Mitochondria can orchestrate sex differences in cell fate of vascular smooth muscle cells from rats.

BACKGROUND: In basal conditions, vascular smooth muscle cells freshly isolated from aortas of male and female rats display marked sex differences in terms of redox balance and susceptibility to ultraviolet radiation-induced cell death. In particular, in the same experimental conditions, cells from male rats are more susceptible to oxidative stress and underwent apoptosis, while cells from female rats underwent premature senescence. In the present work, the mechanism involved in cell fate after ultraviolet radiation exposure is investigated. METHODS: Vascular smooth muscle cells, isolated from the descending aortas of both female and male Sprague-Dawley young rats, were exposed to a single sub-cytotoxic dose of ultraviolet radiation (200 mJ/cm(2)). The distribution and the expression of molecules involved in cell survival and mitochondrial physiology were evaluated by static and flow cytometry using commercial kits and antibodies. Statistical analyses were performed by using Student's t test and two-way ANOVA. RESULTS: After exposure to ultraviolet radiation, an upregulation of survival proteins such as BclxL, survivin and the presence in the nucleus of NF-κB were found in cells from females. Conversely, pro-apoptotic proteins such as Bax, caspase-3, and caspase-9 as well as loss of mitochondrial membrane potential were found in cells from male rats. CONCLUSIONS: Our results suggest that (i) mitochondria, being producers of ROS, can orchestrate sex differences in cell fate of VSMC and (ii) mitochondrial dysfunction may be a significant mechanism by which cardiovascular risk factors lead to the formation of vascular lesions in a sex-specific way.

In vitro expansion of U87-MG human glioblastoma cells under hypoxic conditions affects glucose metabolism and subsequent in vivo growth.

Hypoxia is a characteristic feature of solid tumors leading to the over expression of hypoxia-inducible factor (HIF)-1α protein and therefore to a specific cellular behavior. However, even though the oxygen tension in tumors is low (<5 %), most of the cell lines used in cancer studies are grown under 21 % oxygen tension. This work focuses on the impact of oxygen conditions during in vitro cell culture on glucose metabolism using 1-(13)C-glucose. Growing U87-MG glioma cells under hypoxic conditions leads to a two- to threefold reduction of labeled glutamine and an accumulation of fructose. However, under both hypoxic and normoxic conditions, glucose is used for de novo synthesis of pyrimidine since the (13)C label is found both in the uracil and ribose moieties. Labeling of the ribose ring demonstrates that U87-MG glioma cells use the reversible branch of the non-oxidative pentose phosphate pathway. Interestingly, stereotactic implantation of U87-MG cells grown under normoxia or mild hypoxia within the striatum of nude mice led to differential growth; the cells grown under hypoxia retaining an imprint of the oxygen adaptation as their development is then slowed down.

The effect of patient sex on the incidence of early adverse effects in a population of elderly patients.

Patient sex is known to influence the response to general and regional anaesthesia and recovery after surgery. However, most studies come from analyses carried out on middle-aged patients. As most of the patients admitted to the post-anaesthesia recovery room in our institution are elderly, we took the opportunity to investigate the association between sex and incidence of early adverse events in this older population of patients after major surgery. Consecutive patients undergoing general, orthopaedic, urological and gynaecological surgery, admitted to the recovery room of our institution over a 15-month period, were retrospectively studied. The following adverse events were considered in the analysis: shivering, postoperative nausea and vomiting, hypotension and hypertensive responses, new arrhythmias requiring treatment, acute respiratory failure and desaturation. A total of 1347 patients (mean age 73.3±15.1 years, 61.4% women) were included. Women showed a higher incidence of shivering (relative difference +48%, P=0.0003), postoperative nausea and vomiting (+91%, P<0.0001), hypotension (+32%, P=0.044) and desaturation (+60%, P=0.0030) than men. The incidence of hypertensive response, arrhythmias and acute respiratory failure were not statistically significantly different. The findings of this exploratory study suggest that women have a higher risk of early postoperative adverse events even in a more elderly population.

Modeling nigrostriatal degeneration in organotypic cultures, a new ex vivo model of Parkinson's disease.

Parkinson's disease (PD) is the second most frequent neurodegenerative disorder afflicting 2% of the population older than 65 years worldwide. Recently, brain organotypic slices have been used to model neurodegenerative disorders, including PD. They conserve brain three-dimensional architecture, synaptic connectivity and its microenvironment. This model has allowed researchers a simple and rapid method to observe cellular interactions and mechanisms. In the present study, we developed an organotypic PD model from rat brains that includes all the areas involved in the nigrostriatal pathway in a single slice preparation, without using neurotoxins to induce the dopaminergic lesion. The mechanical transection of the nigrostriatal pathway obtained during slice preparation induced PD-like histopathology. Progressive nigrostriatal degeneration was monitored combining innovative approaches, such as diffusion tensor magnetic resonance imaging (DT-RMI) to follow fiber degeneration and mass spectrometry to quantify striatal dopamine content, together with bright-field and fluorescence microscopy imaging. A substantia nigra dopaminergic cell number decrease was observed by immunohistochemistry against rat tyrosine hydroxylase (TH) reaching 80% after 2 days in culture associated with a 30% decrease of striatal TH-positive fiber density, a 15% loss of striatal dopamine content quantified by mass spectrometry and a 70% reduction of nigrostriatal fiber fractional anisotropy quantified by DT-RMI. In addition, a significant decline of medium spiny neuron density was observed from days 7 to 16. These sagittal organotypic slices could be used to study the early stage of PD, namely dopaminergic degeneration, and the late stage of the pathology with dopaminergic and GABAergic neuron loss. This novel model might improve the understanding of PD and may represent a promising tool to refine the evaluation of new therapeutic approaches.

Perfluorocarbon-loaded lipid nanocapsules as oxygen sensors for tumor tissue pO2 assessment.

The assessment of tumor oxygenation is a crucial factor in cancer therapy and may be carried out using fluorine MRI once fluorine probes have been distributed within the tumor. However, the deposit of those highly fluorinated compounds often jeopardizes anatomical image quality and requires emulsification of the probes. Due to the high density and the high lipophilicity of perfluorocarbons, nanoemulsion of these molecules usually requires high-energy processes. In the present work, we discuss the synthesis and the physico-chemical characterization of perfluorocarbon nanocapsules using a low-energy phase-inversion process. The nanocapsules were tested on a mouse tumor brain model to assess oxygenation.

Gender effect on the relation between diabetes and hospitalization for heart failure.

AIMS: Cardiovascular risk among diabetic patients is at least twice as much the one for non-diabetic individuals and even greater when diabetic women are considered. Heart failure (HF) is a common unfavorable outcome of cardiovascular disease in diabetes. However, since the comparison among sexes of heart failure prevalence in diabetic patients remains limited, this study is aimed at expanding the information about this point. METHODS: We have evaluated the association between diabetes and HF by reviewing the medical records of all subjects discharged from the Internal Medicine and Cardiology Units of all hospitals in the Tuscany region, Italy, during the period January 2002 through December 2008. In particular we sought concomitance of ICD-9-CM codes for diabetes and HF. RESULTS: Patients discharged by Internal Medicine were on average older, more represented by women, and had a lesser number of individuals coded as diabetic (p<0.05 for all). Relative risk for HF (95% CI) was significantly higher in patients with diabetes, irrespective of gender 1.39 (1.36-1.41) in males; 1.40 (1.37-1.42) in females. When the diabetes-HF association was analyzed according to decades of age, a "horse-shoe" pattern was apparent with an increased risk in 40-59 years old in female patients discharged by Internal Medicine. CONCLUSIONS: Although there is not a difference in the overall HF risk between hospitalized male and female diabetic patients, women have an excess risk at perimenopausal age.

Long term predictors of post-partum glucose metabolism in women with gestational diabetes mellitus.

AIMS: The question asked by this study was whether ß-cell function expressed by insulin secretion/sensitivity measured during pregnancy in women with gestational diabetes (GDM) predicts post-partum long-term derangement in glucose metabolism. METHODS: Seventy-four Caucasian women with previous GDM were retested through a 75 g-2-h-OGTT after 8 [6] years (median[interquartile range]) from index pregnancy, measuring at pregnancy and follow-up insulin sensitivity, insulin secretion (1-h-incremental-insulin-area/incremental-glucose-area: ΔAUC60 (I)/ΔAUC60 (G)) as well as the product of Stumvoll-first-phase - secretion x insulin sensitivity (insulin-secretion-sensitivity index (ISSI). RESULTS: At follow-up 47 women were normotelerant to glucose and 27 had altered glucose metabolism (AGM:10 with type 2 diabetes and 17 with IGT). Women progressed to AGM had at their index pregnancy higher mean 2-h-OGTT-glucose area (1.15±0.09 VS. 1.09±0.09 mol l 2-h (-1);p=0.014), and lower ΔAUC60 (I)/ΔAUC60 (median [interquantile range]) (54.4 [51.7] vs. 73.4 [60] pmol mmol (-1)) and ISSI (2 977 [766] vs. 3 708 [1 141]; p<0.05 for both), but similar insulin sensitivity index 2.9 [2.5] VS. 3.2 [2.2] ml min (-1) m (-2);p=NS). Two-h-OGTT-glucose area, or decrease in ΔAUC60 (I)/ΔAUC60 (G) and ISSI were significantly associated with glucose tolerance impairment and with raised adjusted risk for AGM while insulin sensitivity at pregnancy did no predict AGM development. CONCLUSIONS: In this group of women increased post-load plasma glucose and impaired ß-cell function assessed during GDM pregnancy predict long-term post-partum AGM, while insulin sensitivity measured at the same time does not.

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.

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.

Does parity increase insulin resistance during pregnancy?

AIMS: To study the effect of parity on impairment of insulin sensitivity during pregnancy and on the risk of gestational diabetes (GDM). METHODS: We studied the relationship between parity and peripheral insulin sensitivity index (ISI(OGTT)) or GDM in 1880 caucasian women, who underwent a 100-g, 3-h oral glucose tolerance test (OGTT) between the 24th and 28th gestational week and in 75 women who underwent an OGTT in two consecutive pregnancies. A proxy for beta-cell function (basal plasma C peptide/fasting plasma glucose; CP/FPG) was also measured. RESULTS: By univariate analysis parity was related to decreased ISI(OGTT) and to increased CP/FPG in those with parity > 3 and likewise GDM, diagnosed in 124 women (6.58%), was linearly related to parity (P = 0.0034) and strongly age dependent. The relationships between parity and ISI(OGTT), CP/FPG and GDM were no longer significant after adjustment for age, pregestational body mass index (BMI), and weight gain. GDM was significantly related to age and pregestational weight, while ISI(OGTT) and CP/FPG were inversely related to prepregnancy BMI or weight gain. In comparison with the index pregnancy, the subsequent pregnancy was characterized by an increase in actual and prepregnancy BMI, in 2 h area under curve (AUC) glucose and by a decrease in ISI(OGTT) (P = 0.0001). The longer the time interval between pregnancies and the higher the increment in pregestational BMI or in weight gain during the pregnancy, the greater were the ISI(OGTT) decrease and 2-h AUC glucose increase. CONCLUSIONS: Parity is not directly linked to insulin sensitivity deterioration, to CP/FPG increase during pregnancy, or to GDM appearance, although it is linked through the mediation of progressive ageing and weight gain either before or during pregnancy, when there is a sufficiently long time interval between pregnancies.

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.

In vitro antimycotic activity of some plant extracts towards yeast and yeast-like strains.

As part of screening aimed at the selection of novel antimycotic compounds of vegetable origin, leaf extracts of Camellia sinensis L., Cupressus sempervirens L. and Pistacia lentiscus L. and the seed extract of Glycine soja Sieb. et Zucc. were tested against yeast and yeast-like species implicated in human mycoses. Of the extracts only those of C. sinensis (obtained from a commercial preparation of green tea) exhibited broad activity towards Candida glabrata, Clavispora lusitatiae, Cryptococcus laurentii, Filobasidiella neoformans, Issatchenkia orientalis, Saccharomyces cerevisiae and Prototheca wickerhamii strains. MICs ranging from 300 to 4800 microg extract/mL (corresponding to 130-2010 microg/mL total polyphenols) were observed. Concentrations of the C. sinensis extract over 25 000 microg/mL caused a rapid decrease of viable cells of Fil. neoformans and its activity was dose-dependent. Tests carried out using the pure polyphenols present in C. sinensis extract composition, showed that only epicatechin-3-O-gallate (ECG) and epigallocatechin-3-O-gallate (EGCG) possess antimycotic activity.

Long-term taurine supplementation reduces mortality rate in streptozotocin-induced diabetic rats.

Oxidative stress is implicated in the pathogenesis of diabetes mellitus. Taurine and vitamin E+selenium supplementation has some benefits in experimental models of diabetes mellitus. This study evaluates whether taurine and vitamin E+selenium supplementations reduce a hard end-point such as mortality due to diabetes. Streptozotocin-induced diabetic rats were fed with standard diet or taurine (5%, w/w) or vitamin E (500 UI/Kg)+selenium (8 mg/Kg) enriched diets. Taurine significantly decreased mortality rate (p < 0.04), while vitamin E failed to increase survival. In the late phase of the disease, taurine significantly decreased glycaemia, being vitamin E ineffective. No correlation between glycaemia and survival was found. None of supplementations modified body weight. Thus, only taurine decreases the mortality rate and glycaemia. These results encourage new research in the field, since classical hypoglycaemic agents are unable to decrease mortality in diabetic patients.

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.

Taurine administration during lactation modifies hippocampal CA1 neurotransmission and behavioural programming in adult male mice.

Taurine plays a role in neuronal development. In this study, we examined whether postnatal taurine administration influences the long-term consequences induced by mild neonatal stressors (10 min maternal deprivation plus sham injection, applied daily to neonatal mice up to 21 days). At 30 days of age stressed mice showed higher pain threshold both in the tail-flick--which measures mostly the spinal mechanisms of pain--and in the hot-plate test--which reflects mainly the supraspinal mechanisms of pain. The latter effect was prevented completely by neonatal taurine administration, while the tail-flick test was not affected, thus suggesting that spinal pain is not sensitive to taurine treatment. At 140 days of age, mice which were stressed during the neonatal period showed consistent decrease in immobility time in forced swimming test, and taurine did not influence this parameter. At the same age, the fear/anxiety axis, measured with elevated plus maze test, did not show any consistent changes. Electrophysiological experiments in brain slices obtained from adult mice showed that input-output curves in hippocampal CA1 were increased by taurine administration in lactation. Hence, neonatal administration of taurine might permanently modify the functioning of hippocampus, a brain area which is known to be crucial for learning and memory.

Thiolation and nitrosation of cysteines in biological fluids and cells.

Thiols (RSH) are potent nucleophilic agents, the rates of which depend on the pKa of the sulfhydryl. Unlike compounds having other nucleophile moieties (-OH or -NH(2)), RSH are involved in reactions, such as conjugations, redox and exchange reactions. Although protein SH groups (PSH) react like non-protein thiols (NPSH), the biochemistry of proteins is much more complex for reasons such as steric hindrance, charge distribution and accessibility of PSH to the solvent (protein conformation). The reaction rates and types of end-products of PSH vary a lot from protein to protein. The biological problem is even more complex because in all compartments and tissues, there may be specific competition between thiols (namely between GSH and PSH), regulated by the properties of antioxidant enzymes. Moreover, PSH are divided biologically into essential and non-essential and their respective influence in the various biological systems is unknown. It follows that during phenomena eliciting a prompt thiol response (oxidative stress), the antioxidant PSH response and reaction mechanisms vary considerably from case to case. For example, in spite of a relatively low pKa that should guarantee good antioxidant capacity, PSH of albumin has much less propensity to form adducts with conjugating agents than NPSH; moreover, the structural characteristics of the protein prevent albumin from forming protein disulfides when exposed to oxidants (whereas protein-thiol mixed disulfides are formed in relative abundance). On the other hand, proteins with a relatively high reactivity, such rat hemoglobin, have much greater antioxidant capacity than GSH, but although human hemoglobin has a pKa similar to GSH, for structural reasons it has less antioxidant capacity than GSH. When essential PSH are involved in S-thiolation and S-nitrosation reactions, a similar change in biological activity is observed. S-thiolated proteins are a recurrent phenomenon in oxidative stress elicited by reactive oxygen species (ROS). This event may be mediated by disulfides, that exchange with PSH, or by the protein intermediate sulfenic acid that reacts with thiols to form protein-mixed disulfides. During nitrosative stress elicited by reactive nitrogen species (RNS), depending on the oxygen concentration of the system, nitrosation reactions of thiols may also be accompanied by protein S-thiolation. In this review we discuss a number of cell processes and biochemical modifications of enzymes that indicate that S-thiolation and S-nitrosation may occur simultaneously in the same protein in the presence of appropriate interactions between ROS and RNS.

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.

Quantitative MR renography using a calibrated internal signal (ERETIC).

To measure MR renograms, cortical and medullary kidney signal intensity evolution is followed after contrast agent injection. To obtain an accurate quantitative signal measurement, the use of a reference signal is necessary to correct the potential MRI system variations in time. The ERETIC method (Electronic Reference To access In vivo Concentrations) provides an electronic reference signal. It is synthesized as an amplitude modulated RF pulse applied during the acquisition. The ERETIC method was as precise as the external tube reference method but presents major advantages like its free adjustability (shape, location and magnitude) to the characteristics of the organ studied as well as its not taking room inside the magnet. Even though ERETIC showed a very good intrinsic stability, systems' variations still affect its signal in the same way as real NMR signals are affected. This method can be easily implemented on any imaging system with two RF channels.

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.