T1 mapping performance and measurement repeatability: results from the multi-national T1 mapping standardization phantom program (T1MES).

BACKGROUND: The T1 Mapping and Extracellular volume (ECV) Standardization (T1MES) program explored T1 mapping quality assurance using a purpose-developed phantom with Food and Drug Administration (FDA) and Conformité Européenne (CE) regulatory clearance. We report T1 measurement repeatability across centers describing sequence, magnet, and vendor performance. METHODS: Phantoms batch-manufactured in August 2015 underwent 2 years of structural imaging, B0 and B1, and "reference" slow T1 testing. Temperature dependency was evaluated by the United States National Institute of Standards and Technology and by the German Physikalisch-Technische Bundesanstalt. Center-specific T1 mapping repeatability (maximum one scan per week to minimum one per quarter year) was assessed over mean 358 (maximum 1161) days on 34 1.5 T and 22 3 T magnets using multiple T1 mapping sequences. Image and temperature data were analyzed semi-automatically. Repeatability of serial T1 was evaluated in terms of coefficient of variation (CoV), and linear mixed models were constructed to study the interplay of some of the known sources of T1 variation. RESULTS: Over 2 years, phantom gel integrity remained intact (no rips/tears), B0 and B1 homogenous, and "reference" T1 stable compared to baseline (% change at 1.5 T, 1.95 ± 1.39%; 3 T, 2.22 ± 1.44%). Per degrees Celsius, 1.5 T, T1 (MOLLI 5s(3s)3s) increased by 11.4 ms in long native blood tubes and decreased by 1.2 ms in short post-contrast myocardium tubes. Agreement of estimated T1 times with "reference" T1 was similar across Siemens and Philips CMR systems at both field strengths (adjusted R2 ranges for both field strengths, 0.99-1.00). Over 1 year, many 1.5 T and 3 T sequences/magnets were repeatable with mean CoVs < 1 and 2% respectively. Repeatability was narrower for 1.5 T over 3 T. Within T1MES repeatability for native T1 was narrow for several sequences, for example, at 1.5 T, Siemens MOLLI 5s(3s)3s prototype number 448B (mean CoV = 0.27%) and Philips modified Look-Locker inversion recovery (MOLLI) 3s(3s)5s (CoV 0.54%), and at 3 T, Philips MOLLI 3b(3s)5b (CoV 0.33%) and Siemens shortened MOLLI (ShMOLLI) prototype 780C (CoV 0.69%). After adjusting for temperature and field strength, it was found that the T1 mapping sequence and scanner software version (both P < 0.001 at 1.5 T and 3 T), and to a lesser extent the scanner model (P = 0.011, 1.5 T only), had the greatest influence on T1 across multiple centers. CONCLUSION: The T1MES CE/FDA approved phantom is a robust quality assurance device. In a multi-center setting, T1 mapping had performance differences between field strengths, sequences, scanner software versions, and manufacturers. However, several specific combinations of field strength, sequence, and scanner are highly repeatable, and thus, have potential to provide standardized assessment of T1 times for clinical use, although temperature correction is required for native T1 tubes at least.

Quantitative T2 mapping of the mouse heart by segmented MLEV phase-cycled T2 preparation.

PURPOSE: A high-quality, reproducible, multi-slice T2-mapping protocol for the mouse heart is presented. METHODS: A T2-prepared sequence with composite 90° and 180° radiofrequency pulses in a segmented MLEV phase cycling scheme was developed. The T2-mapping protocol was optimized using simulations and evaluated with phantoms. RESULTS: Repeatability for determination of myocardial T2 values was assessed in vivo in n = 5 healthy mice on 2 different days. The average baseline T2 of the left ventricular myocardium was 22.5 ± 1.7 ms. The repeatability coefficient for R2 = 1/T2 for measurements at different days was ΔR2 = 6.3 s(−1). Subsequently, T2 mapping was applied in comparison to late-gadolinium-enhancement (LGE) imaging, to assess 1-day-old ischemia/reperfusion (IR) myocardial injury in n = 8 mice. T2 in the infarcts was significantly higher than in remote tissue, whereas remote tissue was not significantly different from baseline. Infarct sizes based on T2 versus LGE showed strong correlation. To assess the time-course of T2 changes in the infarcts, T2 mapping was performed at day 1, 3, and 7 after IR injury in a separate group of mice (n = 16). T2 was highest at day 3, in agreement with the expected time course of edema formation and resolution after myocardial infarction. CONCLUSION: T2 prepared imaging provides high quality reproducible T2 maps of healthy and diseased mouse myocardium.

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

PURPOSE: 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.

Internalization of paramagnetic phosphatidylserine-containing liposomes by macrophages.

BACKGROUND: Inflammation plays an important role in many pathologies, including cardiovascular diseases, neurological conditions and oncology, and is considered an important predictor for disease progression and outcome. In vivo imaging of inflammatory cells will improve diagnosis and provide a read-out for therapy efficacy. Paramagnetic phosphatidylserine (PS)-containing liposomes were developed for magnetic resonance imaging (MRI) and confocal microscopy imaging of macrophages. These nanoparticles also provide a platform to combine imaging with targeted drug delivery. RESULTS: Incorporation of PS into liposomes did not affect liposomal size and morphology up to 12 mol% of PS. Liposomes containing 6 mol% of PS showed the highest uptake by murine macrophages, while only minor uptake was observed in endothelial cells. Uptake of liposomes containing 6 mol% of PS was dependent on the presence of Ca2+ and Mg2+. Furthermore, these 6 mol% PS-containing liposomes were mainly internalized into macrophages, whereas liposomes without PS only bound to the macrophage cell membrane. CONCLUSIONS: Paramagnetic liposomes containing 6 mol% of PS for MR imaging of macrophages have been developed. In vitro these liposomes showed specific internalization by macrophages. Therefore, these liposomes might be suitable for in vivo visualization of macrophage content and for (visualization of) targeted drug delivery to inflammatory cells.

Targeting of ICAM-1 on vascular endothelium under static and shear stress conditions using a liposomal Gd-based MRI contrast agent.

BACKGROUND: The upregulation of intercellular adhesion molecule-1 (ICAM-1) on the endothelium of blood vessels in response to pro-inflammatory stimuli is of major importance for the regulation of local inflammation in cardiovascular diseases such as atherosclerosis, myocardial infarction and stroke. In vivo molecular imaging of ICAM-1 will improve diagnosis and follow-up of patients by non-invasive monitoring of the progression of inflammation. RESULTS: A paramagnetic liposomal contrast agent functionalized with anti-ICAM-1 antibodies for multimodal magnetic resonance imaging (MRI) and fluorescence imaging of endothelial ICAM-1 expression is presented. The ICAM-1-targeted liposomes were extensively characterized in terms of size, morphology, relaxivity and the ability for binding to ICAM-1-expressing endothelial cells in vitro. ICAM-1-targeted liposomes exhibited strong binding to endothelial cells that depended on both the ICAM-1 expression level and the concentration of liposomes. The liposomes had a high longitudinal and transversal relaxivity, which enabled differentiation between basal and upregulated levels of ICAM-1 expression by MRI. The liposome affinity for ICAM-1 was preserved in the competing presence of leukocytes and under physiological flow conditions. CONCLUSION: This liposomal contrast agent displays great potential for in vivo MRI of inflammation-related ICAM-1 expression.

Contrast-enhanced MRI of murine myocardial infarction - part I.

The use of contrast agents has added considerable value to the existing cardiac MRI toolbox that can be used to study murine myocardial infarction, as it enables detailed in vivo visualization of the molecular and cellular processes that occur in the infarcted and remote tissue. A variety of non-targeted and targeted contrast agents to study myocardial infarction are available and under development. Manganese, which acts as a calcium analogue, can be used to assess cell viability. Traditionally, low-molecular-weight Gd-containing contrast agents are employed to measure infarct size in a late gadolinium enhancement experiment. Gd-based blood-pool agents are used to study the vascular status of the myocardium. The use of targeted contrast agents facilitates more detailed imaging of pathophysiological processes in the acute and chronic infarct. Cell death was visualized by contrast agents functionalized with annexin A5 that binds specifically to phosphatidylserine accessible on dying cells and with an agent that binds to the exposed DNA of dead cells. Inflammation in the myocardium was depicted by contrast agents that target cell adhesion molecules expressed on activated endothelium, by contrast agents that are phagocytosed by inflammatory cells, and by using a probe that targets enzymes excreted by inflammatory cells. Cardiac remodeling processes were visualized with a contrast agent that binds to angiogenic vasculature and with an MR probe that specifically binds to collagen in the fibrotic myocardium. These recent advances in murine contrast-enhanced cardiac MRI have made a substantial contribution to the visualization of the pathophysiology of myocardial infarction, cardiac remodeling processes and the progression to heart failure, which helps to design new treatments. This review discusses the advances and challenges in the development and application of MRI contrast agents to study murine myocardial infarction.

Contrast-enhanced MRI of murine myocardial infarction - part II.

Mouse models are increasingly used to study the pathophysiology of myocardial infarction in vivo. In this area, MRI has become the gold standard imaging modality, because it combines high spatial and temporal resolution functional imaging with a large variety of methods to generate soft tissue contrast. In addition, (target-specific) MRI contrast agents can be employed to visualize different processes in the cascade of events following myocardial infarction. Here, the MRI sequence has a decisive role in the detection sensitivity of a contrast agent. However, a straightforward translation of clinically available protocols for human cardiac imaging to mice is not feasible, because of the small size of the mouse heart and its extremely high heart rate. This has stimulated intense research in the development of cardiac MRI protocols specifically tuned to the mouse with regard to timing parameters, acquisition strategies, and ECG- and respiratory-triggering methods to find an optimal trade-off between sensitivity, scan time, and image quality. In this review, a detailed analysis is given of the pros and cons of different mouse cardiac MR imaging methodologies and their application in contrast-enhanced MRI of myocardial infarction.

Regional contrast agent quantification in a mouse model of myocardial infarction using 3D cardiac T1 mapping.

BACKGROUND: Quantitative relaxation time measurements by cardiovascular magnetic resonance (CMR) are of paramount importance in contrast-enhanced studies of experimental myocardial infarction. First, compared to qualitative measurements based on signal intensity changes, they are less sensitive to specific parameter choices, thereby allowing for better comparison between different studies or during longitudinal studies. Secondly, T1 measurements may allow for quantification of local contrast agent concentrations. In this study, a recently developed 3D T1 mapping technique was applied in a mouse model of myocardial infarction to measure differences in myocardial T1 before and after injection of a liposomal contrast agent. This was then used to assess the concentration of accumulated contrast agent. MATERIALS AND METHODS: Myocardial ischemia/reperfusion injury was induced in 8 mice by transient ligation of the LAD coronary artery. Baseline quantitative T1 maps were made at day 1 after surgery, followed by injection of a Gd-based liposomal contrast agent. Five mice served as control group, which followed the same protocol without initial surgery. Twenty-four hours post-injection, a second T1 measurement was performed. Local ΔR1 values were compared with regional wall thickening determined by functional cine CMR and correlated to ex vivo Gd concentrations determined by ICP-MS. RESULTS: Compared to control values, pre-contrast T1 of infarcted myocardium was slightly elevated, whereas T1 of remote myocardium did not significantly differ. Twenty-four hours post-contrast injection, high ΔR1 values were found in regions with low wall thickening values. However, compared to remote tissue (wall thickening > 45%), ΔR1 was only significantly higher in severe infarcted tissue (wall thickening < 15%). A substantial correlation (r = 0.81) was found between CMR-based ΔR1 values and Gd concentrations from ex vivo ICP-MS measurements. Furthermore, regression analysis revealed that the effective relaxivity of the liposomal contrast agent was only about half the value determined in vitro. CONCLUSIONS: 3D cardiac T1 mapping by CMR can be used to monitor the accumulation of contrast agents in contrast-enhanced studies of murine myocardial infarction. The contrast agent relaxivity was decreased under in vivo conditions compared to in vitro measurements, which needs consideration when quantifying local contrast agent concentrations.

Contrast enhancement by differently sized paramagnetic MRI contrast agents in mice with two phenotypes of atherosclerotic plaque.

Interest in the use of contrast-enhanced MRI to enable in vivo specific characterization of atherosclerotic plaques is increasing. In this study the intrinsic ability of three differently sized gadolinium-based contrast agents to permeate different mouse plaque phenotypes was evaluated with MRI. A tapered cast was implanted around the right carotid artery of apoE(-/-) mice to induce two different plaque phenotypes: a thin cap fibroatheroma (TCFA) and a non-TCFA lesion. Both plaques were allowed to develop over 6 and 9 weeks, leading to an intermediate and advanced lesion, respectively. Signal enhancement in the carotid artery wall, following intravenous injection of Gd-HP-DO3A as well as paramagnetic micelles and liposomes was evaluated. In vivo T(1) -weighted MRI plaque enhancement characteristics were complemented by fluorescence microscopy and correlated to lesion phenotype. The two smallest contrast agents, i.e. Gd-HP-DO3A and micelles, were found to enhance contrast in T(1) -weighted MR images of all investigated plaque phenotypes. Maximum contrast enhancement ranged between 53 and 70% at 6 min after injection of Gd-HP-DO3A with highest enhancement and longest retention in the non-TCFA lesion. Twenty-four hours after injection of micelles maximum contrast enhancement ranged between 24 and 35% in all plaque phenotypes. Administration of the larger liposomes did not cause significant contrast enhancement in the atherosclerotic plaques. Confocal fluorescence microscopy confirmed the MRI-based differences in plaque permeation between micelles and liposomes. Plaque permeation of contrast agents was strongly dependent on size. Our results implicate that, when equipped with targeting ligands, liposomes are most suitable for the imaging of plaque-associated endothelial markers due to low background enhancement, whereas micelles, which accumulate extravascularly on a long timescale, are suited for imaging of less abundant markers inside plaques. Low molecular weight compounds may be employed for target-specific imaging of highly abundant extravascular plaque-associated targets.

Three-dimensional T1 mapping of the mouse heart using variable flip angle steady-state MR imaging.

Cardiac MR T(1) mapping is a promising quantitative imaging tool for the diagnosis and evaluation of cardiomyopathy. Here, we present a new preclinical cardiac MRI method enabling three-dimensional T(1) mapping of the mouse heart. The method is based on a variable flip angle analysis of steady-state MR imaging data. A retrospectively triggered three-dimensional FLASH (fast low-angle shot) sequence (3D IntraGate) enables a constant repetition time and maintains steady-state conditions. 3D T(1) mapping of the complete mouse heart could be achieved in 20 min. High-quality, bright-blood T(1) maps were obtained with homogeneous T(1) values (1764 ± 172 ms) throughout the myocardium. The repeatability coefficient of R(1) (1/T(1) ) in a specific region of the mouse heart was between 0.14 and 0.20 s(-1) , depending on the number of flip angles. The feasibility for detecting regional differences in ΔR(1) was shown with pre- and post-contrast T(1) mapping in mice with surgically induced myocardial infarction, for which ΔR(1) values up to 0.83 s(-1) were found in the infarct zone. The sequence was also investigated in black-blood mode, which, interestingly, showed a strong decrease in the apparent mean T(1) of healthy myocardium (905 ± 110 ms). This study shows that 3D T(1) mapping in the mouse heart is feasible and can be used to monitor regional changes in myocardial T(1), particularly in relation to pathology and in contrast-enhanced experiments to estimate local concentrations of (targeted) contrast agent.

Mouse myocardial first-pass perfusion MR imaging.

A first-pass myocardial perfusion sequence for mouse cardiac MRI is presented. A segmented ECG-triggered acquisition combined with parallel imaging acceleration was used to capture the first pass of a Gd-DTPA bolus through the mouse heart with a temporal resolution of 300-400 msec. The method was applied in healthy mice (N = 5) and in mice with permanent occlusion of the left coronary artery (N = 6). Baseline semiquantitative perfusion values of healthy myocardium showed excellent reproducibility. Infarct regions revealed a significant decrease in the semiquantitative myocardial perfusion values (0.05 ± 0.02) compared to remote myocardium (0.20 ± 0.04). Myocardial areas of decreased perfusion correlated well to infarct areas identified on the delayed-enhancement scans. This protocol is a valuable addition to the mouse cardiac MRI toolbox for preclinical studies of ischemic heart disease.

Switching on the lights for gene therapy.

Strategies for non-invasive and quantitative imaging of gene expression in vivo have been developed over the past decade. Non-invasive assessment of the dynamics of gene regulation is of interest for the detection of endogenous disease-specific biological alterations (e.g., signal transduction) and for monitoring the induction and regulation of therapeutic genes (e.g., gene therapy). To demonstrate that non-invasive imaging of regulated expression of any type of gene after in vivo transduction by versatile vectors is feasible, we generated regulatable herpes simplex virus type 1 (HSV-1) amplicon vectors carrying hormone (mifepristone) or antibiotic (tetracycline) regulated promoters driving the proportional co-expression of two marker genes. Regulated gene expression was monitored by fluorescence microscopy in culture and by positron emission tomography (PET) or bioluminescence (BLI) in vivo. The induction levels evaluated in glioma models varied depending on the dose of inductor. With fluorescence microscopy and BLI being the tools for assessing gene expression in culture and animal models, and with PET being the technology for possible application in humans, the generated vectors may serve to non-invasively monitor the dynamics of any gene of interest which is proportionally co-expressed with the respective imaging marker gene in research applications aiming towards translation into clinical application.