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Abstract Introduction Increasing evidence implicates proteostatic dysfunction as an early event in the development of frontotemporal dementia (FTD). This study aimed to explore potential cerebrospinal fluid (CSF) biomarkers associated with the proteolytic systems in genetic FTD caused by CHMP2B mutation. Methods Combining solid‐phase extraction and parallel reaction monitoring mass spectrometry, a panel of 47 peptides derived from 20 proteins was analyzed in CSF from 31 members of the Danish CHMP2B‐FTD family. Results Compared with family controls, mutation carriers had significantly higher levels of complement C9, lysozyme and transcobalamin II, and lower levels of ubiquitin, cathepsin B, and amyloid precursor protein. Discussion Lower CSF ubiquitin concentrations in CHMP2B mutation carriers indicate that ubiquitin levels relate to the specific disease pathology, rather than all‐cause neurodegeneration. Increased lysozyme and complement proteins may indicate innate immune activation. Altered levels of amyloid precursor protein and cathepsins have previously been associated with impaired lysosomal proteolysis in FTD. Highlights CSF markers of proteostasis were explored in CHMP2B‐mediated frontotemporal dementia (FTD). 31 members of the Danish CHMP2B‐FTD family were included. We used solid‐phase extraction and parallel reaction monitoring mass spectrometry. Six protein levels were significantly altered in CHMP2B‐FTD compared with controls. Lower CSF ubiquitin levels in patients suggest association with disease mechanisms.

Most of the neurodegenerative diseases that afflict humans manifest with the intraneuronal accumulation of toxic proteins that are aggregate-prone. Extensive data in cell and neuronal models support the concept that such proteins, like mutant huntingtin or alpha-synuclein, are substrates for macroautophagy (hereafter autophagy). Furthermore, autophagy-inducing compounds lower the levels of such proteins and ameliorate their toxicity in diverse animal models of neurodegenerative diseases. However, most of these compounds also have autophagy-independent effects and it is important to understand if similar benefits are seen with genetic strategies that upregulate autophagy, as this strengthens the validity of this strategy in such diseases. Here we review studies in vertebrate models using genetic manipulations of core autophagy genes and describe how these improve pathology and neurodegeneration, supporting the validity of autophagy upregulation as a target for certain neurodegenerative diseases.

Abstract Background Dietary inequalities between population groups are common with older and more affluent individuals tending to have healthier diets. Differential responses to health interventions may exacerbate inequalities. Changing what foods are displayed at supermarket checkouts is one intervention that has the potential to change diets. The aim of this study was to assess whether differences in purchases of common checkout foods from supermarkets with different checkout food policies varied according to age group and social grade. Methods We analysed annual household purchase data for 2013–17 from nine leading UK supermarkets, split according to age of the main household shopper and household social grade. Checkout food policies were categorised as clear and consistent, vague or inconsistent, and none. Policies were heterogeneous but all included removal of confectionery and/or chocolate from checkouts. Mixed effects linear regression models were used to assess differences in purchases of common checkout foods (sugary confectionery, chocolate and potato crisps) by checkout food policy and whether these varied by age group or occupational social grade. Results Relative to supermarkets with no checkout food policy, 14% (95% CI: 4–22%) fewer purchases of common checkout foods per household per percentage market share were made in supermarkets with a clear and consistent policy. Adjusted mean numbers of purchases were higher in older age groups than the youngest, but there were no differences between the highest and other social grades. There were significant interactions between checkout food policy and both age group and social grade. In supermarkets with clear and consistent policies, 23% (6–36%), 20% (2–34%), and 23% (7–37%) fewer purchases were made in age groups 45–54, 55–64 and 65+ years respectively, compared to all groups combined. In supermarkets with clear and consistent policies, there were 21% (4–35%), 26% (9–39%) and 21% (3–35%) fewer purchases made by households in the highest two and lowest social grades respectively, compared to all groups combined. Conclusions Households with older main shoppers and those in the most and least affluent social grades may be most responsive to supermarket checkout food policies. As older and more affluent groups tend to have healthier diets overall, it is unlikely that supermarket checkout food policies contribute to greater dietary equity.

Appropriate regulation of autophagy is crucial for clearing toxic proteins from cells. Defective autophagy results in accumulation of toxic protein aggregates that detrimentally affect cellular function and organismal survival. Here, we report that the microRNA miR-1 regulates the autophagy pathway through conserved targeting of the orthologous Tre-2/Bub2/CDC16 (TBC) Rab GTPase-activating proteins TBC-7 and TBC1D15 in Caenorhabditis elegans and mammalian cells, respectively. Loss of miR-1 causes TBC-7/TBC1D15 overexpression, leading to a block on autophagy. Further, we found that the cytokine interferon-β (IFN-β) can induce miR-1 expression in mammalian cells, reducing TBC1D15 levels, and safeguarding against proteotoxic challenges. Therefore, this work provides a potential therapeutic strategy for protein aggregation disorders.

Abstract Major Histocompability Complex I (MHC-I) molecules present cellularly derived peptides to the adaptive immune system. Generally MHC-I is not expressed on healthy post-mitotic neurons in the central nervous system, but it is known to increase upon immune activation such as viral infections and also during neurodegenerative processes. MHC-I expression is known to be regulated by the DNA methyltransferase DNMT1 in non-neuronal cells. Interestingly DNMT1 expression is high in neurons despite these being non-dividing. This suggests a role for DNMT1 in neurons beyond the classical re-methylation of DNA after cell division. We thus investigated whether DNMT1 regulates MHC-I in post-mitotic neurons. For this we used primary cultures of mouse cerebellar granule neurons (CGNs). Our results showed that knockdown of DNMT1 in CGNs caused upregulation of some, but not all subtypes of MHC-I genes. This effect was synergistically enhanced by subsequent IFNγ treatment. Overall MHC-I protein level was not affected by knockdown of DNMT1 in CGNs. Instead our results show that the relative MHC-I expression levels among the different MHC subtypes is regulated by DNMT1 activity. In conclusion, we show that while the mouse H2-D1/L alleles are suppressed in neurons by DNMT1 activity under normal circumstances, the H2-K1 allele is not. This finding is particularly important in two instances. One: in the context of CNS autoimmunity with epitope presentation by specific MHC-I subtypes where this allele specific regulation might become important; and two: in amyotropic lateral sclerosis (ALS) where H2-K but not H2-D protects motor neurons from ALS astrocyte-induced toxicity in a mouse model of ALS.

Abstract Background Food choices are often determined by stimuli from our immediate surroundings, including strategic placement in shops to encourage impulse purchases. One example of this is food in shop checkout areas. Recently a number of UK supermarkets have voluntarily committed to providing healthier checkout foods. The aim of this study was to document the nature of current UK supermarket checkout food policies; determine whether there are any differences in the healthfulness and type of food displayed at checkouts in supermarkets according to the presence or nature of policies; and determine whether supermarkets are adhering to their checkout food policies. Methods Survey of checkout food policies. Cross-sectional observations in 69 supermarkets (covering 14 store formats) in the East of England in Feb-May 2017 of the number and type of checkout foods on each ‘checkout journey’ (each possible route through the checkout area). Checkout foods were categorised as less healthy or healthier, using the UK Food Standard’s Agency’s Nutrient Profile Model, and into food groups. Checkout food policies were categorised as clear and consistent, vague or inconsistent, or absent. Results Checkout food policies differed between store formats in some supermarket groups. Across the 14 store formats included, two had no checkout food policy, six had ‘clear and consistent’ policies, and six ‘vague or inconsistent’ policies. In supermarkets with clear and consistent policies there were a median of 13 products per checkout journey, of which 35% were less healthy. Comparable figures for supermarkets with vague or inconsistent, and absent policies were 15 (57%) and 39 (90%) respectively (ps for trend

Abstract Background Further collection of surveillance data is warranted, particularly in preschool populations, for optimizing future public health promotion strategies. This study aims to describe physical activity (PA) and sedentary behavior (SB) across different settings, including time in and out of daycare, and to determine the proportion of children complying with suggested PA recommendations in a high income country. Methods Valid PA was assessed in 231 children (36.4 ± 1.1 months) with the Actigraph GT3X accelerometer, and information regarding date and time of dropping-off/picking-up children in daycare was provided by parents. Mean total PA (i.e., counts per minute (CPM)), moderate-to-vigorous physical activity (MVPA), SB time, and non-SB time was generated and compared across settings. Post hoc, PA and SB were examined in subgroups of low-active (1st quartile) and high-active (4th quartile) children. Results Overall, boys and girls spent 1.4 ± 0.3 h/day and 1.2 ± 0.4 h/day in MVPA, respectively. Likewise, boys and girls accumulated 6.7 ± 0.8 h and 6.8 ± 0.9 h of SB time per day, respectively. Higher PA levels consistently co-occurred with lower SB time in the daycare setting. Girls accumulated less SB time in daycare than before and after daycare (β = −12.2%, p

Neurons can convert pathogenic T cells to anti-inflammatory FoxA1+ regulatory T cells (Tregs), which can ameliorate EAE, but the molecular mechanism is only partially understood. Liu et al. show that autocrine interferon β signalling induces PDL1 expression in neurons, which is essential for neurons to reprogramme pathogenic T cells to FoxA1+ Tregs.

BackgroundIn response to public concerns and campaigns, some United Kingdom supermarkets have implemented policies to reduce less-healthy food at checkouts. We explored the effects of these policies on purchases of less-healthy foods commonly displayed at checkouts.Methods and findingsWe used a natural experimental design and two data sources providing complementary and unique information. We analysed data on purchases of small packages of common, less-healthy, checkout foods (sugary confectionary, chocolate, and potato crisps) from 2013 to 2017 from nine UK supermarkets (Aldi, Asda, Co-op, Lidl, M&S, Morrisons, Sainsbury's, Tesco, and Waitrose). Six supermarkets implemented a checkout food policy between 2013 and 2017 and were considered intervention stores; the remainder were comparators. Firstly, we studied the longitudinal association between implementation of checkout policies and purchases taken home. We used data from a large (n ≈ 30,000) household purchase panel of food brought home to conduct controlled interrupted time series analyses of purchases of less-healthy common checkout foods from 12 months before to 12 months after implementation. We conducted separate analyses for each intervention supermarket, using others as comparators. We synthesised results across supermarkets using random effects meta-analyses. Implementation of a checkout food policy was associated with an immediate reduction in four-weekly purchases of common checkout foods of 157,000 (72,700-242,800) packages per percentage market share-equivalent to a 17.3% reduction. This decrease was sustained at 1 year with 185,100 (121,700-248,500) fewer packages purchased per 4 weeks per percentage market share-equivalent to a 15.5% reduction. The immediate, but not sustained, effect was robust to sensitivity analysis. Secondly, we studied the cross-sectional association between checkout food policies and purchases eaten without being taken home. We used data from a smaller (n ≈ 7,500) individual purchase panel of food bought and eaten 'on the go'. We conducted cross-sectional analyses comparing purchases of common checkout foods in 2016-2017 from supermarkets with and without checkout food policies. There were 76.4% (95% confidence interval 48.6%-89.1%) fewer annual purchases of less-healthy common checkout foods from supermarkets with versus without checkout food policies. The main limitations of the study are that we do not know where in the store purchases were selected and cannot determine the effect of changes in purchases on consumption. Other interventions may also have been responsible for the results seen.ConclusionsThere is a potential impact of checkout food polices on purchases. Voluntary supermarket-led activities may have public health benefits.

Little is known about the development of dietary patterns during toddlerhood and the relation to growth and health. The study objective was to characterise the development of dietary patterns from 9-36 mo of age and investigate the association to body size, body composition and metabolic risk markers at 36 mo. Food records were filled out at 9, 18 and 36 mo of age (n = 229). Dietary patterns were identified by principal component analysis (PCA). Three dietary patterns were identified: Transition Food, Healthy Food and Traditional Food. The course of development in dietary patterns from 9-36 mo indicated tracking for a relatively large group of participants in the three patterns. Transition Food and Healthy Food were associated with some of the investigated outcomes. Children with lower adherence to the Transition Food pattern than average at 18 and 36 mo irrespectively of intake at 9 mo had higher BMI z-scores at 36 mo. Similar trend was identified for higher fat mass indices. Children with lower adherence to the Healthy Food pattern than average at all three ages compared to children with higher adherence to the Healthy Food pattern at the first two registrations, 9 and 18 mo had higher total cholesterol and LDL. Hence, this could represent undesirable development of dietary patterns in toddlers. In conclusion, development of dietary patterns can be exploratory characterised by PCA and related to potential cardiovascular risk markers in toddlers even within a relatively homogeneous population with a high socioeconomic status. The tracking of dietary patterns from 9 mo of age indicates a need for early and sustained promotion of healthy diets.

Nature Communications 8: Article number: 14709 (2017); Published 24 April 2017; Updated 22 May 2017 The affiliation details for Marco Prinz are incorrect in this Article. The correct affiliation details for this author are given below: Institute of Neuropathology, Faculty of Medicine, University of Freiburg, Freiburg 79106, Germany.

Objective High infancy levels of insulin-like growth factor-I (IGF-I) have been associated with increased linear growth and fat-free mass (FFM) but also with risk of obesity. This paper examines how IGF-I at 9 and 36 months relates to diet and body composition. Design Healthy term infants from the prospective cohort study, SKOT, were examined at 9 and 36 months with anthropometry, bioelectrical impedance (36 months), 7-day food records and blood analysis of IGF-I and IGFBP-3 by chemiluminescent immunometric assay. Results IGF-I at 36 months (n = 229) was positively correlated with 9 months values and values were considerably higher in girls (43%). Children breastfed at 9 months had lower IGF-I concentrations at 9 months but reached the same IGF-I concentrations at 36 months as infants not breastfed at 9 months. IGF-I at 36 months was positively associated with height, weight, BMI, predicted FFM and FFM index (FFM/height (kg/m 2 )). Although there also was a positive association with predicted fat mass (FM) there was no association with FM index (FM/height (kg/m 2 )). Further, a negative association with skin fold thickness was observed. A change in IGF-I from 9–36 months was positively related to FFM and FFM index but not BMI, FM and FM index. No associations were seen between IGF-I and current intake of milk, meat or protein energy percentage, but both fat and saturated fat energy percentage were negatively associated with IGF-I. Conclusion IGF-I concentrations were positively associated with growth but not with adiposity at this age. However, the higher tempo of growth may influence age at adiposity rebound and thereby later risk of obesity. Milk and protein intake at 36 months did not influence IGF-I but there was a negative association with intake of fat and saturated fat. The implications of this finding for development of obesity need further exploration. [ABSTRACT FROM AUTHOR]

Introduction: Increasing evidence implicates proteostatic dysfunction as an early event in the development of frontotemporal dementia (FTD). This study aimed to explore potential cerebrospinal fluid (CSF) biomarkers associated with the proteolytic systems in genetic FTD caused by CHMP2B mutation., Methods: Combining solid-phase extraction and parallel reaction monitoring mass spectrometry, a panel of 47 peptides derived from 20 proteins was analyzed in CSF from 31 members of the Danish CHMP2B -FTD family., Results: Compared with family controls, mutation carriers had significantly higher levels of complement C9, lysozyme and transcobalamin II, and lower levels of ubiquitin, cathepsin B, and amyloid precursor protein., Discussion: Lower CSF ubiquitin concentrations in CHMP2B mutation carriers indicate that ubiquitin levels relate to the specific disease pathology, rather than all-cause neurodegeneration. Increased lysozyme and complement proteins may indicate innate immune activation. Altered levels of amyloid precursor protein and cathepsins have previously been associated with impaired lysosomal proteolysis in FTD., Highlights: CSF markers of proteostasis were explored in CHMP2B -mediated frontotemporal dementia (FTD).31 members of the Danish CHMP2B -FTD family were included.We used solid-phase extraction and parallel reaction monitoring mass spectrometry.Six protein levels were significantly altered in CHMP2B -FTD compared with controls.Lower CSF ubiquitin levels in patients suggest association with disease mechanisms., Competing Interests: K.B. has served as a consultant, at advisory boards, or at data monitoring committees for Abcam, Axon, BioArctic, Biogen, and JOMDD/Shimadzu. Julius Clinical, Lilly, MagQu, Novartis, Ono Pharma, Pharmatrophix, Prothena, Roche Diagnostics, and Siemens Healthineers, and is a co‐founder of Brain Biomarker Solutions in Gothenburg AB (BBS), which is a part of the GU Ventures Incubator Program, outside the work presented in this paper. H.Z. has served at scientific advisory boards and/or as a consultant for Abbvie, Alector, ALZPath, Annexon, Apellis, Artery Therapeutics, AZTherapies, CogRx, Denali, Eisai, Nervgen, Novo Nordisk, Passage Bio, Pinteon Therapeutics, Red Abbey Labs, reMYND, Roche, Samumed, Siemens Healthineers, Triplet Therapeutics, and Wave, has given lectures in symposia sponsored by Cellectricon, Fujirebio, Alzecure, Biogen, and Roche, and is a co‐founder of Brain Biomarker Solutions in Gothenburg AB (BBS), which is a part of the GU Ventures Incubator Program (outside submitted work). The remaining authors (A.T., S.S., A.H.S., P.E., P.R., C.S.M., E.E.H., T.T.N., A.B., and J.E.N.) have no conflicts of interest to declare. Author disclosures are available in the supporting information., (© 2023 The Authors. Alzheimer's & Dementia: Diagnosis, Assessment & Disease Monitoring published by Wiley Periodicals, LLC on behalf of Alzheimer's Association.)

Objective: Parkinson's disease (PD) manifests in motor dysfunction, non-motor symptoms, and eventual dementia (PDD). Neuropathological hallmarks include nigrostriatal neurodegeneration, Lewy body (LB) pathology, and neuroinflammation. Alpha-synuclein (α-syn), a primary component of LBs, is implicated in PD pathogenesis, accumulating, and aggregating in both familial and sporadic PD. However, as α-syn pathology is often comorbid with amyloid-beta (Aβ) plaques and phosphorylated tau (pTau) tangles in PDD, it is still unclear whether α-syn is the primary cause of neurodegeneration in sporadic PDD. We aimed to determine how the absence of α-syn would affect PDD manifestation., Methods: IFN-β knockout (Ifnb -/- ) mice spontaneously develop progressive behavior abnormalities and neuropathology resembling PDD, notably with α-syn + LBs. We generated Ifnb/Snca double knockout (DKO) mice and evaluated their behavior and neuropathology compared with wild-type (Wt), Ifnb -/- , and Snca -/- mice using immunohistochemistry, electron microscopy, immunoblots, qPCR, and modification of neuronal signaling., Results: Ifnb/Snca DKO mice developed all clinical PDD-like behavioral manifestations induced by IFN-β loss. Independently of α-syn expression, lack of IFN-β alone induced Aβ plaques, pTau tangles, and LB-like Aβ + /pTau + inclusion bodies and neuroinflammation. IFN-β loss caused significant elevated glial and neuronal TNF-α and neuronal TNFR1, associated with neurodegeneration. Restoring neuronal IFN-β signaling or blocking TNFR1 rescued caspase 3/t-BID-mediated neuronal-death through upregulation of c-FLIP S and lowered intraneuronal Aβ and pTau accumulation., Interpretation: These findings increase our understanding of PD pathology and suggest that targeting α-syn alone is not sufficient to mitigate disease. Targeting specific aspects of neuroinflammation, such as aberrant neuronal TNF-α/TNFR1 or IFN-β/IFNAR signaling, may attenuate disease. ANN NEUROL 2021;90:789-807., (© 2021 American Neurological Association.)

Appropriate regulation of autophagy is crucial for clearing toxic proteins from cells. Defective autophagy results in accumulation of toxic protein aggregates that detrimentally affect cellular function and organismal survival. Here, we report that the microRNA miR-1 regulates the autophagy pathway through conserved targeting of the orthologous T re-2/ B ub2/ C DC16 (TBC) Rab GTPase-activating proteins TBC-7 and TBC1D15 in Caenorhabditis elegans and mammalian cells, respectively. Loss of miR-1 causes TBC-7/TBC1D15 overexpression, leading to a block on autophagy. Further, we found that the cytokine interferon-β (IFN-β) can induce miR-1 expression in mammalian cells, reducing TBC1D15 levels, and safeguarding against proteotoxic challenges. Therefore, this work provides a potential therapeutic strategy for protein aggregation disorders., Competing Interests: CN, PE, SG, AH, LN, PM, HL, SI, DR, RP No competing interests declared, (© 2019, Nehammer et al.)

Most of the neurodegenerative diseases that afflict humans manifest with the intraneuronal accumulation of toxic proteins that are aggregate-prone. Extensive data in cell and neuronal models support the concept that such proteins, like mutant huntingtin or alpha-synuclein, are substrates for macroautophagy (hereafter autophagy). Furthermore, autophagy-inducing compounds lower the levels of such proteins and ameliorate their toxicity in diverse animal models of neurodegenerative diseases. However, most of these compounds also have autophagy-independent effects and it is important to understand if similar benefits are seen with genetic strategies that upregulate autophagy, as this strengthens the validity of this strategy in such diseases. Here we review studies in vertebrate models using genetic manipulations of core autophagy genes and describe how these improve pathology and neurodegeneration, supporting the validity of autophagy upregulation as a target for certain neurodegenerative diseases., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Neurons reprogramme encephalitogenic T cells (T enc ) to regulatory T cells (T regs ), either FoxP3 + T regs or FoxA1 + T regs . We reported previously that neuronal ability to generate FoxA1 + T regs was central to preventing neuroinflammation in experimental autoimmune encephalomyelitis (EAE). Mice lacking interferon (IFN)-β were defective in generating FoxA1 + T regs in the brain. Here we show that lack of neuronal IFNβ signalling is associated with the absence of programme death ligand-1 (PDL1), which prevents their ability to reprogramme T enc cells to FoxA1 + T regs . Passive transfer-EAE via IFNβ-competent T enc cells to mice lacking IFNβ and active induced-EAE in mice lacking its receptor, IFNAR, in the brain (Nes Cre :Ifnar fl/fl ) result in defective FoxA1 + T regs generation and aggravated neuroinflammation. IFNβ activates neuronal PI3K/Akt signalling and Akt binds to transcription factor FoxA1 that translocates to the nucleus and induces PDL1. Conversely, inhibition of PI3K/Akt, FoxA1 and PDL1 blocked neuronal ability to generate FoxA1 + T regs . We characterize molecular factors central for neuronal ability to reprogramme pathogenic T cells to FoxA1 + T regs preventing neuroinflammation.

Background: Secretion of proteopathic α-synuclein (α-SNC) species from neurons is a suspected driving force in the propagation of Parkinson's disease (PD). We have previously implicated exophagy, the exocytosis of autophagosomes, as a dominant mechanism of α-SNC secretion in differentiated PC12 or SH-SY5Y nerve cells. Here we have examined the regulation of exophagy associated with different forms of nerve cell stress relevant to PD., Results: We identify cJUN-N-terminal kinase (JNK) activity as pivotal in the secretory fate of autophagosomes containing α-SNC. Pharmacological inhibition or genetic (shRNA) knockdown of JNK2 or JNK3 decreases α-SNC secretion in differentiated PC12 and SH-SY5Y cells, respectively. Conversely, expression of constitutively active mitogen-activated protein kinase kinase 7 (MKK7)-JNK2 and -JNK3 constructs augment secretion. The transcriptional activity of cJUN was not required for the observed effects. We establish a causal relationship between increased α-SNC release by exophagy and JNK activation subsequent to lysosomal fusion deficiency (overexpression of Lewy body-localized protein p25α or bafilomycin A1). JNK activation following neuronal ER or oxidative stress was not correlated with exophagy, but of note, we demonstrate that reciprocal signaling between microglia and neurons modulates α-SNC secretion. NADPH oxidase activity of microglia cell lines was upregulated by direct co-culture with α-SNC-expressing PC12 neurons or by passive transfer of nerve cell-conditioned medium. Conversely, inflammatory factors secreted from activated microglia increased JNK activation and α-SNC secretion several-fold in PC12 cells. While we do not identify these factors, we extend our observations by showing that exposure of neurons in monoculture to TNFα, a classical pro-inflammatory mediator of activated microglia, is sufficient to increase α-SNC secretion in a mechanism dependent on JNK2 or JNK3. In continuation hereof, we show that also IFNβ and TGFβ increase the release of α-SNC from PC12 neurons., Conclusions: We implicate stress kinases of the JNK family in the regulation of exophagy and release of α-SNC following endogenous or exogenous stimulation. In a wider scope, our results imply that microglia not only inflict bystander damage to neurons in late phases of inflammatory brain disease but may also be active mediators of disease propagation.

Neurodegenerative diseases have been linked to inflammation, but whether altered immunomodulation plays a causative role in neurodegeneration is not clear. We show that lack of cytokine interferon-β (IFN-β) signaling causes spontaneous neurodegeneration in the absence of neurodegenerative disease-causing mutant proteins. Mice lacking Ifnb function exhibited motor and cognitive learning impairments with accompanying α-synuclein-containing Lewy bodies in the brain, as well as a reduction in dopaminergic neurons and defective dopamine signaling in the nigrostriatal region. Lack of IFN-β signaling caused defects in neuronal autophagy prior to α-synucleinopathy, which was associated with accumulation of senescent mitochondria. Recombinant IFN-β promoted neurite growth and branching, autophagy flux, and α-synuclein degradation in neurons. In addition, lentiviral IFN-β overexpression prevented dopaminergic neuron loss in a familial Parkinson's disease model. These results indicate a protective role for IFN-β in neuronal homeostasis and validate Ifnb mutant mice as a model for sporadic Lewy body and Parkinson's disease dementia., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Aggregation of α-synuclein can be promoted by the tubulin polymerization-promoting protein/p25α, which we have used here as a tool to study the role of autophagy in the clearance of α-synuclein. In NGF-differentiated PC12 catecholaminergic nerve cells, we show that de novo expressed p25α co-localizes with α-synuclein and causes its aggregation and distribution into autophagosomes. However, p25α also lowered the mobility of autophagosomes and hindered the final maturation of autophagosomes by preventing their fusion with lysosomes for the final degradation of α-synuclein. Instead, p25α caused a 4-fold increase in the basal level of α-synuclein secreted into the medium. Secretion was strictly dependent on autophagy and could be up-regulated (trehalose and Rab1A) or down-regulated (3-methyladenine and ATG5 shRNA) by enhancers or inhibitors of autophagy or by modulating minus-end-directed (HDAC6 shRNA) or plus-end-directed (Rab8) trafficking of autophagosomes along microtubules. Finally, we show in the absence of tubulin polymerization-promoting protein/p25α that α-synuclein release was modulated by dominant mutants of Rab27A, known to regulate exocytosis of late endosomal (and amphisomal) elements, and that both lysosomal fusion block and secretion of α-synuclein could be replicated by knockdown of the p25α target, HDAC6, the predominant cytosolic deacetylase in neurons. Our data indicate that unconventional secretion of α-synuclein can be mediated through exophagy and that factors, which increase the pool of autophagosomes/amphisomes (e.g. lysosomal disturbance) or alter the polarity of vesicular transport of autophagosomes on microtubules, can result in an increased release of α-synuclein monomer and aggregates to the surroundings.

Here, we report that activation of different types of tissue macrophages, including microglia, by lipopolysaccharide (LPS) or GM-CSF stimulation correlates with the quantitative redistribution of NADPH oxidase (cyt b(558)) from the plasma membrane to an intracellular stimulus-responsive storage compartment. Cryo-immunogold labeling of gp91(phox) and CeCl(3) cytochemistry showed the presence of gp91(phox) and oxidant production in numerous small (<100 nm) vesicles. Cell homogenization and sucrose gradient centrifugation in combination with transferrin-HRP/DAB ablation showed that more than half of cyt b(558) is present in fractions devoid of endosomal markers, which is supported by morphological evidence to show that the cyt b(558)-containing compartment is distinct from endosomes or biosynthetic organelles. Streptolysin-O-mediated guanosine 5'-3-O-(thio)triphosphate loading of Ra2 microglia caused exocytosis of a major complement of cyt b(558) under conditions where lysosomes or endosomes were not mobilized. We establish phagocytic particles and soluble mediators ATP, TNFα, and CD40L as physiological inducers of cyt b(558) exocytosis to the cell surface, and by shRNA knockdown, we identify Rab27A/B as positive or negative regulators of vesicular mobilization to the phagosome or the cell surface, respectively. Exocytosis was followed by clathrin-dependent internalization of cyt b(558), which could be blocked by a dominant negative mutant of the clathrin-coated pit-associated protein Eps15. Re-internalized cyt b(558) did not reach lysosomes but associated with recycling endosomes and undefined vesicular elements. In conclusion, cyt b(558) depends on clathrin for internalization, and in activated macrophages NADPH oxidase occupies a Rab27A/B-regulated secretory compartment, which allows rapid agonist-induced redistribution of superoxide production in the cell.