Modular Acquisition and Stimulation System for Timestamp-Driven Neuroscience Experiments

Dedicated systems are fundamental for neuroscience experimental protocols that require timing determinism and synchronous stimuli generation. We developed a data acquisition and stimuli generator system for neuroscience research, optimized for recording timestamps from up to 6 spiking neurons and entirely specified in a high-level Hardware Description Language (HDL). Despite the logic complexity penalty of synthesizing from such a language, it was possible to implement our design in a low-cost small reconfigurable device. Under a modular framework, we explored two different memory arbitration schemes for our system, evaluating both their logic element usage and resilience to input activity bursts. One of them was designed with a decoupled and latency insensitive approach, allowing for easier code reuse, while the other adopted a centralized scheme, constructed specifically for our application. The usage of a high-level HDL allowed straightforward and stepwise code modifications to transform one architecture into the other. The achieved modularity is very useful for rapidly prototyping novel electronic instrumentation systems tailored to scientific research.Comment: Preprint submitted to ARC 2015. Extended: 16 pages, 10 figures. The final publication is available at link.springer.co

Pattern matching through Chaos Game Representation: bridging numerical and discrete data structures for biological sequence analysis

BACKGROUND: Chaos Game Representation (CGR) is an iterated function that bijectively maps discrete sequences into a continuous domain. As a result, discrete sequences can be object of statistical and topological analyses otherwise reserved to numerical systems. Characteristically, CGR coordinates of substrings sharing an L-long suffix will be located within 2(-L )distance of each other. In the two decades since its original proposal, CGR has been generalized beyond its original focus on genomic sequences and has been successfully applied to a wide range of problems in bioinformatics. This report explores the possibility that it can be further extended to approach algorithms that rely on discrete, graph-based representations. RESULTS: The exploratory analysis described here consisted of selecting foundational string problems and refactoring them using CGR-based algorithms. We found that CGR can take the role of suffix trees and emulate sophisticated string algorithms, efficiently solving exact and approximate string matching problems such as finding all palindromes and tandem repeats, and matching with mismatches. The common feature of these problems is that they use longest common extension (LCE) queries as subtasks of their procedures, which we show to have a constant time solution with CGR. Additionally, we show that CGR can be used as a rolling hash function within the Rabin-Karp algorithm. CONCLUSIONS: The analysis of biological sequences relies on algorithmic foundations facing mounting challenges, both logistic (performance) and analytical (lack of unifying mathematical framework). CGR is found to provide the latter and to promise the former: graph-based data structures for sequence analysis operations are entailed by numerical-based data structures produced by CGR maps, providing a unifying analytical framework for a diversity of pattern matching problems

Epigallocathechin-O-3-Gallate Inhibits Trypanothione Reductase of Leishmania infantum, Causing Alterations in Redox Balance and Leading to Parasite Death

Leishmania infantum is a protozoan parasite that causes a vector borne infectious disease in humans known as visceral leishmaniasis (VL). This pathology, also caused by L. donovani, presently impacts the health of 500,000 people worldwide, and is treated with outdated anti-parasitic drugs that suffer from poor treatment regimens, severe side effects, high cost and/or emergence of resistant parasites. In previous works we have disclosed the anti-Leishmania activity of (-)-Epigallocatechin 3-O-gallate (EGCG), a flavonoid compound present in green tea leaves. To date, the mechanism of action of EGCG against Leishmania remains unknown. This work aims to shed new light into the leishmanicidal mode of action of EGCG. Towards this goal, we first confirmed that EGCG inhibits L. infantum promastigote proliferation in a concentration-dependent manner. Second, we established that the leishmanicidal effect of EGCG was associated with i) mitochondria depolarization and ii) decreased concentration of intracellular ATP, and iii) increased concentration of intracellular H2O2. Third, we found that the leishmanicidal effect and the elevated H2O2 levels induced by of EGCG can be abolished by PEG-catalase, strongly suggesting that this flavonoid kills L. infantum promastigotes by disturbing their intracellular redox balance. Finally, we gathered in silico and in vitro evidence that EGCG binds to trypanothione reductase (TR), a central enzyme of the redox homeostasis of Leishmania, acting as a competitive inhibitor of its trypanothione substrate.This work was supported by Fundação Carlos Chagas Filho de Amparo a Pesquisa do Estado do Rio de Janeiro (FAPERJ), Conselho Nacional de Desenvolvimento Cientı́fico e Tecnológico (CNPq), Programa Estratégico de Apoio a Pesquisa em Saúde (PAPES/FIOCRUZ);, and Fundação Oswaldo Cruz (FIOCRUZ). EA-A is the ecipient of a research scholarship from Conselho Nacional de Desenvolvimento Cientı́fico e Tecnológico (CNPq). JI was supported by a postdoctoral fellowship from Conselho Nacional de Desenvolvimento Cientı́fico e Tecnológico (CNPq). JI is a recipient of a postdoctoral fellowship from Fundação Oswaldo Cruz (FIOCRUZ). HC is funded by Fundação para a Ciência e Tecnologia (FCT, Portugal) through the “Investigador FCT” contract IF/01244/2015

The Schistosoma mansoni genome encodes thousands of long non-coding RNAs predicted to be functional at different parasite life-cycle stages

Next Generation Sequencing (NGS) strategies, like RNA-Seq, have revealed the transcription of a wide variety of long non-coding RNAs (lncRNAs) in the genomes of several organisms. In the present work we assessed the lncRNAs complement of Schistosoma mansoni, the blood fluke that causes schistosomiasis, ranked among the most prevalent parasitic diseases worldwide. We focused on the long intergenic/intervening ncRNAs (lincRNAs), hidden within the large amount of information obtained through RNA-Seq in S. mansoni (88 libraries). Our computational pipeline identified 7029 canonically-spliced putative lincRNA genes on 2596 genomic loci (at an average 2.7 isoforms per lincRNA locus), as well as 402 spliced lncRNAs that are antisense to protein-coding (PC) genes. Hundreds of lincRNAs showed traits for being functional, such as the presence of epigenetic marks at their transcription start sites, evolutionary conservation among other schistosome species and differential expression across five different life-cycle stages of the parasite. Real-time qPCR has confirmed the differential life-cycle stage expression of a set of selected lincRNAs. We have built PC gene and lincRNA co-expression networks, unraveling key biological processes where lincRNAs might be involved during parasite development. This is the first report of a large-scale identification and structural annotation of lncRNAs in the S. mansoni genome

Physical Interactions With Bacteria and Protozoan Parasites Establish the Scavenger Receptor SSC4D as a Broad-Spectrum Pattern Recognition Receptor

Since the pioneering discoveries, by the Nobel laureates Jules Hoffmann and Bruce Beutler, that Toll and Toll-like receptors can sense pathogenic microorganisms and initiate, in vertebrates and invertebrates, innate immune responses against microbial infections, many other families of pattern recognition receptors (PRRs) have been described. One of such receptor clusters is composed by, if not all, at least several members of the scavenger receptor cysteine-rich (SRCR) superfamily. Many SRCR proteins are plasma membrane receptors of immune cells; however, a small subset consists of secreted receptors that are therefore in circulation. We here describe the first characterization of biological and functional roles of the circulating human protein SSC4D, one of the least scrutinized members of the family. Within leukocyte populations, SSC4D was found to be expressed by monocytes/macrophages, neutrophils, and B cells, but its production was particularly evident in epithelial cells of several organs and tissues, namely, in the kidney, thyroid, lung, placenta, intestinal tract, and liver. Similar to other SRCR proteins, SSC4D shows the capacity of physically binding to different species of bacteria, and this opsonization can increase the phagocytic capacity of monocytes. Importantly, we have uncovered the capacity of SSC4D of binding to several protozoan parasites, a singular feature seldom described for PRRs in general and here demonstrated for the first time for an SRCR family member. Overall, our study is pioneer in assigning a PRR role to SSC4D.This work was funded by National Funds through FCT– Fundação para a Ciência e a Tecnologia, I.P., under the projects SRecognite Infect-ERA/0003/2015 and UIDB/04293/ 2020. Individual funding to JT was provided by FCT through CEECIND/02362/2017. MC, RS, and MS were recipients of studentships from FCT, respectively, SFRH/BD/116791/2016, SFRH/BD/110691/2015, and SFRH/BD/133485/2017. This paper is dedicated to our colleague and friend Rui Appelberg (1960-2020). The authors acknowledge the support of the i3S Scientific Platform BioSciences Screening, member of the national infrastructure PPBI–Portuguese Platform of Bioimaging (PPBI-POCI-01-0145-FEDER-022122) and PT-OPENSCREEN. Tissue sections were kindly provided by Amaro Frutuoso, Department of Complementary Means of Diagnosis and Therapy, Service of Pathology, Hospital Pedro Hispano, Matosinhos

H-Ferritin Is Essential for Macrophages' Capacity to Store or Detoxify Exogenously Added Iron

Macrophages are central cells both in the immune response and in iron homeostasis. Iron is both essential and potentially toxic. Therefore, iron acquisition, transport, storage, and release are tightly regulated, by several important proteins. Cytosolic ferritin is an iron storage protein composed of 24 subunits of either the L- or the H-type chains. H-ferritin differs from L-ferritin in the capacity to oxidize Fe2+ to Fe3+. In this work, we investigated the role played by H-ferritin in the macrophages' ability to respond to immune stimuli and to deal with exogenously added iron. We used mice with a conditional deletion of the H-ferritin gene in the myeloid lineage to obtain bone marrow-derived macrophages. These macrophages had normal viability and gene expression under basal culture conditions. However, when treated with interferon-gamma and lipopolysaccharide they had a lower activation of Nitric Oxide Synthase 2. Furthermore, H-ferritin-deficient macrophages had a higher sensitivity to iron-induced toxicity. This sensitivity was associated with a lower intracellular iron accumulation but a higher production of reactive oxygen species. These data indicate that H-ferritin modulates macrophage response to immune stimuli and that it plays an essential role in protection against iron-induced oxidative stress and cell death.Tis work was fnanced by FEDER - Fundo Europeu de Desenvolvimento Regional funds through the COMPETE2020 - Operacional Programme for Competitiveness and Internationalization (POCI), Portugal 2020, and by Portuguese funds through FCT - Fundação para a Ciência e a Tecnologia/Ministério da Ciência, Tecnologia e Ensino Superior in the framework of the project PTDC/IMI-MIC/1683/2014 (POCI-01-0145-FEDER-016590). PFO and MGA acknowledge FCT for the Investigador FCT 2015. We thank the valuable collaboration of the following i3S Scientifc Platforms: Cell Culture and Genotyping Core Facility (CCGen), [Histology and Electron Microscopy Service (HEMS), and BioSciences Screening], member of the PPBI (PPBI-POCI-01-0145-FEDER-022122)], Animal Facility, and Flow Cytometry Unit (TraCy). We acknowledge Lukas Kuhn (Swiss Institute for Experimental Cancer Research, Lausanne, Switzerland) for kindly providing the frst breeding pairs of Fth1−/− mice. Te authors also acknowledge Marisa Castro, from Departamento de Biologia Molecular from ICBAS, Clara Bento, from i3S, and Edgar Pinto from LAQV – REQUIMTE for technical assistance at diferent stages of the project

Obesity and brain cancer: proteomic analyzes of the influence of the adipocyte secretome on glioma Gl261 cells

Glioma is the most frequent form of malignant brain tumor in the adults and childhood. There is a global tendency toward a higher incidence of gliomas in highly developed and industrialized countries. Simultaneously obesity is reaching epidemic proportions in such developed countries. It has been highly accepted that obesity may play an important role in the biology of several types of cancer. We have developed an in vitro method for the understanding of the influence of obesity on glioma mouse cells (Gl261).info:eu-repo/semantics/publishedVersio

In silico single strand melting curve: a new approach to identify nucleic acid polymorphisms in Totiviridae

CpG-island promoters of developmental genes are unmethylated. DNA methylation state of CpG islands overlapping and surrounding the promoter region of Pax3 (a) and Pax7 (b) genes in myogenic (MB, MT, MF) and non-myogenic samples (ESC). CpG islands are indicated in green and regions analysed by sodium bisulphite sequencing are shown in red. Each circle represents a CpG dinucleotide and its distance to the gene TSS is indicated below. The colour gradient represents the percentage of methylation indicated in the legend. Abbreviations: ESC, embryonic stem cell; MB, myoblast; MT, myotube; MF, myofiber; TSS, transcription start site. c. DNA methylation state of -5 kb distal regulatory region for MyoD was analysed by sodium bisulphite sequencing in ESC and myoblast samples, and represented as above. (PDF 171 kb