other therapeutic agents stored within their porous network. The common material thread throughout all of these constructs, silicon and its associated dielectrics (silicon dioxide, silicon nitride, etc.), can be precisely and accurately machined using the same scalable micro- and nanofabrication protocols that are ubiquitous within the semiconductor industry. These techniques lend themselves to the high throughput production of exquisitely defined and monodispersed nanoscale features that should eliminate architectural randomness as a source of experimental variation thereby potentially leading to more rapid clinical translation.