Repeated intraperitoneal injections of liposomes containing phosphatidic acid and cardiolipin reduce amyloid-β levels in APP/PS1 transgenic mice

© 2015 Elsevier Inc. The accumulation of extracellular amyloid-beta (Aβ) peptide and intracellular neurofibrillary tangles in the brain are two major neuropathological hallmarks of Alzheimer's disease (AD). It is thought that an equilibrium exists between Aβ in the brain and in the peripheral blood and thus, it was hypothesized that shifting this equilibrium towards the blood by enhancing peripheral clearance might reduce Aβ levels in the brain: the 'sink effect'. We tested this hypothesis by intraperitoneally injecting APP/PS1 transgenic mice with small unilamellar vesicles containing either phosphatidic acid or cardiolipin over 3. weeks. This treatment reduced significantly the amount of Aβ in the plasma and the brain levels of Aβ were lighter affected. Nevertheless, this dosing regimen did modulate tau phosphorylation and glycogen synthase kinase 3 activities in the brain, suggesting that the targeting of circulating Aβ may be therapeutically relevant in AD.CIBERNED (an initiative of ISCIII) and the Plan Nacional DGCYT (SAF2009-12249-C02-01) and by an Institutional grant from the 'Fundación Areces'Peer Reviewe

Peripheral amyloid levels present gender differences associated with aging in AβPP/PS1 mice

The accumulation of amyloid-β (Aβ) peptide is one of the major neuropathological hallmarks of Alzheimer's disease (AD). We have analyzed whether the progression of amyloidosis differentially affects males and females along aging in AβPP/PS1 transgenic mice. The levels of peripheral amyloid, Aβ40 and Aβ42, are not modified in either sex until 9 months of age. After that, however, there is an increase in amyloid levels in plasma among females and a decrease among males. These findings could be essential to design gender-specific strategies in other in vivo experiments or even in AD treatments. Supplementary Figure 1. A) Percentage of the cases of AβPP/PS1 transgenic mice (males and females) that show bladder disturbances. In dark affected individuals, in grey no evidence of any disturbance. Image of a healthy bladder (B) and a swollen bladder (C). D) Western blot analysis of some biochemical markers in the mice brains; 6 and 15 months, male, female, transgenic and wild type.This work was supported by grants from the Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED; an initiative of the ISCIII). In addition, work in FW’s lab was supported by grants from the “Plan Nacional”, “Dirección General de Ciencia y Tecnología -DGCYT-” SAF2012-39148-C03-01; CAMS2010/BMD-231-(2010-14) and EU-FP7-2009-CT222887, and an Institutional grant from the “Fundacion Areces”.Peer Reviewe

AMPK activation does not enhance autophagy in neurons in contrast to MTORC1 inhibition: different impact on β-amyloid clearance.

The physiological AKT-MTORC1 and AMPK signaling pathways are considered key nodes in the regulation of anabolism-catabolism, and particularly of macroautophagy/autophagy. Indeed, it is reported that these are altered processes in neurodegenerative proteinopathies such as Alzheimer disease (AD), mainly characterized by deposits of β-amyloid (Aβ) and hyperphosphorylated MAPT. These accumulations disrupt the optimal neuronal proteostasis, and hence, the recovery/enhancement of autophagy has been proposed as a therapeutic approach against these proteinopathies. The purpose of the present study was to characterize the modulation of autophagy by MTORC1 and AMPK signaling pathways in the highly specialized neurons, as well as their repercussions on Aβ production. Using a double transgenic mice model of AD, we demonstrated that MTORC1 inhibition, either in vivo or ex vivo (primary neuronal cultures), was able to reduce amyloid secretion through moderate autophagy induction in neurons. The pharmacological prevention of autophagy in neurons augmented the Aβ secretion and reversed the effect of rapamycin, confirming the anti-amyloidogenic effects of autophagy in neurons. Inhibition of AMPK with compound C generated the expected decrease in autophagy induction, though surprisingly did not increase the Aβ secretion. In contrast, increased activity of AMPK with metformin, AICAR, 2DG, or by gene overexpression did not enhance autophagy but had different effects on Aβ secretion: whereas metformin and 2DG diminished the secreted Aβ levels, AICAR and PRKAA1/AMPK gene overexpression increased them. We conclude that AMPK has a significantly different role in primary neurons than in other reported cells, lacking a direct effect on autophagy-dependent amyloidosis.pre-print832 K

Males vs females: differences in the AB accumulation

Background: The accumulation of extracellular amyloid-beta (Aß) peptide and intracellular neurofibrillary tangles in the brain are two major neuropathological hallmarks of Alzheimer's disease (AD). For the analysis of Aß-peptide aggregation, different mouse models (single or double transgenic mice) have been used to follow the evolution of AD-amyloidosis, and to test potential treatments. So far, cerebellum tissue has not been deeply analyzed to check the amyloidosis in these transgenic models. Besides, sex influence hasn't been systematically studied in these models, even it has been described important gender differences in the evolution of AD in human population. We have checked whether the progression of amyloidosis in a double transgenic mouse, APP/PS1, is susceptible to aging and differentially affects males and females. Methods: Aß levels were measured by ELISA in plasma and tissue samples. Cortex and cerebellum tissue of transgenic males and females from 6 to 15 months of age were processed to be analyzed. In addition, fixed hemibrains brain were coronally sectioned and used to perform immunohistochemistry and immunofluorescence studies.Peer reviewe

Typology Selection of Retaining Walls Based on Multicriteria Decision-Making Methods

This research received no external funding.In civil engineering and construction, in the selection of the most adequate and sustainable alternative, all of the alternatives and selection criteria, such as the requirements of the construction process (which are often overlooked) and the preferences of designers, clients, or contractors, are not always taken into account. The purpose of this article is to suggest a methodology that may allow studying all of the possible alternatives to find the most ideal solution among all of the existing possibilities for the selection of retaining walls to be built in infrastructures in different environments. For this purpose, all typologies of retaining walls and selection criteria (external requirements, construction requirements, characteristics of the natural land and economic criteria) are first identified. Subsequently, a simple methodological method is proposed, allowing the relative importance of each criterion to be established and allowing us to select the most suitable solution for each situation by successively applying different multicriteria decision-making methods. Finally, the methodology developed is applied to two projects in different locations with different constraints. The results obtained provide a set of compromise solutions that remain as best-rank alternatives when the weights of the criteria change. Therefore, the methodology developed can be applied to the selection of typologies of other structures in future projects

Comparison of ultrafiltration and dissolved air flotation efficiencies in industrial units during the papermaking process

The efficiency of an ultrafiltration unit has been studied and compared with a dissolved air flotation system to get water with a suited quality to be reused in the process. The study was done at a paper mill producing light weight coated paper and newsprint paper from 100% recovered paper. Efficiency was analysed by removal of turbidity, cationic demand, total and dissolved chemical oxygen demand, hardness, sulphates and microstickies. Moreover, the performance of the ultrafiltration unit and the membranes were studied deeply, analysing its variability during the filtration process. As expected, the ultrafiltration gave higher removal efficiencies than the dissolved air flotation cell in parameters like turbidity, cationic demand, dissolved chemical oxygen demand and microstickies. The greatest difference in performance between the units concerned cationic demand and dissolved chemical oxygen demand. Ultrafiltration was influenced by the operating time, decreasing the removal efficiency of the dissolved fraction by 75% and of the colloidal fraction by 30% after 312 of running. Membrane autopsy, carried out to identify the cause of poor membrane performance, showed that the active layer was degraded due to the effect of suspended solids

Uncertainty analysis methods to select the optimal alternative in the design of parking facilities

The selection of the preferred alternative in a parking facility project is usually made in a state of uncertainty. Decision-making methods are a useful tool to systematically arrive at a final decision between different alternatives and reduce subjectivity in decision making by creating a series of filters. However, the selection of the appropriate variables to be considered in the analysis may be problematic as well. Performing sensitivity analyses on entry variables is a key feature to ensure that the final choice is stable when initial conditions experience changes. This paper suggests a methodology to select the best alternative when considering parking facilities. The methodology compares the results from two different sensitivity analyses techniques. The changes in preference experienced as the applied weights change through the process are analyzed and the most critical criteria are identified

Characterisation of the ex vivo virulence of Leishmania infantum isolates from Phlebotomus perniciosus from an outbreak of human leishmaniosis in Madrid, Spain

BACKGROUND: Since mid 2009, an outbreak of human leishmaniosis in Madrid, Spain, has involved more than 560 clinical cases. Many of the cases occurred in people who live in areas around a newly constructed green park (BosqueSur). This periurban park provides a suitable habitat for sand flies (the vectors of Leishmania infantum). Indeed, studies of blood meals from sand flies captured in the area showed a strong association between the insect vector, hares or rabbits, and humans in the area. Interestingly, up to 70% of cases have been found in immunocompetent patients (aged between 46-60 years). This study was designed to evaluate the ex vivo virulence of the L. infantum isolates from Phlebotomus perniciosus captured in this area of Madrid. METHODS: Murine macrophages and dendritic cells were infected ex vivo with L. infantum strain BCN150, isolate BOS1FL1, or isolate POL2FL7. At different times after infection, the infection indices, cytokine production (IL-12p40 and IL-10), NO release and arginase activities were evaluated. RESULTS: Using an ex vivo model of infection in murine bone marrow-derived cells, we found that infection with isolates BOS1FL1 and POL2FL7 undermined host immune defence mechanisms in multiple ways. The main factors identified were changes in both the balance of iNOS versus arginase activities and the equilibrium between the production of IL-12 and IL-10. Infection with isolates BOS1FL1 and POL2FL7 also resulted in higher infection rates compared to the BCN150 strain. Infection index values at 24 h were as follows: BCN150-infected cells, 110 for infected MØ and 115 for infected DC; BOS1FL1-infected cells, 300 for infected MØ and 247 for infected DC; and POL2FL7-infected cells, 275 for infected MØ and 292 for infected DC. CONCLUSIONS: Our data indicate that L. infantum isolates captured from this endemic area exhibited high virulence in terms of infection index, cytokine production and enzymatic activities involved in the pathogenesis of visceral leishmaniosis. Altogether, these data provide a starting point for the study of the virulence behaviour of parasites (BOS1FL1 and POL2FL7) isolated from P. perniciosus during the outbreak of human leishmaniosis in Madrid, Spain, and their involvement in infecting immunocompetent hosts.This study was supported by Grants AGL2010-17394 and AGL2013-44100R from the Spanish Ministry of Economy and Competitiveness and was partially funded by EU grant FP7-2011-261504 EDENext and the paper is catalogued by the EDENext Steering Committee as EDENext 276 (http://www.edenext.eu).S

RAB7L1-Mediated Relocalization of LRRK2 to the Golgi Complex Causes Centrosomal Deficits via RAB8A

Mutations in the LRRK2 gene cause autosomal-dominant Parkinson’s disease (PD), and both LRRK2 as well as RAB7L1 have been implicated in increased susceptibility to idiopathic PD. RAB7L1 has been shown to increase membrane-association and kinase activity of LRRK2, and both seem to be mechanistically implicated in the same pathway. Another RAB protein, RAB8A, has been identified as a prominent LRRK2 kinase substrate, and our recent work demonstrates that aberrant LRRK2-mediated phosphorylation of RAB8A leads to centrosomal alterations. Here, we show that RAB7L1 recruits LRRK2 to the Golgi complex, which causes accumulation of phosphorylated RAB8A in a pericentrosomal/centrosomal location as well as centrosomal deficits identical to those observed with pathogenic LRRK2. The centrosomal alterations induced by wildtype LRRK2 in the presence of RAB7L1 depend on Golgi integrity. This is in contrast to pathogenic LRRK2 mutants, which cause centrosomal deficits independent of Golgi integrity or largely independent on RAB7L1 expression. Furthermore, centrosomal alterations in the presence of wildtype LRRK2 and RAB7L1 are at least in part mediated by aberrant LRRK2-mediated RAB8A phosphorylation, as abolished by kinase inhibitors and reduced upon knockdown of RAB8A. These results indicate that pathogenic LRRK2, as well as increased levels of RAB7L1, cause centrosomal deficits in a manner dependent on aberrant RAB8A phosphorylation and centrosomal/pericentrosomal accumulation, suggesting that centrosomal cohesion deficits may comprise a useful cellular readout for a broader spectrum of the disease

Pathogenic LRRK2 regulates centrosome cohesion via Rab10/RILPL1-mediated CDK5RAP2 displacement

Mutations in LRRK2 increase its kinase activity and cause Parkinson's disease. LRRK2 phosphorylates a subset of Rab proteins which allows for their binding to RILPL1. The phospho-Rab/RILPL1 interaction causes deficits in ciliogenesis and interferes with the cohesion of duplicated centrosomes. We show here that centrosomal deficits mediated by pathogenic LRRK2 can also be observed in patient-derived iPS cells, and we have used transiently transfected cell lines to identify the underlying mechanism. The LRRK2-mediated centrosomal cohesion deficits are dependent on both the GTP conformation and phosphorylation status of the Rab proteins. Pathogenic LRRK2 does not displace proteinaceous linker proteins which hold duplicated centrosomes together, but causes the centrosomal displacement of CDK5RAP2, a protein critical for centrosome cohesion. The LRRK2-mediated centrosomal displacement of CDK5RAP2 requires RILPL1 and phospho-Rab proteins, which stably associate with centrosomes. These data provide fundamental information as to how pathogenic LRRK2 alters the normal physiology of a cell.We are grateful to Erich Nigg and Francis Barr for providing a variety of constructs and antibodies, and to Dario Alessi for providing various A549 cell lines and MEF cells. We thank LauraMontosa for excellent technical assistance with confocal microscopy. This work was supported by The Michael J. Fox Foundation for Parkinson's research (to S.H.), intramural funding from Rutgers University (to S.H.), the Spanish Ministry of Economy and Competitiveness (SAF2017-89402-R to S.H.), the BBVA Foundation (to S.H., S.A.C., and R.W. M.), the Spanish Ministry of Education, Culture and Sport (FPU12/04367 to J.M. P., FPU15/05233 to A.J. L.O.), and the Spanish Ministry of Science, Innovation and Universities (EST18/00412 to A.J.L. O.)