Imagerie moléculaire de la neuroinflammation dans la maladie de Parkinson (étude préclinique dans un modèle animal de rat)

Bien que les mécanismes moléculaires précis à l origine de la neurodégénérescence dopaminergique ne soient pas encore totalement connus, un ensemble de preuves épidémiologiques, cliniques et expérimentales indiquent que la neuroinflammation peut avoir un rôle important dans la pathogenèse de la MP. L étude des liens spatio-temporels entre la neuroinflammation et la neurodégénérescence au cours de la MP pourrait améliorer la compréhension du mécanisme physiopathologique et aussi l'accessibilité à un diagnostic précoce et/ou à de nouvelles approches thérapeutiques anti-inflammatoires. Le développement actuel des méthodes non invasives d'imagerie moléculaire permettant la surveillance directe du processus de neuroinflammation devrait être utile à cet effet. La cible moléculaire de choix dans ce domaine est la protéine de 18 kDa translocateur (TSPO), biomarqueur sensible associée à la neuroinflammation, qui est surexprimé dans les microglies activées. Dans le travail présenté ici nous avons réalisé l'évaluation longitudinale des deux mécanismes physiopathologiques en parallèle avec les modifications de la fonction dopaminergique à plusieurs points au cours du temps après lésion à la 6-OHDA chez le rat, modèle qui imite un stade précoce de la MP. Après l'administration unilatérale, intra-striatale de la 6-OHDA, nous avons quantifié l'évolution temporelle de la TSPO, de l immunoréactivité TH et du DAT dans le striatum et la SNC de 3 à 56 jours post-lésion (jpl). L augmentation de la liaison des ligands de la TSPO utilisés, c-à-d [3H]-PK11195 et [125I]-CLINDE, a été observée dans le striatum lésé à 3, 7 et 14 jpl, suivie d'un retour progressif à un niveau basal à 56 jpl. Le profil de liaison dans la SNC a montré une augmentation progressive de la fixation qui débute à 3 jpl, avec un pic à 14 jpl, et diminue progressivement jusqu'à ce que 56 jpl. Dans ce modèle de rongeur de la MP, les processus neuroinflammatoire et neurodégénératif surviennent de façon concomitante. La présence transitoire de l'activation microgliale pourrait être impliquée dans l apparition et l'installation durable de la perte neuronale dopaminergique. Cette étude confirme donc le lien entre la neuroinflammation et de la neurodégénérescence et met aussi l'accent sur l'intérêt du CLINDE comme traceur potentiel de la neuroinflammation in-vivo en fournissant des informations précieuses pour le diagnostic précoce et le suivi longitudinal de la progression de la maladie, avec des applications potentielles chez l'homme. En effet, la détection précoce de la neuroinflammation, de façon antérieure à une perte neuronale cliniquement significative, pourrait devenir un enjeu majeur dans la prise en charge pré-symptomatique de la MP. Dans ce sens, nous mettons en évidence l existence d'une fenêtre thérapeutique, survenant juste après la lésion, qui peut être proposé pour l'introduction de traitements anti-inflammatoires qui viseraient à ralentir le processus neurodégénératif. La poursuite de l exploration des relations entre la neuroinflammation et la neurodégénéréscence in-vivo dans le même modèle animal avec la méthode d imagerie micro-TEP, transposable à l homme, en utilisant en parallèle le [18F]-DPA714 pour la TSPO et le [18F]-LBT999 pour le DAT est en cours.Although the precise molecular mechanisms causing the dopaminergic neurodegeneration are still not totally understood, a body of epidemiological, clinical and experimental evidence indicates that neuroinflammation may have an important role in the pathogenesis of PD. Study of spatio-temporal links between neuroinflammation and neurodegeneration during the course of PD would improve understanding of the physiopathological mechanism and also accessibility to early diagnosis and/or new antiinflammatory therapeutic approaches. The current development of non-invasive molecular imaging methods allowing direct monitoring of the neuroinflammation process should be valuable for this purpose. The molecular target of choice in this field is the 18 kDa translocator protein (TSPO), a sensitive biomarker associated with neuroinflammation, which is over-expressed in activated microglia. In the study presented here we achieved the longitudinal evaluation of both physiopayhological mechanisms in parallel with the modifications of dopaminergic function at several time-points after 6-OHDA lesion in the rat that mimics an early stage of PD. After unilateral intra-striatal 6-OHDA administration, we quantified the temporal evolution of the TSPO, TH immunoreactivity and DAT in the striatum and the SNc from 3 to 56 days post-lesion (dpl). Increased binding of TSPO ligands used, i.e. [3H]PK11195 and [125I]CLINDE, was observed in the lesioned striatum at 3, 7 and 14 dpl, followed by a progressive return to the basal level at 56 dpl. The binding profile in the SNc showed progressive binding beginning at 3 dpl, peaking at 14 dpl, and progressively decreasing until 56 dpl. In this rodent model of PD, the neuroinflammatory and neurodegenerative processes occurred concomitantly. The transitory occurrence of microglial activation could be involved in the advent and the lasting installation of dopaminergic neuron loss. This study supports the link between neuroinflammation and neurodegeneration and emphasizes the interest of CLINDE as potent in vivo tracer of neuroinflammation by providing valuable information for early diagnosis and longitudinal follow-up of disease progression, with potential applications to human patients. Indeed, early detection of neuroinflammation, prior to a clinically significant loss of neurons, could become a major issue in the management of pre-symptomatic PD. To support this idea, we demonstrate the existence of a therapeutic window, occurring just after the lesion, which may be proposed for the introduction of anti-inflammatory treatments that aimed to slow the neurodegenerative process. Further exploration of the relationship between neuroinflammation and neurodegeneration in vivo in the same animal model with the method of micro-PET imaging, transposable to humans, using in parallel the [18F]-DPA714 for TSPO and [18F]-LBT999 for DAT is pending.TOURS-Bibl.électronique (372610011) / SudocSudocFranceF

Blood-brain barrier disruption with focused ultrasound enhances delivery of dopamine transporter tracer (PE2I) into the brain

International audiencePE2I is one of the most selective ligands for dopamine transporter. However it is associated with blood-brain barrier (BBB) permeability limitations. The aim of this study was to investigate the use of ultrasound and microbubbles to increase its delivery through the BBB and by determining the optimal experimental conditions that achieve a transient and safe BBB disruption. First, we stablished the ultrasound conditions that achieved a transient BBB disruption in rats using a non-permeant marker, Evans blue. Hence SonoVue® (450μL/kg) and Evans blue (100mg/kg) were intravenously administered. BBB leakage was obtained using ultrasound insonation through the rat skull at 1.6MPa, PRF 1Hz, duty cycle 1%, burst 10ms during 120sec. BBB disruption was observed in all treated animals (N=4) by histological analysis. The same experimental conditions were applied to enhance brain uptake of PE2I. Biological samples were analyzed using a scintillation counter apparatus. The results showed 50% and 20% increase of 125I-PE2I uptake in the striatum and cerebral cortex, respectively, in the treated rats (N=5) versus control (N=4). Similar enhancements were observed using SonoVue® at half concentration. This innovative method provides a great potential for intracerebral delivery of molecular ligands that could be used for the therapy of brain diseases

In vivo PET quantification of the dopamine transporter in rat brain with [¹⁸F]LBT-999.

INTRODUCTION: We examined whether [(18)F]LBT-999 ((E)-N-(4-fluorobut-2-enyl)2β-carbomethoxy-3β-(4'-tolyl)nortropane) is an efficient positron emission tomography (PET) tracer for the quantification of the dopamine transporter (DAT) in the healthy rat brain. METHODS: PET studies were performed using several experimental designs, i.e. test-retest, co-injection with different doses of unlabelled LBT, displacement with GBR12909 and pre-injection of amphetamine. RESULTS: The uptake of [(18)F]LBT-999 confirmed its specific binding to the DAT. The non-displaceable uptake (BP(ND)) in the striatum, between 5.37 and 4.39, was highly reproducible and reliable, and was decreased by 90% by acute injection of GBR12909. In the substantia nigra/ventral tegmental area (SN/VTA), the variability was higher and the reliability was lower. Pre-injection of amphetamine induced decrease of [(18)F]LBT-999 BP(ND) of 50% in the striatum. CONCLUSIONS: [(18)F]LBT-999 allows the quantification of the DAT in living rat brain with high reproducibility, sensitivity and specificity. It could be used to quantify the DAT in rodent models, thereby allowing to study neurodegenerative and neuropsychiatric diseases

4DGVF segmentation of vector-valued images

International audienceIn this paper, we extend the gradient vector flow field to the vector-valued case for robust variational segmentation of 4D images with active surfaces. Instead of only exploiting scalar edge strength in order to identify vector edges, we propagate both directions and amplitudes of vector gradients computed from the analysis of a structure tensor of the vector-valued image. To reduce contributions from noise in the calculation of the structure tensor, image channels are weighted according to a blind estimator of contrast that take profit of the deformable models framework. The proposed 4DGVF vector field is validated on synthetic image datasets and applied to biological volume delineation in dynamic PET imaging

Variational segmentation of vector-valued images with gradient vector flow

International audienceIn this paper, we extend the gradient vector flow field for robust variational segmentation of vector-valued images. Rather than using scalar edge information, we define a vectorial edge map derived from a weighted local structure tensor of the image that enables the diffusion of the gradient vectors in accurate directions through the 4DGVF equation. To reduce the contribution of noise in the structure tensor, image channels are weighted according to a blind estimator of contrast. The method is applied to biological volume delineation in dynamic PET imaging, and validated on realistic Monte Carlo simulations of numerical phantoms as well as on real images

Regional characterization of energy metabolism in the brain of normal and MPTP-intoxicated mice using new markers of glucose and phosphate transport

The gibbon ape leukemia virus (GALV), the amphotropic murine leukemia virus (AMLV) and the human T-cell leukemia virus (HTLV) are retroviruses that specifically bind nutrient transporters with their envelope glycoproteins (Env) when entering host cells. Here, we used tagged ligands derived from GALV, AMLV, and HTLV Env to monitor the distribution of their cognate receptors, the inorganic phosphate transporters PiT1 and PiT2, and the glucose transporter GLUT1, respectively, in basal conditions and after acute energy deficiency. For this purpose, we monitored changes in the distribution of PiT1, PiT2 and GLUT1 in the cerebellum, the frontal cortex, the corpus callosum, the striatum and the substantia nigra (SN) of C57/BL6 mice after administration of 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridinium (MPTP), a mitochondrial complex I inhibitor which induces neuronal degeneration in the striato-nigral network

Segmentation of Dynamic PET Images with Kinetic Spectral Clustering

International audienceSegmentation is often required for the analysis of dynamic positron emission tomography (PET) images. However, noise and low spatial resolution make it a difficult task and several supervised and unsupervised methods have been proposed in the literature to perform the segmentation based on semi-automatic clustering of the time activity curves of voxels. In this paper we propose a new method based on spectral clustering that does not require any prior information on the shape of clusters in the space in which they are identified. In our approach, the p-dimensional data, where p is the number of time frames, is first mapped into a high dimensional space and then clustering is performed in a low-dimensional space of the Laplacian matrix. An estimation of the bounds for the scale parameter involved in the spectral clustering is derived. The method is assessed using dynamic brain PET images simulated with GATE and results on real images are presented. We demonstrate the usefulness of the method and its superior performance over three other clustering methods from the literature. The proposed approach appears as a promising pre-processing tool before parametric map calculation or ROI-based quantification tasks

Alteration of the Oligodendrocyte Lineage Varies According to the Systemic Inflammatory Stimulus in Animal Models That Mimic the Encephalopathy of Prematurity

Preterm birth before the gestational age of 32 weeks is associated with the occurrence of specific white matter damage (WMD) that can compromise the neurological outcome. These white matter abnormalities are embedded in more global brain damage defining the encephalopathy of prematurity (EoP). A global reduction in white matter volume that corresponds to chronic diffuse WMD is the most frequent form in contemporary cohorts of very preterm infants. This WMD partly results from alterations of the oligodendrocyte (OL) lineage during the vulnerability window preceding the beginning of brain myelination. The occurrence of prenatal, perinatal and postnatal events in addition to preterm birth is related to the intensity of WMD. Systemic inflammation is widely recognised as a risk factor of WMD in humans and in animal models. This review reports the OL lineage alterations associated with the WMD observed in infants suffering from EoP and emphasizes the role of systemic inflammation in inducing these alterations. This issue is addressed through data on human tissue and imaging, and through neonatal animal models that use systemic inflammation to induce WMD. Interestingly, the OL lineage damage varies according to the inflammatory stimulus, i.e., the liposaccharide portion of the E.Coli membrane (LPS) or the proinflammatory cytokine Interleukin-1β (IL-1β). This discrepancy reveals multiple cellular pathways inducible by inflammation that result in EoP. Variable long-term consequences on the white matter morphology and functioning may be speculated upon according to the intensity of the inflammatory challenge. This hypothesis emerges from this review and requires further exploration

Longitudinal mouse-PET imaging: a reliable method for estimating binding parameters without a reference region or blood sampling

International audienceLongitudinal mouse PET imaging is becoming increasingly popular due to the large number of transgenic and disease models available but faces challenges. These challenges are related to the small size of the mouse brain and the limited spatial resolution of microPET scanners, along with the small blood volume making arterial blood sampling challenging and impossible for longitudinal studies. The ability to extract an input function directly from the image would be useful for quantification in longitudinal small animal studies where there is no true reference region available such as TSPO imaging.METHODS:Using dynamic, whole-body 18F-DPA-714 PET scans (60 min) in a mouse model of hippocampal sclerosis, we applied a factor analysis (FA) approach to extract an image-derived input function (IDIF). This mouse-specific IDIF was then used for 4D-resolution recovery and denoising (4D-RRD) that outputs a dynamic image with better spatial resolution and noise properties, and a map of the total volume of distribution (VT) was obtained using a basis function approach in a total of 9 mice with 4 longitudinal PET scans each. We also calculated percent injected dose (%ID) with and without 4D-RRD. The VT and %ID parameters were compared to quantified ex vivo autoradiography using regional correlations of the specific binding from autoradiography against VT and %ID parameters.RESULTS:The peaks of the IDIFs were strongly correlated with the injected dose (Pearson R = 0.79). The regional correlations between the %ID estimates and autoradiography were R = 0.53 without 4D-RRD and 0.72 with 4D-RRD over all mice and scans. The regional correlations between the VT estimates and autoradiography were R = 0.66 without 4D-RRD and 0.79 with application of 4D-RRD over all mice and scans.CONCLUSION:We present a FA approach for IDIF extraction which is robust, reproducible and can be used in quantification methods for resolution recovery, denoising and parameter estimation. We demonstrated that the proposed quantification method yields parameter estimates closer to ex vivo measurements than semi-quantitative methods such as %ID and is immune to tracer binding in tissue unlike reference tissue methods. This approach allows for accurate quantification in longitudinal PET studies in mice while avoiding repeated blood sampling

The Story of the Dopamine Transporter PET Tracer LBT-999: From Conception to Clinical Use

The membrane dopamine transporter (DAT) is involved in a number of brain disorders and its exploration by positron emission tomography (PET) imaging is highly relevant for the early and differential diagnosis, follow-up and treatment assessment of these diseases. A number of carbon-11 and fluor-18 labeled tracers are to date available for this aim, the majority of them being derived from the chemical structure of cocaine. The development of such a tracer, from its conception to its use, is a long process, the expected result being to obtain the best radiopharmaceutical adapted for clinical protocols. In this context, the cocaine derivative (E)-N-(4-fluorobut-2-enyl)2β-carbomethoxy-3β-(4′-tolyl)nortropane, or LBT-999, has passed all the required stages of the development that makes it now a highly relevant imaging tool, particularly in the context of Parkinson's disease. This review describes the different steps of the development of LBT-999 which initially came from its non-fluorinated derivative (E)-N-(3-iodoprop-2-enyl)-2-carbomethoxy-3-(4-methylphenyl) nortropane, or PE2I, because of its high promising properties. [18F]LBT-999 has been extensively characterized in rodent and non-human primate models, in which it demonstrated its capability to explore in vivo the DAT localized at the dopaminergic nerve endings as well as at the mesencephalic cell bodies, in physiological conditions. In lesion-induced rat models of Parkinson's disease, [18F]LBT-999 was able to precisely quantify in vivo the dopaminergic neuron loss, and to assess the beneficial effects of therapeutic approaches such as pharmacological treatment and cell transplantation. Finally recent clinical data demonstrated the efficiency of [18F]LBT-999 in the diagnosis of Parkinson's disease