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NCBI Bookshelf. Created: October 13, ; Last Update: November 30,

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You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser or turn off compatibility mode in Internet Explorer. Magnetic targeting utilises the properties of superparamagnetic iron oxide nanoparticles SPIONs to accumulate particles in specified vasculature regions under an external magnetic field.

Introduction

As the behaviour of circulating particles varies depending on nanoparticle characteristics, magnetic field strength and flow dynamics, we established an improved ex vivo model in order to estimate the magnetic capture of SPIONs in physiological-like settings. We describe here a new, easy to handle ex vivo model of human umbilical artery. Using this model, the magnetic targeting of different types of SPIONs under various external magnetic field gradients and flow conditions was investigated by atomic emission spectroscopy and histology.

Among tested particles, SPION-1 with lauric acid shell had the largest capacity to accumulate at the specific artery segment. Taken together, the umbilical artery model constitutes a time- and cost-efficient, 3R-compliant tool to assess magnetic targeting of SPIONs under flow.

Our further imply the possibility of an efficient in vivo targeting of certain types of SPIONs to superficial arteries. Among the wide variety of nanoparticle systems which are being studied for the purpose of medical applications, magnetic nanoparticles represent a versatile platform that can be potentially utilized both as a diagnostic contrast agent and as a drug delivery system.

The latter application can be greatly improved by active targeting to allow a better control of nanoparticle biodistribution and to enhance their therapeutic efficacy. For magnetic nanoparticles, a promising strategy of drug delivery, which in increased drug paylo in the target tissue, at the same time reducing their systemic dose and toxicity, is based on so-called magnetic drug targeting MDT. In this approach, conjugation of superparamagnetic iron oxide nanoparticles SPIONs with drugs in combination with an external magnetic field is used to target the particles to the diseased vasculature regions as demonstrated by the studies in a rabbit model of cancer 123a mouse model of thrombosis 4a mouse model of cardiac ischemia 5 and several mouse models of cancer 678.

The existing studies mostly utilized the phenomenon of an enhanced permeability of the microvessel endothelium in cancer and inflammatory diseases, which facilitates the extravasation of nano-sized particles 9. Magnetic capture under flow conditions characteristic for larger vessels occurs when the force exerted on the particles by the magnetic field overcomes the particle hydrodynamic drag force.

Hence, the behaviour of magnetic particles in circulation may vary greatly depending on the nanoparticle characteristics, the magnetic field gradients and the flow dynamics. In particular, the effect of particle-particle interactions under the influence of magnetic field gradients is a very complex matter 10 and the extent its contribution to MDT is not yet fully understood.

In physiological conditions, arterial wall is constantly exposed to shear stress induced by the flow of blood and its viscosity, and the patterns of shear stress affect the behaviour of the blood-borne cells and particles Thus far, the experimental attempts to magnetically target medium and large vessels have been very scarce 1213 It is therefore important to investigate the possibility of accumulation of magnetic particles under physiologic-like flow conditions in the experimental models prior to the in vivo MDT application.

In our present report, we describe the development of an ex vivo model for investigations of magnetic accumulation of SPIONs based on the use of human umbilical cord arteries. Normal human umbilical cords contain 2 arteries and are usually disposed of as post-partum waste.

The advantage of these arteries is their size similar to human coronary arteries 15 and the lack of any branches. In order to control the vessel diameter and distensibility, arteries are embedded in a supporting matrix. This new model, allowing improved standardisation, was employed here to investigate the efficacy of magnetic targeting, utilizing 3 types of SPIONs with different physicochemical characteristics. Furthermore, the effects of varying external magnetic field parameters and of the variation of SPION circulation time and flow conditions on the magnetic capture efficacy were analysed.

The umbilical arteries used in this study had diameter of 2. The arteries were isolated from the umbilical cords between day 1 and day 4 post-partum. Only the intact artery fragments were used in order to avoid the leakage of SPIONs and unspecific accumulation in the artery wall. The example images of vessel morphology analysed by histological staining are shown in Suppl.

The phenotypic features and viability of human umbilical artery endothelial cells HUAECs isolated from the characterised arteries are summarized in Suppl. Figs 2 and 3. The arteries used in the present study were routinely isolated from the respective cords on the same day as human umbilical vein endothelial cells HUVECswhich provided an independent control of the presence of viable cells in the umbilical vessels. To provide an in vivo -like mechanical support and control over the artery diameter, an entirely new set-up was established and optimised as described in detail in Methods.

After gel solidification, the artery fragments were perfused with medium at the flow rate of 4.

Among the tested gel densities, the concentration of 0. These conditions were selected as the final set-up, which was subsequently used in the experiments with SPIONs. A Umbilical artery embedded in agarose gel; B Schematic presentation of the experimental set-up for magnetic targeting; C Example image showing the electromagnet, the artery, SPION reservoir, and peristaltic pump. Red arrows indicate the flow direction. The considerable differences in the physicochemical properties of SPIONs resulted in dramatic differences of their magnetic accumulation under flow conditions, as also shown in our in vitro studies The quantitative of the iron content analysis obtained with atomic emission spectroscopy AES were subsequently confirmed qualitatively using histological staining.

Iron accumulation, visualised with Prussian blue staining, is highlighted with arrows. There were no ificant differences in tissue iron content between magnetically targeted and un-targeted artery samples.

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As shown in the Fig. Nanoparticle accumulation reflected by blue staining shows clear differences depending on the SPION type. Iron accumulation, visualized with Prussian blue staining, is highlighted with arrows.

To address the effect of magnetic field parameters on SPION accumulation, a series of experiments was performed with a reduced magnetic field gradient and b increased distance from the tip of the magnet to the artery. In those experiments, SPION-1 were applied, as these particles showed the highest iron peak under the magnetic pole shoe and therefore, we expected to detect the resulting changes in their accumulation with higher sensitivity. The of these experiments are summarised in Fig.

As shown in Fig. Concerning the potential in vivo applications, these data suggest that targeting with external magnet should allow SPION accumulation in the superficial arteries, but can prove difficult in the case of arteries localised in the deeper body regions. Both the broader particle distribution at the decreased magnetic field strength and the clear dependency of MDT efficacy on the distance from the tip of the magnet indicate that a certain threshold gradient exists that must be present to overcome the hydrodynamic forces of the flowing fluid in order to effectively accumulate the magnetic nanoparticles at the arterial wall.

In further experiments, the effects of circulation time and flow rate were evaluated.

In further experiments, flow rate was reduced to 2. This was an unexpected result, which however can be explained by a strong sedimentation of SPION-1 observed at this flow rate. The efficacy of magnetic drug targeting, both in terms of the amounts of delivered drug and the therapeutic outcome has been demonstrated in several studies on tumour-bearing rabbits treated intra-arterially with mitoxantrone-loaded SPIONs 12as well in the mouse models of cancer upon intravenous application of doxorubicin-loaded nanoparticles Chao et al.

Magnetic targeting was furthermore effective in a rat model of myocardial infarction reported by Zhang et al. It must be noted, however, that the above-mentioned in vivo studies applied the external magnetic field gradient to the microvasculature regions characterized by relatively slow flow and in the disease conditions, where enhanced capillary leakage and retention effects strongly support the accumulation of SPIONs. Theoretically, the targeting efficiency is expected to grow with increased magnetic forces, decreased flow rate and reduced vessel diameter Nevertheless, the in silico mathematical simulations based on the models of tubular vessels predict that magnetic accumulation of nanoparticles against the hydrodynamic drag force is not possible These models, however, often ignore dipole-dipole interactions between the magnetized particles.

Electrostatic or steric repulsion due to the surface chemistry also plays an important role The contribution of in situ- formed reversible aggregates during high gradient magnetic separation of magnetic nanoparticles has been described earlier by Moeser et al. In support of their hypothesis, a successful accumulation of circulating SPIONs under external magnets has been demonstrated in several in vitro models of straight 2122 and bifurcating channels 1623 Furthermore, our experimental ex vivo work in bovine arteries 1425 showed that accumulation of flowing SPIONs in the arterial wall is achievable under the guidance of a sufficiently strong external magnet, as confirmed by histology, microCT and magnetorelaxometry 14 Despite these promising initialbovine carotid artery model was suboptimal due to the time-consuming procedures of isolation and closing the multiple branches.

To answer the need for an easy to handle and reliable basic research model system for MDT investigations under arterial flow conditions, we therefore developed a new ex vivo model based on the branch-free human umbilical cord arteries. By using particles differing in multiple physicochemical properties e.

As demonstrated here, SPION-1 had by far the largest capacity to accumulate at the artery segment exposed to strongest magnetic field gradient. Steric repulsion, such as the hindrance provided by a coating of the nanoparticle surface e. However, in the case of SPION-1, a strong tendency to sediment and cluster was observed in cell culture media. It is therefore plausible that, in accordance with the findings of Moeser et al.

Despite the fact that these particles constitute a very good model nanosystem for evaluation of various parameters influencing the magnetic capture under flow conditions, SPION-1 being characterised by the tendency to agglomerate, enhanced cellular uptake and relatively high endothelial toxicity 16are not a candidate for further development and potential clinical use. For clinical applications, especially nanoparticle agglomeration may be a decisive factor limiting their use in patients, as it affects both safety and bioavailability.

This could indicate that due to the bulky polymer shell around the particles, the dipole-dipole interactions cannot easily occur in case of SPION SPION-2, the particle type with good biocompatibility profile 28as well as good colloidal and blood stability 29were successfully targeted to the specific artery region under external magnetic field gradient.

Although the observed peak in the iron content under the tip of the magnet was ificantly smaller than for SPION-1, the difference in particle accumulation between magnetically targeted and untargeted samples was very strongly pronounced. Moreover, it is plausible that in physiologic-like situation, where the SPIONs are administered via bolus injection near the target region, the magnetic accumulation of SPION-2 can be further enhanced due to increased probability of dipole-dipole interactions of magnetized SPIONs. Our present study has several limitations, including the use of cell culture media and diluted SPION suspensions in the flow experiments.

The use of cell culture media instead of blood or a fluid with comparable viscosity in the necessity of increasing the flow rate in order to achieve physiologic arterial shear stress levels. Furthermore, in contrast to the bolus administration commonly applied in vivothe use of diluted nanoparticle suspensions decreases the interactions between SPIONs.

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Both these factors are thus likely to impact the magnetic capture efficacy. Furthermore, as the connectors are a stiff and b slightly narrower than the vessel diameter, some recirculation areas at both inflow and outflow zone can occur. In the future, we aim to perform a computational flow dynamics analysis in our model to better estimate the possible effects of local hemodynamic conditions on the SPION capture. Despite the existing limitations, many valuable studies can be devised using this model, including the modifications of arterial wall geometry to mimic the presence of stenosis, or the implantation of stents to investigate the magnetic capture of SPION-loaded cells.

Collectively, these also imply that a precise positioning of the external magnet should allow an efficient targeting of certain types of SPIONs to superficial arteries in vivobut more sophisticated magnetic field geometries will be necessary in the case of arteries localised in the deeper body regions. Moreover, as the rheological behaviour of blood cells in the arterial flow may strongly affect the capturing efficacy, further extensive investigations in the presence of whole blood will be needed to better characterise the margination and accumulation of magnetic particles under arterial flow conditions.

All compounds used were of pharmaceutical Ph. Trichrome stain reagents were purchased from Merck and hematoxylin from Dako Hamburg, Germany. Lauric acid-coated iron oxide nanoparticles were synthesized using a coprecipitation method as described by Tietze et al.

Background

The precipitate was then washed with 1. The resulting lauric acid-coated particles were washed 10 times with 1. The suspension was then dialysed multiple times against ultrapure water. For the preparation of dextran-coated iron oxide nanoparticles, the synthesis method described by Unterweger et al.

The solution was then dialysed against water, concentrated by ultrafiltration and sterile filtered through 0. The extensive physicochemical characterisation of these particles was reported in the publications 2829313233 and is briefly summarised in the Table 1. Human umbilical arteries were isolated from freshly collected umbilical cords kindly provided by the Dept.