Elsevier

Acta Biomaterialia

Volume 24, 15 September 2015, Pages 266-278
Acta Biomaterialia

Full length article
Self-assembled microbubbles as contrast agents for ultrasound/magnetic resonance dual-modality imaging

https://doi.org/10.1016/j.actbio.2015.06.025Get rights and content

Abstract

In this work, superparamagnetic self-assembled microbubbles (SAMBs) consisting of “Poly(acrylic acid)-Iron oxide nanoparticles-Polyamine” sandwich-like shells and tetradecafluorohexane cores were fabricated by a template-free self-assembly approach. The SAMBs exhibit not only magnetic resonance (MR) T2 imaging functionality, but also ultrasound (US) image contrast, showing great potential as US/MR dual contrast agents. The diameters of the SAMBs can be tuned easily from 450 nm to 1300 nm by changing the precursor ratio, and this size variation directly affects their in vitro MRI and US signals. The SAMBs also exhibit in vivo contrast enhancement capabilities in rat liver with injection through portal vein, for both MR and US imaging. Additionally, the biodistribution of SAMBs over time suggests normal systemic metabolic activity through the spleen. The results show that the Fe content in rat liver reduces to a level of which Fe cannot be detected in 45 days. The SAMBs exhibit no obvious damage to the primary organs of rat during the metabolic process, indicating their good biocompatibility in vivo.

Graphical abstract

Microbubbles bearing “Poly(acrylic acid)-Iron oxide nanoparticles-Polyamine” sandwich-like shells were fabricated by a self-assembly approach for MR/US dual-modal imaging applications.

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Introduction

Iron oxide nanoparticles (IONPs) act as T2-weighted magnetic resonance imaging (MRI) contrast agents based on their superparamagnetic properties and have great potential in biomedical applications [1], [2]. The contrast enhancement provided by IONPs contrast agents is due to the acceleration on the transverse relaxation of the water protons, caused by their interaction with the neighboring contrast agents. This results in a T2 and T2 shortening. Previous studies have revealed that the T1 relaxivity, r2, strongly depends on the volume magnetization of the particle and the radius of the particle [3], [4], [5], [6]. Hence, embedding IONPs into microcapsules will essentially affect T2 and T2 which are related to the spherical structure of the cluster and to the magnetic field distribution around it. Based on the IONP-conjugated microcapsule structure, microbubbles have been proven to endow the composites with unique sonic properties [7]. For better visualization of specific tissues, microbubbles and perfluorocarbon emulsions of various formulations have been developed and applied clinically as US contrast agents. However, they have suffered from instability due to ultrasonic pressures, broad size distributions, and poor circulating acoustic contrast properties [8], [9]. Inorganic nanoparticle/polymer composites and inorganic-material based US contrast agents have recently attracted great attention due to their tunable particle diameters, good compatibility and superior stability over that provided by traditional organic microbubbles under US exposure [10], [11], [12]. Meanwhile, IONP-conjugated microbubbles behave as MRI/US dual-modal contrast agents, thus providing more precise diagnostic molecular imaging that combines the susceptibility of US imaging and the exquisite soft tissue contrast provided by the MRI modality [13], [14].

Dual-modal contrast agents for T2-weighted MR and US imaging can be categorized into two general classes, including IONP-conjugated microbubbles bearing lipid shells [13] and polymer shells [7], [10], [15], [16], [17], [18]. IONP-conjugated microbubbles bearing lipid shells have certain limitations that arise not only due to the tedious synthetic procedure via the thin film hydration method required for their preparation, but also because of their instability that causes them to be potentially toxic [13]. In comparison with magnetic lipid microbubbles, IONP-conjugated microbubbles bearing biodegradable polymer shells are provided with physical rigidity and may be loaded with drugs or other cargos. The biocompatible outer coating provides a biological interface and a potential “scaffold” for targeting species. The only reported method to fabricate IONP-conjugated polymer microbubbles is through the double emulsion method, which builds a platform for dual-mode MRI/US contrast agents [7], [10], [15], [16], [17], [18]. However, only hydrophobic polymers can be selected to form the bubble’s shell during the double emulsion process, and the removal of the residual emulsifier is another concern during the synthesis. Hence, it remains a challenge to develop a facile method to fabricate a multifunctional contrast agent for dual MR/US imaging.

The self-assembly approach is a widely used method to fabricate nanoparticle/polymer composite microcapsules with diverse compositions and sizes [19], [20], [21], [22]. The self-assembly method can be divided into template-based self-assembly and template-free self-assembly strategies. In particular, the layer-by-layer (LBL) self-assembly method is a well-established template-based method for constructing superparamagnetic IONP-based microcapsules [5]. However, the LBL self-assembly method involves a time-consuming process to form a template with a fixed size and also requires the subsequent removal of the template to generate hollow capsules. Recently, a novel self-assembly method known as the polyamine–salt aggregate (PSA) assembly method was proposed by Wong et al. [23]. This self-assembly route involves the formation of polymer aggregates and the subsequent deposition of particles around these aggregates to form microcapsules. The entire process is based on electrostatic interactions between oppositely charged components. In addition to providing a simplified procedure compared to the LBL method, the PSA approach can also be applied to encapsulate various agents and the shell thickness can be conveniently tuned [3], [24], [25], [26], [27], [28], [29]. These features provide the PSA assembly strategy with advantages in fabricating nanoparticle-conjugated microbubbles.

In this study, we developed a novel self-assembly approach to fabricate a multifunctional iron oxide nanoparticle-conjugated microbubble as a contrast agent for dual-modality MR/US imaging. The self-assembled microbubble has a unique triple layered shell constructed by “Poly(acrylic acid)-IONPs-Polyamine” composites. The MR and US imaging properties of self-assembled microbubbles were investigated both in vitro and in vivo. Additionally, metabolic investigations and histological evaluations were performed to characterize the biocompatibility of the microbubbles.

Section snippets

Materials

Anhydrous ferric chloride, anhydrous ethanol, N,-N-dimethyl formamide (DMF), sodium oleate, n-hexane, acetone, 1,2-dichlorobenzene, toluene, dimethyl sulfoxide (DMSO), citric acid monohydrate, trisodium citrate dehydrate, fluorescein sodium salt and glacial acetic acid (AR) were purchased from Sinopharm Chemical Reagent Co., Ltd. Oleic acid (OA, 90%), 1-octadecene (ODE) (90%), a poly(allylamine) solution (Mw = 65000, 20 wt.% in water), poly(acrylic acid) (PAA, Mw = 1800), tetradecafluorohexane (C6F14

Fabrication and characterization of self-assembled microbubble (SAMB)

Scheme 1 shows the approach used to prepare the self-assembled MR/US dual imaging microbubbles with a “sandwich-like” structure. The fabrication of SAMB initially involves the self-assembly of the precursors to form self-assembled microcapsule (SAMC). A gas-filling process is subsequently employed to obtain SAMB. During this overall procedure, the fabrication of SAMC involves three steps. Firstly, cationic polyamines undergo complexation with sodium citrate (Cit) to form polyamine–citrate

Conclusions

A novel self-assembly approach has been developed to fabricate magnetic self-assembled microbubbles bearing “PAA-IONP-Polyamine” sandwich-like shells. The structures, sizes and the magnetic properties of the SAMBs were investigated. It was found that the sizes of SAMBs could be easily tuned by changing the ratio of the polyamine within the precursors, while the different SAMBs contained almost identical magnetite contents. The superparamagnetic SAMBs exhibited MR/US dual imaging contrast

Acknowledgments

The authors thank the financial support of the National Natural Science Foundation of China (Nos. 21174082, 21374061, 81100295), the SJTU SMC-Chen Xing Young Scholars Award, Program of New Century Excellent Talent in University (NCET-13-0360), International Science and Technology Cooperation Project of the Science and Technology Commission of Shanghai Municipality (No. 14520710300) and the Instrumental Analysis Center of the SJTU.

References (37)

  • F. Tranquart et al.

    Clinical use of ultrasound tissue harmonic imaging

    Ultrasound Med. Biol.

    (1999)
  • Q.L. Vuong et al.

    A universal scaling law to predict the efficiency of magnetic nanoparticles as MRI T2-contrast agents

    Adv. Healthcare Mater.

    (2012)
  • M.P. Morales et al.

    Relaxation times of colloidal iron platinum in polymer matrixes

    J. Mater. Chem.

    (2009)
  • M. Hahn et al.

    Nanoparticles as contrast agents for in-vivo bioimaging: current status and future perspectives

    Anal. Bioanal. Chem.

    (2011)
  • F. Kiessling et al.

    Ultrasound microbubbles for molecular diagnosis, therapy, and theranostics

    J. Nucl. Med.

    (2012)
  • B. Xu et al.

    “Two-in-one” fabrication of Fe3O4/MePEG-PLA composite nanocapsules as a potential ultrasonic/MRI dual contrast agent

    Langmuir

    (2011)
  • L.G. Delogu et al.

    Functionalized multiwalled carbon nanotubes as ultrasound contrast agents

    Proc. Natl. Acad. Sci.

    (2012)
  • F.A. Jaffer et al.

    Molecular imaging in the clinical arena

    JAMA

    (2005)
  • View full text