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Supplemental material, sj-pdf-1-jcb-10.1177_0271678X221108841 for Quantitative kinetic modelling and mapping of cerebral glucose transport and metabolism using glucoCESL MRI by Ben R Dickie, Tao Jin, Ping Wang, Rainer Hinz, William Harris, Hervé Boutin, Geoff JM Parker, Laura M Parkes and Julian C Matthews in Journal of Cerebral Blood Flow & Metabolism

Mouse models of Alzheimer’s disease (AD) are valuable but do not fully recapitulate human AD pathology, such as spontaneous Tau fibril accumulation and neuronal loss, necessitating the development of new AD models. The transgenic (TG) TgF344-AD rat has been reported to develop age-dependent AD features including neuronal loss and neurofibrillary tangles, despite only expressing APP and PSEN1 mutations, suggesting an improved modelling of AD hallmarks. Alterations in neuronal networks as well as learning performance and cognition tasks have been reported in this model, but none have combined a longitudinal, multimodal approach across multiple centres, which mimics the approaches commonly taken in clinical studies. We therefore aimed to further characterise the progression of AD-like pathology and cognition in the TgF344-AD rat from young-adults (6 months (m)) to mid- (12 m) and advanced-stage (18 m, 25 m) of the disease.Methods. TgF344-AD rats and wild-type (WT) littermates were imaged at 6 m, 12 m and 18 m with [18F]DPA-714 (TSPO, neuroinflammation), [18F]Florbetaben (Aβ) and [18F]ASEM (α7- nicotinic acetylcholine receptor) and with magnetic resonance spectroscopy (MRS) and with (S)-[18F]THK5117 (Tau) at 15 and 25 m. Behaviour tests were also performed at 6 m, 12 m and 18 m. Immunohistochemistry (CD11b, GFAP, Aβ, NeuN, NeuroChrom) and Tau (S)-[18F]THK5117 autoradiography, immunohistochemistry and Western blot were also performed.Results. [18F]DPA-714 positron emission tomography (PET) showed an increase in neuroinflammation in TG vs wildtype animals from 12 m in the hippocampus (+11%), and at the advanced-stage AD in the hippocampus (+12%), the thalamus (+11%) and frontal cortex (+14%). This finding coincided with strong increases in brain microgliosis (CD11b) and astrogliosis (GFAP) at these time-points as assessed by immunohistochemistry. In vivo [18F]ASEM PET revealed an age-dependent increase uptake in the striatum and pallidum/nucleus basalis of Meynert in WT only, similar to that observed with this tracer in humans, resulting in TG being significantly lower than WT by 18 m. In vivo [18F]Florbetaben PET scanning detected Aβ accumulation at 18 m, and (S)-[18F]THK5117 PET revealed subsequent Tau accumulation at 25m in hippocampal and cortical regions. Aβ plaques were low but detectable by immunohistochemistry from 6 m, increasing further at 12 and 18 m with Tau-positive neurons adjacent to Aβ plaques at 18 m. NeuroChrom (a pan neuronal marker) immunohistochemistry revealed a loss of neuronal staining 5 at the Aβ plaques locations, while NeuN labelling 103 revealed an age-dependent decrease in hippocampal neuron number in both genotypes. Behavioural assessment using the novel object recognition task revealed that both WT & TgF344-AD animals discriminated the novel from familiar object at 3 m and 6 m of age. However, low levels of exploration observed in both genotypes at later time-points resulted in neither genotype successfully completing the task. Deficits in social interaction were only observed at 3 m in the TgF344-AD animals. By in vivo MRS, we showed a decrease in neuronal marker N-acetyl-aspartate in the hippocampus at 18 m (-18% vs age-matched WT, and -31% vs 6 m TG) and increased Taurine in the cortex of TG (+35% vs age-matched WT, and +55% vs 6 m TG).Conclusions. This multi-centre multi-modal study demonstrates, for the first time, alterations in brain metabolites, cholinergic receptors and neuroinflammation in vivo in this model, validated by robust ex vivo approaches. Our data confirm that, unlike mouse models, the TgF344- AD express Tau pathology that can be detected via PET, albeit later than by ex vivo techniques, and is a useful model to assess and longitudinally monitor early neurotransmission dysfunction and neuroinflammation in AD.

A defining pathophysiological hallmark of Alzheimer's disease (AD) is the amyloid plaque; an extracellular deposit of aggregated fibrillar Aβ1-42 peptides. Amyloid plaques are hard, brittle structures scattered throughout the hippocampus and cerebral cortex and are thought to cause hyperphosphorylation of tau, neurofibrillary tangles, and progressive neurodegeneration. Reactive astrocytes and microglia envelop the exterior of amyloid plaques and infiltrate their inner core. Glia are highly mechanosensitive cells and can almost certainly sense the mismatch between the normally soft mechanical environment of the brain and very stiff amyloid plaques via mechanosensing ion channels. Piezo1, a non-selective cation channel, can translate extracellular mechanical forces to intracellular molecular signaling cascades through a process known as mechanotransduction. Here, we utilized an aging transgenic rat model of AD (TgF344-AD) to study expression of mechanosensing Piezo1 ion channels in amyloid plaque-reactive astrocytes. We found that Piezo1 is upregulated with age in the hippocampus and cortex of 18-month old wild-type rats. However, more striking increases in Piezo1 were measured in the hippocampus of TgF344-AD rats compared to age-matched wild-type controls. Interestingly, repeated urinary tract infections with Escherichia coli bacteria, a common comorbidity in elderly people with dementia, caused further elevations in Piezo1 channel expression in the hippocampus and cortex of TgF344-AD rats. Taken together, we report that aging and peripheral infection augment amyloid plaque-induced upregulation of mechanoresponsive ion channels, such as Piezo1, in astrocytes. Further research is required to investigate the role of astrocytic Piezo1 in the Alzheimer's brain, whether modulating channel opening will protect or exacerbate the disease state, and most importantly, if Piezo1 could prove to be a novel drug target for age-related dementia.

AIMS: Although changes in extracellular matrix (ECM) scaffold have been reported previously in AD compared to normal ageing, it is not known how alterations in the numerous components of the perivascular ECM might occur at different stages of AD. This study therefore investigates potential changes in basement membrane-associated ECM molecules in relation to increasing Braak stages. METHODS: Thirty patients were divided into three groups (control subject, subclinical AD and AD patients). ECM levels of collagen IV, perlecan and fibronectin as well as human platelet endothelial cell adhesion molecule (hPECAM) were quantified by immunohistochemistry. Von Willebrand factor staining was measured to assess vessel density. Expression levels were correlated with the presence of amyloid plaques. RESULTS: Collagen IV, perlecan and fibronectin expression was increased in subclinical AD and AD patients when compared to controls, in frontal and temporal cortex, whilst no further increase was detected between subclinical AD and AD. These changes were not associated with an increase in vessel density, which was instead decreased in the temporal cortex of AD patients. In contrast, hPECAM levels remained unchanged. Finally, we found similar pattern in levels of amyloid deposition between the different Braak stages and showed that changes in ECM components correlated with amyloid deposition. CONCLUSION: Present data support the hypothesis that significant ECM changes occur during the early stages of AD. ECM changes affecting brain microvascular functions could therefore drive disease progression and provide potential new early investigational biomarkers in AD.

PURPOSE: Neuroinflammation plays a critical role in various neuropathological conditions, and hence there is renewed interest in the translocator protein (TSPO) as a biomarker of microglial activation and macrophage infiltration in the brain. This is reflected in the large amount of research conducted seeking to replace the prototypical PET radiotracer 11C-R-PK11195 with a TSPO ligand with higher performance. Here we report the in vivo preclinical investigation of the novel TSPO tracer 18F-GE-180 in a rat model of stroke. METHODS: Focal cerebral ischaemia was induced in Wistar rats by 60-min occlusion of the middle cerebral artery (MCAO). Brain damage was assessed 24 h after MCAO by T2 MRI. Rats were scanned with 11C-R-PK11195 and 18F-GE-180 5 or 6 days after MCAO. Specificity of binding was confirmed by injection of unlabelled R-PK11195 or GE-180 20 min after injection of 18F-GE-180. In vivo data were confirmed by ex vivo immunohistochemistry for microglial (CD11b) and astrocytic biomarkers (GFAP). RESULTS: 18F-GE-180 uptake was 24 % higher in the core of the ischaemic lesion and 18 % lower in the contralateral healthy tissue than that of 11C-R-PK11195 uptake (1.5 ± 0.2-fold higher signal to noise ratio). We confirmed this finding using the simplified reference tissue model (BPND = 3.5 ± 0.4 and 2.4 ± 0.5 for 18F-GE-180 and 11C-R-PK11195, respectively, with R 1 = 1). Injection of unlabelled R-PK11195 or GE-180 20 min after injection of 18F-GE-180 significantly displaced 18F-GE-180 (69 ± 5 % and 63 ± 4 %, respectively). Specificity of the binding was also confirmed by in vitro autoradiography, and the location and presence of activated microglia and infiltrated macrophages were confirmed by immunohistochemistry. CONCLUSION: The in vivo binding characteristics of 18F-GE-180 demonstrate a better signal to noise ratio than 11C-R-PK11195 due to both a better signal in the lesion and lower nonspecific binding in healthy tissue. These results provide evidence that 18F-GE-180 is a strong candidate to replace 11C-R-PK11195.