Peter Chhour

Dextran coated bismuth–iron oxide nanohybrid contrast agents for computed tomography and magnetic resonance imaging

Bismuth nanoparticles have been proposed as a novel CT contrast agent, however few syntheses of biocompatible bismuth nanoparticles have been achieved. We herein report the synthesis of composite bismuth-iron oxide nanoparticles (BION) that are based on a clinically approved, dextran-coated iron oxide formulation; the particles have the advantage of acting as contrast agents for both CT and MRI. BION were synthesized and characterized using various analytical methods. BION CT phantom images revealed that the X-ray attenuation of the different formulations was dependent upon the amount of bismuth present in the nanoparticle, while T2-weighted MRI contrast decreased with increasing bismuth content. No cytotoxicity was observed in Hep G2 and BJ5ta cells after 24 hours incubation with BION. The above properties, as well as the yield of synthesis and bismuth inclusion efficiency, led us to select the Bi-30 formulation for in vivo experiments, performed in mice using a micro-CT and a 9.4 T MRI system. X-ray contrast was observed in the heart and blood vessels over a 2 hour period, indicating that Bi-30 has a prolonged circulation half-life. Considerable signal loss in T2-weighted MR images was observed in the liver compared to pre-injection scans. Evaluation of the biodistribution of Bi-30 revealed that bismuth is excreted via the urine, with significant concentrations found in the kidneys and urine. In vitro experiments confirmed the degradability of Bi-30. In summary, dextran coated BION are biocompatible, biodegradable, possess strong X-ray attenuation properties and also can be used as T2-weighted MR contrast agents.

Cyclic Stretch–Induced Oxidative Stress Increases Pulmonary Alveolar Epithelial Permeability

Mechanical ventilation with high tidal volumes has been associated with pulmonary alveolar flooding. Understanding the mechanisms underlying cyclic stretch-induced increases in alveolar epithelial permeability may be important in designing preventive measures for acute lung injury. In this work, we assessed whether cyclic stretch leads to the generation of reactive oxygen species in type I-like alveolar epithelial cells, which increase monolayer permeability via activation of NF-κB and extracellular signal-regulated kinase (ERK). We cyclically stretched type I-like rat primary alveolar epithelial cells at magnitudes of 12, 25, and 37% change in surface area (ΔSA) for 10 to 120 minutes. High levels of reactive oxygen species and of superoxide and NO specifically were detected in cells stretched at 37% ΔSA for 10 to 120 minutes. Exogenous superoxide and NO stimulation increased epithelial permeability in unstretched cells, which was preventable by the NF-κB inhibitor MG132. The cyclic stretch-induced increase in permeability was decreased by the superoxide scavenger tiron and by MG132. Furthermore, tiron had a dramatic protective effect on in vivo lung permeability under mechanical ventilation conditions. Cyclic stretch increased the activation of the NF-κB signaling pathway, which was significantly decreased with the ERK inhibitor U0126. Altogether, our in vitro and in vivo data demonstrate the sensitivity of permeability to stretch- and ventilation-induced superoxide production, suggesting that using antioxidants may be helpful in the prevention and treatment of ventilator-induced lung injury.

Systematic in vitro toxicological screening of gold nanoparticles designed for nanomedicine applications.

Gold nanoparticles (AuNP) are increasingly being applied in the biomedical field as therapeutics, contrast agents, and in diagnostic systems, motivating investigations of their toxicity that might arise from accidental exposure. While other work has investigated the toxicological response to gold nanoparticles for industrial purposes, here we have surveyed formulations that have been developed for biomedical use, are in clinical trials or have been FDA-approved. The AuNP library tested contains a range of shapes, including spheres, rods and shells, that possess a range of coatings, such as silica, citrate, lipoprotein, polymaleic acid, polyethylene glycol, DNA and others. Good cytocompatibility for all formulations was observed after 1 h of incubation. However after 24 h exposure, a nanorod and a spherical DNA coated formulation resulted in toxicity. The coating material was the only factor that influenced toxicity. AuNP exposure seemed to have no effect on cell cytoskeleton deformation and cell spreading. Cell uptake, as measured by computed tomography and ICP-OES, as well as TEM images of cells, confirmed strong AuNP uptake for certain formulations, but there was no correlation with toxicity. No glove translocation occurred, therefore, nitrile gloves are an adequate safety precaution for working with the AuNP studied. In conclusion, the majority of AuNP formulations tested have very low adverse effects.

Nanoparticle Loaded Polymeric Microbubbles as Contrast Agents for Multimodal Imaging

Ultrasound contrast agents are typically microbubbles (MB) with a gas core that is stabilized by a shell made of lipids, proteins, or polymers. The high impedance mismatch between the gas core and an aqueous environment produces strong contrast in ultrasound (US). Poly(lactic acid) (PLA) MB, previously developed in our laboratory, have been shown to be highly echogenic both in vitro and in vivo. Combining US with other imaging modalities such as fluorescence, magnetic resonance imaging (MRI), or computerized tomography (CT) could improve the accuracy of many US applications and provide more comprehensive diagnostic information. Furthermore, our MB have the capacity to house a drug in the PLA shell and create drug-loaded nanoparticles in situ when passing through an ultrasound beam. To create multimodal contrast agents, we hypothesized that the polymer shell of our PLA MB platform could accommodate additional payloads. In this study, we therefore modified our current MB by encapsulating nanoparticles including aqueous or organic quantum dots (QD), magnetic iron oxide nanoparticles (MNP), or gold nanoparticles (AuNP) to create bimodality platforms in a manner that minimally compromised the performance of each individual imaging technique.

Nanodisco balls: control over surface versus core loading of diagnostically active nanocrystals into polymer nanoparticles.

Nanoparticles of complex architectures can have unique properties. Self-assembly of spherical nanocrystals is a high yielding route to such systems. In this study, we report the self-assembly of a polymer and nanocrystals into aggregates, where the location of the nanocrystals can be controlled to be either at the surface or in the core. These nanospheres, when surface decorated with nanocrystals, resemble disco balls, thus the term nanodisco balls. We studied the mechanism of this surface loading phenomenon and found it to be Ca2+ dependent. We also investigated whether excess phospholipids could prevent nanocrystal adherence. We found surface loading to occur with a variety of nanocrystal types including iron oxide nanoparticles, quantum dots, and nanophosphors, as well as sizes (10–30 nm) and shapes. Additionally, surface loading occurred over a range of polymer molecular weights (∼30–3000 kDa) and phospholipid carbon tail length. We also show that nanocrystals remain diagnostically active after loading onto the polymer nanospheres, i.e., providing contrast in the case of magnetic resonance imaging for iron oxide nanoparticles and fluorescence for quantum dots. Last, we demonstrated that a fluorescently labeled protein model drug can be delivered by surface loaded nanospheres. We present a platform for contrast media delivery, with the unusual feature that the payload can be controllably localized to the core or the surface.

Effect of Gold Nanoparticle Size and Coating on Labeling Monocytes for CT Tracking

With advances in cell therapies, interest in cell tracking techniques to monitor the migration, localization, and viability of these cells continues to grow. X-ray computed tomography (CT) is a cornerstone of medical imaging but has been limited in cell tracking applications due to its low sensitivity toward contrast media. In this study, we investigate the role of size and surface functionality of gold nanoparticles for monocyte uptake to optimize the labeling of these cells for tracking in CT. We synthesized gold nanoparticles (AuNP) that range from 15 to 150 nm in diameter and examined several capping ligands, generating 44 distinct AuNP formulations. In vitro cytotoxicity and uptake experiments were performed with the RAW 264.7 monocyte cell line. The majority of formulations at each size were found to be biocompatible, with only certain 150 nm PEG functionalized particles reducing viability at high concentrations. High uptake of AuNP was found using small capping ligands with distal carboxylic acids (11-MUA and 16-MHA). Similar uptake values were found with intermediate sizes (50 and 75 nm) of AuNP when coated with 2000 MW poly(ethylene-glycol) carboxylic acid ligands (PCOOH). Low uptake values were observed with 15, 25, 100, and 150 nm PCOOH AuNP, revealing interplay between size and surface functionality. Transmission electron microscopy (TEM) and CT performed on cells revealed similar patterns of high gold uptake for 50 nm PCOOH and 75 nm PCOOH AuNP. These results demonstrate that highly negatively charged carboxylic acid coatings for AuNP provide the greatest internalization of AuNP in monocytes, with a complex dependency on size.

Use of Nanoparticle Contrast Agents for Cell Tracking with Computed Tomography

Efforts to develop novel cell-based therapies originated with the first bone marrow transplant on a leukemia patient in 1956. Preclinical and clinical examples of cell-based treatment strategies have shown promising results across many disciplines in medicine, with recent advances in immune cell therapies for cancer producing remarkable response rates, even in patients with multiple treatment failures. However, cell-based therapies suffer from inconsistent outcomes, motivating the search for tools that allow monitoring of cell delivery and behavior in vivo. Noninvasive cell imaging techniques, also known as cell tracking, have been developed to address this issue. These tools can allow real-time, quantitative, and long-term monitoring of transplanted cells in the recipient, providing insight on cell migration, distribution, viability, differentiation, and fate, all of which play crucial roles in treatment efficacy. Understanding these parameters allows the optimization of cell choice, delivery route, and dosage for therapy and advances cell-based therapy for specific clinical uses. To date, most cell tracking work has centered on imaging modalities such as MRI, radionuclide imaging, and optical imaging. However, X-ray computed tomography (CT) is an emerging method for cell tracking that has several strengths such as high spatial and temporal resolution, and excellent quantitative capabilities. The advantages of CT for cell tracking are enhanced by its wide availability and cost effectiveness, allowing CT to become one of the most popular clinical imaging modalities and a key asset in disease diagnosis. In this review, we will discuss recent advances in cell tracking methods using X-ray CT in various applications, in addition to predictions on how the field will progress.

CD163+ macrophages promote angiogenesis and vascular permeability accompanied by inflammation in atherosclerosis

Intake of hemoglobin by the hemoglobin-haptoglobin receptor CD163 leads to a distinct alternative non–foam cell antiinflammatory macrophage phenotype that was previously considered atheroprotective. Here, we reveal an unexpected but important pathogenic role for these macrophages in atherosclerosis. Using human atherosclerotic samples, cultured cells, and a mouse model of advanced atherosclerosis, we investigated the role of intraplaque hemorrhage on macrophage function with respect to angiogenesis, vascular permeability, inflammation, and plaque progression. In human atherosclerotic lesions, CD163+ macrophages were associated with plaque progression, microvascularity, and a high level of HIF1&agr; and VEGF-A expression. We observed irregular vascular endothelial cadherin in intraplaque microvessels surrounded by CD163+ macrophages. Within these cells, activation of HIF1&agr; via inhibition of prolyl hydroxylases promoted VEGF-mediated increases in intraplaque angiogenesis, vascular permeability, and inflammatory cell recruitment. CD163+ macrophages increased intraplaque endothelial VCAM expression and plaque inflammation. Subjects with homozygous minor alleles of the SNP rs7136716 had elevated microvessel density, increased expression of CD163 in ruptured coronary plaques, and a higher risk of myocardial infarction and coronary heart disease in population cohorts. Thus, our findings highlight a nonlipid-driven mechanism by which alternative macrophages promote plaque angiogenesis, leakiness, inflammation, and progression via the CD163/HIF1&agr;/VEGF-A pathway.

Nanoparticles for Cardiovascular Imaging with CT

Complications involved with the progression of atherosclerosis in the coronary arteries are responsible for the majority of deaths related to cardiovascular disease. Clinically, X-ray computed tomography (CT) plays a key role in the assessment of atherosclerosis but current iodinated contrast agents relied upon for CT applications suffer from fast clearance, biocompatibility issues, and nonspecific uptake. Nanotechnologies have presented solutions to these problems by offering precise control over the properties of novel contrast agents. In this chapter, we discuss nanoparticle CT contrast agents that have been used for cardiovascular imaging. We cover iodinated and inorganic nanoparticle structures including nano-emulsions, liposomes, micelles, dendrimers, solid metal core nanoparticles, and others and discuss their applications for vascular and target specific imaging. Tremendous progress has been made in the field over the past decade and we expect that the next decade will see the clinical translation of nanoparticle formulations among many additional technological advances.

Wulff in a cage gold nanoparticles as contrast agents for computed tomography and photoacoustic imaging

Nanostructures have potential for use in biomedical applications such as sensing, imaging, therapeutics, and drug delivery. Among nanomaterials, gold nanostructures are of considerable interest for biomedical research, owing to their bio-inertness, controllable surface chemistry, X-ray opacity, and optical properties. Gold nanocages are particularly attractive for imaging and therapeutic applications, because they strongly absorb light in the near infra-red region which has high light transmission in tissue. However, the X-ray attenuation of nanocages is relatively low due to their hollow structure. In this study, for the first time, we sought to combine the attractive optical properties of nanoshells with the high payloads of solid nanoparticles and investigated their biomedical applications. Here, we report the engineering of Wulff in a cage nanoparticles via converting gold Wulff-shaped seeds into gold-silver core-shell structures and then performing a galvanic replacement reaction. The structure of these nanoparticles was determined using transition electron microscopy. This morphological transformation of gold nanoparticles shaped as truncated octahedrons into a complex Wulff in a cage nanoparticles during the reaction resulted in extensive changes in their optical properties that made these unique structures a potential contrast agent for photoacoustic imaging. We found that the Wulff in a cage nanoparticles had no adverse effects on the viabilities of J774A.1, Renca, and HepG2 cells at any of the concentrations tested. In vitro and in vivo experiments showed robust signals in both photoacoustic imaging and computed tomography. To the best of our knowledge, this is the first report of Wulff in a cage nanoparticles serving as a platform for multiple imaging modalities. This unique multifunctional nanostructure, which integrates the competencies of both core and shell structures, allows their use as contrast agents for photoacoustic imaging, computed tomography and as a potential agent for photothermal therapy.