Amyotrophic Lateral Sclerosis/parkinsonism-dementia Complex Research Articles

Aberrant CHCHD2-associatedmitochondriopathy in Kii ALS/PDC astrocytes.

Amyotrophic Lateral Sclerosis/Parkinsonism-Dementia Complex (ALS/PDC), a rare and complex neurological disorder, is predominantly observed in the Western Pacific islands, including regions of Japan, Guam, and Papua. This enigmatic condition continues to capture medical attention due to affected patients displaying symptoms that parallel those seen in either classical amyotrophic lateral sclerosis (ALS) or Parkinson's disease (PD). Distinctly, postmortem examinations of the brains of affected individuals have shown the presence of α-synuclein aggregates and TDP-43, which are hallmarks of PD and classical ALS, respectively. These observations are further complicated by the detection of phosphorylated tau, accentuating the multifaceted proteinopathic nature of ALS/PDC. The etiological foundations of this disease remain undetermined, and genetic investigations have yet to provide conclusive answers. However, emerging evidence has implicated the contribution of astrocytes, pivotal cells for maintaining brain health, to neurodegenerative onset, and likely to play a significant role in the pathogenesis of ALS/PDC. Leveraging advanced induced pluripotent stem cell technology, our team cultivated multiple astrocyte lines to further investigate the Japanese variant of ALS/PDC (Kii ALS/PDC). CHCHD2 emerged as asignificantly dysregulated gene when disease astrocytes were compared to healthy controls. Our analyses also revealed imbalances in the activation of specific pathways: those associated with astrocytic cilium dysfunction, known to be involved in neurodegeneration, and those related to major neurological disorders, including classical ALS and PD. Further in-depth examinations revealed abnormalities in the mitochondrial morphology and metabolic processes of the affected astrocytes. A particularly striking observation was the reduced expression of CHCHD2 in the spinal cord, motor cortex, and oculomotor nuclei of patients with Kii ALS/PDC. In summary, our findings suggest a potential reduction in the support Kii ALS/PDC astrocytes provide to neurons, emphasizing the need to explore the role of CHCHD2 in maintaining mitochondrial health and its implications for the disease.

Ultrafine particulate matter pollution and dysfunction of endoplasmic reticulum Ca2+ store: A pathomechanism shared with amyotrophic lateral sclerosis motor neurons?

Increased risk of neurodegenerative diseases has been envisaged for air pollution exposure. On the other hand, environmental risk factors, including air pollution, have been suggested for Amyotrophic Lateral Sclerosis (ALS) pathomechanism. Therefore, the neurotoxicity of ultrafine particulate matter (PM0.1) (PM < 0.1 μm size) and its sub-20 nm nanoparticle fraction (NP20) has been investigated in motor neuronal-like cells and primary cortical neurons, mainly affected in ALS. The present data showed that PM0.1 and NP20 exposure induced endoplasmic reticulum (ER) stress, as occurred in cortex and spinal cord of ALS mice carrying G93A mutation in SOD1 gene. Furthermore, NSC-34 motor neuronal-like cells exposed to PM0.1 and NP20 shared the same proteomic profile on some apoptotic factors with motor neurons treated with the L-BMAA, a neurotoxin inducing Amyotrophic Lateral Sclerosis/Parkinson–Dementia Complex (ALS/PDC). Of note ER stress induced by PM0.1 and NP20 in motor neurons was associated to pathological changes in ER morphology and dramatic reduction of organellar Ca2+ level through the dysregulation of the Ca2+-pumps SERCA2 and SERCA3, the Ca2+-sensor STIM1, and the Ca2+-release channels RyR3 and IP3R3. Furthermore, the mechanism deputed to ER Ca2+ refilling (e.g. the so called store operated calcium entry-SOCE) and the relative currents ICRAC were also altered by PM0.1 and NP20 exposure. Additionally, these carbonaceous particles caused the exacerbation of L-BMAA-induced ER stress and Caspase-9 activation. In conclusion, this study shows that PM0.1 and NP20 induced the aberrant expression of ER proteins leading to dysmorphic ER, organellar Ca2+ dysfunction, ER stress and neurotoxicity, providing putative correlations with the neurodegenerative process occurring in ALS.

Tau filaments from amyotrophic lateral sclerosis/parkinsonism-dementia complex adopt the CTE fold.

The amyotrophic lateral sclerosis/parkinsonism-dementia complex (ALS/PDC) of the island of Guam and the Kii peninsula of Japan is a fatal neurodegenerative disease of unknown cause that is characterized by the presence of abundant filamentous tau inclusions in brains and spinal cords. Here, we used electron cryo-microscopy to determine the structures of tau filaments from the cerebral cortex of three cases of ALS/PDC from Guam and eight cases from Kii, as well as from the spinal cord of two of the Guam cases. Tau filaments had the chronic traumatic encephalopathy (CTE) fold, with variable amounts of Type I and Type II filaments. Paired helical tau filaments were also found in three Kii cases and tau filaments with the corticobasal degeneration fold in one Kii case. We identified a new Type III CTE tau filament, where protofilaments pack against each other in an antiparallel fashion. ALS/PDC is the third known tauopathy with CTE-type filaments and abundant tau inclusions in cortical layers II/III, the others being CTE and subacute sclerosing panencephalitis. Because these tauopathies are believed to have environmental causes, our findings support the hypothesis that ALS/PDC is caused by exogenous factors.

Guam ALS-PDC is a distinct double-prion disorder featuring both tau and Aβ prions

The amyotrophic lateral sclerosis-parkinsonism dementia complex (ALS-PDC) of Guam is an endemic neurodegenerative disease that features widespread tau tangles, occasional α-synuclein Lewy bodies, and sparse β-amyloid (Aβ) plaques distributed in the central nervous system. Extensive studies of genetic or environmental factors have failed to identify a cause of ALS-PDC. Building on prior work describing the detection of tau and Aβ prions in Alzheimer's disease (AD) and Down syndrome brains, we investigated ALS-PDC brain samples for the presence of prions. We obtained postmortem frozen brain tissue from 26 donors from Guam with ALS-PDC or no neurological impairment and 71 non-Guamanian donors with AD or no neurological impairment. We employed cellular bioassays to detect the prion conformers of tau, α-synuclein, and Aβ proteins in brain extracts. In ALS-PDC brain samples, we detected high titers of tau and Aβ prions, but we did not detect α-synuclein prions in either cohort. The specific activity of tau and Aβ prions was increased in Guam ALS-PDC compared with sporadic AD. Applying partial least squares regression to all biochemical and prion infectivity measurements, we demonstrated that the ALS-PDC cohort has a unique molecular signature distinguishable from AD. Our findings argue that Guam ALS-PDC is a distinct double-prion disorder featuring both tau and Aβ prions.

The biotoxin BMAA promotes dysfunction via distinct mechanisms in neuroblastoma and glioblastoma cells.

Chronic exposure to the Cyanobacteria biotoxin Beta-methylamino-L-alanine (BMAA) has been associated with development of a sporadic form of ALS called Amyotrophic Lateral Sclerosis/Parkinsonism-Dementia Complex (ALS/PDC), as observed within certain Indigenous populations of Guam and Japan. Studies in primate models and cell culture have supported the association of BMAA with ALS/PDC, yet the pathological mechanisms at play remain incompletely characterized, effectively stalling the development of rationally-designed therapeutics or application of preventative measures for this disease. In this study we demonstrate for the first time that sub-excitotoxic doses of BMAA modulate the canonical Wnt signaling pathway to drive cellular defects in human neuroblastoma cells, suggesting a potential mechanism by which BMAA may promote neurological disease. Further, we demonstrate here that the effects of BMAA can be reversed in cell culture by use of pharmacological modulators of the Wnt pathway, revealing the potential value of targeting this pathway therapeutically. Interestingly, our results suggest the existence of a distinct Wnt-independent mechanism activated by BMAA in glioblastoma cells, highlighting the likelihood that neurological disease may result from the cumulative effects of distinct cell-type specific mechanisms of BMAA toxicity.

Degradation of cyanobacterial neurotoxin β-N-methylamino-L-alanine (BMAA) using ozone process: influencing factors and mechanism.

β-N-methylamino-L-alanine (BMAA), which has been considered as an environmental factor that caused amyotrophic lateral sclerosis/parkinsonism-dementia complex (ALS/PDC) or Alzheimer's disease, could be produced by a variety of genera cyanobacteria. BMAA is widely present in water sources contaminated by cyanobacteria and may threaten human health through drinking water. Although oxidants commonly used in drinking water plants such as chlorine, ozone, hydrogen peroxide, and hydroxyl radicals have been shown to effectively degrade BMAA, there are limited studies on the mechanism of BMAA degradation by different oxidants, especially ozone. This work systematically explored the effectiveness of BMAA ozonation degradation, investigated the effect of the operating parameters on the effectiveness of degradation, and speculated on the pathways of BMAA decomposition. The results showed that BMAA could be quickly eliminated by ozone, and the removal rates of BMAA were nearly 100% in pure water, but the removal rates were reduced in actual water. BMAA was primarily degraded by direct oxidation of ozone molecules in acidic and near-neutral conditions, and indirect oxidation of •OH accounted for the main part under strong alkaline conditions. The pH value had a significant effect on the decomposition of BMAA, and the degradation rate of BMAA was fastest at near-neutral pH value. The degradation rates of TOC were significantly lower than that of BMAA, indicating that by-products were generated during the degradation process. Three by-products ([M-H]+ = 105, 90, and 88) were identified by UPLC-MS/MS, and the degradation pathways of BMAA were proposed. The production of by-products was attributed to the fracture of the C-N bonds. This work is helpful for the in-depth understanding on the mechanism and demonstration of the feasibility of the oxidation of BMAA by the ozone process. HIGHLIGHTS: • The reaction of ozonation BMAA was easy to occur. • The degradation rate was fast under near-neutral conditions. • Direct oxidation under neural conditions was the main pathway for ozone degradation of BMAA. • Three products were detected, and the reaction path was inferred.

Metals and Metalloids Increase in Cycas micronesica Seed Gametophyte Tissue in Shaded Growth Conditions.

Exposure to environmental toxins may be partly responsible for mammal neurodegenerative disorders. Consumption of seeds from Guam’s cycad tree has been linked to the disorder known as amyotrophic lateral sclerosis-parkinsonism dementia complex (ALS-PDC). The unambiguous identification of causal agents of ALS-PDC has been elusive. We have examined the levels of eight metals and metalloids in cycad seeds as a function of the ambient shade in which the plants were grown. Of these metals, the data strongly suggest that aluminum (Al) and selenium (Se) are present in washed flour prepared from southern Guam’s cycad seed tissues at elevated levels, especially when the trees are grown in shade. Previous authors have speculated that Al and Se are involved in various ALS outcomes, and our results support this interpretation.

Matrix Effect of Diverse Biological Samples Extracted with Different Extraction Ratios on the Detection of β-N-Methylamino-L-Alanine by Two Common LC-MS/MS Analysis Methods.

Neurotoxin β-N-methylamino-L-alanine (BMAA) is hypothesized as an important pathogenic factor for neurodegenerative diseases such as amyotrophic lateral sclerosis/parkinsonism-dementia complex (ALS-PDC). Comparative study on the accuracy of BMAA analyzed by the regular LC-MS/MS methods is still limited for different biological matrices. In this study, a free-BMAA sample of cyanobacterium and BMAA-containing positive samples of diatom, mussel, scallop, and oyster were extracted with varied extraction ratios (ER) ranging from 1:20 to 1:2000. These extracts were then purified by MCX cartridges. After SPE purification, these different biological samples were analyzed by two common LC-MS/MS analysis methods, a direct analysis without derivatization by a hydrophilic interaction liquid chromatography (HILIC)-MS/MS and pre-column 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) derivatization combined with a C18 column. The results suggested that the recoveries of BMAA spiked in the cyanobacterial sample were close to 100% in the total soluble form extracts with the ER of 1:100 (g/mL) and the precipitated bound form extracts with the ER of 1:500. The recommended ER for the precipitated bound form of BMAA in diatoms and the total soluble form of BMAA in mollusks are 1:500 and 1:50, respectively. The quantitative results determined by the AQC derivatization method were lower than those determined by the direct analysis of the HILIC method in diatom and mollusk samples. The results of the HILIC method without the derivatization process were closer to the true value of BMAA in cyanobacteria. This work contributes to the performance of the solid-phase extraction (SPE) purification protocol and the accuracy of BMAA analysis by LC-MS/MS in diverse biological samples.

Towards the Selection of Single‐Stranded DNA Molecular Recognition Element Against Cyanotoxin L‐BMAA

Global warming, anthropogenic pressure and eutrophication contribute to the formation and persistence of cyanobacterial blooms in aquatic environments. Cyanotoxins produced during these blooms represent a threat to ecosystems, commercial industries and human health. B‐N‐methylamino‐L‐alanine (L‐BMAA) is a neurotoxic amino acid that has been linked to neurodegenerative diseases such as Amyotrophic Lateral Sclerosis/Parkinsonism Dementia Complex (ALS/PDC), Alzheimer’s disease (AD) and Amyotrophic Lateral Sclerosis (ALS). Research has shown that L‐BMAA can act as a glutamate receptor antagonist and can incorporate into proteins occasioning protein misfolding that leads to detrimental processes in brain tissues. Currently, the detection of L‐BMAA is costly, time‐consuming and labor‐intensive. Many research groups around the world are working towards the development of bioanalytical methods to improve the detection of these toxins, including the development of molecular biorecognition elements (MREs) that can provide analyte specificity in biosensing applications. Among MREs, aptamers are well‐known to possess an extraordinary target recognition due to the formation of unique three‐dimensional structures capable of binding specifically to their respective targets.We aim to perform an in vitro selection of a single‐stranded DNA (ssDNA) aptamer that can act as an MRE against L‐BMAA, using the systematic evolution of ligands by exponential enrichment (SELEX) technique. We have worked on the optimization of incubation conditions, PCR amplification parameters and the generation of ssDNA using two different ssDNA libraries, with randomized regions of varying lengths. To facilitate the separation of bound and unbound ssDNA after incubation with the target, we modified L‐BMAA with a biotinylated linker to immobilize it in streptavidin‐coated magnetic beads. For each library, we have identified the appropriate annealing temperature, primer concentrations and number of cycles to favor the amplification of the main product while minimizing by‐product formation. The most challenging step of the SELEX procedure has been the generation and purification of ssDNA in sufficient amounts to continue to the next round of selection. To achieve this, we have tried four main methods: exonuclease digestion, size separation derived from uneven primers, asymmetric PCR and alkaline denaturation using sodium hydroxide. Hitherto, we have completed the first round of selection with both libraries, by performing large‐scale PCR reactions and alkaline denaturation followed by ethanol precipitation to generate the ssDNA. We are working in the first round of negative selection to discard those sequences with affinity to the immobilization linker and enrich only those that bind exclusively to L‐BMAA. Given that our target is a small molecule, our goal is to conduct a stringent SELEX process to obtain a highly specific MRE that can be incorporated into an environmental sensing device.

Longitudinal Assessment of Behaviour and Associated Bio-Markers Following Chronic Consumption of β-Sitosterol β-D-Glucoside in Rats: A Putative Model of Parkinson's Disease.

The consumption of cycad (Cycas circinalis) seeds has been linked to the development of Amyotrophic Lateral Sclerosis-Parkinsonism Dementia Complex (ALS-PDC) in humans. ALS-PDC is a clinically variable disease presenting as a combination of symptoms typical of PD and/or ALS. Chronic consumption of β-sitosterol β-D-glucoside (BSSG), a component of the cycad seed, by rats (Rattus norvegicus) has been previously reported to initiate a progressive pathology that develops over several months and manifests as behavioural and histopathological changes that resemble characteristic features of Parkinson’s disease. As part of an independent multi-site validation study, we have tried to replicate and further characterize the BSSG model with a focus on motor function, and associated immunohistochemical markers. Beginning at 3 months of age, male CD® (Sprague Dawley) rats (N = 80) were dosed orally with either a flour pellet or a flour pellet containing BSSG (3 mg) daily (5×/week) for 16 weeks consistent with previous reports of the model. Following BSSG intoxication, separate cohorts of animals (n = 10/treatment) were exposed to a behavioural test battery at 16, 24, 32, or 40 weeks post-initial BSSG feeding. The test battery consisted of the open field test, cylinder test, and ultrasonic vocalization (USV) assessment. No changes in behaviour were observed at any time point. Following behavioural testing, animals were processed for immunohistochemical markers of substantia nigra integrity. Immunohistochemistry of brain tissue revealed no differences in the microglial marker, Iba1, or the dopaminergic integrity marker, tyrosine hydroxylase (TH), in the substantia nigra at any assessment point. The absence of any group differences in behaviour and immunhistochemistry indicates an inability to replicate previous reports. Further investigation into the sources of variability in the model is necessary prior to further utilization of the BSSG model in preclinical studies.

Na+/Ca2+ exchanger isoform 1 takes part to the Ca2+-related prosurvival pathway of SOD1 in primary motor neurons exposed to beta-methylamino-l-alanine

BackgroundThe cycad neurotoxin beta-methylamino-l-alanine (L-BMAA), one of the environmental trigger factor for amyotrophic lateral sclerosis/Parkinson-dementia complex (ALS/PDC), may cause neurodegeneration by disrupting organellar Ca2+ homeostasis. Through the activation of Akt/ERK1/2 pathway, the Cu,Zn-superoxide dismutase (SOD1) and its non-metallated form, ApoSOD1, prevent endoplasmic reticulum (ER) stress-induced cell death in motor neurons exposed to L-BMAA. This occurs through the rapid increase of intracellular Ca2+ concentration ([Ca2+]i) in part flowing from the extracellular compartment and in part released from ER. However, the molecular components of this mechanism remain uncharacterized.MethodsBy an integrated approach consisting on the use of siRNA strategy, Western blotting, confocal double- labeling immunofluorescence, patch-clamp electrophysiology, and Fura 2-/SBFI-single-cell imaging, we explored in rat motor neuron-enriched cultures the involvement of the plasma membrane proteins Na+/Ca2+ exchanger (NCX) and purinergic P2X7 receptor as well as that of the intracellular cADP-ribose (cADPR) pathway, in the neuroprotective mechanism of SOD1.ResultsWe showed that SOD1-induced [Ca2+]i rise was prevented neither by A430879, a P2X7 receptor specific antagonist or 8-bromo-cADPR, a cell permeant antagonist of cADP-ribose, but only by the pan inhibitor of NCX, CB-DMB. The same occurred for the ApoSOD1. Confocal double labeling immunofluorescence showed a huge expression of plasmalemmal NCX1 and intracellular NCX3 isoforms. Furthermore, we identified NCX1 reverse mode as the main mechanism responsible for the neuroprotective ER Ca2+ refilling elicited by SOD1 and ApoSOD1 through which they promoted translocation of active Akt in the nuclei of a subset of primary motor neurons. Finally, the activation of NCX1 by the specific agonist CN-PYB2 protected motor neurons from L-BMAA-induced cell death, mimicking the effect of SOD1.ConclusionCollectively, our data indicate that SOD1 and ApoSOD1 exert their neuroprotective effect by modulating ER Ca2+ content through the activation of NCX1 reverse mode and Akt nuclear translocation in a subset of primary motor neurons.7n_RSb6LJnzKN1GMcoyFawVideo

An immigrant family with Kii amyotrophic lateral sclerosis/parkinsonism-dementia complex.

Amyotrophic lateral sclerosis/parkinsonism-dementia complex (ALS/PDC) is a unique endemic on Guam island of the USA, the Kii Peninsula of Japan, and Papua state of Indonesia. The pathomechanism of ALS/PDC remains to be solved, although interaction between some environmental factors and genetic background is plausible. This is the first autopsy-proven immigrant family of ALS/PDC of the Kii Peninsula. A daughter and her father immigrated to the high incident area from outside the Kii Peninsula. The father developed ALS 18years later after immigration, and his daughter also developed ALS 65years after immigration. They showed pure ALS phenotype without parkinsonism and dementia. The daughter was diagnosed neuropathologically with Kii ALS/PDC with multiple proteinopathies: tauopathy, α-synucleinopathy, and TDP-43 proteinopathy. Gene analysis of familial ALS-related genes, including C9orf72, showed no mutation. The findings in an immigrant family established that certain environmental factors play a critical role in the pathogenesis of Kii ALS/PDC.

Elimination of β-N-methylamino-l-alanine (BMAA) during UV/chlorine process: Influence factors, transformation pathway and DBP formation

As a new cyanobacterial neurotoxin generated by cyanobacteria, BMAA was closely related to amyotrophic lateral sclerosis-parkinsonism dementia complex (ALS/PDC). In this study, the degradation of BMAA by UV/chlorine process was investigated under the impacts of chlorine dosage, NOM dosage, pH and alkalinity. Results showed that only 10% of BMAA was removed by UV irradiation and 46.8% by chlorination in 5 min, however, 98.6% of BMAA was removed by UV/chlorine process in 5 min. The reaction rates were increased under alkaline conditions, but all achieved complete degradation in 5 min. Besides, HCO3− had slight inhibition, while NOM had significant inhibition on the degradation of BMAA. Furthermore, based on the detected degradation products of BMAA during UV/chlorine process, the possible degradation pathways were concluded. Overall, outcomes of this study exhibited that the use of the UV/chlorine process for BMAA degradation was appropriate in practical applications.

Nodding syndrome, an epidemic young-onset epilepsy-dementia complex in Uganda

Nodding syndrome (NS) is an enigmatic recurrent epidemic neurological disease that affects children in East Africa. The illness begins with nodding of the head and grand mal seizures that may lead to death after several years. The most recent outbreaks of NS occurred in northern Uganda and South Sudan. We describe the clinicopathologic spectrum of NS in Uganda. Ten children or young adults with NS were studied at autopsy and the neuropathological findings correlated with the onset, duration and progression of their neurological illness. All cases had epilepsy with grand mal seizures. Three cases had a clinical course that was predominantly characterized by epilepsy. Seven patients had progressive frontotemporal dementia. Two of the patients with dementia also had progressive quadriparesis. In all cases, the brain revealed tau pathology. In cases with an epilepsy-predominate course, the tau pathology was largely limited to the anterior frontal lobes but cases with dementia had more widespread cortical and subcortical tau pathology. In some cases, the histologic pattern was reminiscent of progressive supranuclear palsy. There are some interesting parallels between NS and the amyotrophic lateral sclerosis/Parkinson-dementia complex (ALS/PDC). The similarities are the presence of geographical isolates of disease manifesting in indigenous populations with familial clusters but no clear heritability. Both disorders appear to be related to an unknown environmental factor and both diseases appear to be fading over time in the respective geographical locations. One of the major open questions is whether ALS occurs in NS. This question will be addressed in future clinical studies and postmortem examination of the spinal cord. We propose that NS is a unique epilepsy-dementia complex in East Africa.LEARNING OBJECTIVESThis presentation will enable the learner to: 1.Describe the clinicopathologic features of a nodding syndrome.2.Compare the pathology of NS to ALS/PDC and related disease

Presence of ß-N-methylamino-L-alanine in cyanobacteria and aquatic organisms from waters of Northern Poland; BMAA toxicity studies

BMAA (ß-N-methylamino-L-alanine) was originally found in the seeds of cycad Cycas micronesica in the 1960s. Some years later it was discovered that the amino acid is genuinely produced by endosymbiotic cyanobacteria. Further research has proven the neurotoxic activity of BMAA, leading to neurodegenerative disease diagnosed as amyotrophic lateral sclerosis/parkinsonism-dementia complex (ALS/PDC). The aim of the present work was to examine the occurrence of BMAA in samples from Polish waterbodies. Both, the field cyanobacterial samples and the isolated cyanobacterial strains were analyzed. Also mussel and fish samples were checked for the BMAA accumulation. The additional goal was to assess the biological activity of BMAA in in vivo and in vitro assays. In waters of Northern Poland, BMAA was detected in cyanobacteria from Synechococcales, Oscillatoriales and Nostocales orders. The free and protein-bound forms of BMAA were detected in 9 and 4 (of 37) environmental samples, respectively. Both forms of BMAA were also identified in 2 out of 21 cyanobacterial strains isolated from Polish waterbodies. Our analyses of cyanobacterial material did not confirm the presence of soluble protein-bound form of BMAA. The amino acid was detected neither in the tissues of fish nor in the mussels. Biological activity of BMAA was tested with the application of hippocampal neural cell line HT22 and crustaceans: Thamnocephalus platyurus, Artemia franciscana and Daphnia magna. Among them, only D. magna assay revealed toxic effects of BMAA. The results of our research did not demonstrate the widespread production of BMAA by cyanobacteria from Northern Poland waters.

Microbial BMAA elicits mitochondrial dysfunction, innate immunity activation, and Alzheimer\u2019s disease features in cortical neurons

BackgroundAfter decades of research recognizing it as a complex multifactorial disorder, sporadic Alzheimer’s disease (sAD) still has no known etiology. Adding to the myriad of different pathways involved, bacterial neurotoxins are assuming greater importance in the etiology and/or progression of sAD. β-N-Methylamino-l-alanine (BMAA), a neurotoxin produced by some microorganisms namely cyanobacteria, was previously detected in the brains of AD patients. Indeed, the consumption of BMAA-enriched foods has been proposed to induce amyotrophic lateral sclerosis-parkinsonism-dementia complex (ALS-PDC), which implicated this microbial metabolite in neurodegeneration mechanisms.MethodsFreshly isolated mitochondria from C57BL/6 mice were treated with BMAA and O2 consumption rates were determined. O2 consumption and glycolysis rates were also measured in mouse primary cortical neuronal cultures. Further, mitochondrial membrane potential and ROS production were evaluated by fluorimetry and the integrity of mitochondrial network was examined by immunofluorescence. Finally, the ability of BMAA to activate neuronal innate immunity was quantified by addressing TLRs (Toll-like receptors) expression, p65 NF-κB translocation into the nucleus, increased expression of NLRP3 (Nod-like receptor 3), and pro-IL-1β. Caspase-1 activity was evaluated using a colorimetric substrate and mature IL-1β levels were also determined by ELISA.ResultsTreatment with BMAA reduced O2 consumption rates in both isolated mitochondria and in primary cortical cultures, with additional reduced glycolytic rates, decrease mitochondrial potential and increased ROS production. The mitochondrial network was found to be fragmented, which resulted in cardiolipin exposure that stimulated inflammasome NLRP3, reinforced by decreased mitochondrial turnover, as indicated by increased p62 levels. BMAA treatment also activated neuronal extracellular TLR4 and intracellular TLR3, inducing p65 NF-κB translocation into the nucleus and activating the transcription of NLRP3 and pro-IL-1β. Increased caspase-1 activity resulted in elevated levels of mature IL-1β. These alterations in mitochondrial metabolism and inflammation increased Tau phosphorylation and Aβ peptides production, two hallmarks of AD.ConclusionsHere we propose a unifying mechanism for AD neurodegeneration in which a microbial toxin can induce mitochondrial dysfunction and activate neuronal innate immunity, which ultimately results in Tau and Aβ pathology. Our data show that neurons, alone, can mount inflammatory responses, a role previously attributed exclusively to glial cells.

Damaging effects of BMAA on retina neurons and Müller glial cells

B–N-methylamino-L-alanine (BMAA), a cyanotoxin produced by most cyanobacteria, has been proposed to cause long term damages leading to neurodegenerative diseases, including Amyotrophic Lateral Sclerosis/Parkinsonism Dementia complex (ALS/PDC) and retinal pathologies. Previous work has shown diverse mechanisms leading to BMAA-induced degeneration; however, the underlying mechanisms of toxicity affecting retina cells are not fully elucidated. We here show that BMAA treatment of rat retina neurons in vitro induced nuclear fragmentation and cell death in both photoreceptors (PHRs) and amacrine neurons, provoking mitochondrial membrane depolarization. Pretreatment with the N-Methyl-D-aspartate (NMDA) receptor antagonist MK-801 prevented BMAA-induced death of amacrine neurons, but not that of PHRs, implying activation of NMDA receptors participated only in amacrine cell death. Noteworthy, BMAA stimulated a selective axonal outgrowth in amacrine neurons, simultaneously promoting growth cone destabilization. BMAA partially decreased the viability of Müller glial cells (MGC), the main glial cell type in the retina, induced marked alterations in their actin cytoskeleton and impaired their capacity to protect retinal neurons. BMAA also induced cell death and promoted axonal outgrowth in differentiated rat pheochromocytoma (PC12) cells, implying these effects were not limited to amacrine neurons. These results suggest that BMAA is toxic for retina neurons and MGC and point to the involvement of NMDA receptors in amacrine cell death, providing new insight into the mechanisms involved in BMAA neurotoxic effects in the retina.

Expression of Mutant Ubiquitin and Proteostasis Impairment in Kii Amyotrophic Lateral Sclerosis/Parkinsonism-Dementia Complex Brains.

Kii amyotrophic lateral sclerosis/parkinsonism-dementia complex (ALS/PDC) is a progressive neurodegenerative disorder that is endemic to the Kii peninsula of Japan. The disorder is clinically characterized by a variable combination of parkinsonism, dementia, and motor neuron symptoms. Despite extensive investigations, the etiology and pathogenesis of ALS/PDC remain unclear. At the neuropathological level, Kii ALS/PDC is characterized by neuronal loss and tau-dominant polyproteinopathy. Here, we report the accumulation of several proteins involved in protein homeostasis pathways, that is, the ubiquitin-proteasome system and the autophagy-lysosome pathway, in postmortem brain tissue from a number of Kii ALS/PDC cases (n = 4). Of particular interest is the presence of a mutant ubiquitin protein (UBB+1), which is indicative of disrupted ubiquitin homeostasis. The findings suggest that abnormal protein aggregation is linked to impaired protein homeostasis pathways in Kii ALS/PDC.

A Locomotor Activity Study of the Effects of L‐Serine to Neurodegeneration Impacts of Beta‐Methylamino‐L‐alanine in Drosophila melanogaster

Amyotrophic Lateral Sclerosis‐Parkinsonism Dementia Complex (ALS‐PDC) is a neurodegenerative disease that described by loss of motor function, tremors, and dementia due to neurodegeneration. The disease is thought to be caused by Beta‐Methylamino‐L‐Alanine (BMAA), a neurotoxin from blue‐green algae. Since it has a structure that is similar to glutamate, BMAA act as an agonist to the glutamate receptor N‐methyl‐D‐aspartate which lead to excitotoxicity and degenerate neurons. L‐serine is an essential amino acid that has shown to interfere with the effect of BMAA in vivo studies. The purpose of this research is to further test the novel impact of L‐serine to BMAA‐induced fly in long‐term study. The interaction of BMAA and L‐serine was analyzed through circadian rhythm activity and viability of wild‐type Drosophila melanogaster in ALS‐PDC model. We used two Drosophila Activity Monitor (DAM) systems to measure activity of 64 fruit flies (Gender and Age‐Match). The experimental groups were fed with L‐serine, BMAA, BMAA+L‐serine and the control group was fed with standard fly agar. Fly activity was recorded by the number of fly pass the IR beam in the monitor for 10‐day (12L/12D at 22°C). After 10 days, circadian rhythm data showed BMAA flies increased activity over the first few days, and decreased gradually to the end of the period. BMAA‐induced flies (n = 12) does not show pattern of regular activity in L/D as compared to Control (n = 15) and L‐serine (n = 16). The co‐fed BMAA+L‐serine (n = 7) showed a similar pattern as the Control. For daytime vs nighttime activity (mean ± s.e.m), Control (n = 15) shown regular activity (day: 137 ± 13%, night: 21 ± 5%) as compared to BMAA (n = 12) that shown high activity in dark cycle (day: 162 ± 10%, night: 155 ± 21%). Moreover, the cofed L‐serine + BMAA (n = 7) exhibited normal daytime activity but not nighttime activity (day: 138 ± 14%, night: 167 ± 31%). The data from DAM shows the abnormal activity of flies in long term when inducing with BMAA such as abrupt sleep due to high activity during dark phase. L‐serine shows to reduce down the high activity when cofed with BMAA. This supports our hypothesis of L‐serine ameliorates the effects of BMAA in fly. This preliminary data is a starting point for further research in drug discovery for ALS‐PDC.Support or Funding InformationAcknowledgementsThis work was funded by the Fresno State Undergraduate Research Grant Award (2019–2020), and Howard P. Kubo, M.D. Award for Outstanding Achievement in Biomedical Science.

Research in Motor Neuron Diseases Caused by Natural Substances: Focus on Pathological Mechanisms of Neurolathyrism

Diseases of the motor-conducting system that cause moving disability affect socio-economic activity as well as human dignity. Neurolathyrism, konzo, and amyotrophic lateral sclerosis-parkinsonism-dementia complex (ALS-PDC) have attracted researchers to study the pathology of motor neuron (MN) diseases such as ALS. I have been studying neurolathyrism, which is caused by overconsumption of a legume grass pea (Lathyrys sativus L.). Among people who consume the legume as a food staple, many developed life-long paraparesis in their legs. β-N-oxalyl-l-α,β- diaminopropionic (l-β-ODAP; BOAA), contained in this plant, is a neurotoxic analog of l-glutamic acid. We have clarified that in addition to the causal involvement of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) type glutamatergic receptor in MN death, a toxic role of group I metabotropic glutamate receptors as well as transient receptor potential channels were involved in the MN insult by l-β-ODAP using primary MN culture. We have also established a neurolathyrism rat model by repeated, peripheral l-β-ODAP treatment to newborn rats under mild stress. Rats showing hind-leg paraparesis with an incidence rate of around 25% were useful to study the in vivo pathology of MN disease. MNs of these rats were greatly decreased at their lumbo/sacral segments at various ages. Intra-parenchymal hemorrhage was consistently observed in paraparetic rats but not in cripple-free, treated rats. MN were depleted even at an acute period around bleeding spots, suggesting catastrophic neuro-vascular-glial interaction in this MN disease. Summaries of konzo and ALS-PDCs studies are also introduced.