Kandimalla, Karunya K.

AbstractINTRODUCTIONUnderstanding the relationship between amyloid beta (Aβ) positron emission tomography (PET) and Aβ cerebrospinal fluid (CSF) biomarkers will define their potential utility in Aβ treatment. Few population‐based or neuropathologic comparisons have been reported.METHODSParticipants 50+ years with Aβ PET and Aβ CSF biomarkers (phosphorylated tau [p‐tau]181/Aβ42, n = 505, and Aβ42/40, n = 54) were included from the Mayo Clinic Study on Aging. From these participants, an autopsy subgroup was identified (n = 47). The relationships of Aβ PET and Aβ CSF biomarkers were assessed cross‐sectionally in all participants and longitudinally in autopsy data.RESULTSCross‐sectionally, more participants were Aβ PET+ versus Aβ CSF− than Aβ PET− versus Aβ CSF+ with an incremental effect when using Aβ PET regions selected for early Aβ deposition. The sensitivity for the first detection of Thal phase ≥ 1 in longitudinal data was higher for Aβ PET (89%) than p‐tau181/Aβ42 (64%).DISCUSSIONAβ PET can detect earlier cortical Aβ deposition than Aβ CSF biomarkers. Aβ PET+ versus Aβ CSF− findings are several‐fold greater using regional Aβ PET analyses and in peri‐threshold‐standardized uptake value ratio participants.Highlights Amyloid beta (Aβ) positron emission tomography (PET) has greater sensitivity for Aβ deposition than Aβ cerebrospinal fluid (CSF) in early Aβ development. A population‐based sample of participants (n = 505) with PET and CSF tests was used. Cortical regions showing early Aβ on Aβ PET were also used in these analyses. Neuropathology was used to validate detection of Aβ by Aβ PET and Aβ CSF biomarkers.

Background: A strong body of evidence suggests that cerebrovascular pathologies augment the onset and progression of Alzheimer’s disease (AD). One distinctive aspect of this cerebrovascular dysfunction is the degeneration of brain pericytes—often overlooked supporting cells of blood-brain barrier endothelium. Objective: The current study investigates the influence of pericytes on gene and protein expressions in the blood-brain barrier endothelium, which is expected to facilitate the identification of pathophysiological pathways that are triggered by pericyte loss and lead to blood-brain barrier dysfunction in AD. Methods: Bioinformatics analysis was conducted on the RNA-Seq expression counts matrix (GSE144474), which compared solo-cultured human blood-brain barrier endothelial cells against endothelial cells co-cultured with human brain pericytes in a non-contact model. We constructed a similar cell culture model to verify protein expression using western blots. Results: The insulin resistance and ferroptosis pathways were found to be enriched. Western blots of the insulin receptor and heme oxygenase expressions were consistent with those observed in RNA-Seq data. Additionally, we observed more than 5-fold upregulation of several genes associated with neuroprotection, including insulin-like growth factor 2 and brain-derived neurotrophic factor. Conclusions: Results suggest that pericyte influence on blood-brain barrier endothelial gene expression confers protection from insulin resistance, iron accumulation, oxidative stress, and amyloid deposition. Since these are conditions associated with AD pathophysiology, they imply mechanisms by which pericyte degeneration could contribute to disease progression.

Background: Age is the most common risk factor for Alzheimer’s disease (AD), a neurodegenerative disorder characterized by the hallmarks of toxic amyloid-β (Aβ) plaques and hyperphosphorylated tau tangles. Moreover, sub-physiological brain insulin levels have emerged as a pathological manifestation of AD. Objective: Identify age-related changes in the plasma disposition and blood-brain barrier (BBB) trafficking of Aβ peptides and insulin in mice. Methods: Upon systemic injection of 125I-Aβ40, 125I-Aβ42, or 125I-insulin, the plasma pharmacokinetics and brain influx were assessed in wild-type (WT) or AD transgenic (APP/PS1) mice at various ages. Additionally, publicly available single-cell RNA-Seq data [GSE129788] was employed to investigate pathways regulating BBB transport in WT mice at different ages. Results: The brain influx of 125I-Aβ40, estimated as the permeability-surface area product, decreased with age, accompanied by an increase in plasma AUC. In contrast, the brain influx of 125I-Aβ42 increased with age, accompanied by a decrease in plasma AUC. The age-dependent changes observed in WT mice were accelerated in APP/PS1 mice. As seen with 125I-Aβ40, the brain influx of 125I-insulin decreased with age in WT mice, accompanied by an increase in plasma AUC. This finding was further supported by dynamic single-photon emission computed tomography (SPECT/CT) imaging studies. RAGE and PI3K/AKT signaling pathways at the BBB, which are implicated in Aβ and insulin transcytosis, respectively, were upregulated with age in WT mice, indicating BBB insulin resistance. Conclusion: Aging differentially affects the plasma pharmacokinetics and brain influx of Aβ isoforms and insulin in a manner that could potentially augment AD risk.

Objective The objective is to investigate cellular mechanisms mediating the internalization of beta‐amyloid 40 (Aβ40) in part of the neurovascular unit, neurons, and blood‐brain barrier (BBB) endothelial cells. Methods Localization of fluorescein labeled Aβ40 (F‐Aβ40) and other endocytotic markers in wild type (WT) mouse brain slices, PC12 cells, rat primary hippocampal (RPH) neurons, and bovine brain microvascular endothelial (BBME) cells was investigated using laser confocal or fluorescence microscopy. The uptake of fluorophores in these cells was quantified by flow cytometry. Results Laser confocal micrographs of WT mouse brain slices treated with F‐Aβ40 demonstrated selective accumulation of the protein in cortical and hippocampal neurons via non‐saturable, energy independent, and non‐endocytotic pathways. A significant portion of internalized Aβ40 is located outside of the endosomal or lysosomal compartments, which may accumulate without degradation. In contrast, BBME cells exhibit energy dependent uptake of Aβ40, and accumulate the protein in acidic cell organelles, indicative of endocytotic uptake. Conclusion The difference in internalization of Aβ40 between neurons and BBB endothelial cells may provide clues to understanding how various cells can regulate Alzheimer's disease (AD) proteins and help explain the vulnerability of cortical and hippocampal neurons to AD toxicity.

Background Microbial conversion of dietary substrate to bioactive molecules represents a potential regulatory mechanism by which gut microbes can alter intestinal physiology. Tryptamine, a monoamine structurally similar to serotonin (5‐HT), can be generated via decarboxylation of tryptophan by the gut bacteria but its physiological effect is not known. Aim To determine the effect of bacterial metabolite tryptamine on colonic secretion. Methods We mounted proximal colon mucosa‐submucosa preparations from germ free (GF), humanized (HM; ex‐GF colonized with human gut bacteria for 4–5 weeks), conventionally raised WT and 5‐HT4 receptor knockout (5‐HT4R −/− ) male mice in the Ussing chamber. The change in short circuit current (ΔIsc) was determined in response to application of increasing concentrations of tryptamine (3μM – 3mM) on the submucosal (basolateral) side either alone or in the presence of 5‐HT3R or 5‐HT4R antagonist, ondansetron (100nM) and GR‐113808 (30nM) respectively, with and without TTX (500nM). Colonoids (3‐dimensional epithelial organoids derived from colon) from GF and HM were used for swelling assay and ELISA based cAMP measurement. Results Tryptamine application significantly increased Isc in both GF and HM mice (n=6, P < 0.05, two‐way ANOVA with Bonferroni post‐hoc). Tryptamine mediated ΔIsc response was not significantly affected by amiloride, an inhibitor of epithelial sodium channels and Na + /H + exchangers, which are primarily responsible for absorption but was significantly decreased by 5‐Nitro‐2‐(3‐phenylpropylamino) benzoic acid (NPPB; anion blocker), a calcium‐sensitive chloride channel and sodium bicarbonate exchanger inhibitor responsible for anionic secretion in both GF and HM mice (n=4, P < 0.05, two‐way ANOVA with Bonferroni post‐hoc). Tryptamine evoked ΔIsc was not inhibited by 5‐HT3R (n=4, P > 0.05) but completely blocked by 5‐HT4R antagonist and was absent in 5HT4R knock out mice suggesting that the effect of tryptamine is mediated via 5HT4R. The increase in Isc in response to tryptamine and its inhibition by 5‐HT4R antagonist was also seen in presence of TTX (neuronal blocker; n=4, one‐way ANOVA , P < 0.05) suggesting that the epithelial 5‐HT4R alone can mediate the effect of tryptamine. Application of tryptamine (1mM) for 30 minutes led to a significant increase in the surface area of fluorescein labelled colonoids (n=5, one‐way ANOVA , P < 0.05), similar to forskolin, when compared to vehicle treated and 5HT4R antagonist pre‐treated GF and HM colonoids. Colonoids incubated with tryptamine (1mM) for 60 minutes had significantly higher cAMP levels (normalized to forskolin) compared to vehicle treated GF and HM colonoids, confirming that tryptamine activates 5‐HT4 GPCR (n=3, student's t‐test, P < 0.05). Conclusion Tryptamine, a tryptophan‐derived microbial metabolite, increases colonic secretion via 5HT4 GPCR activation. The identification of a bacterial metabolite activating a host GPCR to alter GI function represents an important advance in the field, and provides a distinct example of small molecule mediated host‐microbe interaction. Engineered probiotics producing tryptamine could be used as therapeutics in diseases associated with constipation. Support or Funding Information NIH DK111850, DK114007 (PCK) This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .