Mitochondrial axonal transport in vivo: development of a new assay in zebrafish to study the effects of Familial Alzheimerâs Disease Presenilin 2 mutations

Alzheimer's disease (AD) is the most common form of neurodegenerative disease, affecting more than 35 million people worldwide. Although the majority of cases is caused by a combination of risk factors (genetic and environmental), a small percentage is genetically inherited and is called familial Alzheimer's disease (FAD). FAD is caused by mutations in three genes: APP, which encodes for the amyloid precursor protein, and presenilins genes (PSEN1 and PSEN2), encoding for presenilin 1 and 2 (PS1, PS2), respectively. Mutations in these genes result in an increased formation of the product of APP cleavage, the amyloid Aβ peptide: in AD the levels of Aβ42, the most aggregation-prone, 42 amino acids long, form of Aβ, are increased, leading to the formation of amyloid plaques in patients' brains. According to the "amyloid hypothesis", Aβ accumulation triggers a cascade that leads to progressive synaptic and neuronal injury and eventually to cell death. In this scenario, presenilins play a key role. Presenilins are membrane spanning proteins known to most as key components of the γ-secretase, the enzymatic complex located in plasma membrane and internal membranes (Golgi and endoplasmic reticulum, ER) responsible for the cleavage of many proteins, including NOTCH and APP. However, they also have γ-secretase-independent functions: in particular PS2, but not PS1, has been recently described to be able to modulate ER-mitochondria tethering and their Ca2+ cross-talk. The overexpression of FAD-linked PS2 mutants increases both the tethering and the Ca2+ transfer between these two organelles. Itâs worth mentioning that alterations in Ca2+ signalling are emerging as an attractive hypothesis to explain AD pathogenesis: many cellular processes are dependent on a proper Ca2+ signalling, and any dysfunction is expected to have consequences on neuronal physiology. Among these processes, mitochondrial axonal transport represents a very interesting feature: mitochondria are key organelles for cells, whose functions influence several cell life aspects as well as cell death. Mitochondrial axonal transport is finely regulated by Ca2+: increases in cytosolic Ca2+ levels arrest mitochondrial movements along axons, to ensure their deliver to metabolically active sites, such as activated synapses. The previously described FAD-PS2 mediated alterations in Ca2+ handling and in the physical interaction between ER and mitochondria could potentially modify axonal mitochondrial transport, and an altered mitochondrial distribution has been reported within neuronal cells in different AD models. Zebrafish has emerged as an attractive vertebrate system to study human diseases: in particular, its genome is extensively annotated and orthologs of genes involved in FAD have been already identified. Mitochondrial axonal transport analysis is strongly facilitated by the transparency of embryos and by the availability of tools to manipulate gene expression. We here generated a method to study PS2-dependent effects on mitochondrial dynamics in vivo. Firstly, by genetically targeting the fluorescent protein Kaede to mitochondria in Rohon-Beard (RB) sensory neurons, we have characterized mitochondrial axonal transport properties during development. We found that the percentage of mobile mitochondria is reduced from one to three days post fertilization (dpf) embryos; mitochondrial average speed, however, increases during development. The directionality of transport is strongly biased towards the anterograde one, reflecting a net translocation of mitochondria from the cell body to peripheral arbors. Nocodazole, a strong inhibitor of microtubule polymerization, totally disrupts mitochondrial axonal transport, validating the sensitivity of our assay. Interestingly, the mitochondrial axonal transport parameters, measured with our approach, are similar to those reported in recent in vivo studies. We found that psen2 knockdown (KD), but not psen1 KD, reduces the percentage of motile organelles, compared to controls. This phenotype is recovered by rescue experiments, in which PS2 was re-introduced, demonstrating a specific PS2-dependent effect. On the other side, human PS2 expression in zebrafish leads to different results: human PS2 (both wild-type, wt, and mutated) is expressed at the level of intracellular membranes (mainly in the ER) in the soma and through the axon of RB neurons but, while overexpression of wt PS2 does not affect mitochondrial axonal transport, that of the PS2-T122R mutant increases the percentage of motile mitochondria. Moreover, we verified whether intra-mitochondrial Ca2+ rises could be involved as signal in the regulation of mitochondrial axonal transport, as previously proposed: the KD of the mitochondrial calcium uniporter (Mcu), the protein responsible for mitochondrial Ca2+ uptake, however, is without effect on either the speed and the percentage of motile mitochondria in RB neurons. Mitochondrial morphology and density were also investigated and both features resulted unchanged upon the different treatments, suggesting that the observed alterations in mitochondrial axonal transport induced by PS2 are due to neither an imbalance in mitochondrial fusion and fission nor an abnormal mitochondrial biogenesis. Further investigations are required to better understand the mechanism through which PS2 alters mitochondrial axonal transport, as well as the role played by this alteration in AD pathogenesis

SPLICS: a split green fluorescent protein-based contact site sensor for narrow and wide heterotypic organelle juxtaposition

Contact sites are discrete areas of organelle proximity that coordinate essential physiological processes across membranes, including Ca2+ signaling, lipid biosynthesis, apoptosis, and autophagy. However, tools to easily image inter-organelle proximity over a range of distances in living cells and in vivo are lacking. Here we report a split-GFP-based contact site sensor (SPLICS) engineered to fluoresce when organelles are in proximity. Two SPLICS versions efficiently measured narrow (8\u201310 nm) and wide (40\u201350 nm) juxtapositions between endoplasmic reticulum and mitochondria, documenting the existence of at least two types of contact sites in human cells. Narrow and wide ER\u2013mitochondria contact sites responded differently to starvation, ER stress, mitochondrial shape modifications, and changes in the levels of modulators of ER\u2013mitochondria juxtaposition. SPLICS detected contact sites in soma and axons of D. rerio Rohon Beard (RB) sensory neurons in vivo, extending its use to analyses of organelle juxtaposition in the whole anim

Tau localises within mitochondrial sub-compartments and its caspase cleavage affects ER-mitochondria interactions and cellular Ca2+ handling

Intracellular neurofibrillary tangles (NFT) composed by tau and extracellular amyloid beta (A\u3b2) plaques accumulate in Alzheimer's disease (AD) and contribute to neuronal dysfunction. Mitochondrial dysfunction and neurodegeneration are increasingly considered two faces of the same coin and an early pathological event in AD. Compelling evidence indicates that tau and mitochondria are closely linked and suggests that tau-dependent modulation of mitochondrial functions might be a trigger for the neurodegeneration process; however, whether this occurs either directly or indirectly is not clear. Furthermore, whether tau influences cellular Ca2+ handling and ER-mitochondria cross-talk is yet to be explored. Here, by focusing on wt tau, either full-length (2N4R) or the caspase 3-cleaved form truncated at the C-terminus (2N4R\u394C20), we examined the above-mentioned aspects. Using new genetically encoded split-GFP-based tools and organelle-targeted aequorin probes, we assessed: i) tau distribution within the mitochondrial sub-compartments; ii) the effect of tau on the short- (8-10\u202fnm) and the long- (40-50\u202fnm) range ER-mitochondria interactions; and iii) the effect of tau on cytosolic, ER and mitochondrial Ca2+ homeostasis. Our results indicate that a fraction of tau is found at the outer mitochondrial membrane (OMM) and within the inner mitochondrial space (IMS), suggesting a potential tau-dependent regulation of mitochondrial functions. The ER Ca2+ content and the short-range ER-mitochondria interactions were selectively affected by the expression of the caspase 3-cleaved 2N4R\u394C20 tau, indicating that Ca2+ mis-handling and defects in the ER-mitochondria communications might be an important pathological event in tau-related dysfunction and thereby contributing to neurodegeneration. Finally, our data provide new insights into the molecular mechanisms underlying tauopathies

Kajian Perbedaan Konsentrasi Larutan Garam Pada Perendaman Rgh Dan Vaksin Terhadap Kelulushidupan Dan Pertumbuhan Benih Ikan Lele Sangkuriang (Clarias Gariepinus)

Berdasarkan data KKP (2013), pencapaian produksi ikan lele pada tahun 2013 mampu melampaui target. Ikan lele merupakan ikan yang mudah dibudidayakan, sehingga banyak dilakukan penelitian agar didapatkan benih lele dengan pertumbuhan dan kelulushidupan yang lebih baik, serta tahan terhadap serangan penyakit. Penelitian ini bertujuan untuk mengetahui pengaruh konsentrasi larutan garam yang berbeda dan konsentrasi yang terbaik pada perendaman rGH dan vaksin terhadap kelulushidupan dan pertumbuhan benih lele sangkuriang. Penelitian ini dilaksanakan pada tanggal 13 November 2014 – 15 Februari 2015 di Satuan Kerja Pembenihan dan Budidaya Ikan Air Tawar (SATKER PBIAT), Siwarak, Ungaran, Semarang. Ikan uji yang digunakan adalah benih lele sangkuriang umur 12 hari. Metode yang digunakan adalah eksperimental dengan rancangan acak lengkap (RAL) 4 perlakuan dan 3 ulangan, yaitu: (A) perlakuan tanpa larutan garam, (B) konsentrasi 0,5%, (C) konsentrasi 1,0% dan (D) konsentrasi 1,5%. Pemeliharaan ikan dilakukan selama 42 hari. Variabel data yang diamati meliputi kelulushidupan, SGR, panjang mutlak, FCR, EPP dan kelulushidupan setelah uji tantang. Analisa data dengan menggunakan anova untuk mengetahui pengaruh perlakuan yang berbeda nyata, apabila hasil yang didapatkan berbeda nyata, maka dilanjutkan dengan uji duncan untuk mengetahui perlakuan yang terbaik. Hasil penelitian menunjukkan perlakuan terbaik adalah perlakuan C, dengan nilai kelulushidupan (87,00±1,00%), SGR (7,79±0,03%), nilai panjang mutlak (6,75±0,15cm), nilai FCR (0,71±0,01) dan nilai EPP (140,28±1,25%), sedangkan untuk kelulushidupan setelah uji tantang didapatkan hasil yang tidak berbeda nyata, dimana: A (93,33±5,77%), B (96,67±5,77%), C (96,67±5,77%) dan D (96,67±5,77%). Kesimpulan dari penelitian ini ialah pemberian konsentrasi larutan garam yang berbeda berpengaruh sangat nyata terhadap kelulushidupan, SGR, panjang mutlak, FCR dan EPP, namun tidak berpengaruh nyata terhadap kelulushidupan setelah uji tantang. Konsentrasi larutan garam terbaik pada penelitian ini adalah 1,0%. Based on data from KKP (2013), the achievement of the production of catfish in 2013 was able of exceeding the target. Catfish is a fish that easily cultivated, so a lot of research done to get catfish\u27s seed with better survival and growth, and it can resistant to attack of deseases. This research was aimed to find out the effect of different salt solution concentrations and the best concentration from immersion of rGH and vaccine for survival rate and growth of sangkuriang catfish\u27s seed. This research was conducted on November 13th, 2014 – February 15th, 2015 at Satuan Kerja Pembenihan dan Budidaya Ikan Air Tawar (SATKER PBIAT), Siwarak, Ungaran, Semarang. The fish that used for this research is sangkuriang catfish\u27s seed aged 12 days. Completely randomized design (CRD) was used in this research with four treatments and three replication, which treatments are: (A) without salt solution concentration, (B) Using concentration 0.5%, (C) Concentration 1.0% and (D) Concentration 1.5%. The Fishes are maintained for 42 days. Observational variable are survival rate, SGR, absolute length, FCR, EPP and survival rate after challenge test. Data analysis using anova to know the effect of treatment is significantly different, if the result is significantly different, then continue with duncan test to know the best treatment. The best result is treatment C, with survival rate (87.00±1.00%), SGR (7.79±0.03%), absolute length (6.75±0.15cm), FCR (0.71±0.01), EPP (140.28±1.25%), and for survival rate after challenge test are not significantly different, where: treatment A (93.33±5.77%), B (96.67±5.77%), C (96.67±5.77%) and D (96.67±5.77%). The conclusion of this research is giving of salt solution with different concentration take significantly effect for survival rate, SGR, absolute length, FCR and EPP, but did not take significantly effect for survival rate after challenge test. The best salt solution concentration is 1.0%

Não Ver, (Não) Ser Visto, Per-Tecer: narrativas de alunos com deficiência visual na universidade

El objetivo de este artículo es conocer las narrativas de alumnos con deficiencia visual en una universidad pública de Goiás, para discutir su experiencia y los procesos de inclusión-exclusión. Como método de investigación realizamos una sesión de grupo operativo, conforme la teoría y técnica del psicólogo social argentino Enrique Pichon-Rivière. Como resultados, desarrollamos tres categorías de análisis: no ver, (no) ser visto y per-tejer. En la primera categoría citamos las principales vicisitudes relativas a lo no ver en la Universidad, que se refieren principalmente a las barreras arquitectónicas y la dificultad de accesibilidad al material didáctico y procedimientos de enseñanza de los profesores. En la segunda categoría discutimos la invisibilidad del estudiante con deficiencia visual en la Universidad, que pasa a no ser visto por sus colegas. En la última, discutimos las tramas que deben ser realizadas entre videntes y no videntes para que la inclusión pueda efectivamente ser construida en la Universidad.O objetivo deste artigo é conhecer as narrativas de alunos com deficiência visual em uma universidade pública de Goiás, para discutir sua experiência e os processos de inclusão-exclusão. Como método de investigação realizamos uma sessão de grupo operativo, conforme a teoria e técnica do psicólogo social argentino Enrique Pichon-Rivière. Como resultados, elaboramos três categorias de análise: não ver, (não) ser visto e per-tecer. Na primeira categoria elencamos os principais percalços relacionados ao não ver na Universidade, que se referem principalmente às barreiras arquitetônicas e a dificuldade de acessibilidade ao material didático e procedimentos de ensino dos professores. Na segunda categoria, (não) ser visto, discutimos a invisibilidade do estudante com deficiência visual na Universidade, que passa a não ser visto por seus colegas, bem como por alguns professores. Na última categoria, per-tecer, discutimos as tessituras que devem ser realizadas entre videntes e não videntes para que a inclusão possa efetivamente ser construída na Universidade.Le but de cet article est de connaître les récits des étudiants ayant une déficience visuelle dans une université publique de Goiás, afin de discuter de leur expérience et des processus d'inclusion-exclusion. Comme méthode de recherche, nous avons réalisé une session de groupe opératif, selon la théorie et la technique du psychologue social argentin Enrique Pichon-Rivière. En conséquence, nous avons développé trois catégories d'analyse: ne pas voir, (ne pas) être vu et per-tecter. Dans la première catégorie, nous citons les principales vicissitudes liées au fait de ne pas voir à l'Université, qui se réfèrent principalement aux barrières architecturales et à la difficulté d'accès au matériel didactique et aux procédures d'enseignement des professeurs. Dans la deuxième catégorie, nous abordons l'invisibilité de l'étudiant malvoyant à l'université, qui n'est pas vu par ses collègues. Dans la dernière catégorie, nous discutons des tissages qui doivent être faits entre les étudiants voyants et non voyants pour que l'inclusion puisse être effectivement construite à l'Université.The aim of this article is to know the narratives of students with visual impairment in a public university in Goiás, to discuss their experience and the inclusion-exclusion processes. As a research method, we carried out an operative group session, according to the theory and technique of the Argentine social psychologist Enrique Pichon-Rivière. As a result, we developed three categories of analysis: not seeing, (not) be seeing and per-weaving. In the first category, we list the main difficulties related to not see at the University, which mainly refer to architectural barriers and the difficulty of accessing teaching material and procedures of professors. In the second category, we discuss the invisibility of the visually impaired student at the University, who is not seen by colleagues. In the last one, we discuss the weave that must be performed between sighted and non-sighted so that inclusion can be effectively built at the University

Search for heavy resonances decaying to two Higgs bosons in final states containing four b quarks

A search is presented for narrow heavy resonances X decaying into pairs of Higgs bosons (H) in proton-proton collisions collected by the CMS experiment at the LHC at root s = 8 TeV. The data correspond to an integrated luminosity of 19.7 fb(-1). The search considers HH resonances with masses between 1 and 3 TeV, having final states of two b quark pairs. Each Higgs boson is produced with large momentum, and the hadronization products of the pair of b quarks can usually be reconstructed as single large jets. The background from multijet and t (t) over bar events is significantly reduced by applying requirements related to the flavor of the jet, its mass, and its substructure. The signal would be identified as a peak on top of the dijet invariant mass spectrum of the remaining background events. No evidence is observed for such a signal. Upper limits obtained at 95 confidence level for the product of the production cross section and branching fraction sigma(gg -> X) B(X -> HH -> b (b) over barb (b) over bar) range from 10 to 1.5 fb for the mass of X from 1.15 to 2.0 TeV, significantly extending previous searches. For a warped extra dimension theory with amass scale Lambda(R) = 1 TeV, the data exclude radion scalar masses between 1.15 and 1.55 TeV

Search for neutral long-lived particles in pp collisions at √s = 13 TeV that decay into displaced hadronic jets in the ATLAS calorimeter

A search for decays of pair-produced neutral long-lived particles (LLPs) is presented using 139 fb−1 of proton-proton collision data collected by the ATLAS detector at the LHC in 2015–2018 at a centre-of-mass energy of 13 TeV. Dedicated techniques were developed for the reconstruction of displaced jets produced by LLPs decaying hadronically in the ATLAS hadronic calorimeter. Two search regions are defined for different LLP kinematic regimes. The observed numbers of events are consistent with the expected background, and limits for several benchmark signals are determined. For a SM Higgs boson with a mass of 125 GeV, branching ratios above 10% are excluded at 95% confidence level for values of c times LLP mean proper lifetime in the range between 20 mm and 10 m depending on the model. Upper limits are also set on the cross-section times branching ratio for scalars with a mass of 60 GeV and for masses between 200 GeV and 1 TeV. [Figure not available: see fulltext.

Measurement and interpretation of same-sign W boson pair production in association with two jets in pp collisions at s = 13 TeV with the ATLAS detector

This paper presents the measurement of fducial and diferential cross sections for both the inclusive and electroweak production of a same-sign W-boson pair in association with two jets (W±W±jj) using 139 fb−1 of proton-proton collision data recorded at a centre-of-mass energy of √s = 13 TeV by the ATLAS detector at the Large Hadron Collider. The analysis is performed by selecting two same-charge leptons, electron or muon, and at least two jets with large invariant mass and a large rapidity diference. The measured fducial cross sections for electroweak and inclusive W±W±jj production are 2.92 ± 0.22 (stat.) ± 0.19 (syst.)fb and 3.38±0.22 (stat.)±0.19 (syst.)fb, respectively, in agreement with Standard Model predictions. The measurements are used to constrain anomalous quartic gauge couplings by extracting 95% confdence level intervals on dimension-8 operators. A search for doubly charged Higgs bosons H±± that are produced in vector-boson fusion processes and decay into a same-sign W boson pair is performed. The largest deviation from the Standard Model occurs for an H±± mass near 450 GeV, with a global signifcance of 2.5 standard deviations

Search for pair production of squarks or gluinos decaying via sleptons or weak bosons in final states with two same-sign or three leptons with the ATLAS detector

A search for pair production of squarks or gluinos decaying via sleptons or weak bosons is reported. The search targets a final state with exactly two leptons with same-sign electric charge or at least three leptons without any charge requirement. The analysed data set corresponds to an integrated luminosity of 139 fb−1 of proton-proton collisions collected at a centre-of-mass energy of 13 TeV with the ATLAS detector at the LHC. Multiple signal regions are defined, targeting several SUSY simplified models yielding the desired final states. A single control region is used to constrain the normalisation of the WZ + jets background. No significant excess of events over the Standard Model expectation is observed. The results are interpreted in the context of several supersymmetric models featuring R-parity conservation or R-parity violation, yielding exclusion limits surpassing those from previous searches. In models considering gluino (squark) pair production, gluino (squark) masses up to 2.2 (1.7) TeV are excluded at 95% confidence level

Studies of new Higgs boson interactions through nonresonant HH production in the b¯bγγ fnal state in pp collisions at √s = 13 TeV with the ATLAS detector

A search for nonresonant Higgs boson pair production in the b ¯bγγ fnal state is performed using 140 fb−1 of proton-proton collisions at a centre-of-mass energy of 13 TeV recorded by the ATLAS detector at the CERN Large Hadron Collider. This analysis supersedes and expands upon the previous nonresonant ATLAS results in this fnal state based on the same data sample. The analysis strategy is optimised to probe anomalous values not only of the Higgs (H) boson self-coupling modifer κλ but also of the quartic HHV V (V = W, Z) coupling modifer κ2V . No signifcant excess above the expected background from Standard Model processes is observed. An observed upper limit µHH < 4.0 is set at 95% confdence level on the Higgs boson pair production cross-section normalised to its Standard Model prediction. The 95% confdence intervals for the coupling modifers are −1.4 < κλ < 6.9 and −0.5 < κ2V < 2.7, assuming all other Higgs boson couplings except the one under study are fxed to the Standard Model predictions. The results are interpreted in the Standard Model efective feld theory and Higgs efective feld theory frameworks in terms of constraints on the couplings of anomalous Higgs boson (self-)interactions