In vitro and in vivo evaluation of superparamagnetic iron oxide nanoparticles coated by bisphosphonates: the effects of electrical charge and molecule length.

Physicochemical coating properties are often considered to be determining factors for in vivo characteristics of superparamagnetic iron oxide nanoparticles, used as contrast agent in Magnetic Resonance Imaging (MRI). To investigate the electrical charge (modified by zero, one or two ammonium groups) and the molecule length (3, 5 or 7 methylene chains) effects of bisphosphonate-type coatings, we assessed the complement activation, in vivo plasma and tissue relaxation time alterations of intravenously injected small iron oxide nanoparticles (<25 nm) on male healthy Wistar rats. The presence of ammonium groups induces a weak activation of the complement whatever the size and the concentration of particles, whereas hydroxyethylenebisphosphonate (HEBP)-coated particles are poor complement activators only at the lowest concentration. In vivo, HEBP-coated nanoparticles have the greatest prolonged relaxation time effects, despite their higher negative electrical charge, contrary to two ammonium bearing coatings. No significant differences were observed between mono-ammonium molecular coatings.

A starch-based microparticulate system dedicated to diagnostic and therapeutic nuclear medicine applications.

The aim of this work was to develop a new microparticulate system able to form a complex with radionuclides with a high yield of purity for diagnostic or therapeutic applications. Owing to its properties potato starch was chosen as starting material and modified by oxidization and coupling of a ligand (polyamine) enabling modified starch to chelate radionuclides. The choice of suitable experiments was based on a combination of a Rechtschaffner experimental design and a surface response design to determine the influence of experimental parameters and to optimize the final product. Starch-based microparticle formulations from the experimental plans were compared and characterized through particle size analysis, scanning electron microscopy, elemental analysis and, for the most promising formulations, by in vitro labeling stability studies and determination of free polyamine content or in vivo imaging studies. The mechanism of starch-based microparticle degradation was identified by means of size measurements. The results of the Rechtschaffner design showed the positive qualitative effect of the temperature and the duration of coupling reaction whereas surface response analysis clearly showed that, by increasing the oxidization level and starch concentration, the nitrogen content in the final product is increased. In vitro and in vivo characterization led to identification of the best formulation. With a size around 30 µm, high radiochemical purity (over 95%) and a high signal-to-noise ratio (over 600), the new starch-based microparticulate system could be prepared as ready-to-use kits and sterilized without modification of its characteristics, and thus meet the requirement for in vivo diagnostic and therapeutic applications.

Susceptibility gradient quantization by MRI signal response mapping (SIRMA) to dephaser.

PURPOSE: Susceptibility effects are a very efficient source of contrast in magnetic resonance imaging. However, detection is hampered by the fact the induced contrast is negative. In this work, the SIgnal Response MApping (SIRMA) to dephaser method is proposed to map susceptibility gradient to improve visualization. METHODS: In conventional gradient echo acquisitions, the echo formation of susceptibility affected spins is shifted in k-space, the shift being proportional to the susceptibility gradient. Susceptibility gradients map can be produced by measuring this induced shifts. The SIRMA method measures these shifts from a series of dephased images collected with additional incremental dephasers. These additional dephasers correspond either to a slice refocusing gradient offset or to a reconstruction window off-centering. The signal intensity profile as a function of the additional dephaser was determined on a pixel-by-pixel basis from the ensemble of dephased images. Susceptibility affected voxels presented a signal response profile maximum shifted compared to nonaffected voxels ones. Shift magnitude and sign were measured for each pixel to determine susceptibility gradients and produce a susceptibility gradient map. RESULTS: In vitro experiments demonstrated the ability of the method to map gradient inhomogeneities induced by a cylinder. Quantization accuracy was evaluated comparing SIRMA images and simulations performed on the well-characterized air filled cylinder model. Performances of the SIRMA method, evaluated in vitro on cylinders filled with various superparamagnetic iron oxide SPIO concentrations, showed limited influence of acquisition parameters. Robustness of the method was then assessed in vivo after an infusion of SPIO-loaded nanocapsules into the rat brain using a convection-enhanced drug delivery approach. The region of massive susceptibility gradient induced by the SPIO-loaded nanocapsules was clearly delineated on SIRMA maps and images were compared to T2* weighted images, Susceptibility Gradient Map (SGM), and histological Perl's staining slice. The potential for quantitative evaluation of SPIO distribution volume was demonstrated. CONCLUSIONS: The proposed method is a promising technique for a wide range of applications especially in molecular or cellular imaging with respect to its quantitative nature and its computational simplicity.

In vivo evaluation of lipid nanocapsules as a promising colloidal carrier for paclitaxel.

Paclitaxel-loaded lipid nanocapsules (PX-LNC) exhibit interesting in vitro characteristics with improved antitumoral activity compared with free PX formulation. Biodistribution studies were realized with the use of (14)C-trimyristin ((14)C-TM) or (14)C-phosphatidylcholine ((14)C-PC) whereas antitumoral activity of PX-LNC formulations was based on the animal survival in a chemically induced hepatocellular carcinoma (HCC) model in Wistar rats. Blood concentration-time profiles for both labeled (14)C-TM-LNC and (14)C-PC-LNC were similar; the t(1/2) and MRT values (over 2h and close to 3h, respectively, for both formulations) indicated the long circulating properties of the LNC carrier with a slow distribution and elimination phase. Survival curves of paclitaxel treated groups showed a statistical significant difference compared to the control survival curve (P=0.0036 and 0.0408). Animals treated with 4x 70 mg/m(2) of PX-LNC showed the most significant increase in mean survival times compared to the controls (IST(mean) 72%) and cases of long-term survivors were preferentially observed in the PX-LNC treated group (37.5%; 3/8). These results demonstrate the great interest to use LNC as drug delivery system for paclitaxel, permitting with an equivalent therapeutic efficiency to avoid the use of excipients such as polyoxyethylated castor oil for its formulation.

Evaluating SPIO-labelled cell MR efficiency by three-dimensional quantitative T2* MRI.

An in vitro MR-assay for superparamagnetic iron oxide (SPIO) particle cell labelling assessment via three-dimensional quantitative T(2) (*) MR microscopy was proposed. On high-resolution images, and due to the high susceptibility difference between the particles and the surrounding medium, SPIO internalized in cells induces signal loss which may be counted and measured on T(2) (*) maps. The increase in both labelled cell percentage and the average perturbation volume with an added amount of iron in the incubation medium proved that intracellular iron uptake is dependent upon the initial concentration of incubation iron. It also proved that the observed increases in total cellular iron uptake measured by inductively coupled plasma optical emission spectroscopy are due to both an increase in the iron mass per cell and also an increase in labelled cell concentration. MR results were compared with Prussian blue staining histology. The sensitivity of the MR methodology was then used to distinguish labelling differences for two different types of particle coating. The MRI-assay we proposed is a compulsory tool to optimize labelling efficiency in order to improve in vivo cell detection. Key parameters for detection, such as the percentage of cell labelling, the effect on the image for a given amount of internalized iron and labelling distribution among a cell population, are easily obtained. The comparison of different contrast agents for labelling one cell type, the assessment of one type of contrast agent for labelling different cell types and/or the evaluation of labelling strategies, are possible without having recourse to classical methods, and provide improved accuracy, since the principle is based on intracellular relaxivity.

Volumetric assessment of myocardial viability in rats using 3D double contrast enhanced T1 and T2-weighted MRI.

OBJECTIVE: Volumetric evaluation of the myocardial viability post-infarction in rats using 3D in vivo MR imaging at 7 T using injection of an extracellular paramagnetic contrast agent and intravascular superparamagnetic iron oxide nanoparticles in the same imaging session. MATERIALS AND METHODS: Five hours after induction of permanent myocardial infarction in rats (n=6), 3D in vivo T1- and T2-weighted MR Imaging was performed prior to and after Gd-DOTA injection (0.2 mmol/kg) and prior to and after nanoparticle injection (5 mg Fe/kg) to assess infarct size and myocardial viability. RESULTS: 3D MR Imaging using a successive contrast agent injection showed a difference of infarct size after Gd-DOTA injection on T1-weighted images compared to the one measured on T2-weighted images after Gd-DOTA and nanoparticle injection. CONCLUSION: The use of 3D T1- and T2-weighted MR Imaging using a double contrast agents protocol made possible the accurate characterization of myocardial infarction volume and allowed the detection of myocardial viability post-infarction in rats.

Prenatal evaluation of kidney function in mice using dynamic contrast-enhanced magnetic resonance imaging.

Glomerular differentiation starts as soon as embryonic stage 12 in mice and suggests that kidneys may be functional at this stage. Dynamic contrast-enhanced magnetic resonance microscopy, a noninvasive imaging technique, was used to assess renal function establishment in utero. Indeed, in adults (n = 3), an intravenous injection of gadolinium-DOTA induced in a first step a massive and rapid drop in kidney signal intensity followed, in a second step, by a drop in bladder signal intensity. The delay in signal changes between kidney and bladder reflected glomerular filtration. Pregnant mice underwent anatomical and dynamic contrast-enhanced magnetic resonance microscopy on postcoital days 12-13 (n = 2), 13-14 (n = 1), 14-15 (n = 3), 15-16 (n = 2), 16-17 (n = 3), 17-18 (n = 3), and 18-19 (n = 1). Kidneys and bladder were unambiguously depicted prior to contrast agent injection on stage 15-16 embryos. Contrast agent injection allowed kidney, detection as early as stage 12-13 but not bladder. Kinetics of signal changes demonstrated that glomerular filtration is established at embryonic stage 15-16 in mice. Thus, anatomical and dynamic contrast-enhanced magnetic resonance microscopy may be a powerful noninvasive method for in vivo prenatal developmental and functional studies.

Characterization and detection of experimental rat gliomas using magnetic resonance imaging.

Two different experimental rat brain tumours (F98 glioma and 9L glioma) were characterized using T1 and T2, apparent diffusion coefficient (ADC) and magnetization transfer ratio (MTR). Even though both tumours appeared homogenous at the early stage of growth, significant differences were measured for all parametric images between tumours and normal brain tissue. Irrespective of the sequence used, tumour lesion/normal parenchyma contrast for the non-infiltrative 9L was twice that of the infiltrative F98 glioma. The use of spin preparation via an inversion pulse in a fast spin echo sequence increases contrast by a factor of 20-30.

High field magnetic resonance imaging evaluation of superparamagnetic iron oxide nanoparticles in a permanent rat myocardial infarction.

RATIONALE AND OBJECTIVES: The purpose of this study was to evaluate superparamagnetic iron oxide (SPIO) nanoparticles to discriminate infarcted from normal tissue after myocardial infarction using high field MR imaging (7 tesla). MATERIALS AND METHODS: Permanent myocardial infarction was induced in rats. SPIO nanoparticles (1 mg Fe/kg) were assessed with T1-weighted gradient echo sequence to visualize the myocardial infarction 48 hours after ligature (n = 6). Furthermore, MR Imaging was performed using a T2-weighted RARE sequence and nanoparticles were injected (5 or 10 mg Fe/kg) on 36 rats 5, 24 or 48 hours after infarction. RESULTS: No changes in contrast between normal and infarcted myocardium was observed after nanoparticle injection on T1-weighted images. However, nanoparticles induced a significant contrast increase between normal and infarcted myocardium on T2-weighted images whatever the delay between infarction and imaging (2.99 +/- 1.66 preinjection vs. 7.82 +/- 1.96 after SPIO injection at a dose of 5 mg Fe/kg 5 hours postinfarction, P = 0.0001). CONCLUSIONS: Nanoparticle injection made it possible to discriminate normal from infarcted myocardium on T2-weighted images. However, the high magnetic field prevented the visualization of the T1 effect of SPIO nanoparticles.

In utero time-course assessment of mouse embryo development using high resolution magnetic resonance imaging.

Magnetic resonance microscopy, a non-invasive imaging technique was used for a longitudinal follow-up of mouse embryonic development in utero and for the assessment of embryonic kidney function using 50 nm magnetite dextran particles. Even though the morphologic proton images obtained were still far from classical histological slices quality, an in-plan resolution of 195 microm was achieved for a slice thickness of 800 microm. Mouse embryos sub-structures such as the fourth ventricle, the mesencephalic vesicle, the aorta or the liver can be revealed as early as E11/12. Heart, diaphragm, spinal cord, third, fourth and lateral ventricles were unambiguously seen at E13/14; whereas skeleton, tail, kidney and digit can only be seen from E15/16. Kidney and bladder were certainly identified from E16 on. MR microscopy offers a possibility for in utero phenotyping of mice and can therefore be a powerful tool for post-genomic applications.

Therapeutic efficacy of 5-fluorouracil-loaded microspheres on rat glioma: a magnetic resonance imaging study.

The aim of this work was to assess the therapeutic efficacy of an intratumoral bolus injection of 5-fluorouracil (FU) compared to that of drug loaded in biodegradable microspheres, for the treatment of brain tumour. Experiments were carried out using a fast-growing C6-glioma rat model. The therapeutic protocols were performed 12 days after the injection of glioma cells. At this stage, the tumours were installed and the mean volume was 13 +/- 2 microl as measured by proton magnetic resonance (MR) imaging. This technique was used for the follow-up of the tumour volume with respect to time and therapy. In terms of rat survival, both therapies induced a significant 50% increase in animal life span (p < 0.05) compared to animals receiving no drug or unloaded microspheres. Whilst no cure was observed, analysis of the MR images showed that the local and sustained delivery of FU slowed the tumour development in the vicinity of the microspheres by a factor of 3, compared with the bolus intratumoral injection.

In vivo quantitative microimaging of rat spinal cord at 7T.

In vivo T(2), ADC, and MT properties of the GM and WM of the rat spinal cord were measured at 7T in the cervical region. The GM T(2), T(2GM) = 43.2 +/- 1.0 msec is significantly reduced compared to the WM T(2), T(2WM) = 57.0 +/- 1.6 msec. Diffusion is anisotropic for both GM and WM, with a larger ADC value along the cord axis (ADC(GM//) = 1.05 +/- 0.09 10(-9) m(2)sec(-1) and ADC(WM//) = 1.85 +/- 0.18 10(-9) m(2)sec(-1)) than perpendicular to this plane (ADC(GM)( perpendicular) approximately 0.50 * 10(-9) m(2)sec(-1) and ADC(WM)( perpendicular) approximately 0.18 * 10(-9) m(2)sec(-1)). The MT properties do not significantly differ between the WM and the GM, but allow one to distinguish the thin CSF layer from the WM. DWI with the sensitizing gradient perpendicular to the cord axis leads to the best contrast between GM and WM in the cervical region.

High-field quantitative transverse relaxation time, magnetization transfer and apparent water diffusion in experimental rat brain tumour.

The potential of quantitative parameter images of transverse relaxation time T(2), apparent diffusion coefficient (ADC) and magnetization transfer ratio (MTR) to characterize experimental brain tumours was studied. Necrosis or haemorrhage can be detected using either MTR, ADC or T(2) (necrosis-MTR reduced by 35%, ADC and T(2) increased respectively by 170% and 100% compared with normal brain tissue; haemorrhage-MTR increased by 60%, ADC and T(2) decreased by 40% and 20%, respectively). Normal brain tissue can only be distinguished from tumour on T(2) and MTR parameter images. However, for small tumours (10 microl), the best contrast is observed with MTR, ca. 30%, whereas for T(2) the contrast is ca. 10%.

Error assessment due to coronary stents in flow-encoded phase contrast MR angiography: a phantom study.

Phase contrast magnetic resonance imaging (PC MRI) is a promising method for assessing coronary flow. MR angiography images in the presence of coronary stents display artifacts because of the metal present in the stent. Using a flow phantom, the goal of this in vitro study was to assess quantitatively the effects of flow dephasing caused by magnetic susceptibility in velocity measurements in a region where the artifact is not visualized in a magnitude image. The results showed that for high velocities, significant errors in measurements exist around the stent, outside the susceptibility artifact visible on a magnitude image. J. Magn. Reson. Imaging 1999;10:899-902.

[MRI quantification of regional variations of left ventricular parietal stress in normal subjects]. / Quantification par imagerie par résonance magnétique des variations régionales de la contrainte pariétale du ventricule gauche chez le sujet normal.

The object of this study was to analyse regional variations in end systolic left ventricular wall stress in normal subjects using three-dimensional magnetic resonance imaging (MRI) with excellent spatial resolution. Eight to 12 contiguous short axis sections of the left ventricle were acquired from the apex to the base in apnoea with a rapid echo-gradient sequence in 15 healthy volunteers. The end systolic wall stress was calculated by three methods: Grossman's formula (CR) using the wall thickness and radius of curvature, Janz's formula (CS) using the surfaces, and a three-dimensional approach (C3D) providing a precise calculation of the radius of curvature. The values of wall stress obtained by CS and CR were lower (p < 0.001) at the apex (3.2 and 3.3 10(3) newton/m2 respectively) than at the base (6.9 and 7.1 10(3) newton/m2). There was no difference between the base and apex with the C3D method (8.0 and 9.0 10(3) newton/m2 respectively, NS). The same results were observed at the inferior, lateral, anterior and septal segments with an increase at the base using the CS and CR formulae, the C3D remaining homogeneous in the left ventricle except for the interventricular septum. The lateral wall stress was significantly lower with respect to the interventricular septum in all sections from the apex to the base, irrespective of the method of calculation used. The differences in regional wall stress from the base to the apex reported in healthy subjects seem to be related to an underestimation of left ventricular wall thickness and an underestimation of the radius of curvature rather than to a physiological phenomenon.

Introduction of the closed cranial window technique in gerbils and verification by observation of the effects of specific drugs.

The exact mechanisms of cerebral arterial hypoxia are not perfectly defined. Our purpose is to adapt and validate, with drugs well known in rats and rabbits, a closed cranial window technique in gerbils. The method was used with seventeen gerbils to measure diameter changes of the pial arterioles under normoxia (after the topical application of agonists and antagonists of ATP-sensitive and Ca2+-dependent potassium channels), as well as under hypoxia. In normoxia, aprikalim (10(-6) M), a direct activator of ATP-sensitive potassium channels, increases the diameter of pial arterioles by 10+/-2% (N = 17). This effect is inhibited by glibenclamide (10(-6) M), but not affected by iberiotoxin (10(-6) M), a specific inhibitor of Ca2+-dependent potassium channels. The adenosine-induced dilation by 19+/-5% (N = 17) is reduced by 59+/-16% with iberiotoxin, by 33+/-23% with glibenclamide and inhibited by theophylline (10(-5) M). In hypoxia (15% O2), pial arteriole diameters are increased by 24+/-5% (N = 17) and partially decreased by the application of glibenclamide and iberiotoxin to 59+/-11% and 54+/-5%, respectively. These data are similar to those obtained in other species and validate the closed cranial window technique on gerbils. They indicate that, as for rats and rabbits, both ATP-sensitive and Ca2+-dependent potassium channels are present in gerbil pial vessels and play a role in hypoxia.

Regional assessment of wall curvature and wall stress in left ventricle with magnetic resonance imaging.

Left ventricular functional abnormalities are associated with regional increases of wall stress and modifications of wall curvature. This study describes the integration of the short-axis and long-axis wall curvatures for determining peak systolic wall stress. Quantification was realized with cine magnetic resonance imaging (MRI) from the location of the endocardial and epicardial borders of the left ventricle on pairs of consecutive short-axis sections. Fifteen normal volunteers were subjected to cine MRI, and different methods of calculating peak systolic wall stress were compared. A short-axis analysis showed a 55 +/- 13% increase of the circumferential mean of the peak systolic wall stress between apical and basal sections. Regarding the curvature, no significant increase of wall stress was observed except on the septal wall (31 +/- 18%). Short-axis studies proved to be insufficient for determining the regional variations of left ventricular wall stress and for providing normal reference values for the location of abnormal regions in patients.

Three-dimensional coronary artery MR imaging using prospective real-time respiratory navigator and linear phase shift processing: comparison with conventional coronary angiography.

Respiratory gating with navigator echo is a recent technique to detect diaphragm position in 3D magnetic resonance (MR) coronary angiography. The purpose of our study was to image proximal coronary arteries and to detect significant stenoses in patients with coronary artery diseases and to compare with contrast enhanced angiography results. Twenty patients with coronary artery diseases who were referred for conventional angiography underwent magnetic resonance angiography (MRA). Three-dimensional gradient echo volumes were acquired using cardiac and respiratory gating and fat suppression. Using reformatted oblique planes and maximum intensity projection technique, visualization coronary segments and detection of significant coronary stenoses were made. Eighty-three coronary segments were analyzed. The sensitivity and specificity were 65% and 93%, respectively. The corresponding positive and negative predictive values were 69% and 91%. This study shows the ability to image correctly coronary arteries and to identify proximal stenoses, but image quality need to be improved for an efficiency detection of coronary artery stenoses in clinical practice.

Noninvasive assessment of the infarct-related coronary artery blood flow velocity using phase-contrast magnetic resonance imaging after coronary angioplasty.

This study assesses infarct-related coronary artery blood flow velocity using phase-contrast magnetic resonance imaging (MRI) in patients with reperfused acute myocardial infarction (AMI) and compares these results with flow measurements obtained nonsimultaneously by intracoronary Doppler ultrasound. MRI examination was performed in 17 patients with AMI within 1 to 4 days (mean 2.5 days) after direct or rescue coronary angioplasty using a 0.014-in Doppler guidewire. MRI was performed on a 1.5-T clinical imager. The fast gradient echo segmented k-space phase-contrast pulse sequence was employed during breath-hold. The MRI and Doppler parameters of average peak velocity and maximum peak velocity were measured. Mean phase contrast MRI average peak velocity was 13.3+/-10.7 cm/s, and mean phase-contrast MRI maximum peak velocity was 27+/-16.6 cm/s. Mean Doppler average peak velocity was 17.1+/-5.1 cm/s, and mean Doppler maximum peak velocity was 35.5+/-10.1 cm/s. At the same anatomic levels, phase-contrast MRI average peak velocity correlated significantly to Doppler average peak velocity (r = 0.52; p<0.006) and Doppler maximum peak velocity (r = 0.42; p<0.03). Phase-contrast MRI velocity measurements were correlated with the same heterogeneity of Thrombolysis In Myocardial Infarction 3 flow velocity observed during Doppler examination. Thus, by comparing phase-contrast MRI with invasive intracoronary Doppler flow measurements, the measured MRI values showed significant correlation with Doppler data. Phase-contrast MRI has the potential to noninvasively quantify coronary flow velocity and to evaluate quality of reperfusion in patients with AMI after reperfused therapy.

Thallium-201 right lung/heart ratio during exercise in patients with coronary artery disease: relation to thallium-201 myocardial single-photon emission tomography, rest and exercise left ventricular function and coronary angiography.

The aim of this study was to correlate lung thallium-201 uptake on exercise with 201Tl single-photon emission tomography (SPET) myocardial perfusion imaging, rest and exercise equilibrium radionuclide angiographic and coronary angiographic findings in patients with coronary artery disease (CAD) using a simple, reproducible lung/heart (L/H) ratio that would be easy to use in clinical practice. L/H ratio was defined on the anterior planar image obtained during exercise 201Tl SPET acquisition as the mean counts per pixel in an entire right lung field region of interest divided by the mean counts per pixel in the hottest myocardial wall region of interest. We studied 103 patients. Fifty-nine patients (group I) with <5% likelihood of CAD were used as a reference group. In 44 CAD patients (group II), L/H ratio was compared with 201Tl SPET, radionuclide angiographic and coronary angiographic variables. The group I L/H ratio of 0.35+/-0.05 (mean +/-1 SD) was significantly lower (P<0.001) than the group II L/H ratio of 0. 45+/-0.10. An L/H ratio >0.45 (mean + 2 SD in group I) was considered abnormal. In group II, L/H ratio showed a significant correlation with stress and rest 201Tl perfusion defect size (r = 0. 39 and r = 0.42, P<0.01, respectively), but not with extent of ischaemic myocardium. The mean L/H ratio was 0.41+/-0.10 in patients with one-vessel disease (n = 15), 0.46+/-0.08 in those with two-vessel disease (n = 17) and 0.47+/-0.12 in those with three-vessel disease (n = 12), but no significant difference was found between the three subgroups. L/H ratio showed a significant inverse relation with rest and exercise left ventricular ejection fraction (r = -0.37 and r = -0.50, P<0.05 and P<0.001, respectively). Using stepwise multiple regression analysis, exercise left ventricular ejection fraction and previous history of hypertension were the sole two variables independently predictive of the L/H ratio. In conclusion, although lung thallium uptake is usually found to correlate with extent and severity of CAD, increased L/H ratio should primarily be considered as a marker of exercise-induced left ventricular systolic and perhaps diastolic dysfunction, probably independent of the underlying cardiac disease.