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The effects of Alzheimer's disease (AD) and ageing on blood–brain barrier (BBB) breakdown are investigated in TgF344-AD and wild-type rats aged 13, 18 and 21 months. Permeability surface area products of the BBB to water (PSw) and gadolinium-based contrast agent (PSg) were measured in grey matter using multiflip angle multiecho dynamic contrast-enhanced MRI. At 13 months of age, there was no significant difference in PSw between TgF344-AD and wild-types (p = 0.82). Between 13 and 18 months, PSw increased in TgF344-AD rats (p = 0.027), but not in wildtypes (p = 0.99), leading to significantly higher PSw in TgF344-AD rats at 18 months, as previously reported (p = 0.012). Between 18 and 21 months, PSw values increased in wild-types (p = 0.050), but not in TgF344-AD rats (p = 0.50). These results indicate that BBB water permeability is affected by both AD pathology and ageing, but that changes occur earlier in the presence of AD pathology. There were no significant genotype or ageing effects on PSg (p > 0.05). In conclusion, we detected increases in BBB water permeability with age in TgF344-AD and wild-type rats, and found that changes occurred at an earlier age in rats with AD pathology.

In the brain, REST (Repressor Element-1 Silencing Transcription factor) is a key regulator of neuron cell-specific gene expression. Nuclear translocation of neuronal REST has been shown to be neuroprotective in a healthy ageing context. In contrast, inability to upregulate nuclear REST is thought to leave ageing neurons vulnerable to neurodegenerative stimuli, such as Alzheimer's disease (AD) pathology. Hippocampal and cortical neurons are known to be particularly susceptible to AD-associated neurodegeneration. However, REST expression has not been extensively characterised in the healthy ageing brain. Here, we examined the spatiotemporal immunolocalisation of REST in the brains of healthy ageing wild-type Fischer-344 and transgenic Alzheimer's disease rats (TgF344-AD). Nuclear expression of REST increased from 6 months to 18 months of age in the hippocampus, frontal cortex and subiculum of wild-type rats, but not in TgF344-AD rats. No changes in REST were measured in more posterior cortical regions or in the thalamus. Interestingly, levels of the presynaptic marker synaptophysin, a known gene target of REST, were lower in CA1 hippocampal neurons of 18-month TgF344-AD rats compared to 18-month wild-types, suggesting that elevated nuclear REST may protect against synapse loss in the CA1 of 18-month wild-type rats. High REST expression in ageing wild-type rats did not, however, protect against axonal loss nor against astroglial reactivity in the hippocampus. Taken together, our data confirm that changes in nuclear REST expression are context-, age- and brain region-specific. Moreover, key brain structures involved in learning and memory display elevated REST expression in healthy ageing wild-type rats but not TgF344-AD rats.

The sigma 1 receptor (S1R) is widely expressed in the CNS and is mainly located on the endoplasmic reticulum. The S1R is involved in the regulation of many neurotransmission systems and, indirectly, in neurodegenerative diseases. The S1R may therefore represent an interesting neuronal biomarker in neurodegenerative diseases such as Parkinson’s (PD) or Alzheimer’s diseases (AD). Here we present the characterisation of the S1R-specific 18F-labelled tracer 18F-IAM6067 in two animal models and in human brain tissue. Methods: Wistar rats were used for PET-CT imaging (60 min dynamic acquisition) and metabolite analysis (1, 2, 5, 10, 20, 60 min post-injection). To verify in vivo selectivity, haloperidol, BD1047 (S1R ligand), CM398 (S2R ligand) and SB206553 (5HT2B/C antagonist) were administrated for pre-saturation studies. Excitotoxic lesions induced by intra-striatal injection of AMPA were also imaged by 18F-IAM6067 PET-CT to test the sensitivity of the methods in a well-established model of neuronal loss. Tracer brain uptake was also verified by autoradiography in rats and in a mouse model of PD (intrastriatal 6-hydroxydopamine (6-OHDA) unilateral lesion). Finally, human cortical binding was investigated by autoradiography in three groups of subjects (control subjects with Braak ≤2, and AD patients, Braak >2 & ≤4 and Braak >4 stages). Results: We demonstrate that despite rapid peripheral metabolism of 18F-IAM6067, radiolabelled metabolites were hardly detected in brain samples. Brain uptake of 18F-IAM6067 showed differences in S1R anatomical distribution, namely from high to low uptake: pons-raphe, thalamus medio-dorsal, substantia nigra, hypothalamus, cerebellum, cortical areas and striatum. Pre-saturation studies showed 79-90% blockade of the binding in all areas of the brain indicated above except with the 5HT2B/C antagonist SB206553 and S2R ligand CM398 which induced no significant blockade, indicating good specificity of 18F-IAM6067 for S1Rs. No difference between ipsi- and contralateral sides of the brain in the mouse model of PD was detected. AMPA lesion induced a significant 69% decrease in 18F-IAM6067 uptake in the globus pallidus matching the neuronal loss as measured by NeuN, but only a trend to decrease (-16%) in the caudate putamen despite a significant 91% decrease in neuronal count. Moreover, no difference in the human cortical binding was shown between AD groups and controls.

It has become increasingly evident that neuroinflammation plays a critical role in the pathophysiology of Alzheimer’s disease (AD) and other neurodegenerative disorders. Increased glial cell activation is consistently reported in both rodent models of AD and in AD patients. Moreover, recent genome wide association studies have revealed multiple genes associated with inflammation and immunity are significantly associated with an increased risk of AD development (e.g. TREM2). Noninvasive in vivo detection and tracking of neuroinflammation is necessary to enhance our understanding of the contribution of neuroinflammation to the initiation and progression of AD. Importantly, accurate methods of quantifying neuroinflammation may aid early diagnosis and serve as an output for therapeutic monitoring and disease management. This review details current in vivo imaging biomarkers of neuroinflammation being explored and summarizes both pre-clinical and clinical results from molecular imaging studies investigating the role of neuroinflammation in AD, with a focus on positron emission tomography and magnetic resonance spectroscopy (MRS).

International audience; IntroductionSynchrotron Radiation X-ray Phase Computed Tomography (SR-PCT) of brain tissue can reveal different image contrast depending on sample preparation. Indeed, we have shown that ethanol dehydration of brain samples reveals myelin as a hyper-intense signal while maintaining optimal detection of packed proteinaceous structures such as amyloid plaques [1]. The present work builds on this versatile “virtual histology” tool to characterize amyloid deposits in brain samples from Alzheimer’s disease (AD) animal models and patients.MethodsMono, bi-, and triple transgenic lines of mouse (PDAPP, APP-PS1, 3xTg [2]) were studied, along with one line of transgenic rats [3]. Human samples of frontal cortex were obtained from 6 sporadic AD cases and 1 familial AD case (Cardiobiotec, CRB-HCL, Lyon, France). All samples were scanned after dehydration in ethanol on beamlines ID19 and ID17 at ESRF. In line propagation imaging was performed at 26keV, voxel size of 6.5μm and 3m free space propagation on ID19 (34 keV, 6.5µm and 11m on ID17). Images were reconstructed using Paganin’s method [4].Results/DiscussionAll transgenic animal brains (Fig. 1) and human AD (Fig. 2) cases exhibited amyloid signals in the form of bright spots. Myelin-based hyper-intensities enabled a detailed visualisation of white-matter tracts and subsequent structural connectivity. Additionally, vessels appeared hypo-intense in perfused brains from animals (Fig.1) and hyper-intense in human autopsic brains (Fig. 2), probably because of the presence of blood-related iron in non-perfused tissue. Concurrent immunofluorescence analyses of these observations are in progress.ConclusionsWhole-brain mouse acquisitions were performed faster (3-10 min) than previously described phase contrast approaches [5]. Human amyloid plaques were evidenced for the first time by SR-PCT. Dehydration-added myelin contrast may be used to highlight unexplored associations between amyloid deposition and myelin alterations.References[1] Rositi et al. Abstract #233, presented in European Molecular Imaging Meeting, Mar 18-20, Tübingen, Germany; 2015.[2] Mucke et al. J Neurosci. 2000 ; Blanchard et al. Exp Neurol 2003 ; Oddo et al. Neuron. 2003.[3] Cohen RM, et al. J Neurosci. 2013.[4] Paganin et al. J Microsc. 2002.[5] Noda-Saita et al. Neuroscience 2006 ; Connor et al. NeuroImage 2009 ; Pinzer et al. NeuroImage 2012 ; Astolfo et al. J Synchrotron Radiat. 2016.AcknowledgementWe are grateful to our collaborators (Nicolas Rama, Corinne Perrin, Karen Silva) for their help in gathering animal and human brain samples.

Mixed leukocyte (white blood cells [WBCs]) trafficking using positron emission tomography (PET) is receiving growing interest to diagnose and monitor inflammatory conditions. PET, a high sensitivity molecular imaging technique, allows precise quantification of the signal produced from radiolabelled moieties. We have evaluated a new method for radiolabelling WBCs with either zirconium-89 ((89) Zr) or copper-64 ((64) Cu) for PET imaging. Chitosan nanoparticles (CNs) were produced by a process of ionotropic gelation and used to deliver radiometals into WBCs. Experiments were carried out using mixed WBCs freshly isolated from whole human blood. WBCs radiolabelling efficiency was higher with [(89) Zr]-loaded CN (76.8 ± 9.6% (n = 12)) than with [(64) Cu]-loaded CN (26.3 ± 7.0 % (n = 7)). [(89) Zr]-WBCs showed an initial loss of 28.4 ± 5.8% (n = 2) of the radioactivity after 2 h. This loss was then followed by a plateau as (89) Zr remains stable in the cells. [(64) Cu]-WBCs showed a loss of 85 ± 6% (n = 3) of the radioactivity after 1 h, which increased to 96 ± 6% (n = 3) loss after 3 h. WBC labelling with [(89) Zr]-loaded CN showed a fast kinetic of leukocyte association, high labelling efficiency and a relatively good retention of the radioactivity. This method using (89) Zr has a potential application for PET imaging of inflammation.

Permanent focal cortical ischemia was induced in mice by electrocoagulation of the middle cerebral artery. At different time intervals after the injury, the volume of infarction was assessed together with an analysis of neuronal death. Morphological studies of ischemic brains and detection of nucleosomal DNA ladder within ipsilateral cortices might implicate a component of this neuronal loss to apoptosis as well as necrosis. Furthermore, we used the TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling) procedure to detect in situ DNA fragmentation. The localization and the proportion of apoptotic cells in the ischemic mouse brain would indicate that apoptosis contributes largely to the cellular loss induced by cerebral ischemia.