Vascular dysfunction occurs prior to the onset of amyloid pathology and Aß plaque deposits colocalize with endothelial cells in the hippocampus of female APPswe/PSEN1dE9 mice.

Increasing evidence shows that cardiovascular diseases (CVDs) are associated with an increased risk of cognitive impairment and Alzheimer's diseases (AD). It is unknown whether systemic vascular dysfunction occurs prior to the development of AD, if this occurs in a sex-dependent manner, and whether endothelial cells play a role in the deposition of amyloid beta (Aß) peptides. We hypothesized that vascular dysfunction occurs prior to the onset of amyloid pathology, thus escalating its progression. Furthermore, endothelial cells from female mice will present with an exacerbated formation of Aß peptides due to an exacerbated pressure pulsatility. To test this hypothesis, we used a double transgenic mouse model of early-onset AD (APPswe/PSEN1dE9). We evaluated hippocampus-dependent recognition memory and the cardiovascular function by echocardiography and direct measurements of blood pressure through carotid artery catheterization. Vascular function was evaluated in resistance arteries, morphometric parameters in the aortas, and immunofluorescence in the hippocampus and aortas. We observed that endothelial dysfunction occurred prior to the onset of amyloid pathology irrespective of sex. However, during the onset of amyloid pathology, only female APP/PS1 mice had vascular stiffness in the aorta. There was elevated Aß deposition which colocalized with endothelial cells in the hippocampus from female APP/PS1 mice. Overall, these data showed that vascular abnormalities may be an early marker, and potential mediator of AD, but exacerbated aortic stiffness and pressure pulsatility after the onset of amyloid pathology may be associated with a greater burden of Aß formation in hippocampal endothelial cells from female but not male APP/PS1 mice.

A Single-Short Partial Reprogramming of the Endothelial Cells decreases Blood Pressure via attenuation of EndMT in Hypertensive Mice.

Background: Small artery remodeling and endothelial dysfunction are hallmarks of hypertension. Growing evidence supports a likely causal association between cardiovascular diseases and the presence of endothelial-to-mesenchymal transition (EndMT), a cellular transdifferentiation process in which endothelial cells (ECs) partially lose their identity and acquire additional mesenchymal phenotypes. EC reprogramming represents an innovative strategy in regenerative medicine to prevent deleterious effects induced by cardiovascular diseases. Methods: Using a partial reprogramming of ECs, via overexpression of Oct-3/4, Sox-2, and Klf-4 (OSK) transcription factors, we aimed to bring ECs back to a youthful phenotype in hypertensive mice. Primary ECs were infected with lentiviral vectors (LV) containing the specific EC marker cadherin 5 (Cdh5) and the fluorescent reporter enhanced green fluorescence protein (EGFP) with empty vector (LVCO) or with OSK (LV-OSK). Confocal microscopy and western blotting analysis were used to confirm the OSK overexpression. Cellular migration, senescence, and apoptosis were evaluated. Human aortic ECs (HAoECs) from male and female normotensive and hypertensive patients were analyzed after OSK or control treatments for their endothelial nitric oxide synthase (eNOS) levels, nitric oxide (NO), and genetic profile. Male and female normotensive (BPN/3J) and hypertensive (BPH/2J) mice were treated with an intravenous (i.v.) injection of LVCO or LV-OSK and evaluated 10 days post-infection. The blood pressure, cardiac function, vascular reactivity of small arteries, in vivo EGFP signal and EndMT inhibition were analyzed. Results: OSK overexpression induced partial EC reprogramming in vitro , and these cells showed endothelial progenitor cell (EPC)-like features with lower migratory capability. OSK treatment of hypertensive BPH/2J mice normalized blood pressure and resistance arteries hypercontractility, via the attenuation of EndMT and elastin breaks. EGFP signal was detected in vivo in the prefrontal cortex of both BPN/3J and BPH/2J-treated mice, but OSK induced angiogenesis only in male BPN/3J mice. OSK-treated human ECs from hypertensive patients showed high eNOS activation and NO production, with low ROS formation. Single-cell RNA analysis showed that OSK alleviated EC senescence and EndMT, restoring their phenotypes in human ECs from hypertensive patients. Conclusion: Overall, these data indicate that OSK treatment and EC reprogramming can decrease blood pressure and reverse hypertension-induced vascular damage.

Dimensional crossover in Kardar-Parisi-Zhang growth.

Two-dimensional (2D) Kardar-Parisi-Zhang (KPZ) growth is usually investigated on substrates of lateral sizes L_{x}=L_{y}, so that L_{x} and the correlation length (ξ) are the only relevant lengths determining the scaling behavior. However, in cylindrical geometry, as well as in flat rectangular substrates L_{x}≠L_{y} and, thus, the surfaces can become correlated in a single direction, when ξ∼L_{x}≪L_{y}. From extensive simulations of several KPZ models, we demonstrate that this yields a dimensional crossover in their dynamics, with the roughness scaling as W∼t^{ß_{2D}} for t≪t_{c} and W∼t^{ß_{1D}} for t≫t_{c}, where t_{c}∼L_{x}^{1/z_{2D}}. The height distributions (HDs) also cross over from the 2D flat (cylindrical) HD to the asymptotic Tracy-Widom Gaussian orthogonal ensemble (Gaussian unitary ensemble) distribution. Moreover, 2D to one-dimensional (1D) crossovers are found also in the asymptotic growth velocity and in the steady-state regime of flat systems, where a family of universal HDs exists, interpolating between the 2D and 1D ones as L_{y}/L_{x} increases. Importantly, the crossover scalings are fully determined and indicate a possible way to solve 2D KPZ models.

Enhancing diagnostic accuracy of multiple myeloma through ML-driven analysis of hematological slides: new dataset and identification model to support hematologists.

Multiple Myeloma (MM) is a hematological malignancy characterized by the clonal proliferation of plasma cells within the bone marrow. Diagnosing MM presents considerable challenges, involving the identification of plasma cells in cytology examinations on hematological slides. At present, this is still a time-consuming manual task and has high labor costs. These challenges have adverse implications, which rely heavily on medical professionals' expertise and experience. To tackle these challenges, we present an investigation using Artificial Intelligence, specifically a Machine Learning analysis of hematological slides with a Deep Neural Network (DNN), to support specialists during the process of diagnosing MM. In this sense, the contribution of this study is twofold: in addition to the trained model to diagnose MM, we also make available to the community a fully-curated hematological slide dataset with thousands of images of plasma cells. Taken together, the setup we established here is a framework that researchers and hospitals with limited resources can promptly use. Our contributions provide practical results that have been directly applied in the public health system in Brazil. Given the open-source nature of the project, we anticipate it will be used and extended to diagnose other malignancies.

Ligand-modified nanoparticle surfaces influence CO electroreduction selectivity.

Improving the kinetics and selectivity of CO2/CO electroreduction to valuable multi-carbon products is a challenge for science and is a requirement for practical relevance. Here we develop a thiol-modified surface ligand strategy that promotes electrochemical CO-to-acetate. We explore a picture wherein nucleophilic interaction between the lone pairs of sulfur and the empty orbitals of reaction intermediates contributes to making the acetate pathway more energetically accessible. Density functional theory calculations and Raman spectroscopy suggest a mechanism where the nucleophilic interaction increases the sp2 hybridization of CO(ad), facilitating the rate-determining step, CO* to (CHO)*. We find that the ligands stabilize the (HOOC-CH2)* intermediate, a key intermediate in the acetate pathway. In-situ Raman spectroscopy shows shifts in C-O, Cu-C, and C-S vibrational frequencies that agree with a picture of surface ligand-intermediate interactions. A Faradaic efficiency of 70% is obtained on optimized thiol-capped Cu catalysts, with onset potentials 100 mV lower than in the case of reference Cu catalysts.

The O-GlcNAc dichotomy: when does adaptation become pathological?

O-Linked attachment of ß-N-acetylglucosamine (O-GlcNAc) on serine and threonine residues of nuclear, cytoplasmic, and mitochondrial proteins is a highly dynamic and ubiquitous post-translational modification that impacts the function, activity, subcellular localization, and stability of target proteins. Physiologically, acute O-GlcNAcylation serves primarily to modulate cellular signaling and transcription regulatory pathways in response to nutrients and stress. To date, thousands of proteins have been revealed to be O-GlcNAcylated and this number continues to grow as the technology for the detection of O-GlcNAc improves. The attachment of a single O-GlcNAc is catalyzed by the enzyme O-GlcNAc transferase (OGT), and their removal is catalyzed by O-GlcNAcase (OGA). O-GlcNAcylation is regulated by the metabolism of glucose via the hexosamine biosynthesis pathway, and the metabolic abnormalities associated with pathophysiological conditions are all associated with increased flux through this pathway and elevate O-GlcNAc levels. While chronic O-GlcNAcylation is well associated with cardiovascular dysfunction, only until recently, and with genetically modified animals, has O-GlcNAcylation as a contributing mechanism of cardiovascular disease emerged. This review will address and critically evaluate the current literature on the role of O-GlcNAcylation in vascular physiology, with a view that this pathway can offer novel targets for the treatment and prevention of cardiovascular diseases.

Hot-wiring dynein-2 establishes roles for IFT-A in retrograde train assembly and motility.

Intraflagellar transport (IFT) trains, built around IFT-A and IFT-B complexes, are carried by opposing motors to import and export ciliary cargo. While transported by kinesin-2 on anterograde IFT trains, the dynein-2 motor adopts an autoinhibitory conformation until it needs to be activated at the ciliary tip to power retrograde IFT. Growing evidence has linked the IFT-A complex to retrograde IFT; however, its roles in this process remain unknown. Here, we use CRISPR-Cas9-mediated genome editing to disable the dynein-2 autoinhibition mechanism in Caenorhabditis elegans and assess its impact on IFT with high-resolution live imaging and photobleaching analyses. Remarkably, this dynein-2 "hot-wiring" approach reignites retrograde motility inside IFT-A-deficient cilia without triggering tug-of-war events. In addition to providing functional evidence that multiple mechanisms maintain dynein-2 inhibited during anterograde IFT, our data establish key roles for IFT-A in mediating motor-train coupling during IFT turnaround, promoting retrograde IFT initiation, and modulating dynein-2 retrograde motility.

Universality class of the special adsorption point of two-dimensional lattice polymers.

In recent work [Rodrigues et al., Phys. Rev. E 100, 022121 (2019)10.1103/PhysRevE.100.022121], evidence was found that the surface adsorption transition of interacting self-avoiding trails (ISATs) placed on the square lattice displays a nonuniversal behavior at the special adsorption point (SAP) where the collapsing polymers adsorb. In fact, different surface exponents ϕ^{(s)} and 1/δ^{(s)} were found at the SAP depending on whether the surface orientation is horizontal (HS) or diagonal (DS). Here, we revisit these systems and study other ones, through extensive Monte Carlo simulations, considering much longer trails than previous works. Importantly, we demonstrate that the different exponents observed in the reference above are due to the presence of a surface-attached-globule (SAG) phase in the DS system, which changes the multicritical nature of the SAP and is absent in the HS case. By considering a modified horizontal surface (mHS), on which the trails are forbidden from having two consecutive steps, resembling the DS situation, a stable SAG phase is found in the phase diagram, and both DS and mHS systems present similar 1/δ^{(s)} exponents at the SAP, namely, 1/δ^{(s)}≈0.44, whereas 1/δ^{(s)}≈0.34 in the HS case. Intriguingly, while ϕ^{(s)}≈1/δ^{(s)} is found for the DS and HS scenarios, as expected, in the mHS case ϕ^{(s)} is about 10% smaller than 1/δ^{(s)}. These results strongly indicate that at least two universality classes exist for the SAPs of adsorbing ISATs on the square lattice.

Intracellular peptides in SARS-CoV-2-infected patients.

Intracellular peptides (InPeps) generated by the orchestrated action of the proteasome and intracellular peptidases have biological and pharmacological significance. Here, human plasma relative concentration of specific InPeps was compared between 175 patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and 45 SARS-CoV-2 non-infected patients; 2,466 unique peptides were identified, of which 67% were InPeps. The results revealed differences of a specific group of peptides in human plasma comparing non-infected individuals to patients infected by SARS-CoV-2, following the results of the semi-quantitative analyses by isotope-labeled electrospray mass spectrometry. The protein-protein interactions networks enriched pathways, drawn by genes encoding the proteins from which the peptides originated, revealed the presence of the coronavirus disease/COVID-19 network solely in the group of patients fatally infected by SARS-CoV-2. Thus, modulation of the relative plasma levels of specific InPeps could be employed as a predictive tool for disease outcome.

Development of an Enhanced High-Yield Influenza Vaccine Backbone in Embryonated Chicken Eggs.

Vaccination is an efficient approach to preventing influenza virus infections. Recently, we developed influenza A and B virus vaccine backbones that increased the yield of several vaccine viruses in Madin-Darby canine kidney (MDCK) and African green monkey kidney (Vero) cells. These vaccine backbones also increased viral replication in embryonated chicken eggs, which are the most frequently used platform for influenza vaccine manufacturing. In this study, to further increase the viral titers in embryonated chicken eggs, we introduced random mutations into the 'internal genes' (i.e., all influenza viral genes except those encoding the hemagglutinin and neuraminidase proteins) of the influenza A virus high-yield virus backbone we developed previously. The randomly mutated viruses were sequentially passaged in embryonated chicken eggs to select variants with increased replicative ability. We identified a candidate that conferred higher influenza virus growth than the high-yield parental virus backbone. Although the observed increases in virus growth may be considered small, they are highly relevant for vaccine manufacturers.

Step-by-Step Approach to Build Multiple Reaction Monitoring (MRM) Profiling Instrument Acquisition Methods for Class-based Lipid Exploratory Analysis by Mass Spectrometry.

Multiple reaction monitoring (MRM) profiling is a strategy for the exploratory analysis of small molecules and lipids by direct sample injection, ie, without the use of chromatographic separation. It is based on instrument methods that comprise a list of ion transitions (MRMs), in which the precursor ion is the expected ionized m/z of the lipid at its species level, ie, the description of lipid class and number of carbon and double bonds in the fatty acid chain(s), and the product ion is a fragment expected for the lipid class or for the fatty acid neutral loss. The Lipid Maps database is expanding constantly, and therefore the MRM-profiling methods associated with this database need to be continuously updated. Here, we provide a comprehensive overview and the key references for the MRM-profiling methodology and workflow, followed by a step-by-step approach to build MRM-profiling instrument acquisition methods for class-based lipid exploratory analysis based on the Lipid Maps database. The detailed workflow includes (1) importing the list of lipids from the database; (2) for a given class, combining isomeric lipids described at full structural level into 1 entry to obtain the neutral mass at species level; (3) attributing the standard Lipid Maps abbreviated nomenclature for the lipid at its species level; (4) predicting the ionized precursor ions; and (5) adding the expected product ion. We also describe how to simulate the precursor ion for the suspect screening of modified lipids using lipid oxidation and their expected product ions as an example. After determining the MRMs, information about collision energy, dwell time, and other instrument parameters are added to finalize the acquisition method. As an example of final method output, we describe the format for Agilent MassHunter v.B.06 and provide the parameters in which optimization can be performed by lipid class using one or more lipid standards.

Specialized Pro-resolving Mediator Improves Vascular Relaxation via Formyl Peptide Receptor-2.

BACKGROUND: The resolution of inflammation is an active phenomenon important for switching off inflammatory processes once the harmful stimuli are removed and facilitate the return to homeostasis. Specialized pro-resolving mediators (SPMs), such as lipoxin A4, resolvin D1, and resolvin E1, derived from ω-3 or ω-6 polyunsaturated fatty acids, are crucial for the resolution of inflammation. We hypothesized that SPMs are decreased in hypertension which contributes to the acetylcholine-induced contraction in resistance arteries, which are well known to be mediated by leukotrienes and prostaglandins. Moreover, treatment with SPMs will decrease this contraction via formyl peptide receptor-2 (FPR-2) in resistance arteries from spontaneously hypertensive rats (SHR). METHODS AND RESULTS: We performed a comprehensive eicosanoid lipid panel analysis, and our data showed for the first time that precursors of SPMs are decreased in SHR, limiting the production of SPMs and resolution of inflammation in vivo. This phenomenon was associated with an increase in lipid peroxidation in resistance arteries. Although SPMs did not abolish acetylcholine-induced contraction, these lipid mediators improved endothelial function in arteries from SHR via FPR-2 activation at nanomolar concentrations. SPMs also buffered TNF-α-induced reactive oxygen species generation in endothelial cells from C57Bl/6 mice. CONCLUSIONS: We suggest that FPR-2 and SPMs could be revealed as a new target or therapeutic agent to improve vascular function in arteries from hypertensive rats.

One-point height fluctuations and two-point correlators of (2+1) cylindrical KPZ systems.

While the one-point height distributions (HDs) and two-point covariances of (2+1) Kardar-Parisi-Zhang (KPZ) systems have been investigated in several recent works for flat and spherical geometries, for the cylindrical one the HD was analyzed for few models and nothing is known about the spatial and temporal covariances. Here, we report results for these quantities, obtained from extensive numerical simulations of discrete KPZ models, for three different setups yielding cylindrical growth. Beyond demonstrating the universality of the HD and covariances, our results reveal other interesting features of this geometry. For example, the spatial covariances measured along the longitudinal and azimuthal directions are different, with the former being quite similar to the curve for flat (2+1) KPZ systems, while the latter resembles the Airy_{2} covariance of circular (1+1) KPZ interfaces. We also argue (and present numerical evidence) that, in general, the rescaled temporal covariance A(t/t_{0}) decays asymptotically as A(x)∼x^{-λ[over ¯]} with an exponent λ[over ¯]=ß+d^{*}/z, where d^{*} is the number of interface sides kept fixed during the growth (being d^{*}=1 for the systems analyzed here). Overall, these results complete the picture of the main statistics for the (2+1) KPZ class.

Protonated and Sodiated Cyclophosphamide Fragmentation Pathways Evaluation by Infrared Multiple Photon Dissociation Spectroscopy.

Cyclophosphamide (CP or CTX) is a widely used antineoplastic agent, and the evaluation of its efficacy and its impacts on the environment are dependent on tandem mass spectrometry (MSn) techniques. Because there is no dedicated experimental study to characterize the actual molecular nature of the CP fragments upon collision-induced dissociation, this work evaluated the chemical structure of the fragments of protonated and sodiated CP and CP protonation sites by infrared multiple photon dissociation spectroscopy supported by density functional theory calculations. This study allowed us to propose a new fragment structure and confirm the nature of multiple fragments, including those relevant for transitions used for CP quantitative and qualitative analyses. Our results also show that there is no spectroscopic evidence that can rule out the existence of aziridinium fragments, making it clear that further studies on the nature of iminium/aziridinium fragments in the gas phase are necessary.

Full-scale network analysis reveals properties of the FV protein structure organization.

Blood coagulation is a vital process for humans and other species. Following an injury to a blood vessel, a cascade of molecular signals is transmitted, inhibiting and activating more than a dozen coagulation factors and resulting in the formation of a fibrin clot that ceases the bleeding. In this process, the Coagulation factor V (FV) is a master regulator, coordinating critical steps of this process. Mutations to this factor result in spontaneous bleeding episodes and prolonged hemorrhage after trauma or surgery. Although the role of FV is well characterized, it is unclear how single-point mutations affect its structure. In this study, to understand the effect of mutations, we created a detailed network map of this protein, where each node is a residue, and two residues are connected if they are in close proximity in the three-dimensional structure. Overall, we analyzed 63 point-mutations from patients and identified common patterns underlying FV deficient phenotypes. We used structural and evolutionary patterns as input to machine learning algorithms to anticipate the effects of mutations and anticipated FV-deficiency with fair accuracy. Together, our results demonstrate how clinical features, genetic data and in silico analysis are converging to enhance treatment and diagnosis of coagulation disorders.

A graph-based machine learning framework identifies critical properties of FVIII that lead to hemophilia A.

Introduction: Blood coagulation is an essential process to cease bleeding in humans and other species. This mechanism is characterized by a molecular cascade of more than a dozen components activated after an injury to a blood vessel. In this process, the coagulation factor VIII (FVIII) is a master regulator, enhancing the activity of other components by thousands of times. In this sense, it is unsurprising that even single amino acid substitutions result in hemophilia A (HA)-a disease marked by uncontrolled bleeding and that leaves patients at permanent risk of hemorrhagic complications. Methods: Despite recent advances in the diagnosis and treatment of HA, the precise role of each residue of the FVIII protein remains unclear. In this study, we developed a graph-based machine learning framework that explores in detail the network formed by the residues of the FVIII protein, where each residue is a node, and two nodes are connected if they are in close proximity on the FVIII 3D structure. Results: Using this system, we identified the properties that lead to severe and mild forms of the disease. Finally, in an effort to advance the development of novel recombinant therapeutic FVIII proteins, we adapted our framework to predict the activity and expression of more than 300 in vitro alanine mutations, once more observing a close agreement between the in silico and the in vitro results. Discussion: Together, the results derived from this study demonstrate how graph-based classifiers can leverage the diagnostic and treatment of a rare disease.

Why copper catalyzes electrochemical reduction of nitrate to ammonia.

Electrochemical reduction of nitrate (NO3RR) has drawn significant attention in the scientific community as an attractive route for ammonia synthesis as well as alleviating environmental concerns for nitrate pollution. To improve the efficiency of this process, the development of catalyst materials that exhibit high activity and selectivity is of paramount importance. Copper and copper-based catalysts have been widely investigated as potential catalyst materials for this reaction both computationally and experimentally. However, less attention has been paid to understanding the reasons behind such high activity and selectivity. Herein, we use Density Functional Theory (DFT) to identify reactivity descriptors guiding the identification of active catalysts for the NO3RR, establish trends in activity, and explain why copper is the most active and selective transition metal for the NO3RR to ammonia among ten different transition metals, namely Au, Ag, Cu, Pt, Pd, Ni, Ir, Rh, Ru, and Co. Furthermore, we assess NO3RR selectivity by taking into account the competition between the NO3RR and the hydrogen evolution reaction. Finally, we propose various approaches for developing highly active catalyst materials for the NO3RR.

Biallelic variants in DNA2 cause poikiloderma with congenital cataracts and severe growth failure reminiscent of Rothmund-Thomson syndrome.

Rothmund-Thomson syndrome (RTS) is a rare, heterogeneous autosomal recessive genodermatosis, with poikiloderma as its hallmark. It is classified into two types: type I, with biallelic variants in ANAPC1 and juvenile cataracts, and type II, with biallelic variants in RECQL4, increased cancer risk and no cataracts. We report on six Brazilian probands and two siblings of Swiss/Portuguese ancestry presenting with severe short stature, widespread poikiloderma and congenital ocular anomalies. Genomic and functional analysis revealed compound heterozygosis for a deep intronic splicing variant in trans with loss of function variants in DNA2, with reduction of the protein levels and impaired DNA double-strand break repair. The intronic variant is shared by all patients, as well as the Portuguese father of the European siblings, indicating a probable founder effect. Biallelic variants in DNA2 were previously associated with microcephalic osteodysplastic primordial dwarfism. Although the individuals reported here present a similar growth pattern, the presence of poikiloderma and ocular anomalies is unique. Thus, we have broadened the phenotypical spectrum of DNA2 mutations, incorporating clinical characteristics of RTS. Although a clear genotype-phenotype correlation cannot be definitively established at this moment, we speculate that the residual activity of the splicing variant allele could be responsible for the distinct manifestations of DNA2-related syndromes.

Dent's disease: An unusual cause of kidney failure.

Dent's disease is an X-linked recessive disease characterized by proximal tubulopathy with low-molecular weight proteinuria, hypercalciuria, nephrolithiasis, nephrocalcinosis, and kidney failure. It is mainly caused by mutations in the CLCN5 or OCRL1 genes, and only ~ 250 families have been identified with these mutations. We present a 31-year-old male referred to a nephrology consultation due to elevated serum creatinine and a history of nephrolithiasis. Complementary evaluation revealed protein/creatinine ratio of 1.9 g/g and albumin/creatinine ratio of 0.5 g/g, hypercalciuria and medullary nephrocalcinosis. These findings raised the suspicion of Dent's disease, which was confirmed by genetic testing. A missense mutation in the CLCN5 gene (c.810C>G, p.(Ser270Arg)), not previously reported in populational databases, was identified. During the evaluation of the patient, it came to our attention that a first-degree male cousin was being followed in our kidney transplantation unit. Given the unknown etiology of his chronic kidney disease, genetic testing was performed, identifying the same mutation. This case highlights the importance of considering the diagnosis of Dent's disease in the setting of a male patient with chronic kidney disease of unknown etiology, low-molecular-weight proteinuria, hypercalciuria, and nephrocalcinosis. Despite progression to end-stage kidney failure in a significant portion of male patients, there are no reports of recurrence after kidney transplantation.