Introduction Neutrophil accumulation is a well-established feature of Alzheimer’s disease (AD) and has been linked to cognitive impairment by modulating disease-relevant neuroinflammatory and vascular pathways. Neutrophils express high levels of the oxidant-generating enzyme myeloperoxidase (MPO), however there has been controversy regarding the cellular source and localisation of MPO in the AD brain. Materials and methods We used immunostaining and immunoassays to quantify the accumulation of neutrophils in human AD tissue microarrays and in the brains of APP/PS1 mice. We also used multiplexed immunolabelling to define the presence of NETs in AD. Results There was an increase in neutrophils in AD brains as well as in the murine APP/PS1 model of AD. Indeed, MPO expression was almost exclusively confined to S100A8-positive neutrophils in both human AD and murine APP/PS1 brains. The vascular localisation of neutrophils in both human AD and mouse models of AD was striking and driven by enhanced neutrophil adhesion to small vessels. We also observed rare infiltrating neutrophils and deposits of MPO around plaques. Citrullinated histone H3, a marker of neutrophil extracellular traps (NETs), was also detected in human AD cases at these sites, indicating the presence of extracellular MPO in the vasculature. Finally, there was a reduction in the endothelial glycocalyx in AD that may be responsible for non-productive neutrophil adhesion to the vasculature. Conclusion Our report indicates that vascular changes may drive neutrophil adhesion and NETosis, and that neutrophil-derived MPO may lead to vascular oxidative stress and be a relevant therapeutic target in AD.
Platelet-derived growth factor-BB (PDGF-BB):PDGF receptor-β (PDGFRβ) signalling in brain pericytes is critical to the development, maintenance and function of a healthy blood-brain barrier (BBB). Furthermore, BBB impairment and pericyte loss in Alzheimer’s disease (AD) is well documented. We found that PDGF-BB:PDGFRβ signalling components were altered in human AD brains, with a marked reduction in vascular PDGFB. We hypothesised that reduced PDGF-BB:PDGFRβ signalling in pericytes may impact on the BBB. We therefore tested the effects of PDGF-BB on primary human brain pericytes in vitro to define pathways related to BBB function. Using pharmacological inhibitors, we dissected distinct aspects of the PDGF-BB response that are controlled by extracellular signal-regulated kinase (ERK) and Akt pathways. PDGF-BB promotes the proliferation of pericytes and protection from apoptosis through ERK signalling. In contrast, PDGF-BB:PDGFRβ signalling through Akt augments pericyte-derived inflammatory secretions. It may therefore be possible to supplement PDGF-BB signalling to stabilise the cerebrovasculature in AD.
Traditional neuroanatomy immunohistology studies involve low-content analyses of a few antibodies of interest, typically applied and compared across sequential tissue sections. The efficiency, consistency, and ultimate insights of these studies can be substantially improved using high-plex immunofluorescence labelling on a single tissue section to allow direct comparison of many markers. Here we present an expanded and efficient multiplexed fluorescence-based immunohistochemistry (MP-IHC) approach that improves throughput with sequential labelling of up to 10 antibodies per cycle, with no limitation on the number of cycles, and maintains versatility and accessibility by using readily available commercial reagents and standard epifluorescence microscopy imaging. We demonstrate this approach by cumulatively screening up to 100 markers on formalin-fixed paraffin-embedded sections of human olfactory bulb sourced from neurologically normal (no significant pathology), Alzheimer’s (AD), and Parkinson’s disease (PD) patients. This brain region is involved early in the symptomology and pathophysiology of AD and PD. We also developed a spatial pixel bin analysis approach for unsupervised analysis of the high-content anatomical information from large tissue sections. Here, we present a comprehensive immunohistological characterisation of human olfactory bulb anatomy and a summary of differentially expressed biomarkers in AD and PD using the MP-IHC labelling and spatial protein analysis pipeline.
Olfactory deficits are an early and prevalent non-motor symptom of Huntington's disease (HD). In other neurodegenerative diseases where olfactory deficits occur, such as Alzheimer's disease and Parkinson's disease, pathological protein aggregates (tau, β-amyloid, α-synuclein) accumulate in the anterior olfactory nucleus (AON) of the olfactory bulb (OFB). Therefore, in this study we determined whether aggregates are also present in HD OFBs; 13 HD and five normal human OFBs were stained for mutant huntingtin (mHtt), tau, β-amyloid, TDP-43, and α-synuclein. Our results show that mHtt aggregates detected with 1F8 antibody are present within all HD OFBs, and mHtt aggregate load in the OFB does not correlate with Vonsattel grading scores. The majority of the aggregates were located in the AON and in similar abundance in each anatomical segment of the AON. No mHtt aggregates were found in controls; 31% of HD cases also contained tau neurofibrillary tangles within the AON. This work demonstrates HD pathology in the OFB and indicates that disease-specific protein aggregation in the AON is a common feature of neurodegenerative diseases that show olfactory deficits.
Gene expression studies of human post-mortem brain tissue are useful for understanding the pathogenesis of neurodegenerative disease. These studies rely on the assumption that RNA quality is consistent between disease and neurologically normal cases; however, previous studies have suggested that RNA quality may be affected by neurodegenerative disease. Here, we compared RNA quality in human post-mortem brain tissue between neurologically normal cases (n = 14) and neurodegenerative disease cases (Alzheimer’s disease n = 10; Parkinson’s disease n = 11; and Huntington’s disease n = 9) in regions affected by pathology and regions that are relatively devoid of pathology. We identified a statistically significant decrease in RNA integrity number (RIN) in Alzheimer’s disease tissue relative to neurologically normal tissue (mixed effects model, p = 0.04). There were no statistically significant differences between neurologically normal cases and Parkinson’s disease or Huntington’s disease cases. Next, we investigated whether total RNA quality affected mRNA quantification, by correlating RIN with qPCR threshold cycle (CT). CT values for all six genes investigated were strongly correlated with RIN (p < 0.05, Pearson correlation); this effect was only partially mitigated by normalization to RPL30. Our results indicate that RNA quality is decreased in Alzheimer’s disease tissue. We recommend that RIN should be considered when this tissue is used in gene expression analyses.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.