Native PLGA nanoparticles attenuate Aß-seed induced tau aggregation under in vitro conditions: potential implication in Alzheimer's disease pathology.

Evidence suggests that beta-amyloid (Aß)-induced phosphorylation/aggregation of tau protein plays a critical role in the degeneration of neurons and development of Alzheimer's disease (AD), the most common cause of dementia affecting the elderly population. Many studies have pursued a variety of small molecules, including nanoparticles conjugated with drugs to interfere with Aß and/or tau aggregation/toxicity as an effective strategy for AD treatment. We reported earlier that FDA approved PLGA nanoparticles without any drug can attenuate Aß aggregation/toxicity in cellular/animal models of AD. In this study, we evaluated the effects of native PLGA on Aß seed-induced aggregation of tau protein using a variety of biophysical, structural and spectroscopic approaches. Our results show that Aß1-42 seeds enhanced aggregation of tau protein in the presence and absence of heparin and the effect was attenuated by native PLGA nanoparticles. Interestingly, PLGA inhibited aggregation of both 4R and 3R tau isoforms involved in the formation of neurofibrillary tangles in AD brains. Furthermore, Aß seed-induced tau aggregation in the presence of arachidonic acid was suppressed by native PLGA. Collectively, our results suggest that native PLGA nanoparticles can inhibit the Aß seed-induced aggregation of different tau protein isoforms highlighting their therapeutic implication in the treatment of AD.

Detection of ß-amyloid aggregates/plaques in 5xFAD mice by labelled native PLGA nanoparticles: implication in the diagnosis of alzheimer's disease.

Evidence suggests that increased level/aggregation of ß-amyloid (Aß) peptide, together with enhanced phosphorylation/aggregation of tau protein, play a critical role in the development of Alzheimer's disease (AD), the leading cause of dementia in the elderly. At present, AD diagnosis is based primarily on cognitive assessment, neuroimaging, and immunological assays to detect altered levels/deposition of Aß peptides and tau protein. While measurement of Aß and tau in the cerebrospinal fluid/blood can indicate disease status, neuroimaging of aggregated Aß and tau protein in the brain using positron emission tomography (PET) enable to monitor the pathological changes in AD patients. With advancements in nanomedicine, several nanoparticles, apart from drug-delivery, have been used as a diagnostic agent to identify more accurately changes in AD patients. Recently, we reported that FDA approved native PLGA nanoparticles can interact with Aß to inhibit its aggregation/toxicity in cellular and animal models of AD. Here, we reveal that fluorescence labelled native PLGA following acute intracerebellar injection can identify majority of the immunostained Aß as well as Congo red labelled neuritic plaques in the cortex of 5xFAD mice. Labelling of plaques by PLGA is apparent at 1 h, peak around 3 h and then start declining by 24 h after injection. No fluorescent PLGA was detected in the cerebellum of 5xFAD mice or in any brain regions of wild-type control mice following injection. These results provide the very first evidence that native PLGA nanoparticles can be used as a novel nano-theragnostic agent in the treatment as well as diagnosis of AD pathology.

Administration of Kainic Acid Differentially Alters Astrocyte Markers and Transiently Enhanced Phospho-tau Level in Adult Rat Hippocampus.

Kainic acid (KA), an analogue of the excitatory neurotransmitter glutamate, when administered systemically can trigger seizures and neuronal loss in a manner that mirrors the neuropathology of human mesial temporal lobe epilepsy (mTLE), which affects ∼50 million people globally. Evidence suggests that changes in astrocytes which precede neuronal damage play an important role in the degeneration of neurons and/or development of seizures in TLE pathogenesis. Additionally, a role for microtubule associated tau protein, involved in various neurodegenerative diseases including Alzheimer's disease, has also been suggested in the development of seizure and/or neurodegeneration in TLE pathogenesis. At present, possible alterations of different subtypes of astrocytes and their association, if any, with tau protein in TLE remain unclear. In this study, we evaluated alterations of different subtypes of astrocytes and phospho-/cleaved-tau levels in KA-treated rat model of TLE. Our results reveal that levels/expression of various astrocyte markers such as GFAP, vimentin, S100B, Aldh1L1, but not GS, are increased in the hippocampus of KA-treated rats. The levels/expression of both A1(C3+) and A2(S100A10+)-like astrocytes are also increased in KA-treated rats. Concurrently, the total (Tau1 and Tau5) and phospho-tau (AT270 and PHF1) levels are transiently enhanced following KA administration. Furthermore, the level/expression of cleaved-tau, which is apparent in a subset of GFAP-, S100B- and A2-positive astrocytes, are increased in KA-treated rats. These results, taken together, suggest a differential role for various astrocytic subpopulations and tau protein in the development of seizure and/or loss of neurons in KA model of TLE and possibly in human mTLE pathogenesis.

Mental health disorders in English newspapers of India: A retrospective study.

BACKGROUND: In recent years there has been significant coverage of mental health in Indian newspapers; the media can play a significant role in perpetuating as well as reducing stigma towards people with mental illness. This paper analyses the content, context and type of newspaper coverage of various mental health disorders in English language newspapers in India between 2016 and 2021. METHODS: A detailed analysis was performed on a sample of articles about mental illness in a range of English language Indian newspapers. RESULTS: Depression was the most prevalent topic amongst the articles followed by anxiety, schizophrenia and bipolar disorder. Our study describes a wide range of use of mental health disorders in various newspapers. All diagnoses were generally described in a criminal context like homicide, sexual assault and other crimes. Over time newspaper coverage of mental illness has become less stigmatising. Further exploration of non-English medium newspapers is required to fully understand the extent of the role of print media in perpetuating unhelpful stereotypes of people with mental illness in India.

Significance of native PLGA nanoparticles in the treatment of Alzheimer's disease pathology.

Alzheimer's disease (AD) is believed to be triggered by increased levels/aggregation of ß-amyloid (Aß) peptides. At present, there is no effective disease-modifying treatment for AD. Here, we evaluated the therapeutic potential of FDA-approved native poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles on Aß aggregation and in cellular/animal models of AD. Our results showed that native PLGA can not only suppress the spontaneous aggregation but can also trigger disassembly of preformed Aß aggregates. Spectroscopic studies, molecular dynamics simulations and biochemical analyses revealed that PLGA, by interacting with the hydrophobic domain of Aß1-42, prevents a conformational shift towards the ß-sheet structure, thus precluding the formation and/or triggering disassembly of Aß aggregates. PLGA-treated Aß samples can enhance neuronal viability by reducing phosphorylation of tau protein and its associated signaling mechanisms. Administration of PLGA can interact with Aß aggregates and attenuate memory deficits as well as Aß levels/deposits in the 5xFAD mouse model of AD. PLGA can also protect iPSC-derived neurons from AD patients against Aß toxicity by decreasing tau phosphorylation. These findings provide unambiguous evidence that native PLGA, by targeting different facets of the Aß axis, can have beneficial effects in mouse neurons/animal models as well as on iPSC-derived AD neurons - thus signifying its unique therapeutic potential in the treatment of AD pathology.

Native PLGA nanoparticles regulate APP metabolism and protect neurons against ß-amyloid toxicity: Potential significance in Alzheimer's disease pathology.

Biodegradable poly(lactic-co-glycolic acid)(PLGA) nanoparticles have been used extensively in delivering drugs to target tissues due to their excellent biocompatibility. Evidence suggests that PLGA-conjugated drugs/agents can attenuate pathology in cellular/animal models of Alzheimer's disease (AD), which is initiated by increased level/aggregation of amyloid ß (Aß) peptide generated from amyloid precursor protein (APP). The beneficial effects were attributed to conjugated-drugs rather than to PLGA nanoparticles. Interestingly, we recently reported that PLGA without any drug/agent (native PLGA) can suppress Aß aggregation/toxicity. However, very little is known about the internalization, subcellular localization or effects of PLGA in neurons. In this study, using primary mouse cortical neurons, we first showed that native PLGA is internalized by an energy-mediated clathrin-dependent/-independent pathway and is localized in endosomal-lysosomal-autophagic vesicles. By attenuating internalization, PLGA can protect neurons against Aß-mediated toxicity. Additionally, PLGA treatment altered expression profiles of certain AD-associated genes and decreased the levels of APP, its cleaved products α-/ß-CTFs and Aß peptides in mouse as well as iPSC-derived neurons from control and AD patients. Collectively, these results suggest that native PLGA not only protects neurons against Aß-induced toxicity but also influences the expression of AD-related genes/proteins - highlighting PLGA's implication in normal and AD-related pathology.

Development of a portable hypoxia chamber for ultra-high dose rate laser-driven proton radiobiology applications.

BACKGROUND: There is currently significant interest in assessing the role of oxygen in the radiobiological effects at ultra-high dose rates. Oxygen modulation is postulated to play a role in the enhanced sparing effect observed in FLASH radiotherapy, where particles are delivered at 40-1000 Gy/s. Furthermore, the development of laser-driven accelerators now enables radiobiology experiments in extreme regimes where dose rates can exceed 109 Gy/s, and predicted oxygen depletion effects on cellular response can be tested. Access to appropriate experimental enviroments, allowing measurements under controlled oxygenation conditions, is a key requirement for these studies. We report on the development and application of a bespoke portable hypoxia chamber specifically designed for experiments employing laser-driven sources, but also suitable for comparator studies under FLASH and conventional irradiation conditions. MATERIALS AND METHODS: We used oxygen concentration measurements to test the induction of hypoxia and the maintenance capacity of the chambers. Cellular hypoxia induction was verified using hypoxia inducible factor-1α immunostaining. Calibrated radiochromic films and GEANT-4 simulations verified the dosimetry variations inside and outside the chambers. We irradiated hypoxic human skin fibroblasts (AG01522B) cells with laser-driven protons, conventional protons and reference 225 kVp X-rays to quantify DNA DSB damage and repair under hypoxia. We further measured the oxygen enhancement ratio for cell survival after X-ray exposure in normal fibroblast and radioresistant patient- derived GBM stem cells. RESULTS: Oxygen measurements showed that our chambers maintained a radiobiological hypoxic environment for at least 45 min and pathological hypoxia for up to 24 h after disconnecting the chambers from the gas supply. We observed a significant reduction in the 53BP1 foci induced by laser-driven protons, conventional protons and X-rays in the hypoxic cells compared to normoxic cells at 30 min post-irradiation. Under hypoxic irradiations, the Laser-driven protons induced significant residual DNA DSB damage in hypoxic AG01522B cells compared to the conventional dose rate protons suggesting an important impact of these extremely high dose-rate exposures. We obtained an oxygen enhancement ratio (OER) of 2.1 ± 0.1 and 2.5 ± 0.1 respectively for the AG01522B and patient-derived GBM stem cells for X-ray irradiation using our hypoxia chambers. CONCLUSION: We demonstrated the design and application of portable hypoxia chambers for studying cellular radiobiological endpoints after exposure to laser-driven protons at ultra-high dose, conventional protons and X-rays. Suitable levels of reduced oxygen concentration could be maintained in the absence of external gassing to quantify hypoxic effects. The data obtained provided indication of an enhanced residual DNA DSB damage under hypoxic conditions at ultra-high dose rate compared to the conventional protons or X-rays.

Unconjugated PLGA nanoparticles attenuate temperature-dependent ß-amyloid aggregation and protect neurons against toxicity: implications for Alzheimer's disease pathology.

Conversion of ß-amyloid (Aß) peptides from soluble random-coil to aggregated protein enriched with ß-sheet-rich intermediates has been suggested to play a role in the degeneration of neurons and development of Alzheimer's disease (AD) pathology. Aggregation of Aß peptide can be prompted by a variety of environmental factors including temperature which can influence disease pathogenesis. Recently, we reported that FDA-approved unconjugated poly (D,L-lactide-co-glycolide) (PLGA) nanoparticles can have beneficial effects in cellular and animal models of AD by targeting different facets of the Aß axis. In this study, using biochemical, structural and spectroscopic analyses, we evaluated the effects of native PLGA on temperature-dependent Aß aggregation and its ability to protect cultured neurons from degeneration. Our results show that the rate of spontaneous Aß1-42 aggregation increases with a rise in temperature from 27 to 40 °C and PLGA with 50:50 resomer potently inhibits Aß aggregation at all temperatures, but the effect is more profound at 27 °C than at 40 °C. It appears that native PLGA, by interacting with the hydrophobic domain of Aß1-42, prevents a conformational shift towards ß-sheet structure, thus precluding the formation of Aß aggregates. Additionally, PLGA triggers disassembly of matured Aß1-42 fibers at a faster rate at 40 °C than at 27 °C. PLGA-treated Aß samples can significantly enhance viability of cortical cultured neurons compared to neurons treated with Aß alone by attenuating phosphorylation of tau protein. Injection of native PLGA is found to influence the breakdown/clearance of Aß peptide in the brain. Collectively, these results suggest that PLGA nanoparticles can inhibit Aß aggregation and trigger disassembly of Aß aggregates at temperatures outside the physiological range and can protect neurons against Aß-mediated toxicity thus validating its unique therapeutic potential in the treatment of AD pathology.

Overview of the Neuroprotective Effects of the MAO-Inhibiting Antidepressant Phenelzine.

Phenelzine (PLZ) is a monoamine oxidase (MAO)-inhibiting antidepressant with anxiolytic properties. This multifaceted drug has a number of pharmacological and neurochemical effects in addition to inhibition of MAO, and findings on these effects have contributed to a body of evidence indicating that PLZ also has neuroprotective/neurorescue properties. These attributes are reviewed in this paper and include catabolism to the active metabolite ß-phenylethylidenehydrazine (PEH) and effects of PLZ and PEH on the GABA-glutamate balance in brain, sequestration of reactive aldehydes, and inhibition of primary amine oxidase. Also discussed are the encouraging findings of the effects of PLZ in animal models of stroke, spinal cord injury, traumatic brain injury, and multiple sclerosis, as well other actions such as reduction of nitrative stress, reduction of the effects of a toxin on dopaminergic neurons, potential anticonvulsant actions, and effects on brain-derived neurotrophic factor, neural cell adhesion molecules, an anti-apoptotic factor, and brain levels of ornithine and N-acetylamino acids.

Effects of Specific Inhibitors for CaMK1D on a Primary Neuron Model for Alzheimer's Disease.

Alzheimer's disease (AD) is the most common cause of dementia worldwide. Despite extensive research and targeting of the main molecular components of the disease, beta-amyloid (Aß) and tau, there are currently no treatments that alter the progression of the disease. Here, we examine the effects of two specific kinase inhibitors for calcium/calmodulin-dependent protein kinase type 1D (CaMK1D) on Aß-mediated toxicity, using mouse primary cortical neurons. Tau hyperphosphorylation and cell death were used as AD indicators. These specific inhibitors were found to prevent Aß induced tau hyperphosphorylation in culture, but were not able to protect cells from Aß induced toxicity. While inhibitors were able to alter AD pathology in cell culture, they were insufficient to prevent cell death. With further research and development, these inhibitors could contribute to a multi-drug strategy to combat AD.

Implications of exosomes derived from cholesterol-accumulated astrocytes in Alzheimer's disease pathology.

Amyloid ß (Aß) peptides generated from the amyloid precursor protein (APP) play a critical role in the development of Alzheimer's disease (AD) pathology. Aß-containing neuronal exosomes, which represent a novel form of intercellular communication, have been shown to influence the function/vulnerability of neurons in AD. Unlike neurons, the significance of exosomes derived from astrocytes remains unclear. In this study, we evaluated the significance of exosomes derived from U18666A-induced cholesterol-accumulated astrocytes in the development of AD pathology. Our results show that cholesterol accumulation decreases exosome secretion, whereas lowering cholesterol increases exosome secretion, from cultured astrocytes. Interestingly, exosomes secreted from U18666A-treated astrocytes contain higher levels of APP, APP-C-terminal fragments, soluble APP, APP secretases and Aß1-40 than exosomes secreted from control astrocytes. Furthermore, we show that exosomes derived from U18666A-treated astrocytes can lead to neurodegeneration, which is attenuated by decreasing Aß production or by neutralizing exosomal Aß peptide with an anti-Aß antibody. These results, taken together, suggest that exosomes derived from cholesterol-accumulated astrocytes can play an important role in trafficking APP/Aß peptides and influencing neuronal viability in the affected regions of the AD brain.

Mimosine functionalized gold nanoparticles (Mimo-AuNPs) suppress ß-amyloid aggregation and neuronal toxicity.

Evidence suggests that increased level/aggregation of beta-amyloid (Aß) peptides initiate neurodegeneration and subsequent development of Alzheimer's disease (AD). At present, there is no effective treatment for AD. In this study, we reported the effects of gold nanoparticles surface-functionalized with a plant-based amino acid mimosine (Mimo-AuNPs), which is found to cross the blood-brain barrier, on the Aß fibrillization process and toxicity. Thioflavin T kinetic assays, fluorescence imaging and electron microscopy data showed that Mimo-AuNPs were able to suppress the spontaneous and seed-induced Aß1-42 aggregation. Spectroscopic studies, molecular docking and biochemical analyses further revealed that Mimo-AuNPs stabilize Aß1-42 to remain in its monomeric state by interacting with the hydrophobic domain of Aß1-42 (i.e., Lys16 to Ala21) there by preventing a conformational shift towards the ß-sheet structure. Additionally, Mimo-AuNPs were found to trigger the disassembly of matured Aß1-42 fibers and increased neuronal viability by reducing phosphorylation of tau protein and the production of oxyradicals. Collectively, these results reveal that the surface-functionalization of gold nanoparticles with mimosine can attenuate Aß fibrillization and neuronal toxicity. Thus, we propose Mimo-AuNPs may be used as a potential treatment strategy towards AD-related pathologies.

High energy implementation of coil-target scheme for guided re-acceleration of laser-driven protons.

Developing compact ion accelerators using intense lasers is a very active area of research, motivated by a strong applicative potential in science, industry and healthcare. However, proposed applications in medical therapy, as well as in nuclear and particle physics demand a strict control of ion energy, as well as of the angular and spectral distribution of ion beam, beyond the intrinsic limitations of the several acceleration mechanisms explored so far. Here we report on the production of highly collimated ([Formula: see text] half angle divergence), high-charge (10s of pC) and quasi-monoenergetic proton beams up to [Formula: see text] 50 MeV, using a recently developed method based on helical coil targetry. In this concept, ions accelerated from a laser-irradiated foil are post-accelerated and conditioned in a helical structure positioned at the rear of the foil. The pencil beam of protons was produced by guided post-acceleration at a rate of [Formula: see text] 2 GeV/m, without sacrificing the excellent beam emittance of the laser-driven proton beams. 3D particle tracing simulations indicate the possibility of sustaining high acceleration gradients over extended helical coil lengths, thus maximising the gain from such miniature accelerating modules.

Proteasomal Inhibition Redirects the PrP-Like Shadoo Protein to the Nucleus.

The Shadoo protein (Sho) exhibits homology to the hydrophobic region of the cellular isoform of prion protein (PrPC). As prion-infected brains gradually accumulate infectivity-associated isoforms of prion protein (PrPSc), levels of mature endogenous Sho become reduced. To study the regulatory effect of the proteostatic network on Sho expression, we investigated the action of lactacystin, MG132, NH4Cl, and 3-methyladenine (3-MA) in two cell culture models. In primary mixed neuronal and glial cell cultures (MNGCs) from transgenic mice expressing wild-type Sho from the PrP gene promoter (Tg.Sprn mice), lactacystin- and MG132-mediated inhibition of proteasomal activity shifted the repertoire of Sho species towards unglycosylated forms appearing in the nuclei; conversely, the autophagic modulators NH4Cl and 3-MA did not affect Sho or PrPC glycosylation patterns. Mouse N2a neuroblastoma cells expressing Sho under control of a housekeeping gene promoter treated with MG132 or lactacystin also showed increased nuclear localization of unglycosylated Sho. As two proteasomal inhibitors tested in two cell paradigms caused redirection of Sho to nuclei at the expense of processing through the secretory pathway, our findings define a balanced shift in subcellular localization that thereby differs from the decreases in net Sho species seen in prion-infected brains. Our data are indicative of a physiological pathway to access Sho functions in the nucleus under conditions of impaired proteasomal activity. We also infer that these conditions would comprise a context wherein Sho's N-terminal nucleic acid-binding RGG repeat region is brought into play.

Attenuation of the effects of oxidative stress by the MAO-inhibiting antidepressant and carbonyl scavenger phenelzine.

Phenelzine (ß-phenylethylhydrazine) is a monoamine oxidase (MAO)-inhibiting antidepressant with anxiolytic properties. It possesses a number of important pharmacological properties which may alter the effects of oxidative stress. After conducting a comprehensive literature search, the authors of this review paper aim to provide an overview and discussion of the mechanisms by which phenelzine may attenuate oxidative stress. It inhibits γ-aminobutyric acid (GABA) transaminase, resulting in elevated brain GABA levels, inhibits both MAO and primary amine oxidase and, due to its hydrazine-containing structure, reacts chemically to sequester a number of reactive aldehydes (e.g. acrolein and 4-hydroxy-2-nonenal) proposed to be implicated in oxidative stress in a number of neurodegenerative disorders. Phenelzine is unusual in that it is both an inhibitor of and a substrate for MAO, the latter action producing at least one active metabolite, ß-phenylethylidenehydrazine (PEH). This metabolite inhibits GABA transaminase, is a very weak inhibitor of MAO but a strong inhibitor of primary amine oxidase, and sequesters aldehydes. Phenelzine may ameliorate the effects of oxidative stress by reducing formation of reactive metabolites (aldehydes, hydrogen peroxide, ammonia/ammonia derivatives) produced by the interaction of MAO with biogenic amines, by sequestering various other reactive aldehydes and by inhibiting primary amine oxidase. In PC12 cells treated with the neurotoxin MPP+, phenelzine has been reported to reduce several adverse effects of MPP+. It has also been reported to reduce lipid peroxidative damage induced in plasma and platelet proteins by peroxynitrite. In animal models, phenelzine has a neuroprotective effect in global ischemia and in cortical impact traumatic brain injury. Recent studies reported in the literature on the possible involvement of acrolein in spinal cord injury and multiple sclerosis indicate that phenelzine can attenuate adverse effects of acrolein in these models. Results from studies in our laboratories on effects of phenelzine and PEH on primary amine oxidase (which catalyzes formation of toxic aldehydes and is overexpressed in Alzheimer's disease), on sequestration of the toxic aldehyde acrolein, and on reduction of acrolein-induced toxicity in mouse cortical neurons are also reported.

Kainate Receptor Activation Enhances Amyloidogenic Processing of APP in Astrocytes.

Kainic acid (KA) is an analogue of the excitatory neurotransmitter glutamate that, when injected systemically into adult rats, can trigger seizures and progressive neuronal loss in a manner that mirrors the neuropathology of human mesial temporal lobe epilepsy. However, biomolecular mechanisms responsible for the neuronal loss that occurs as a consequence of this treatment remains elusive. We have recently reported that toxicity induced by KA can partly be mediated by astrocyte-derived amyloid ß (Aß) peptides, which are critical in the development of Alzheimer's disease (AD). Nonetheless, little is known how KA can influence amyloid precursor protein (APP) levels and processing in astrocytes. Thus, in the present study using human U-373 astrocytoma and rat primary astrocytes, we evaluated the role of KA on APP metabolism. Our results revealed that KA treatment increased the levels of APP and its cleaved products (α-/ß-CTFs) in cultured U-373 astrocytoma and primary astrocytes, without altering the cell viability. The cellular and secretory levels of Aß1-40/Aß1-42 were markedly increased in KA-treated astrocytes. We also demonstrated that the steady-state levels of APP-secretases were not altered but the activity of γ-secretase is enhanced in KA-treated U-373 astrocytoma. Furthermore, using selective receptor antagonists, we showed that the effects of KA is mediated by activation of kainate receptors and not NMDA or AMPA receptors. These results suggest that KA can enhance amyloidogenic processing of APP by activating its own receptor leading to increased production/secretion of Aß-related peptides from activated astrocytes which may contribute to the pathogenesis of temporal lobe epilepsy.

Effects of cholesterol transport inhibitor U18666A on APP metabolism in rat primary astrocytes.

Amyloid ß (Aß) peptides generated from the amyloid precursor protein (APP) play an important role in the degeneration of neurons and development of Alzheimer's disease (AD). Current evidence indicates that high levels of cholesterol-which increase the risk of developing AD-can influence Aß production in neurons. However, it remains unclear how altered level/subcellular distribution of cholesterol in astrocytes can influence APP metabolism. In this study, we evaluated the effects of cholesterol transport inhibitor U18666A-a class II amphiphile that triggers redistribution of cholesterol within the endosomal-lysosomal (EL) system-on APP levels and metabolism in rat primary cultured astrocytes. Our results revealed that U18666A increased the levels of the APP holoprotein and its cleaved products (α-/ß-/η-CTFs) in cultured astrocytes, without altering the total levels of cholesterol or cell viability. The cellular levels of Aß1-40 were also found to be markedly increased, while secretory levels of Aß1-40 were decreased in U18666A-treated astrocytes. We further report a corresponding increase in the activity of the enzymes regulating APP processing, such as α-secretase, ß-secretase, and γ-secretase as a consequence of U18666A treatment. Additionally, APP-cleaved products are partly accumulated in the lysosomes following cholesterol sequestration within EL system possibly due to decreased clearance. Interestingly, serum delipidation attenuated enhanced levels of APP and its cleaved products following U18666A treatment. Collectively, these results suggest that cholesterol sequestration within the EL system in astrocytes can influence APP metabolism and the accumulation of APP-cleaved products including Aß peptides, which can contribute to the development of AD pathology.

Predominant expression of Alzheimer's disease-associated BIN1 in mature oligodendrocytes and localization to white matter tracts.

BACKGROUND: Genome-wide association studies have identified BIN1 within the second most significant susceptibility locus in late-onset Alzheimer's disease (AD). BIN1 undergoes complex alternative splicing to generate multiple isoforms with diverse functions in multiple cellular processes including endocytosis and membrane remodeling. An increase in BIN1 expression in AD and an interaction between BIN1 and Tau have been reported. However, disparate descriptions of BIN1 expression and localization in the brain previously reported in the literature and the lack of clarity on brain BIN1 isoforms present formidable challenges to our understanding of how genetic variants in BIN1 increase the risk for AD. METHODS: In this study, we analyzed BIN1 mRNA and protein levels in human brain samples from individuals with or without AD. In addition, we characterized the BIN1 expression and isoform diversity in human and rodent tissue by immunohistochemistry and immunoblotting using a panel of BIN1 antibodies. RESULTS: Here, we report on BIN1 isoform diversity in the human brain and document alterations in the levels of select BIN1 isoforms in individuals with AD. In addition, we report striking BIN1 localization to white matter tracts in rodent and the human brain, and document that the large majority of BIN1 is expressed in mature oligodendrocytes whereas neuronal BIN1 represents a minor fraction. This predominant non-neuronal BIN1 localization contrasts with the strict neuronal expression and presynaptic localization of the BIN1 paralog, Amphiphysin 1. We also observe upregulation of BIN1 at the onset of postnatal myelination in the brain and during differentiation of cultured oligodendrocytes. Finally, we document that the loss of BIN1 significantly correlates with the extent of demyelination in multiple sclerosis lesions. CONCLUSION: Our study provides new insights into the brain distribution and cellular expression of an important risk factor associated with late-onset AD. We propose that efforts to define how genetic variants in BIN1 elevate the risk for AD would behoove to consider BIN1 function in the context of its main expression in mature oligodendrocytes and the potential for a role of BIN1 in the membrane remodeling that accompanies the process of myelination.