Multifunctional Magnetic-fluorescent Nanocomposites for Biomedical Applications

Nanotechnology is a fast-growing area, involving the fabrication and use of nano-sized materials and devices. Various nanocomposite materials play a number of important roles in modern science and technology. Magnetic and fluorescent inorganic nanoparticles are of particular importance due to their broad range of potential applications. It is expected that the combination of magnetic and fluorescent properties in one nanocomposite would enable the engineering of unique multifunctional nanoscale devices, which could be manipulated using external magnetic fields. The aim of this review is to present an overview of bimodal “two-in-one” magnetic-fluorescent nanocomposite materials which combine both magnetic and fluorescent properties in one entity, in particular those with potential applications in biotechnology and nanomedicine. There is a great necessity for the development of these multifunctional nanocomposites, but there are some difficulties and challenges to overcome in their fabrication such as quenching of the fluorescent entity by the magnetic core. Fluorescent-magnetic nanocomposites include a variety of materials including silica-based, dye-functionalised magnetic nanoparticles and quantum dots-magnetic nanoparticle composites. The classification and main synthesis strategies, along with approaches for the fabrication of fluorescent-magnetic nanocomposites, are considered. The current and potential biomedical uses, including biological imaging, cell tracking, magnetic bioseparation, nanomedicine and bio- and chemo-sensoring, of magnetic-fluorescent nanocomposites are also discussed

Novel contrast agents and strategies for MR molecular imaging

Molecular imaging is a rapidly emerging field of research, which can be broadly defined as the in vivo characterization and measurement of biological processes at the cellular and molecular level. In contrast to conventional clinical diagnostic imaging, this novel field focuses on the in vivo visualization of molecular and cellular processes that underlie disease, thereby allowing its early detection and characterization. For this purpose, conventional imaging modalities are combined with the use of so-called targeted contrast agents that are intended to specifically bind to receptors, which are abundantly expressed at diseased sites. The objective of the present thesis was to develop novel methods for magnetic resonance (MR) molecular imaging of apoptosis (programmed cell death) and angiogenesis (the formation of new blood vessels from pre-existing vessels), as these processes play a key role in the etiology as well as treatment of various disorders with a high prevalence, such as cancer and cardiovascular disease. Magnetic resonance imaging (MRI) was used as this imaging modality provides excellent soft tissue contrast at a high spatial resolution throughout the whole body without using ionizing radiation. MRI however suffers from relatively low sensitivity, and therefore potent contrast agents were designed that contained a high quantity of contrast-generating material such as iron oxide crystals or gadolinium ions. Furthermore, novel molecular imaging methods were introduced and applied to examine important aspects of biomarker-specific MRI. Chapter 1 gives a general introduction to contrast agents for MRI, in which basics such as T1 and T2 (*) relaxation, relaxivities (r1 and r2 (*)) and contrast-generating mechanisms, as well as contrast agent applications are discussed. Moreover, the main molecular pathways of apoptosis and angiogenesis are described. The following three chapters concentrate on molecular imaging of the apoptotic process. An important hallmark of apoptosis is the appearance of the phospholipid phosphatidylserine (PS) in the outer leaflet of the cell membrane, which can be exploited as a diagnostic marker for apoptosis by using annexin A5 conjugates. Chapter 2 and 3 contain in vitro studies on annexin A5-functionalized contrast agents, whereas an in vivo application is described in Chapter 4. In Chapter 2 three different types of lipid-based annexin A5-functionalized MR contrast agents were designed with either a micellular or liposomal morphology. MR contrast was generated by incorporation of T1-reducing paramagnetic gadolinium lipids in the lipid (bi)layer or by encapsulation of T2 (*)-reducing superparamagnetic iron oxide particles in the micellular core. Bimodality was obtained by additional inclusion of fluorescent labels, i.e. fluorescent lipids or quantum dots. The resulting annexin A5-functionalized paramagnetic micelles (~ 10 nm), superparamagnetic micelles (~ 10 nm), and paramagnetic liposomes (~ 100 nm) all demonstrated high specificity for apoptotic Jurkat cells, and could be easily detected with MRI and fluorescence microscopy. Their differences in size, magnetic and fluorescent properties provide the possibility to choose the optimal contrast agent for future in vivo applications. Chapter 3 showed specific association of commercially available annexin A5-functionalized polysaccharide-coated iron oxide particles to apoptotic Jurkat cells. Importantly, it was shown that cell membrane association led to internalization of these particles when coincubated with the apoptotic stimulus, which is also expected to occur in vivo. In the present study, marginal differences were observed between the r2 */r2 ratios of membraneassociated and internalized iron oxide particles, which suggested that both T2- and especially T2 (*)-weighted imaging sequences are suitable for their detection. In Chapter 4 annexin A5-functionalized paramagnetic micellular contrast agents were applied for the non-invasive MRI-based detection of PS-exposing cells in atherosclerotic lesions. Both in vivo MRI as well as ex vivo near-infrared fluorescence imaging (NIRF) showed higher signal enhancement in aortas of apoE-/- mice at 24 hours post-injection of the annexin A5-micelles compared with untargeted control micelles. Confocal fluorescence microscopy confirmed association of the target-specific contrast agent with macrophages and apoptotic cells. These results may be of great value for future diagnostics of atherosclerotic lesion type, as the presence of PS-exposing cells is believed to strongly correlate with plaque vulnerability. The last part of this thesis deals with the pharmacokinetic behavior of liposomal contrast agents as presented in Chapter 2. These nanoparticles were coated with poly(ethylene glycol) (PEG), which is known to increase their circulation half-life and should allow for extensive accumulation at the targeted site. However, the resulting target-to-background ratio may be reduced due to circulating agents that are also detected in the MR images. Therefore, in Chapter 5 we incorporated biotin in the bilayer of the paramagnetic liposomes and successfully introduced a novel strategy to rapidly clear these liposomes from the blood circulation in C57BL/6 mice through a so-called avidin chase. Avidininduced clearance was confirmed both with dynamic in vivo MRI and by determination of the Gd content in blood samples ex vivo. In Chapter 6 the avidin chase was applied to remove non-bound RGD-biotin-liposomes from the blood pool in B16F10 tumor-bearing C57BL/6 mice. These liposomes target the avß3 integrin, which is strongly expressed in angiogenic blood vessels. The avidin chase demonstrated that besides target-associated contrast agent, the circulating contrast agent contributed significantly to the MR contrast enhancement in the tumor as well. These results were confirmed by ex vivo fluorescence microscopy. Consequently, this clearance methodology may be used to increase the specificity of molecular MRI of tumor angiogenesis. Finally, Chapter 7 concludes with a general discussion on the preceding chapters, followed by some thoughts on future perspectives of the work presented

Current applications of nanotechnology for magnetic resonance imaging of apoptosis

Apoptosis, or programmed cell death, is a morphologically and biochemically distinct form of cell death, which together with proliferation plays an important role in tissue development and homeostasis. Insufficient apoptosis is important in the pathology of various disorders such as cancer and autoimmune diseases, whereas a high apoptotic activity is associated with myocardial infarction, neurodegenerative diseases, and advanced atherosclerotic lesions. Consequently, apoptosis is recognized as an important therapeutic target, which should be either suppressed, e.g., during an ischemic cardiac infarction, or promoted, e.g., in the treatment of cancerous lesions. Imaging tools to address location, amount, and time course of apoptotic activity non-invasively in vivo are therefore of great clinical use in the evaluation of such therapies. This chapter reviews current literature and new developments in the application of nanoparticles for non-invasive apoptosis imaging. Focus is on functionalized nanoparticle contrast agents for MRimaging and bimodal nanopartide agents that combine magnetic and fluorescent properties

Annexin A5-conjugated quantum dots with a paramagnetic lipidic coating for the multimodal detection of apoptotic cells

Apoptosis, or programmed cell death, plays an important role in the etiol. of a variety of diseases, including cancer. Visualization of apoptosis would allow both early detection of therapy efficiency and evaluation of disease progression. To that aim the authors developed a novel annexin A5-conjugated bimodal nanoparticle. The nanoparticle is composed of a quantum dot that is encapsulated in a paramagnetic micelle to enable its use both for optical imaging and MRI. Multiple recombinant human annexin A5 protein mols. were covalently coupled to the nanoparticle for targeting. In this study the specificity of the annexin A5-conjugated nanoparticles for apoptotic cells was demonstrated both with fluorescence microscopy and MRI, which confirms its potential for the detection of apoptosis with both imaging modalities in vivo

MRI of ICAM-1 Upregulation After Stroke: the Importance of Choosing the Appropriate Target-Specific Particulate Contrast Agent

Magnetic resonance imaging (MRI) with targeted contrast agents provides a promising means for diagnosis and treatment monitoring after cerebrovascular injury. Our goal was to demonstrate the feasibility of this approach to detect the neuroinflammatory biomarker intercellular adhesion molecule-1 (ICAM-1) after stroke and to establish a most efficient imaging procedure. We compared two types of ICAM-1-functionalized contrast agent: T (1)-shortening gadolinium chelate-containing liposomes and T (2) (()*())-shortening micron-sized iron oxide particles (MPIO). Binding efficacy and MRI contrast effects were tested in cell cultures and a mouse stroke model. Both ICAM-1-targeted agents bound effectively to activated cerebrovascular cells in vitro, generating significant MRI contrast-enhancing effects. Direct in vivo MRI-based detection after stroke was only achieved with ICAM-1-targeted MPIO, although both contrast agents showed similar target-specific vascular accumulation. Our study demonstrates the potential of in vivo MRI of post-stroke ICAM-1 upregulation and signifies target-specific MPIO as most suitable contrast agent for molecular MRI of cerebrovascular inflammation

MRI of ICAM-1 upregulation after stroke: the importance of choosing the appropriate target-specific particulate contrast agent

Item does not contain fulltextPURPOSE: Magnetic resonance imaging (MRI) with targeted contrast agents provides a promising means for diagnosis and treatment monitoring after cerebrovascular injury. Our goal was to demonstrate the feasibility of this approach to detect the neuroinflammatory biomarker intercellular adhesion molecule-1 (ICAM-1) after stroke and to establish a most efficient imaging procedure. PROCEDURES: We compared two types of ICAM-1-functionalized contrast agent: T 1-shortening gadolinium chelate-containing liposomes and T2(*)-shortening micron-sized iron oxide particles (MPIO). Binding efficacy and MRI contrast effects were tested in cell cultures and a mouse stroke model. RESULTS: Both ICAM-1-targeted agents bound effectively to activated cerebrovascular cells in vitro, generating significant MRI contrast-enhancing effects. Direct in vivo MRI-based detection after stroke was only achieved with ICAM-1-targeted MPIO, although both contrast agents showed similar target-specific vascular accumulation. CONCLUSIONS: Our study demonstrates the potential of in vivo MRI of post-stroke ICAM-1 upregulation and signifies target-specific MPIO as most suitable contrast agent for molecular MRI of cerebrovascular inflammation