17 research outputs found
Novel contrast agents and strategies for MR molecular imaging
No full textMolecular 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
MRI contrast agents : current status and future perspectives
No full textA review. Magnetic Resonance Imaging (MRI) is increasingly used in clin. diagnostics, for a rapidly growing no. of indications. The MRI technique is non-invasive and can provide information on the anatomy, function and metab. of tissues in vivo. MRI scans of tissue anatomy and function make use of the two hydrogen atoms in water to generate the image. Apart from differences in the local water content, the basic contrast in the MR image mainly results from regional differences in the intrinsic relaxation times T1 and T2, each of which can be independently chosen to dominate image contrast. However, the intrinsic contrast provided by the water T1 and T2 and changes in their values brought about by tissue pathol. are often too limited to enable a sensitive and specific diagnosis. For that reason increasing use is made of MRI contrast agents that alter the image contrast following i.v. injection. The degree and location of the contrast changes provide substantial diagnostic information. Certain contrast agents are predominantly used to shorten the T1 relaxation time and these are mainly based on low-mol. wt. chelates of the gadolinium ion (Gd3+). The most widely used T2 shortening agents are based on iron oxide (FeO) particles. Depending on their chem. compn., mol. structure and overall size, the in vivo distribution vol. and pharmacokinetic properties vary widely between different contrast agents and these largely det. their use in specific diagnostic tests. This review describes the current status, as well as recent and future developments of MRI contrast agents with focus on applications in oncol. First the basis of MR image contrast and how it is altered by contrast agents will be discussed. After some considerations on bioavailability and pharmacokinetics, specific applications of contrast agents will be presented according to their specific purposes, starting with non-specific contrast agents used in classical contrast enhanced magnetic resonance angiog. (MRA) and dynamic contrast enhanced MRI. Next targeted contrast agents, which are actively directed towards a specific mol. target using an appropriate ligand, functional contrast agents, mainly used for functional brain and heart imaging, smart contrast agents, which generate contrast as a response to a change in their phys. environment as a consequence of some biol. process, and finally cell labeling agents will be presented. To conclude some future perspectives are discussed
In Vivo Molecular MRI of ICAM-1 Expression on Endothelium and Leukocytes from Subacute to Chronic Stages After Experimental Stroke
No full textMolecular MRI allows in vivo detection of vascular cell adhesion molecules expressed on inflamed endothelium, which enables detection of specific targets for anti-neuroinflammatory treatment. We explored to what extent MR contrast agent targeted to intercellular adhesion molecule-1 (ICAM-1) could detect endothelial- and leukocyte-associated ICAM-1 expression at different stages after experimental stroke. Furthermore, we assessed potential interfering effects of ICAM-1-targeted contrast agent on post-stroke lesion growth. Micron-sized particles of iron oxide (MPIO) functionalized with control IgG (IgG-MPIO) or anti-ICAM-1 antibody (αICAM-1-MPIO) were administrated at 1, 2, 3, 7, and 21 days after unilateral transient middle cerebral artery occlusion in mice, followed by in vivo MRI and postmortem immunohistochemistry. αICAM-1-MPIO induced significant contrast effects in the lesion core on post-stroke days 1, 2, and 3, and in the lesion borderzone and contralesional tissue on post-stroke day 2. αICAM-1-MPIO were confined to ICAM-1-positive vessels and occasionally co-localized with leukocytes. On post-stroke day 21, abundant leukocyte-associated αICAM-1-MPIO was immunohistochemically detected in the lesion core. However, MRI-based detection of αICAM-1-MPIO-labeled leukocytes was confounded by pre-contrast MRI hypointensities, presumably caused by phagocytosed blood remains. IgG-MPIO did not induce significant MRI contrast effects at 1 h after injection. Lesion development was not affected by injection of αICAM-1-MPIO or IgG-MPIO. αICAM-1-MPIO are suitable for in vivo MRI of ICAM-1 expression on vascular endothelium and leukocytes at different stages after stroke. Development of clinically applicable MPIO may offer unique opportunities for MRI-based diagnosis of neuroinflammation and identification of anti-inflammatory targets in acute stroke patients
Magnetic resonance imaging of local and remote vascular remodelling after experimental stroke
No full textThe pattern of vascular remodelling in relation to recovery after stroke remains largely unclear. We used steady-state contrast-enhanced magnetic resonance imaging to assess the development of cerebral blood volume and microvascular density in perilesional and exofocal areas from (sub)acutely to chronically after transient stroke in rats. Microvascular density was verified histologically after infusion with Evans Blue dye. At day 1, microvascular cerebral blood volume and microvascular density were reduced in and around the ischemic lesion (intralesional borderzone: microvascular cerebral blood volume = 72 ± 8%; microvascular density = 76 ± 8%) (P < 0.05), while total cerebral blood volume remained relatively unchanged. Perilesional microvascular cerebral blood volume and microvascular density subsequently normalized (day 7) and remained relatively stable (day 70). In remote ipsilateral areas in the thalamus and substantia nigra - not part of the ischemic lesion - microvascular density gradually increased between days 1 and 70 (thalamic ventral posterior nucleus: microvascular density = 119 ± 9%; substantia nigra: microvascular density = 122 ± 8% (P < 0.05)), which was confirmed histologically. Our data indicate that initial microvascular collapse, with maintained collateral flow in larger vessels, is followed by dynamic revascularization in perilesional tissue. Furthermore, progressive neovascularization in non-ischemic connected areas may offset secondary neuronal degeneration and/or contribute to non-neuronal tissue remodelling. The complex spatiotemporal pattern of vascular remodelling, involving regions outside the lesion territory, may be a critical endogenous process to promote post-stroke brain reorganization
Lipid-based nanoparticles for contrast-enhanced MRI and molecular imaging
No full textIn the field of MR imaging and especially in the emerging field of cellular and molecular MR imaging, flexible strategies to synthesize contrast agents that can be manipulated in terms of size and composition and that can be easily conjugated with targeting ligands are required. Furthermore, the relaxivity of the contrast agents, especially for molecular imaging applications, should be very high to deal with the low sensitivity of MRI. Lipid-based nanoparticles, such as liposomes or micelles, have been used extensively in recent decades as drug carrier vehicles. A relatively new and promising application of lipidic nanoparticles is their use as multimodal MR contrast agents. Lipids are amphiphilic molecules with both a hydrophobic and a hydrophilic part, which spontaneously assemble into aggregates in an aqueous environment. In these aggregates, the amphiphiles are arranged such that the hydrophobic parts cluster together and the hydrophilic parts face the water. In the low concentration regime, a wide variety of structures can be formed, ranging from spherical micelles to disks or liposomes. Furthermore, a monolayer of lipids can serve as a shell to enclose a hydrophobic core. Hydrophobic iron oxide particles, quantum dots or perfluorocarbon emulsions can be solubilized using this approach. MR-detectable and fluorescent amphiphilic molecules can easily be incorporated in lipidic nanoparticles. Furthermore, targeting ligands can be conjugated to lipidic particles by incorporating lipids with a functional moiety to allow a specific interaction with molecular markers and to achieve accumulation of the particles at disease sites. In this review, an overview of different lipidic nanoparticles for use in MRI is given, with the main emphasis on Gd-based contrast agents. The mechanisms of particle formation, conjugation strategies and applications in the field of contrast-enhanced, cellular and molecular MRI are discussed
Uric Acid Is Protective After Cerebral Ischemia/Reperfusion in Hyperglycemic Mice
No full textHyperglycemia at stroke onset is associated with poor long-term clinical outcome in numerous studies. Hyperglycemia induces intracellular acidosis, lipid peroxidation, and peroxynitrite production resulting in the generation of oxidative and nitrosative stress in the ischemic tissue. Here, we studied the effects of acute hyperglycemia on in vivo intercellular adhesion molecule-1 (ICAM-1) expression, neutrophil recruitment, and brain damage after ischemia/reperfusion in mice and tested whether the natural antioxidant uric acid was protective. Hyperglycemia was induced by i.p. administration of dextrose 45 min before transient occlusion of the middle cerebral artery. Magnetic resonance imaging (MRI) was performed at 24 h to measure lesion volume. A group of normoglycemic and hyperglycemic mice received an i.v. injection of micron-sized particles of iron oxide (MPIOs), conjugated with either anti-ICAM-1 antibody or control IgG, followed by T2*w MRI. Neutrophil infiltration was studied by immunofluorescence and flow cytometry. A group of hyperglycemic mice received an i.v. infusion of uric acid (16 mg/kg) or the vehicle starting after 45 min of reperfusion. ICAM-1-targeted MPIOs induced significantly larger MRI contrast-enhancing effects in the ischemic brain of hyperglycemic mice, which also showed more infiltrating neutrophils and larger lesions than normoglycemic mice. Uric acid reduced infarct volume in hyperglycemic mice but it did not prevent vascular ICAM-1 upregulation and did not significantly reduce the number of neutrophils in the ischemic brain tissue. In conclusion, hyperglycemia enhances stroke-induced vascular ICAM-1 and neutrophil infiltration and exacerbates the brain lesion. Uric acid reduces the lesion size after ischemia/reperfusion in hyperglycemic mice
Paramagnetic and fluorescent liposomes for target-specific imaging and therapy of tumor angiogenesis
No full textAngiogenesis is essential for tumor growth and metastatic potential and for that reason considered an important target for tumor treatment. Noninvasive imaging technologies, capable of visualizing tumor angiogenesis and evaluating the efficacy of angiostatic therapies, are therefore becoming increasingly important. Among the various imaging modalities, magnetic resonance imaging (MRI) is characterized by a superb spatial resolution and anatomical soft-tissue contrast. Revolutionary advances in contrast agent chemistry have delivered versatile angiogenesis-specific molecular MRI contrast agents. In this paper, we review recent advances in the preclinical application of paramagnetic and fluorescent liposomes for noninvasive visualization of the molecular processes involved in tumor angiogenesis. This liposomal contrast agent platform can be prepared with a high payload of contrast generating material, thereby facilitating its detection, and is equipped with one or more types of targeting ligands for binding to specific molecules expressed at the angiogenic site. Multimodal liposomes endowed with contrast material for complementary imaging technologies, e.g., MRI and optical, can be exploited to gain important preclinical insights into the mechanisms of binding and accumulation at angiogenic vascular endothelium and to corroborate the in vivo findings. Interestingly, liposomes can be designed to contain angiostatic therapeutics, allowing for image-supervised drug delivery and subsequent monitoring of therapeutic efficacy
Nanoparticulate assemblies of amphiphiles and diagnostically active materials for multimodality imaging
No full textNo abstract
Annexin A5-conjugated quantum dots with a paramagnetic lipidic coating for the multimodal detection of apoptotic cells
No full textApoptosis, 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
