BackgroundIncreased levels of the pathogenic amyloid β-peptide (Aβ), released from its precursor by the transmembrane protease γ-secretase, are found in Alzheimer disease (AD) brains. Interestingly, monoamine oxidase B (MAO-B) activity is also increased in AD brain, but its role in AD pathogenesis is not known. Recent neuroimaging studies have shown that the increased MAO-B expression in AD brain starts several years before the onset of the disease. Here, we show a potential connection between MAO-B, γ-secretase and Aβ in neurons.MethodsMAO-B immunohistochemistry was performed on postmortem human brain. Affinity purification of γ-secretase followed by mass spectrometry was used for unbiased identification of γ-secretase-associated proteins. The association of MAO-B with γ-secretase was studied by coimmunoprecipitation from brain homogenate, and by in-situ proximity ligation assay (PLA) in neurons as well as mouse and human brain sections. The effect of MAO-B on Aβ production and Notch processing in cell cultures was analyzed by siRNA silencing or overexpression experiments followed by ELISA, western blot or FRET analysis. Methodology for measuring relative intraneuronal MAO-B and Aβ42 levels in single cells was developed by combining immunocytochemistry and confocal microscopy with quantitative image analysis.ResultsImmunohistochemistry revealed MAO-B staining in neurons in the frontal cortex, hippocampus CA1 and entorhinal cortex in postmortem human brain. Interestingly, the neuronal staining intensity was higher in AD brain than in control brain in these regions. Mass spectrometric data from affinity purified γ-secretase suggested that MAO-B is a γ-secretase-associated protein, which was confirmed by immunoprecipitation and PLA, and a neuronal location of the interaction was shown. Strikingly, intraneuronal Aβ42 levels correlated with MAO-B levels, and siRNA silencing of MAO-B resulted in significantly reduced levels of intraneuronal Aβ42. Furthermore, overexpression of MAO-B enhanced Aβ production.ConclusionsThis study shows that MAO-B levels are increased not only in astrocytes but also in pyramidal neurons in AD brain. The study also suggests that MAO-B regulates Aβ production in neurons via γ-secretase and thereby provides a key to understanding the relationship between MAO-B and AD pathogenesis. Potentially, the γ-secretase/MAO-B association may be a target for reducing Aβ levels using protein–protein interaction breakers.Electronic supplementary materialThe online version of this article (doi:10.1186/s13195-017-0279-1) contains supplementary material, which is available to authorized users.
Insulin signalling deficiencies and insulin resistance have been directly linked to the progression of neurodegenerative disorders like Alzheimer's disease. However, to date little is known about the underlying molecular mechanisms or insulin state and distribution in the brain under pathological conditions. Here, we report that insulin is accumulated and retained as oligomers in hyperphosphorylated tau-bearing neurons in Alzheimer's disease and in several of the most prevalent human tauopathies. The intraneuronal accumulation of insulin is directly dependent on tau hyperphosphorylation, and follows the tauopathy progression. Furthermore, cells accumulating insulin show signs of insulin resistance and decreased insulin receptor levels. These results suggest that insulin retention in hyperphosphorylated tau-bearing neurons is a causative factor for the insulin resistance observed in tauopathies, and describe a novel neuropathological concept with important therapeutic implications.
Subcellular distribution of mitochondria in neurons is crucial for meeting the energetic demands, as well as the necessity to buffer Ca2+ within the axon, dendrites and synapses. Mitochondrial impairment is an important feature of Parkinson disease (PD), in which both familial parkinsonism genes DJ‐1 and PINK1 have a great impact on mitochondrial function. We used differentiated human dopaminergic neuroblastoma cell lines with stable PINK1 or DJ‐1 knockdown to study live motility of mitochondria in neurites. The frequency of anterograde and retrograde mitochondrial motility was decreased in PINK1 knockdown cells and the frequency of total mitochondrial motility events was reduced in both cell lines. However, neither the distribution nor the size of mitochondria in the neurites differed from the control cells even after downregulation of the mitochondrial fission protein, Drp1. Furthermore, mitochondria from PINK1 knockdown cells, in which motility was most impaired, had increased levels of GSK3βSer9 and higher release of mitochondrial Ca2+ when exposed to CCCP‐induced mitochondrial uncoupling. Further analysis of the ER‐mitochondria contacts involved in Ca2+ shuttling showed that PINK1 knockdown cells had reduced contacts between the two organelles. Our results give new insight on how PINK1 and DJ‐1 influence mitochondria, thus providing clues to novel PD therapies.
Background and Objectives:ATN (β-Amyloid, Tau, Neurodegeneration) system categorizes individuals based on their core Alzheimer’s disease (AD) biomarkers. An important potential future use for ATN is therapeutic decision-making in clinical practice once disease-modifying treatments, e.g., anti-amyloid, become widely available. In this cross-sectional study, we applied ATN and estimated potential eligibility for anti-amyloid treatment in a real-life memory clinic with biomarker assessments integrated into the routine diagnostic procedure and all specialized resources available for the implementation of novel treatments.Methods:We included all consecutive patients at the Karolinska University Hospital Memory clinic in Solna, Stockholm, Sweden, who had their first diagnostic visit in April 2018–February 2021, informed consent for the clinic research database, and available clinical and biomarker (CSF, imaging) data. ATN classification was based on CSF Aβ42 (or Aβ42/40; A), CSF phosphorylated tau (T), and medial temporal lobe atrophy (N). For CSF markers, we applied laboratory cut-offs and data-driven cut-offs for comparison (determined with Gaussian mixture modelling). Anti-amyloid treatment eligibility was assessed following the published recommendations for aducanumab (AD dementia or MCI with no evidence of non-AD etiology, appropriate level of cognition, AD-consistent CSF profile).Results:Study population consisted of 410 patients (52% subjective cognitive impairment, 23% mild cognitive impairment MCI, 25% any dementia; age 59±7 years, 56% women). Regardless of biomarker cut-offs, most patients were A−T−N− (54–57%). A+ prevalence was 17–30% (higher with data-driven cut-offs). Up to 13% of all patients (27% of those with MCI and 28% of those with dementia) were potentially eligible for anti-amyloid treatment when AD-consistent CSF was defined as any A+ profile. When A+T+ profile was required, treatment was targeted more to the dementia than MCI stage (eligibility up to 14% in MCI, 22% in dementia). The opposite applied to earlier stage intervention (A+T−N−; eligibility up to 12% in MCI, 2% in dementia).Discussion:In a memory clinic setting with all necessary infrastructure and national guidelines in place for dementia diagnostic examination (“best-case scenario”), most patients did not meet the eligibility criteria for anti-amyloid treatment. Continuing the development of disease-modifying treatments with different mechanisms of action is a priority.
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