Biology subjects

Rodrigues, S.

Publications and source records attributed to Rodrigues, S..

18 recordsLinked to original sources

bioRxiv2026

Geometric characterization of the HSV - 1 glycoprotein B - amyloid β interaction in Alzheimer's disease using Forman-Ricci curvature

Alzheimer's disease is characterized by the accumulation and aggregation of amyloid-{beta}(A{beta}), but the molecular mechanisms linking environmental and infectious factors to A$\beta$ conformational changes remain incompletely understood. Herpes simplex virus type 1 (HSV-1) has been proposed as a potential contributor to AD pathology, and interactions between the viral glycoprotein B (gB) and A$\beta$ may influence the conformational behaviour of the peptide. Molecular dynamics (MD) simulations provide atomic-scale information on such interactions, but conventional structural descriptors may not fully capture changes in the organization of residue interaction networks. Here, we introduce a graph-geometric framework based on Forman-Ricci curvature to characterize the evolution of residue interaction networks during MD simulations. Each simulation frame is represented as a residue interaction graph based on C--C contacts, and residue-wise curvature profiles are analysed across time. We apply the framework to A{beta}1-42 in isolation and in complex with HSV-1 gB. Conventional MD analyses indicate stable association of the simulated complex, favourable interaction energetics, and conformational changes in A{beta}, including a transition from -helical structure toward {beta}-turn-rich conformations over the simulated timescale. Forman-Ricci curvature reveals pronounced and spatially localized remodelling of the A{beta} residue interaction network in the complex, with the strongest changes concentrated in the C-terminal region. These regions also exhibit reduced temporal curvature fluctuations and progressively distinct geometric behaviour throughout the simulation. Hierarchical clustering further identifies cooperative groups of residues with coordinated curvature dynamics, including a prominent C-terminal domain. Together, these results demonstrate that Forman-Ricci curvature provides a complementary description of biomolecular dynamics by capturing changes in the geometric organization of residue interaction networks that are not directly represented by conventional structural descriptors. The framework provides a general computational approach for studying network-level structural remodelling in protein molecular dynamics and offers a quantitative perspective on the conformational consequences of HSV-1 gB--A{beta} association.

bioinformatics↗

bioRxiv2025

Prophage-marine diazotroph interplay shapes both biofilm structure and nitrogen release

Marine environments are frequently oligotrophic, characterized by low amount of bioassimilable Nitrogen sources. At the global scale, the microbial fixation of N2, or diazotrophy, represents the primary source of fixed nitrogen in pelagic marine ecosystems, playing a key role in supporting primary production and driving the export of organic matter to the deep ocean. However, given the high energetic cost of N2 fixation, the active release of fixed nitrogen by diazotrophs appears counterintuitive, suggesting the existence of alternative, passive release pathways, that remain understudied to date. Here, we show that the marine Non Cyanobacterial Diazotroph Vibrio diazotrophicus is endowed with a prophage belonging to the Myoviridae family, whose expression is induced under anoxic and biofilm-forming conditions. We demonstrate that this prophage can spontaneously excise from the genome of its host and that it forms intact and infective phage particles. Moreover, phage-mediated host cell lysis leads to increased biofilm production as compared to a prophage-free derivative mutant, and to increased release of Dissolved Organic Carbon and of ammonium. Altogether, we provide evidences that viruses may play a previously unrecognized role in oceanic ecosystem dynamics by structuring micro-habitats suitable for diazotrophy and by contributing to the recycling of (in-)organic matter. ImportanceDiazotrophs are key players in ocean functioning by providing fixed nitrogen to ecosystems and fueling primary production. However, from a physiological point-of-view, the active release of nitrogenous compounds by diazotrophs is paradoxical, since they would invest in an energy-intensive process and supply nutrient to non-sibling cells, with the risk of being outcompeted. Therefore, alternative ways leading to the release of fixed nitrogen must exist. Here, we show that the marine Non Cyanobacterial Diazotroph Vibrio diazotrophicus possesses one prophage, whose activation leads to cell death, to increased biofilm production and to the release of Dissolved Organic Compounds and of ammonium. Taken together, our results provide evidences that marine phage-diazotroph interplay leads to the creation of micro-habitats suitable for diazotrophy like biofilm and to nutrient cycling, and contribute to better understanding of the role of viruses in marine ecosystems.

microbiology↗

bioRxiv2025

A phenomenological interpretation of multiple bursting patterns in Lateral Habenula neurons

The Lateral Habenula (LHb) is a small brain structure specialized in encoding aversive signals. Bursting activity in the LHb has been consistently linked to mood regulation, with increased bursting activity proposed to promote depressive behaviors. Bursting is a complex dynamic process that has been extensively studied and modeled in other neuronal contexts. However, at the LHb this type of activity has typically been described only as transient periods of high frequency firing. Here, to provide a deeper understanding of LHb bursting, we analyzed this activity from the perspective of dynamical systems. Ex vivo, LHb neurons display a variety of bursting patterns, characterized at one extreme by a dominating square-wave type and in other by parabolic type, plus transitional forms referred to as triangular bursting. Notably, these bursting patterns, which reflect different LHb output modes, can occur within the same neuron, suggesting that they may correspond to distinct dynamic states of the same LHb neuron. To capture these complex behaviors, we propose an idealized multiple-timescale dynamical model. This model successfully reproduces the three main bursting patterns observed in experimental data. Furthermore, we identify a special point in the parameter space, termed the saddle-node homoclinic bifurcation, which acts as an organizing center demarcating the boundary between the two primary bursting patterns and around which the third pattern appear. Our model suggests that LHb bursting activity is structured around distinct dynamic states with potentially diverse and unexplored impacts on mood regulation. By providing new insights into the dynamic principles underlying LHb bursting, this framework may advance our understanding of its biological significance.

neuroscience↗

bioRxiv2024

Medieval genomes from eastern Iberia illuminate the role of Morisco mass deportations in dismantling a long-standing genetic bridge with North Africa

BackgroundThe Islamic influence on the Iberian Peninsula left an enduring legacy culturally and linguistically, however the demographic impact is less well understood. This study aims to explore the dynamics of gene flow and population structure in eastern Iberia from the early to late Medieval period through ancient DNA. ResultsOur comprehensive genomic analysis uncovered gene flow from various Mediterranean regions into Iberia before the Islamic period, supporting a pre-existing pan-Mediterranean homogenization phenomenon during the Roman Empire. North African ancestry is present but sporadic in late antiquity genomes but becomes consolidated during the Islamic period. We uncovered one of the earliest dated Islamic burials in Spain, which showed high levels of inbreeding. For the first time we also prove the persistence of North African ancestry in a Christian cemetery until the 17th century, in addition to evidence of slave traffick from the Maghreb. ConclusionsThis study reveals the complex interaction between political events and cultural shifts that influenced the population of eastern Iberia. It highlights the existence of a slave trade and underscores the lasting impact of historical events, such as the Expulsion of the Moriscos in 1609 CE, on the regions genetic and cultural landscape through mass population displacement and replacement.

evolutionary biology↗

bioRxiv2024

1-Aminocyclopropane-1-carboxylic acid oxidase determines the fate of ethylene biosynthesis in a tissue-specific way to fine-tune development and stress resilience

Ethylene is involved in several developmental processes and responses towards (a)biotic stress. In seed plants, ethylene is produced from its precursor 1-aminocyclopropane-1-carboxylic acid (ACC) by the enzyme ACC-oxidase (ACO). Despite its key role in ethylene synthesis, the ACO gene family has not yet been fully characterized. We investigated the five ACO members of Arabidopsis thaliana and revealed a tissue-and developmentally specific expression pattern. Furthermore, the five ACO enzymes each have a different capacity to produce ethylene. Combined, this allows for a precise spatial and temporal regulation of ethylene synthesis. At the sub-cellular level, we uncovered that ACOs reside in the cytosol, where ethylene is likely synthesized, but surprisingly also in the nucleus. Using reverse genetics of single and higher order aco mutants we revealed a high degree of gene redundancy and minimal phenotypes. A lack of ethylene synthesis by knocking out all five ACOs did not impair normal vegetative and generative development but did influence processes associated with high rates of ethylene production such as petal abscission. This suggests that ethylene is not a prime regulator of development, but more a moderator. We also showed that the inability to synthesize ethylene impairs some abiotic (nutrient deficiency and metal toxicity) and biotic (Botrytis cinerea) stress responses, similar as plants insensitive towards ethylene, corroborating the role of ethylene in mediating stress responses. In conclusion, the ACO gene family enables plants to fine-tune their ethylene synthesis rates, but a lack their off is not crucial for normal development and stress survival.

plant biology↗

bioRxiv2023

Discovering Secondary Protein Structures via Local Euler Curvature

Protein structure analysis and classification, which is fundamental for predicting protein function, still poses formidable challenges in the fields of molecular biology, mathematics, physics and computer science. In the present work we exploit recent advances in computational topology to define a new intrinsic unsupervised topological fingerprint for proteins. These fingerprints, computed via Local Euler Curvature (LECs), identify secondary protein structures, such as Helices and Sheets, by capturing their distinctive topological signatures. Using an extensive protein residue database, the proposed computational framework not only distinguishes between structural classes via unsupervised clustering but also achieves remarkable accuracy in classifying proteins structures through supervised machine learning classifier. We also show that the internal structure of LEC space embeds the information about the secondary structure of proteins. Beyond its immediate implications for the advancement of critical application areas such as drug design and biotechnology, our approach opens a fascinating avenue towards characterizing the multiscale structures of diverse biopolymers based solely on their geometric and topological attributes.

biochemistry↗

bioRxiv2023

Neutralization Escape by SARS-CoV-2 Omicron Subvariant BA.2.86

The continued evolution of SARS-CoV-2 may lead to evasion of vaccine immunity and natural immunity. A highly mutated Omicron variant BA.2.86 has recently been identified with over 30 amino acid changes in Spike compared with BA.2 and XBB.1.5. As of September 4, 2023, BA.2.86 has been identified in 37 sequences from 10 countries, which is likely an underestimate due to limited surveillance. The ability of BA.2.86 to evade NAbs compared with other currently circulating Omicron variants remains unknown. Our data show that NAb responses to BA.2.86 were lower than to BA.2 but were comparable or slightly higher than to the current circulating recombinant variants XBB.1.5, XBB.1.16, EG.5, EG.5.1, and FL.1.5.1.

microbiology↗

bioRxiv2023

Distinct mesenchymal cell states mediate prostate cancer progression

Alterations in tumor stroma influence prostate cancer progression and metastatic potential. However, the molecular underpinnings of this stromal-epithelial crosstalk are largely unknown. Here, we compare mesenchymal cells from four genetically engineered mouse models (GEMMs) of prostate cancer representing different stages of the disease to their wild-type (WT) counterparts by single-cell RNA sequencing (scRNA-seq) and, ultimately, to human tumors with comparable genotypes. We identified 8 transcriptionally and functionally distinct stromal populations responsible for common and GEMM-specific transcriptional programs. We show that stromal responses are conserved in mouse models and human prostate cancers with the same genomic alterations. We noted striking similarities between the transcriptional profiles of the stroma of murine models of advanced disease and those of of human prostate cancer bone metastases. These profiles were then used to build a robust gene signature that can predict metastatic progression in prostate cancer patients with localized disease and is also associated with progression-free survival independent of Gleason score. Taken together, this offers new evidence that stromal microenvironment mediates prostate cancer progression, further identifying tissue-based biomarkers and potential therapeutic targets of aggressive and metastatic disease.

cancer biology↗

bioRxiv2023

Monitoring Alzheimer's Disease via Ultraweak Photon Emission

The present study takes on an innovative experiment involving detection of ultraweak photon emission (UPE) from the hippocampus of male rat brains and finds significant correlations between Alzheimers disease (AD), memory decline, oxidative stress, and the intensity of UPE emitted spontaneously from the hippocampus. These remarkable findings opens up novel methods for screening, detecting, diagnosing and classifying neurodegenerative diseases (and associated sydromes), such as in AD. This also paves the way towards novel advanced brain-computer interfaces (BCIs) photonic chip for the detection of UPE from brains neural tissue. The envisaged BCI photonic chip (BCIPC) would be minimally invasive, cheap, high-speed, scalable, would provide high spatiotemporal resolution of brains activity and would provide short- and long-term screening of clinical patho-neurophysiological signatures, which could be monitored by a smart wristwatch or smartphone via a wireless connection. Background & aimLiving cells spontaneously emit biophotons, or UPE, during the process of metabolic reactions, and these UPE in tissues may be altered in pathological conditions. These compelling observations led us to hypothesise that AD (a severe neuropathological disorder) can be screened via UPE. This is substantiated by previous studies showing that oxidative stress occurs prior to the formation of amyloid plaques and neurofibrillary tangles (i.e. the neuropathological hallmarks of AD). Indeed, oxidative stress is a critical factor contributing to the initiation and progression of AD. Moreover, earlier research have evidenced the association between UPE and oxidative stress of biological tissue. These combined observations set us to investigate whether UPE intensity of the hippocampus in a pathological state, induced by intracerebroventricular (ICV) injection of streptozotocin (STZ), can be correlated with memory, oxidative stress, Acetylcholinesterase (AChE) as a novel screening strategy for AD. Material & methodsThirty-two adult male rats were divided into four groups: Control, Sham, STZ, and STZ+Donp (n=8). Specifically, for inducing sporadic AD (sAD), STZ was injected on days 1 and 3. One week after the second ICV injection, the intraperitoneal (IP) use of donepezil was initiated and continued for two weeks. After treatment, spatial and recognition memory were evaluated from days 24 to 29 of the experiment using the Morris water maze (MWM) and novel object recognition (NOR) test, respectively. Finally, the rats were euthanased by cervical dislocate in day 30. Anesthetic drugs disrupt neural communication from chemical neurotransmitter receptor inhibition. UPE related to cells activity so anesthesia intervention must be considered. Then, their brains were removed and the hippocampus dissected. The Right hippocampus was evaluated in terms of UPE via a Photomultiplier tubes (PMT) device. Moreover, in left hippocampus we measured malondialdehyde (MDA) by the TBARS assay and heat via calorimeter ELIZA device. Acetylcholinesterase (AChE) activity was also scrutinized via acetylthiocholine reaction via the Ellman method. Results & discussionSTZ injection impaired learning and memory function compared with the sham and control groups. The results of the MWM test indicated a decrease in the time used to find the hidden platform in the donepezil-treated group during training days, while in the STZ group, no significant reduction in this time was observed. In the probe trial, the donepezil-treated group showed a significant increase in target quadrant time in comparison with the STZ group (p<0.05). Furthermore, the object recognition test demonstrated that the donepezil-treated group spent more time recognizing new objects in the testing phase (p<0.05). Whereas, in the STZ group, there was no significant difference in spent time for identifying the objects. Ex vivo detection of UPE from the hippocampus of rats showed that the sham group had higher UPE than the Control group (p<0.05). The STZ injection significantly increased UPE and MDA concentrations in the hippocampus than in the Sham and Control groups (p<0.0001). Correlation analysis of results reveal that the emission intensity is associated with the MDA concentration (r = 0.855). Hippocampus AChE activity also significantly increased in STZ-injected groups. Treatment with donepezil decreased MDA concentration, UPE intensity, and activity of AChE in comparison with the STZ group (p<0.05). UPE intensity was linked with AChE activity as evidenced by Pearson correlation analysis between UPE intensity and AChE activity (r = 0.779). Conclusion: The hippocampus UPE increases in STZ-induced sAD and is associated with the redox state of the tissue. Donepezil decreases the UPE and improves the oxidative stress induced by STZ injection. Since oxidative stress is one of the primary hallmarks in the progression of AD, then it stands to reason that the Brains UPE emission can be used as a novel methodology for screening AD. Moreover, UPE could be used to monitor recovery from neurodegenerative diseases upon suitable future therapeutic treatments, as suggested by our experiment involving donepezil. Our findings, encourages further research and suggests the development of a minimally invasive BCI photonic chip (with similar quantum efficiency as PMT) for screening and diagnosing AD.

neuroscience↗

bioRxiv2023

Emergence of High-Order Functional Hubs in the Human Brain

Network theory is often based on pairwise relationships between nodes, which is not necessarily realistic for modeling complex systems. Importantly, it does not accurately capture non-pairwise interactions in the human brain, often considered one of the most complex systems. In this work, we develop a multivariate signal processing pipeline to build high-order networks from time series and apply it to resting-state functional magnetic resonance imaging (fMRI) signals to characterize high-order communication between brain regions. We also propose connectivity and signal processing rules for building uniform hypergraphs and argue that each multivariate interdependence metric could define weights in a hypergraph. As a proof of concept, we investigate the most relevant three-point interactions in the human brain by searching for high-order "hubs" in a cohort of 100 individuals from the Human Connectome Project. We find that, for each choice of multivariate interdependence, the high-order hubs are compatible with distinct systems in the brain. Additionally, the high-order functional brain networks exhibit simultaneous integration and segregation patterns qualitatively observable from their high-order hubs. Our work hereby introduces a promising heuristic route for hypergraph representation of brain activity and opens up exciting avenues for further research in high-order network neuroscience and complex systems.

neuroscience↗

bioRxiv2023

Waning Immunity Against XBB.1.5 Following Bivalent mRNA Boosters

The SARS-CoV-2 Omicron variant has continued to evolve. XBB is a recombinant between two BA.2 sublineages, XBB.1 includes the G252V mutation, and XBB.1.5 includes the G252V and F486P mutations. XBB.1.5 has rapidly increased in frequency and has become the dominant virus in New England. The bivalent mRNA vaccine boosters have been shown to increase neutralizing antibody (NAb) titers to multiple variants, but the durability of these responses remains to be determined. We assessed humoral and cellular immune responses in 30 participants who received the bivalent mRNA boosters and performed assays at baseline prior to boosting, at week 3 after boosting, and at month 3 after boosting. Our data demonstrate that XBB.1.5 substantially escapes NAb responses but not T cell responses after bivalent mRNA boosting. NAb titers to XBB.1 and XBB.1.5 were similar, suggesting that the F486P mutation confers greater transmissibility but not increased immune escape. By month 3, NAb titers to XBB.1 and XBB.1.5 declined essentially to baseline levels prior to boosting, while NAb titers to other variants declined less strikingly.

immunology↗

bioRxiv2022

Genetic and physiological insights into the diazotrophic activity of a non-cyanobacterial marine diazotroph

Nitrogen (N2) fixation, or diazotrophy, supports a large part of primary production in oceans. Culture-independent approaches highlighted the presence in abundance of marine non-cyanobacterial diazotrophs (NCD) but their ecophysiology remains elusive, mostly because of the low number of isolated NCD and because of the lack of available genetic tools for these isolates. Here, a dual genetic and functional approach allowed unveiling the ecophysiology of a marine NCD affiliated to the species Vibrio diazotrophicus. Physiological characterization of the first marine NCD mutant obtained so far was performed using a soft-gellan assay, demonstrating that a {Delta}nifH mutant in not able to grow in nitrogen-deprived media. Furthermore, we demonstrated that V. diazotrophicus produces a thick biofilm under diazotrophic conditions, suggesting biofilm production as an adaptive response of this NCD to cope with the inhibition of nitrogen-fixation by molecular oxygen. Finally, the genomic signature of V. diazotrophicus is essentially absent from metagenomic data of Tara Ocean expeditions, despite having been isolated from various marine environments. We think that the genetically tractable V. diazotrophicus strain used in this study may serve as an ideal model to study the ecophysiology of these overlooked procaryotic group.

microbiology↗

bioRxiv2022

Geometry of spiking activity in early visual cortex: a Topological Data Analytic approach

In the brain, spiking patterns live in a high-dimensional space of neurons and time. Thus, determining the intrinsic structure of this space presents a theoretical and experimental challenge. To address this challenge, we introduce a new framework for applying topological data analysis (TDA) to spike train data and use it to determine the geometry of spiking patterns in the visual cortex. Key to our approach is a parameterized family of distances based on the timing of spikes that quantifies the dissimilarity between neuronal responses. We applied TDA to visually driven single-unit and multiple single-unit spiking activity in macaque V1 and V2. TDA across timescales reveals a common geometry for spiking patterns in V1 and V2 which, among simple models, is most similar to that of a low-dimensional space endowed with Euclidean or hyperbolic geometry with modest curvature. Remarkably, the inferred geometry depends on timescale, and is clearest for the timescales that are important for encoding contrast, orientation, and spatial correlations.

neuroscience↗

bioRxiv2022

Interaction mechanism between the HSV-1 glycoprotein B and the antimicrobial peptide Amyloid-β

Unravelling the mystery of Alzheimers Disease (AD) requires urgent resolution given the worldwide increase of the aging population. There is a growing concern that the current leading AD hypothesis, the amyloid cascade hypothesis, does not stand up to validation with respect to emerging new data. Indeed, several paradoxes are being discussed in the literature, for instance, both the deposition of the Amyloid-Beta peptide (A{beta}) and the intracellular neurofibrillary tangles (NFTs) could occur within the brain without any cognitive pathology. Thus, these paradoxes suggest that something more fundamental is at play in the onset of the disease and other key and related pathomechanisms have to be investigated. The present study follows our previous investigations on the infectious hypothesis, which posits that some pathogens are linked to late onset AD. Our studies also build upon the shattering finding that A{beta} is a powerful antimicrobial agent capable of inhibiting pathogens as observed in in vitro experiments. Herein, we ask what are the molecular mechanisms in play when A{beta} neutralizes infectious pathogens? To answer this question, we probed at nanoscale lengths with FRET (Forster Resonance Energy Transfer), the interaction between A{beta} peptides and glycoprotein B (responsible of virus-cell binding) within the HSV-1 virion. We concluded that there is indeed a close interaction, likely nonspecific or semi-specific, between the two types of molecules, which participate in virus neutralization.

neuroscience↗

bioRxiv2021

Redundant functions of the SLC5A transporters Rumpel Kumpel and Bumpel in ensheathing glial cells

Neuronal processing is energy demanding, and relies on sugar metabolism as an energy source. To provide a constant metabolite supply neurons and glial cells express many glucose and lactate transporters of the solute carrier (SLC) 5A family. Here we dissect the partially redundant functions of three highly related glia specific Drosophila genes encoding SLC5A proteins, Rumpel, Bumpel and Kumpel. While knockdown of rumpel causes several behavioral phenotypes, they are less prominent in rumpel mutants. bumpel and kumpel mutants are viable and fertile, lacking discernible phenotypes. However, in bumpel kumpel double mutants and to an even greater extent in rumpel bumpel kumpel triple mutants oogenesis is disrupted at the onset of the vitollegenic phase. This indicates at least partially redundant functions between these genes. Rescue experiments exploring this effect indicate that oogenesis can be affected by CNS glial cells. Moreover, expression of heterologous mammalian SLC5A transporter proteins, with known transport properties, suggest that Bumpel and/or Kumpel transport glucose or lactate. Overall, our results imply a redundancy in SLC5A nutrient sensing functions in Drosophila glial cells, affecting ovarian development and behavior.

neuroscience↗

bioRxiv2021

Drosophila beta-Heavy-Spectrin is required in polarized ensheathing glia that forma diffusion-barrier around the neuropil

In the central nervous system (CNS), functional tasks are often allocated to distinct compartments. This is also evident in the insect CNS where synapses and dendrites are clustered in distinct neuropil regions. The neuropil is separated from neuronal cell bodies by ensheathing glia, which as we show using dye injection experiments forms an internal diffusion barrier. We find that ensheathing glial cells are polarized with a basolateral plasma membrane rich in phosphatidylinositol-(3,4,5)-triphosphate (PIP3) and the Na+/K+-ATPase Nervana2 (Nrv2) that abuts an extracellular matrix formed at neuropil-cortex interface. The apical plasma membrane is facing the neuropil and is rich in phosphatidylinositol-(4,5)-bisphosphate (PIP2) that is supported by a sub-membranous {beta}Heavy-Spectrin cytoskeleton. {beta}Heavy-spectrin mutant larvae affect ensheathing glial cell polarity with delocalized PIP2 and Nrv2 and exhibit an abnormal locomotion which is similarly shown by ensheathing glia ablated larvae. Thus, polarized glia compartmentalizes the brain and is essential for proper nervous system function.

neuroscience↗

bioRxiv2021

Whole human genome 5'-mC methylation analysis using long read nanopore sequencing

DNA methylation is a type of epigenetic modification that affects gene expression regulation and is associated with several human diseases. Microarray and short read sequencing technologies are often used to study 5-methylcytosine (5-mC) modification of CpG dinucleotides in the human genome. Although both technologies produce trustable results, the evaluation of the methylation status of CpG sites suffers from the potential side effects of DNA modification by bisulfite and the ambiguity of mapping short reads in repetitive and highly homologous genomic regions, respectively. Nanopore sequencing is an attractive alternative for the study of 5-mC since the long reads produced by this technology allow to resolve those genomic regions more easily. Moreover, it allows direct sequencing of native DNA molecules using a fast library preparation procedure. In this work we show that 10X coverage depth nanopore sequencing, using DNA from a human cell line, produces 5-mC methylation frequencies consistent with those obtained by methylation microarray and digital restriction enzyme analysis of methylation. In particular, the correlation of methylation values ranged from 0.73 to 0.90 using an average genome sequencing coverage depth <2X or a minimum read support of 17X for each CpG site, respectively. We also showed that a minimum of 5 reads per CpG yields strong correlations (>0.89) between sequencing runs and an almost uniform variation in methylation frequencies of CpGs across the entire value range. Furthermore, nanopore sequencing was able to correctly display methylation frequency patterns according to genomic annotations, including a majority of unmethylated and methylated sites in the CpG islands and inter-CpG island regions, respectively. These results demonstrate that low coverage depth nanopore sequencing is a fast, reliable and unbiased approach to the study of 5-mC in the human genome.

genomics↗

bioRxiv2021

In situ maturated early-stage human induced pluripotent stem cell-derived cardiomyocytes improve cardiac function by enhancing segmentary contraction in infarcted rats

The scant ability of cardiomyocytes to proliferate makes heart regeneration one of the biggest challenges of science. Current therapies do not contemplate heart re-muscularization. In this scenario, stem cell-based approaches have been proposed to overcome the lack of regeneration. We hypothesize PluriCell hiPSC-derived cardiomyocytes (hiPSC-CMs) could enhance rats cardiac function after myocardial infarction (MI). Animals were subjected to permanent occlusion of the Left-Ventricle (LV) anterior descending coronary artery (LAD). Seven days after MI, Early-stage hiPSC-CMs were injected intramyocardially. Rats were subjected to Echocardiography pre- and post-treatment. Thirty days after injections, treated rats displayed 6.2% human cardiac grafts, which were characterized molecularly. Left ventricle ejection fraction (LVEF) was improved by 7.8% in cell-injected rats, while placebo controls showed an 18.2% deterioration. Also, cell-treated rats displayed a 92% and 56% increase in radial and circumferential strains, respectively. Human cardiac grafts maturate in situ, preserving proliferation with 10% Ki67 and 3% PHH3 positive nuclei. Grafts were perfused by host vasculature with no evidence for immune rejection nor ectopic tissue formations. Our findings support PluriCell hiPSC-CMs as an alternative therapy to treat MI. The next steps of preclinical development include efficacy studies in large animals on the path to clinical-grade regenerative therapy targeting human patients.

physiology↗