Microbeam Radiation Therapy Opens a Several Days' Vessel Permeability Window for Small Molecules in Brain Tumor Vessels.

PURPOSE: Synchrotron microbeam radiation therapy (MRT), based on an inhomogeneous geometric and microscopic irradiation pattern of the tissues with high-dose and high-dose-rate x-rays, enhances the permeability of brain tumor vessels. This study attempted to determine the time and size range of the permeability window induced by MRT in the blood-brain (tumor) barrier. METHODS AND MATERIALS: Rats-bearing 9L gliomas were exposed to MRT, either unidirectional (tumor dose, 406 Gy) or bidirectional (crossfired) (2 × 203 Gy). We measured vessel permeability to molecules of 3 sizes (Gd-DOTA, Dotarem, 0.56 kDa; gadolinium-labeled albumin, ∼74 kDa; and gadolinium-labeled IgG, 160 kDa) by daily in vivo magnetic resonance imaging, from 1 day before to 10 days after irradiation. RESULTS: An equivalent tumor dose of bidirectional MRT delivered from 2 orthogonal directions increased tumor vessel permeability for the smallest molecule tested more effectively than unidirectional MRT. Bidirectional MRT also affected the permeability of normal contralateral vessels to a different extent than unidirectional MRT. Conversely, bidirectional MRT did not modify the permeability of normal or tumor vessels for both larger molecules (74 and 160 kDa). CONCLUSIONS: High-dose bidirectional (cross-fired) MRT induced a significant increase in tumor vessel permeability for small molecules between the first and the seventh day after irradiation, whereas permeability of vessels in normal brain tissue remained stable. Such a permeability window could facilitate an efficient and safe delivery of intravenous small molecules (≤0.56 kDa) to tumoral tissues. A permeability window was not achieved by molecules larger than gado-grafted albumin (74 kDa). Vascular permeability for molecules between these 2 sizes has not been determined.

Neurovascular multiparametric MRI defines epileptogenic and seizure propagation regions in experimental mesiotemporal lobe epilepsy.

OBJECTIVE: Improving the identification of the epileptogenic zone and associated seizure-spreading regions represents a significant challenge. Innovative brain-imaging modalities tracking neurovascular dynamics during seizures may provide new disease biomarkers. METHODS: With use of a multi-parametric magnetic resonance imaging (MRI) analysis at 9.4 Tesla, we examined, elaborated, and combined multiple cellular and cerebrovascular MRI read-outs as imaging biomarkers of the epileptogenic and seizure-propagating regions. Analyses were performed in an experimental model of mesial temporal lobe epilepsy (MTLE) generated by unilateral intra-hippocampal injection of kainic acid (KA). RESULTS: In the ipsilateral epileptogenic hippocampi, tissue T1 and blood-brain barrier (BBB) permeability to gadolinium were increased 48-72 hours post-KA, as compared to sham and contralateral hippocampi. BBB permeability endured during spontaneous focal seizures (4-6 weeks), along with a significant increase of apparent diffusion coefficient (ADC) and blood volume fraction (BVf). Simultaneously, ADC and BVf were augmented in the contralateral hippocampus, a region characterized by electroencephalographic seizure spreading, discrete histological neurovascular cell modifications, and no tissue sclerosis. We next asked whether combining all the acquired MRI parameters could deliver criteria to classify the epileptogenic from the seizure-spreading and sham hippocampi in these experimental conditions and over time. To differentiate sham from epileptogenic areas, the automatic multi-parametric classification provided a maximum accuracy of 97.5% (32 regions) 48-72 hours post-KA and of 100% (60 regions) at spontaneous seizures stage. To differentiate sham, epileptogenic, and seizure-spreading areas, the accuracies of the automatic classification were 93.1% (42 regions) 48-72 hours post-KA and 95% (80 regions) at spontaneous seizure stage. SIGNIFICANCE: Combining multi-parametric MRI acquisition and machine-learning analyses delivers specific imaging identifiers to segregate the epileptogenic from the contralateral seizure-spreading hippocampi in experimental MTLE. The potential clinical value of our findings is critically discussed.

In vivo characterization of physiological and metabolic changes related to isocitrate dehydrogenase 1 mutation expcression by multiparametric MRI and MRS in a rat model with orthotopically grafted human-derived glioblastoma cell lines.

The physiological mechanism induced by the isocitrate dehydrogenase 1 (IDH1) mutation, associated with better treatment response in gliomas, remains unknown. The aim of this preclinical study was to characterize the IDH1 mutation through in vivo multiparametric MRI and MRS. Multiparametric MRI, including the measurement of blood flow, vascularity, oxygenation, permeability, and in vivo MRS, was performed on a 4.7 T animal MRI system in rat brains grafted with human-derived glioblastoma U87 cell lines expressing or not the IDH1 mutation by the CRISPR/Cas9 method, and secondarily characterized with additional ex vivo HR-MAS and histological analyses. In univariate analyses, compared with IDH1-, IDH1+ tumors exhibited higher vascular density (p < 0.01) and better perfusion (p = 0.02 for cerebral blood flow), but lower vessel permeability (p < 0.01 for time to peak (TTP), p = 0.04 for contrast enhancement) and decreased T1 map values (p = 0.02). Using linear discriminant analysis, vascular density and TTP values were found to be independent MRI parameters for characterizing the IDH1 mutation (p < 0.01). In vivo MRS and ex vivo HR-MAS analysis showed lower metabolites of tumor aggressiveness for IDH1+ tumors (p < 0.01). Overall, the IDH1 mutation exhibited a higher vascularity on MRI, a lower permeability, and a less aggressive metabolic profile. These MRI features may prove helpful to better pinpoint the physiological mechanisms induced by this mutation.

In vivo γ-aminobutyric acid increase as a biomarker of the epileptogenic zone: An unbiased metabolomics approach.

OBJECTIVE: Following surgery, focal seizures relapse in 20% to 50% of cases due to the difficulty of delimiting the epileptogenic zone (EZ) by current imaging or electrophysiological techniques. Here, we evaluate an unbiased metabolomics approach based on ex vivo and in vivo nuclear magnetic resonance spectroscopy (MRS) methods to discriminate the EZ in a mouse model of mesiotemporal lobe epilepsy (MTLE). METHODS: Four weeks after unilateral injection of kainic acid (KA) into the dorsal hippocampus of mice (KA-MTLE model), we analyzed hippocampal and cortical samples with high-resolution magic angle spinning (HRMAS) magnetic resonance spectroscopy (MRS). Using advanced multivariate statistics, we identified the metabolites that best discriminate the injected dorsal hippocampus (EZ) and developed an in vivo MEGAPRESS MRS method to focus on the detection of these metabolites in the same mouse model. RESULTS: Multivariate analysis of HRMAS data provided evidence that γ-aminobutyric acid (GABA) is largely increased in the EZ of KA-MTLE mice and is the metabolite that best discriminates the EZ when compared to sham and, more importantly, when compared to adjacent brain regions. These results were confirmed by capillary electrophoresis analysis and were not reversed by a chronic exposition to an antiepileptic drug (carbamazepine). Then, using in vivo noninvasive GABA-edited MRS, we confirmed that a high GABA increase is specific to the injected hippocampus of KA-MTLE mice. SIGNIFICANCE: Our strategy using ex vivo MRS-based untargeted metabolomics to select the most discriminant metabolite(s), followed by in vivo MRS-based targeted metabolomics, is an unbiased approach to accurately define the EZ in a mouse model of focal epilepsy. Results suggest that GABA is a specific biomarker of the EZ in MTLE.

Monochromatic minibeams radiotherapy: from healthy tissue-sparing effect studies toward first experimental glioma bearing rats therapy.

PURPOSE: The purpose of this study was to evaluate high-dose single fraction delivered with monochromatic X-rays minibeams for the radiotherapy of primary brain tumors in rats. METHODS AND MATERIALS: Two groups of healthy rats were irradiated with one anteroposterior minibeam incidence (four minibeams, 123 Gy prescribed dose at 1 cm depth in the brain) or two interleaved incidences (54 Gy prescribed dose in a 5 × 5 × 4.8 mm(3) volume centered in the right hemisphere), respectively. Magnetic resonance imaging (MRI) follow-up was performed over 1 year. T2-weighted (T2w) images, apparent diffusion coefficient (ADC), and blood vessel permeability maps were acquired. F98 tumor bearing rats were also irradiated with interleaved minibeams to achieve a homogeneous dose of 54 Gy delivered to an 8 × 8 × 7.8 mm(3) volume centered on the tumor. Anatomic and functional MRI follow-up was performed every 10 days after irradiation. T2w images, ADC, and perfusion maps were acquired. RESULTS: All healthy rats were euthanized 1 year after irradiation without any clinical alteration visible by simple examination. T2w and ADC measurements remain stable for the single incidence irradiation group. Localized Gd-DOTA permeability, however, was observed 9 months after irradiation for the interleaved incidences group. The survival time of irradiated glioma bearing rats was significantly longer than that of untreated animals (49 ± 12.5 days versus 23.3 ± 2 days, p < 0.001). The tumoral cerebral blood flow and blood volume tend to decrease after irradiation. CONCLUSIONS: This study demonstrates the sparing effect of minibeams on healthy tissue. The increased life span achieved for irradiated glioma bearing rats was similar to the one obtained with other radiotherapy techniques. This experimental tumor therapy study shows the feasibility of using X-ray minibeams with high doses in brain tumor radiotherapy.

Highly constrained backprojection for improving dynamic 3He MR ventilation imaging in rats.

The highly constrained backprojection algorithm (HYPR) has recently been shown to allow accelerated acquisition in various fields of MRI, including angiography, perfusion and diffusion imaging as well as hyperpolarized gas imaging. Increase in temporal resolution is of particular interest in the case of small animal ventilation imaging due to the high respiration rate. In the present study, the two-dimensional HYPR technique and its iterative version (I-HYPR) were applied to (3)He ventilation imaging in rats. Two imaging protocols were used for two separate groups of animals. A single inspiration protocol consisted of (3)He imaging of the lungs during gas inflow and a following apnea. A multiple inspiration protocol involved spontaneous breathing of (3)He contained in a gas reservoir. Series of HYPR frames with four-fold increase in the temporal resolution were obtained in the case of the single inspiration experiment. For the multiple inspiration protocol, series of HYPR images corresponding to four different echo times were obtained and were used to reconstruct T(2)(*) maps at the inspiration and the expiration phases of the breathing cycle. The feasibility of using the two-dimensional HYPR technique for different (3)He ventilation protocols in small animals is demonstrated. Image quality and signal kinetics representations are compared for two variants of the HYPR algorithm.

Spatially resolved assessment of serotonin-induced bronchoconstrictive responses in the rat lung using 3He ventilation MRI under spontaneous breathing conditions.

The effect on lung ventilation of bronchoconstriction induced by serotonin (intravenous injection of 50 microg/kg of serotonin) was imaged using a hyperpolarized (3)He MR ventilation protocol in spontaneously breathing rats. Lung function maps assessing airflow obstruction, a key feature in clinical pneumology, were derived from dynamic image series acquired after inhalation of (3)He gas. Dynamic ventilation (3)He MR images spanning a respiratory cycle were obtained using a retrospective cine image reconstruction procedure. Two parameters, defined as signal amplitude and maximum signal decay rate, were derived from the time course of the MR signal for each pixel. These parameters were averaged over regions of interest placed in the lower part of the right and left lungs and submitted to statistical analysis. A homogeneous and significant decrease of signal amplitude and maximum signal decay rate after serotonin injection was observed for each rat on the parametric color maps. The data indicate that dynamic ventilation HP (3)He MRI can be used to assess, in a spatially resolved manner, the ventilation function of spontaneously breathing rats and the effects of agents eliciting short-lasting bronchoconstriction.

Longitudinal 3He and proton imaging of magnetite biodistribution in a rat model of instilled nanoparticles.

Epidemiological and toxicological studies have provided evidence that accidentally inhaled nanosize ultrafine particles can induce chronic or acute health damage. MRI, being noninvasive, is able to assess the biodistribution and clearance of magnetically labeled nanoparticles induced by instillation or inhalation. We report 3He and proton MRI follow-up of lung, liver, spleen, and kidney distribution of USPIO (ultrasmall superparamagnetic iron oxide) in a rat model. The sensitivity of the imaging technique to various concentrations of instilled magnetite suspension was first assessed in vivo (n=12). A 2-week longitudinal imaging study was then performed on animals (n=7) instilled with a 0.5 mg magnetite solution. Hypointense and void signal regions associated with intrapulmonary USPIO were observed in the 3He ventilation images throughout the study, whereas no USPIO-related proton signal intensity changes were found. Intrapulmonary magnetite nanoparticle confinement was confirmed by ex vivo iron assay and histological analysis. This study demonstrates that combined 3He and proton MRI enables noninvasive assessment of the distribution and clearance of magnetically labeled instilled nanoparticles.

Effects of ozone exposure in rat lungs investigated with hyperpolarized 3 He MRI.

PURPOSE: To investigate the effects of subchronic ozone exposure on rat lung ventilation using hyperpolarized (HP) (3)He MRI. MATERIALS AND METHODS: A total of 24 Sprague-Dawley rats, distributed in one control group and four groups exposed to 0.5 ppm ozone concentration for two days or six days, either continuously (22 hours/day) or alternatingly (12 hours/day). A three-step MRI protocol was designed and applied to each animal, including: 1) (3)He gas distribution images acquired at inspiratory capacity, 2) measurements of intrapulmonary (3)He diffusion coefficients, and 3) dynamic ventilation acquisitions performed during lung filling with (3)He. RESULTS: No differentiation between animals exposed to ozone and control animals was observed from the ventilation images obtained at inspiratory capacity. The (3)He diffusion coefficients were not statistically different from one group to another. Ventilation defects, appearing as delayed lung filling regions and heterogeneous lung filling, were observed in the dynamic lung ventilation image series. The percentage of animals with ventilation defects in the control, two-day, and six-day exposed groups were equal to 20%, 43% and 75%, respectively. In the subgroup of the animals exposed six days for 12 hours per day, the percentage of animals exhibiting ventilation defects was equal to 85%. CONCLUSION: Heterogeneous obstructive patterns in an experimental animal model of subchronic ozone exposure were observed using HP (3)He MRI.

Measurement of nonlinear pO2 decay in mouse lungs using 3He-MRI.

Spatial and temporal variations in oxygen partial pressure (pO(2)) during breath-hold can be exploited to obtain important regional parameters of lung function. In the course of apnea, the oxygen concentration is known to decay exponentially. Therefore, the initial pO(2) (p(0)) can be used to represent local ventilation, and the oxygen depletion time constant can characterize perfusion. The protocol, based on a nonlinear model of pO(2) decay, was validated in six healthy mice. Parametric maps of p(0) and oxygen depletion time constant were obtained for pure (3)He and (3)He/air mixture. The mean measured values of p(0) were 77 +/- 9 mbar for the pure (3)He insufflation and 107 +/- 5 mbar for (3)He/air mixture, in agreement with the predefined p(0) values: 75 +/- 15 mbar and 123 +/- 15 mbar, respectively. The mean measured oxygen depletion time constants were 6.5 +/- 0.2 s for pure (3)He and 7.1 +/- 0.8 s for the (3)He/air mixture, in agreement with physiology.

Retrospective cine 3He ventilation imaging under spontaneous breathing conditions: a non-invasive protocol for small-animal lung function imaging.

A non-invasive and free-breathing hyperpolarized (HP) (3)He imaging protocol for small animals was implemented and validated on rats for lung function imaging. Animals were allowed to breathe a mixture of air and (3)He from a mask and a gas reservoir fitted to their heads. Radial imaging sequences were used, and MRI signal intensity changes were monitored for retrospective cine image reconstruction. The ventilation cycle of the animals was imaged with a 100 ms temporal resolution. The sliding window imaging technique was applied to reconstruct 5 ms time-shifted image series covering the complete breathing cycle. Image series were processed to extract quantitative ventilation parameters such as the gas arrival time. The reproducibility and the non-invasiveness of this ventilation imaging protocol were evaluated by multiple acquisitions on the same animals.

Alveolar oxygen partial pressure and oxygen depletion rate mapping in rats using 3He ventilation imaging.

A hyperpolarized 3He ventilation imaging protocol was implemented to assess alveolar pO2 values and the oxygen depletion rate in rats. The imaging protocol, which is based on spiral k-space sampling, was designed to acquire a high signal-to-noise ratio (SNR) T1-weighted ventilation series of images in a single breath-hold. Simulations were performed to estimate the accuracy and dependence of the pO2 imaging protocol on the image SNR and the RF flip-angle determination. The imaging protocol was validated in vitro in phantoms and in vivo in rats. Imaging sessions were carried out for different inhaled O2 concentrations ranging from 20% to 40%. Parametric maps of alveolar pO2 and oxygen depletion rate were generated from the series of images. For each investigated animal, the differences in measured alveolar pO2 values are in agreement with the changes in inhaled O2 concentration. The oxygen depletion rates, ranging between 0.7 and 8.0 mbar s-1, are in close agreement with the published values for healthy rats.

Quantitative measurements of regional lung ventilation using helium-3 MRI in a methacholine-induced bronchoconstriction model.

PURPOSE: To demonstrate ventilation changes in an animal model of methacholine-induced bronchoconstriction using hyperpolarized (HP) helium-3 (He-3) MRI. MATERIALS AND METHODS: Bronchoconstriction was induced in 11 healthy rats using an intravenous injection of methacholine. The He-3 was laser-polarized using a custom-built system. MRI studies were performed on a 2-Tesla bore magnet. Coronal dynamic ventilation images were obtained using a single inhalation of the laser-polarized He-3 gas before and after methacholine injection. Ventilation image series were processed on a pixel-by-pixel basis to generate three regional ventilation parameters: gas flow rate, filling time, and maximum gas volume. Student's paired t-test was used for analysis. RESULTS: Ventilation image series with a temporal resolution of 5 msec were obtained before and after methacholine challenge. Quantitative regional gas dynamic information demonstrated statistically significant differences between the baseline and constricted states. Following methacholine injection, the mean flow values were significantly lower for the right lung (RL) (P = 0.006) and left lung (LL) (P = 0.024), while the mean filling time was found to be greater (RL: P = 0.08, LL: P = 0.021). Gas volume values at maximum inspiration were found to be significantly lower after methacholine (RL: P = 0.002; LL: P = 0.036). CONCLUSION: He-3 MRI demonstrated and quantified regional ventilation changes in bronchoconstriction conditions in rats.

Dynamic 3He imaging for quantification of regional lung ventilation parameters.

Dynamic ventilation imaging using laser-polarized (3)He has a promising potential for elucidating the physiology and physiopathology of the lungs. In this study, a methodological approach is proposed for the assessment and quantification of local ventilation parameters. High-temporal-resolution coronal ventilation image series were obtained with a projection-reconstruction (PR) sequence combined with the sliding-window technique. After image series were processed, parametric pixel-by-pixel maps of the gas arrival time, filling time constant, inflation rate, and gas volume were generated. The acquisition technique and the signal processing procedure, which are referred to collectively as sliding pulmonary imaging for respiratory overview (SPIRO), were tested in vivo in healthy rat lungs using a contrast media injector for controlled (3)He flow and volume injection in the animal lungs. The same protocol was applied to broncho-constriction animal models using intravenous injection of methacholine solution. Inflation rate values measured in the lungs were found to decrease with increasing doses of injected methacholine solution. This study demonstrates that it is possible to obtain quantitative regional gas dynamic information using the SPIRO technique in a single polarized gas inspiration.

Helium3 polarization using spin exchange technique: application to simultaneous pulmonary ventilation/perfusion imaging in small animals.

RATIONALE AND OBJECTIVES: To develop a simple and robust helium3 polarization system dedicated to small animal imaging. To demonstrate the potential of helium3 imaging for pulmonary ventilation and perfusion studies. METHODS: A home-built polarization system based on spin-exchange technique was used. This system was applied for magnetic resonance imaging ventilation studies on rats using a 2-T magnet. Projection-reconstruction sequences combined with the sliding-window technique were used for acquisition of high temporal resolution ventilation images. RESULTS: Helium3 polarization levels up to 25% were obtained. Simultaneous ventilation and lung perfusion images were acquired with intravenous injection of superparamagnetic contrast agents. Dose effects were investigated using several contrast agent concentration values. CONCLUSIONS: A tabletop helium3 polarization system was realized. This equipment, which is easy to operate, allows the production of polarized gas appropriate to the requirements of small animal studies. This polarization system was used successfully on a ventilation/perfusion imaging study using intravascular contrast agent.