A set of 2000x2000 raw confocal fluorescence images of MOAB2-labelled cortex from APPswe/PS1dE9 mouse models of Alzheimer's disease, un-annotated intensity scaled 8-bit images, sparsely annotated pixel labels and reference segmentation examples. Pixels are annotated as signal (red 0xFFFF0000) and background (blue 0xFF0000FF). Images were captured as stitched 12-bit greyscale single-plane images and cropped to size. Image acquisition was performed at 561nm excitation and 615nm emission wavelengths using a Perkin Elmer Ultraview VOX imaging system with Nikon Eclipse Ti inverted microscope, 20x objective lens, x-y motorised stage, Hamamatsu ORCA-R2 C10600-10B CCD camera and Volocity 6.3 software package. These images are shared as examples for the ImageSURF segmentation plugin for ImageJ2/FIJI (https://doi.org/10.5281/zenodo.818109).
Cerebral blood flow (CBF) is important for the maintenance of brain function and its dysregulation has been implicated in Alzheimer's disease (AD). Microglia associations with capillaries suggest they may play a role in the regulation of CBF or the blood-brain-barrier (BBB). We explored the relationship between microglia and pericytes, a vessel-resident cell type that has a major role in the control of CBF and maintenance of the BBB, discovering a spatially distinct subset of microglia that closely associate with pericytes. We termed these pericyte-associated microglia (PEM). PEM are present throughout the brain and spinal cord in NG2DsRed × CX
Abstract Background Dementia prevention and drug development is hindered by the lack of low‐cost population‐level tests to help identify preclinical Alzheimer’s disease (AD) in the community. Recent evidence suggests that precise analysis of hand movements may detect motor changes indicative of preclinical AD. The objective was to evaluate how TAS Test, a new online automated hand movement test, predicts preclinical AD biomarkers (plasma ptau181 and subtle episodic memory decline) in a cognitively asymptomatic cohort of older adults. Method Participants completed TAS Test online at home using their own computer without researcher assistance: a series of 10‐30 second index finger‐key and index finger‐thumb tapping tests recorded with a keyboard and/or webcam. Movement features including frequency, rhythm, pauses and accuracy were extracted. Participants also completed online tests of episodic memory, CANTAB Paired Associate Learning (CANTAB). A subset provided blood samples for ptau181 analysis. Accuracy of linear regression models comprising hand motor features to predict PAL scores and ptau181 levels, adjusted for confounding, was compared to null models (with only confounders: age, gender, education level, anxiety and depression) using R2adj and AIC. ΔAIC > 2 denotes statistical difference. Result 1,228 adults (mean (SD) age, 65.8 (7.4) years; 73.0% female) completed TAS Test and CANTAB; 459 underwent ptau181 analysis. All keyboard tests improved prediction of asymptomatic episodic memory decline; the 3 step‐ (ΔAIC = 11.2; R2adj = 8.1%) and alternate‐ key (ΔAIC = 3.3; R2adj = 7.5%) tests ranked highest and were the only keyboard tests to improve prediction of ptau181 (3 step ΔAIC = 7.0; R2adj = 17.8%; alternate key ΔAIC = 3.4; R2adj = 17.4%). All finger‐thumb motor measures improved predictions of CANTAB scores and ptau181 levels; the highest performing tests were dominant hand tapping (CANTAB ΔAIC = 2.9; R2adj = 8.2%; ptau181 ΔAIC = 2.4; R2adj = 12.9%) and both hands dual‐task tapping (CANTAB ΔAIC = 3.0; R2adj = 6.8%; ptau181 ΔAIC = 8.7; R2adj = 11.9%). Conclusion TAS Test provides an accessible brief home‐based test for identifying preclinical AD risk. This novel approach holds potential as a pre‐screening/enrichment tool for identifying at risk cohorts for further investigation.
Abstract Background The majority of individuals with Alzheimer’s disease (AD) exhibit vascular damage in the brain including reduced cerebral blood flow (CBF) and blood‐brain‐barrier (BBB) breakdown. Growing evidence suggests pericytes, a vessel‐residing cell able to modulate CBF and BBB integrity, are a major contributor to vascular dysfunction in AD. The close spatial relationship between microglia and capillaries led us to explore the hypothesis that microglia and pericytes functionally interact to influence vessel function in the healthy brain and that this interaction may become pathological in AD. Methods We first characterized the spatial and functional relationship between microglia, capillaries and pericytes in the healthy central nervous system (CNS) in fixed tissue and using in vivo two‐photon microscopy through cranial windows in adult NG2DsRed x CX3CR1 +/GFP mice. We then assessed the spatial relationship between microglia, capillaries and pericytes in the human superior frontal gyrus (SFG) of 12 control and 11 AD cases. Results We discovered a subset of microglia dynamically interacting with pericytes in the healthy CNS, which we termed pe ricyte‐associated m icroglia (PEM). PEM are present throughout the capillary tree, often maintain their position for at least 28 days and frequently were found to associate with pericytes lacking astroglial endfeet coverage. The loss of PEM alters capillary width beneath pericytes, but deletion of the microglial fractalkine receptor (CX3CR1), which regulates the recruitment of microglia to sites of neuroinflammation, does not reduce the proportion of microglia that are PEM. In AD, we found the proportion of microglia associated with capillaries and pericytes declines in the superior frontal gyrus (SFG) and that this decline is exacerbated by the APOE ε3/ε4 genotype. This reduction occurs despite an overall increase in pericyte numbers in AD cases. Conclusion We identify microglia specifically associating with pericytes in the healthy CNS and find these associations are reduced in AD. The changing relationship between microglia, capillaries and pericytes may be a novel mechanism contributing to vascular dysfunction in AD.
Abstract Background The failure of some clinical trials using drugs that remove brain amyloid to prevent progression of dementia suggest that neurodegenerative disease may not progress in a linear fashion and threshold levels of one pathology may initiate alterations that are unable to be reversed upon removal of the initiating pathological process. We used the inducible Tg4510 mouse tauopathy model to determine whether neurodegeneration could be halted following removal of transgene expression in mice aged to 18 months Method Pathology was induced in mice from 8 weeks until 18 months of age (n = 20 equal male/female; both tau transgenic and non‐transgenic) by removal of doxycycline (dox) diet. Dox diet was returned to half the animals from 18‐23 months to inhibit pathology. Levels of serum Nfl were measured using SIMOA assay throughout disease progression and p‐tau181 along with other serum biomarkers were measured at endpoint. Immunohistochemical assays were performed to quantitate numbers of neurons (NeuN) and tau load (PHF‐tau) in hippocampus. Result Serum Nfl increased over aging in all mice and was significantly higher in tau transgenics relative to non‐transgenics at 18 months (p<0.05). Between 18 and 23 months, the rate of serum Nfl increase was significantly(p<0.05) reduced in male transgenic mice on dox compared to female transgenic mice, so that by 23 months levels were not different from non‐transgenic mice. Similarly, the numbers of neurons in the hippocampus of male mice on dox at 23 months was significantly higher than their female counterparts (p<0.05). When looking at levels of serum p‐tau181 at 23 months, there was significantly (p<0.05) higher levels in male mice on dox than female mice Conclusion These data suggest that aged female mice have ongoing neurodegeneration following removal of pathology induction that is similar to mice with continued pathology induction. The differences in serum p‐tau‐181 may indicate that female mice fail to clear pathological tau from the brain. Further studies will examine the relationship between histopathology and blood biomarkers and determine whether female mice on dox have a higher tau pathology load than their male counterparts.
Images of amyloid-β pathology characteristic of Alzheimer's disease are difficult to consistently and accurately segment, due to diffuse deposit boundaries and imaging variations.We evaluated the performance of ImageSURF, our open-source ImageJ plugin, which considers a range of image derivatives to train image classifiers. We compared ImageSURF to standard image thresholding to assess its reproducibility, accuracy and generalizability when used on fluorescence images of amyloid pathology.ImageSURF segments amyloid-β images significantly more faithfully, and with significantly greater generalizability, than optimized thresholding.In addition to its superior performance in capturing human evaluations of pathology images, ImageSURF is able to segment image sets of any size in a consistent and unbiased manner, without requiring additional blinding, and can be retrospectively applied to existing images. The training process yields a classifier file which can be shared as supplemental data, allowing fully open methods and data, and enabling more direct comparisons between different studies.
Two cell lines, U-937 and MOLT-4, were used to investigate the toxicity, DNA incorporation, and effect on mitochondria of 1-(2'-fluoro-2'-deoxy-beta-D-arabinofuranosyl)-5-iodouracil (FIAU) and its putative metabolite 1-(2'-fluoro-2'-deoxy-beta-D-arabinofuranosyl)-uracil (FAU). After 72-hr incubation, the IC50 values for FIAU were 6.4 microM for U-937 cells and 26 microM for MOLT-4 cells. IC50 values for FAU were 10-fold higher in both cell lines. Incubation for 24 hr with 10 microM [2-14C]FIAU led to 2.1% and 0.93% replacement of thymidine in DNA of U-937 and MOLT-4 cells, respectively. The predominant radioactive species measurable in DNA was FIAU. A similar incubation with [2-14C]FAU resulted in 4-fold lower DNA incorporation of a single radioactive species that coeluted with 1-(2'-fluoro-2'-deoxy-beta-D-arabinofuranosyl)-5-methyluracil (FMAU). There was no evidence of a selective repair process after DNA incorporation of FIAU or FAU (FMAU). Increased intracellular concentrations of FIAU triphosphate and incorporation into DNA were associated with an increase in cellular toxicity. Continuous exposure to a clinically achievable concentration of FIAU, 0.44 microM, produced a constant DNA incorporation of 0.80% and 0.11% for U-937 and MOLT-4 cells, respectively. FIAU was not readily metabolized to FAU or iodouracil by human liver in vitro. Compared with 2',3'-dideoxycytidine as a positive control, after 12 days of continuous exposure of U-937 and MOLT-4 cells to FIAU there was no evidence of increased lactate production. These data negate several possible mechanisms (DNA chain termination, DNA polymerase inhibition, one form of selective mitochondrial poisoning, and FAU-mediated toxicity) and provide clues for possible mechanisms (FIAU triphosphate concentration and DNA incorporation). Further work is needed to develop a complete explanation for the delayed hepatic toxicity observed in the investigational clinical trials of FIAU.
Abstract Metabolic changes are observed in patients with both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Although regulation of metabolic processes in the CNS is predominantly carried out within the hypothalamus, extra-hypothalamic CNS areas contain metabolic hormone receptors, including those for leptin (LEPR), insulin (INSR), and neuropeptide Y (NPY), indicating that they may play a role in biological processes underlying pathogenic disease processes. The status of these hormones within regions vulnerable in ALS/FTD is not well described. This study sought to determine whether the expression of these hormones and their receptors is altered in pathology-rich regions in cases of human FTD (superior frontal gyrus and insular cortex) and ALS (primary motor cortex and lumbar spinal cord) with TDP-43 pathology compared to matched healthy controls. LEPR mRNA was increased within the superior frontal gyrus of FTD cases and within primary motor cortex and lumbar spinal cord of ALS cases; INSR mRNA was increased in superior frontal gyrus and insular cortex of FTD cases. NPY protein was decreased in primary motor cortex and lumbar spinal cord of ALS cases. Our results demonstrate that metabolic hormones undergo complex alterations in ALS and FTD and suggest that these hormones could play critical roles in the pathogenesis of these diseases.
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