Cryo-EM Map Anisotropy Can Be Attenuated by Map Post-Processing and a New Method for Its Estimation.

One of the most important challenges in cryogenic electron microscopy (cryo-EM) is the substantial number of samples that exhibit preferred orientations, which leads to an uneven coverage of the projection sphere. As a result, the overall quality of the reconstructed maps can be severely affected, as manifested by the presence of anisotropy in the map resolution. Several methods have been proposed to measure the directional resolution of maps in tandem with experimental protocols to address the problem of preferential orientations in cryo-EM. Following these works, in this manuscript we identified one potential limitation that may affect most of the existing methods and we proposed an alternative approach to evaluate the presence of preferential orientations in cryo-EM reconstructions. In addition, we also showed that some of the most recently proposed cryo-EM map post-processing algorithms can attenuate map anisotropy, thus offering alternative visualization opportunities for cases affected by moderate levels of preferential orientations.

Flutter auricular en neonatologia. reporte de caso / Atrial flutter in neonatology. Case Report

El flutter auricular es la taquicardia confinada al atrio más frecuente en la edad neonatal, su adecuado tratamiento requiere un diagnóstico precoz. La posibilidad de presentar efectos adversos con los fármacos antiarrítmicos en recién nacidos es alta, por lo que se debe evitar el manejo empírico de la taquicardia en éste grupo etario. Se presenta el caso de un recién nacido con flutter auricular en corazón sano, que respondió favorablemente a las medidas farmacológicas convencionales, con buena evolución a mediano plazo.

Atrial flutter is the atrial confined tachycardia more frequent in the neonatal age, its adequate treatment requires an early diagnosis. The possibility of presenting adverse effects with antiarrhythmic drugs in newborns is high, so the empirical management of tachycardia should be avoided in this age group. We present the case of a newborn with atrial flutter in healthy heart, who responded favorably to conventional pharmacological measures, with good evolution in the medium term.

Patrones morfológicos de la enfermedad glomerular en México: Reporte de un centro de referencia en nefropatología / Morphological patterns of glomerular disease in Mexico: Report of a reference center in nephropathology

RESUMEN Introducción: En México, la enfermedad renal crónica (ERC) representa un gran problema de salud y las glomerulopatías (GP) representan la tercera causa de ERC. Objetivo: Describir, desde una base de datos de biopsias renales (BR) nativas, los diferentes patrones morfológicos de GP en México. Métodos: Se analizaron registros de BR de riñón nativo en un centro de referencia en nefropatología, todas las BR fueron evaluadas por una única nefropatóloga (AVP). El diagnóstico final en cada caso se basó en parámetros clínicos e histopatológicos. Resultados: Fueron revisadas 2084 BR, con edad de 34.4 ± 17.6 años. 1085 BR (52.1%) en género femenino; el síndrome nefrótico fue más frecuente en hombres (p<0.001) y síndrome nefrítico fue más frecuente en mujeres (p<0.001). GP primarias y nefropatías túbulo-intersticiales fueron más diagnosticadas en hombres (p<0.01). Nefritis lúpica (NL) fue la GP secundaria más reportada. La glomeruloesclerosis focal y segmentaria (GEFS) fue la GP primaria diagnosticada con mayor frecuencia en ambos géneros. Vasculitis por inmunoglobulina A fue la enfermedad vascular más frecuentemente detectada. Síndrome nefrótico fue la indicación más frecuente de BR (42.9%), seguido de: síndrome nefrítico (23.9%), proteinuria aislada (16.4%), daño renal agudo (8.7%), alteraciones urinarias asintomáticas (6.2%) y ERC (1.8%). Conclusiones: La GP primarias con mayor frecuencia fueron GEFS. Las GP secundarias más frecuentemente reportadas fueron NL, predominantemente en mujeres. Se observó nefropatía IgA con mayor frecuencia en comparación con otras series publicadas en México. Hubo diferencias significativas en la presentación de GP en relación con el género y la edad del paciente.

ABSTRACT Introduction: In Mexico, chronic kidney disease (CKD) represents a major health problem, and glomerulopathies (GP) represent the third leading cause of CKD. Aim: From a database of native kidney biopsies (KB), describe the different morphological patterns of GP in Mexico. Methods: Records of native KB in a nephropathology referral center were evaluated by a single nephropathologist. The final diagnosis in each case was based on clinical parameters and histopathological findings. Results: 2084 KB were analyzed, patients were 34.4±17.6 years of age, there were 1085 KB (52.1%) in females; nephrotic syndrome was most frequent in males (p<0.001), and nephritic syndrome was more frequent in females (p <0.001). Primary GP and túbulo-interstitial diseases were most diagnosed in males (p <0.01). Lupus nephritis (LN) was the most-reported secondary GP. Focal and segmental glomerulosclerosis (FSGS) was the primary GP most often diagnosed in both genders. The most frequently detected vascular disease was immunoglobulin A vasculitis. Nephrotic syndrome was the most frequent indication for KB (42.9%), followed by: nephritic syndrome (23.9%), isolated proteinuria (16.4%), acute kidney injury (8.7%), asymptomatic urinary alterations (6.2%), and CKD (1.8%). Conclusions: The most frequently observed primary GP was FSGS, and LN was the most frequent secondary GP, predominantly in females, and IgA nephropathy was observed more frequently in comparison with other series published in Mexico. There were significant differences in GP presentation in relation to patient sex and age.

Automatic whole blood cell analysis from blood smear using label-free multi-modal imaging with deep neural networks.

Whole blood cell analysis is widely used in medical applications since its results are indicators for diagnosing a series of diseases. In this work, we report automatic whole blood cell analysis from blood smear using label-free multi-modal imaging with deep neural networks. First, a commercial microscope equipped with our developed Phase Real-time Microscope Camera (PhaseRMiC) obtains both bright-field and quantitative phase images. Then, these images are automatically processed by our designed blood smear recognition networks (BSRNet) that recognize erythrocytes, leukocytes and platelets. Finally, blood cell parameters such as counts, shapes and volumes can be extracted according to both quantitative phase images and automatic recognition results. The proposed whole blood cell analysis technique provides high-quality blood cell images and supports accurate blood cell recognition and analysis. Moreover, this approach requires rather simple and cost-effective setups as well as easy and rapid sample preparations. Therefore, this proposed method has great potential application in blood testing aiming at disease diagnostics.

Patch-to-Seq and Transcriptomic Analyses Yield Molecular Markers of Functionally Distinct Brainstem Serotonin Neurons.

Acute regulation of CO2 and pH homeostasis requires sensory feedback from peripheral (carotid body) and central (central) CO2/pH sensitive cells - so called respiratory chemoreceptors. Subsets of brainstem serotonin (5-HT) neurons in the medullary raphe are CO2 sensitive or insensitive based on differences in embryonic origin, suggesting these functionally distinct subpopulations may have unique transcriptional profiles. Here, we used Patch-to-Seq to determine if the CO2 responses in brainstem 5-HT neurons could be correlated to unique transcriptional profiles and/or unique molecular markers and pathways. First, firing rate changes with hypercapnic acidosis were measured in fluorescently labeled 5-HT neurons in acute brainstem slices from transgenic, Dahl SS (SSMcwi) rats expressing T2/ePet-eGFP transgene in Pet-1 expressing (serotonin) neurons (SS ePet1-eGFP rats). Subsequently, the transcriptomic and pathway profiles of CO2 sensitive and insensitive 5-HT neurons were determined and compared by single cell RNA (scRNAseq) and bioinformatic analyses. Low baseline firing rates were a distinguishing feature of CO2 sensitive 5-HT neurons. scRNAseq of these recorded neurons revealed 166 differentially expressed genes among CO2 sensitive and insensitive 5-HT neurons. Pathway analyses yielded novel predicted upstream regulators, including the transcription factor Egr2 and Leptin. Additional bioinformatic analyses identified 6 candidate gene markers of CO2 sensitive 5-HT neurons, and 2 selected candidate genes (CD46 and Iba57) were both expressed in 5-HT neurons determined via in situ mRNA hybridization. Together, these data provide novel insights into the transcriptional control of cellular chemoreception and provide unbiased candidate gene markers of CO2 sensitive 5-HT neurons. Methodologically, these data highlight the utility of the patch-to-seq technique in enabling the linkage of gene expression to specific functions, like CO2 chemoreception, in a single cell to identify potential mechanisms underlying functional differences in otherwise similar cell types.

Multivalent interactions essential for lentiviral integrase function.

A multimer of retroviral integrase (IN) synapses viral DNA ends within a stable intasome nucleoprotein complex for integration into a host cell genome. Reconstitution of the intasome from the maedi-visna virus (MVV), an ovine lentivirus, revealed a large assembly containing sixteen IN subunits1. Herein, we report cryo-EM structures of the lentiviral intasome prior to engagement of target DNA and following strand transfer, refined at 3.4 and 3.5 Å resolution, respectively. The structures elucidate details of the protein-protein and protein-DNA interfaces involved in lentiviral intasome formation. We show that the homomeric interfaces involved in IN hexadecamer formation and the α-helical configuration of the linker connecting the C-terminal and catalytic core domains are critical for MVV IN strand transfer activity in vitro and for virus infectivity. Single-molecule microscopy in conjunction with photobleaching reveals that the MVV intasome can bind a variable number, up to sixteen molecules, of the lentivirus-specific host factor LEDGF/p75. Concordantly, ablation of endogenous LEDGF/p75 results in gross redistribution of MVV integration sites in human and ovine cells. Our data confirm the importance of the expanded architecture observed in cryo-EM studies of lentiviral intasomes and suggest that this organization underlies multivalent interactions with chromatin for integration targeting to active genes.

SERPINE1 rs6092 Variant Is Related to Plasma Coagulation Proteins in Patients with Severe COVID-19 from a Tertiary Care Hospital.

An impaired coagulation process has been described in patients with severe or critical coronavirus disease (COVID-19). Nevertheless, the implication of coagulation-related genes has not been explored. We aimed to evaluate the impact of F5 rs6025 and SERPINE1 rs6092 on invasive mechanical ventilation (IMV) requirement and the levels of coagulation proteins among patients with severe COVID-19. Four-hundred fifty-five patients with severe COVID-19 were genotyped using TaqMan assays. Coagulation-related proteins (P-Selectin, D-dimer, P-selectin glycoprotein ligand-1, tissue plasminogen activator [tPA], plasminogen activator inhibitor-1, and Factor IX) were assessed by cytometric bead arrays in one- and two-time determinations. Accordingly, SERPINE1 rs6092, P-Selectin (GG 385 pg/mL vs. AG+AA 632 pg/mL, p = 0.0037), and tPA (GG 1858 pg/mL vs. AG+AA 2546 pg/mL, p = 0.0284) levels were different. Patients carrying the CT F5-rs6025 genotype exhibited lower levels of factor IX (CC 17,136 pg/mL vs. CT 10,247 pg/mL, p = 0.0355). Coagulation proteins were also different among IMV patients than non-IMV. PSGL-1 levels were significantly increased in the late stage of COVID-19 (>10 days). The frequencies of F5 rs6025 and SERPINE1 rs6092 variants were not different among IMV and non-IMV. The SERPINE1 rs6092 variant is related to the impaired coagulation process in patients with COVID-19 severe.

Incremental PCA algorithm for fringe pattern demodulation.

This work proposes a new algorithm for demodulating fringe patterns using principal component analysis (PCA). The algorithm is based on the incremental implantation of the singular value decomposition (SVD) technique for computing the principal values associated with a set of fringe patterns. Instead of processing an entire set of interferograms, the proposed algorithm proceeds in an incremental way, processing sequentially one (as minimum) interferogram at a given time. The advantages of this procedure are twofold. Firstly, it is not necessary to store the whole set of images in memory, and, secondly, by computing a phase quality parameter, it is possible to determine the minimum number of images necessary to accurately demodulate a given set of interferograms. The proposed algorithm has been tested for synthetic and experimental interferograms showing a good performance.

TNFRSF1B and TNF Variants Are Associated With Differences in Levels of Soluble Tumor Necrosis Factor Receptors in Patients With Severe COVID-19.

BACKGROUND: The impact of genetic variants in the expression of tumor necrosis factor-α (TNF-α) and its receptors in coronavirus disease 2019 (COVID-19) severity has not been previously explored. We evaluated the association of TNF (rs1800629 and rs361525), TNFRSF1A (rs767455 and rs1800693), and TNFRSF1B (rs1061622 and rs3397) variants with COVID-19 severity, assessed as invasive mechanical ventilation (IMV) requirement, and the plasma levels of soluble TNF-α, TNFR1, and TNFR2 in patients with severe COVID-19. METHODS: The genetic study included 1353 patients. Taqman assays were used to assess the genetic variants. ELISA was used to determine soluble TNF-α, TNFR1, and TNFR2 in plasma samples from 334 patients. RESULTS: Patients carrying TT (TNFRSF1B rs3397) exhibited lower PaO2/FiO2 levels than those with CT + CC genotypes. Differences in plasma levels of TNFR1 and TNFR2 were observed according to the genotype of TNFRSF1B rs1061622, TNF rs1800629, and rs361525. According to the studied genetic variants, there were no differences in the soluble TNF-α levels. Higher soluble TNFR1 and TNFR2 levels were detected in patients with COVID-19 requiring IMV. CONCLUSIONS: Genetic variants in TNF and TNFRSFB1 influence the plasma levels of soluble TNFR1 and TNFR2, implicated in COVID-19 severity.

Enhancement of Cryo-EM maps by a multiscale tubular filter.

We present an approach to enhance cryo-electron microscopy (cryo-EM) postprocessed maps based on a multiscale tubular filter. The method determines a tubularness measure locally by the analysis of the eigenvalues of the Hessian matrix. This information is used to enhance elongated local structures and to attenuate blob-like and plate-like structures. The approach, thus, introduces a priori information in the reconstructions to improve their interpretability and analysis at high-resolution. The proposed method has been tested with simulated and real cryo-EM maps including recent reconstructions of the SARS-CoV-2. Our results show that our methods can improve obtained reconstructions.

A panoramic view and swot analysis of artificial intelligence for achieving the sustainable development goals by 2030: progress and prospects.

The17 Sustainable Development Goals (SDGs) established by the United Nations Agenda 2030 constitute a global blueprint agenda and instrument for peace and prosperity worldwide. Artificial intelligence and other digital technologies that have emerged in the last years, are being currently applied in virtually every area of society, economy and the environment. Hence, it is unsurprising that their current role in the pursuance or hampering of the SDGs has become critical. This study aims at providing a snapshot and comprehensive view of the progress made and prospects in the relationship between artificial intelligence technologies and the SDGs. A comprehensive review of existing literature has been firstly conducted, after which a series SWOT (Strengths, Weaknesses, Opportunities and Threats) analyses have been undertaken to identify the strengths, weaknesses, opportunities and threats inherent to artificial intelligence-driven technologies as facilitators or barriers to each of the SDGs. Based on the results of these analyses, a subsequent broader analysis is provided, from a position vantage, to (i) identify the efforts made in applying AI technologies in SDGs, (ii) pinpoint opportunities for further progress along the current decade, and (iii) distill ongoing challenges and target areas for important advances. The analysis is organized into six categories or perspectives of human needs: life, economic and technological development, social development, equality, resources and natural environment. Finally, a closing discussion is provided about the prospects, key guidelines and lessons learnt that should be adopted for guaranteeing a positive shift of artificial intelligence developments and applications towards fully supporting the SDGs attainment by 2030.

Advances in Xmipp for Cryo-Electron Microscopy: From Xmipp to Scipion.

Xmipp is an open-source software package consisting of multiple programs for processing data originating from electron microscopy and electron tomography, designed and managed by the Biocomputing Unit of the Spanish National Center for Biotechnology, although with contributions from many other developers over the world. During its 25 years of existence, Xmipp underwent multiple changes and updates. While there were many publications related to new programs and functionality added to Xmipp, there is no single publication on the Xmipp as a package since 2013. In this article, we give an overview of the changes and new work since 2013, describe technologies and techniques used during the development, and take a peek at the future of the package.

Differential Genomic Profile in TERT, DSP, and FAM13A Between COPD Patients With Emphysema, IPF, and CPFE Syndrome.

Background: Genetic association studies have identified single nucleotide polymorphisms (SNPs) associated with lasting lung diseases such as Chronic Obstructive Pulmonary Disease (COPD) and Idiopathic Pulmonary Fibrosis (IPF), as well as the simultaneous presentation, known as Combined Pulmonary Fibrosis and Emphysema (CPFE) Syndrome. It is unknown if these diseases share genetic variants previously described in an independent way. This study aims to identify common or differential variants between COPD, IPF, and CPFE. Materials and methods: The association analysis was carried out through a case-control design in a Mexican mestizo population (n = 828); three patients' groups were included: COPD smokers (COPD-S, n = 178), IPF patients (n = 93), and CPFE patients (n = 16). Also, two comparison groups were analyzed: smokers without COPD (SWOC, n = 367) and healthy subjects belonging to the Mexican Pulmonary Aging Cohort (PAC, n = 174). Five SNPs in four genes previously associated to interstitial and obstructive diseases were selected: rs2609255 (FAM13A), rs2736100 (TERT), rs2076295 (DSP) rs5743890, and rs111521887 (TOLLIP). Genotyping was performed by qPCR using predesigned Taqman probes. Results: In comparing IPF vs. PAC, significant differences were found in the frequency of the rs260955 G allele associated with the IPF risk (OR = 1.68, p = 0.01). Also, the genotypes, GG of rs260955 (OR = 2.86, p = 0.01) and TT of rs2076295 (OR = 1.79, p = 0.03) were associated with an increased risk of IPF; after adjusting by covariables, only the rs260955 G allele remain significant (p = 0.01). For the CPFE vs. PAC comparison, an increased CPFE risk was identified since there is a difference in the rs2736100 C allele (OR = 4.02, p < 0.01; adjusted p < 0.01). For COPD-S, the rs2609255 TG genotype was associated with increased COPD risk after adjusting by covariables. Conclusion: The rs2736100 C allele is associated with decreased IPF risk and confers an increased risk for CPFE. Also, the rs2076295 TT genotype is associated with increased IPF risk, while the GG genotype is associated with CFPE susceptibility. The rs2609255 G allele and GG genotype are associated with IPF susceptibility, while the TG genotype is present in patients with emphysema.

DeepEMhancer: a deep learning solution for cryo-EM volume post-processing.

Cryo-EM maps are valuable sources of information for protein structure modeling. However, due to the loss of contrast at high frequencies, they generally need to be post-processed to improve their interpretability. Most popular approaches, based on global B-factor correction, suffer from limitations. For instance, they ignore the heterogeneity in the map local quality that reconstructions tend to exhibit. Aiming to overcome these problems, we present DeepEMhancer, a deep learning approach designed to perform automatic post-processing of cryo-EM maps. Trained on a dataset of pairs of experimental maps and maps sharpened using their respective atomic models, DeepEMhancer has learned how to post-process experimental maps performing masking-like and sharpening-like operations in a single step. DeepEMhancer was evaluated on a testing set of 20 different experimental maps, showing its ability to reduce noise levels and obtain more detailed versions of the experimental maps. Additionally, we illustrated the benefits of DeepEMhancer on the structure of the SARS-CoV-2 RNA polymerase.

Robust weighted principal components analysis demodulation algorithm for phase-shifting interferometry.

We present an asynchronous phase-shifting demodulation approach based on the principal component analysis demodulation method that is robust to typical problems as turbulence, vibrations, and temporal instabilities of the optical setup. The method brings together a two-step and a phase-shifting asynchronous demodulation method to share their benefits while reducing their intrinsic limitations. Thus, the proposed approach is based on a two-fold process. First, the modulating phase is estimated from a two-step demodulation approach. Second, this information is used to compute weights to each phase-shifted pattern of the interferogram sequence, which are used in a novel weighted principal component demodulation approach. The proposed technique has been tested with simulated and real interferograms affected by turbulence and vibrations providing very satisfactory results in challenging cases.

Local computational methods to improve the interpretability and analysis of cryo-EM maps.

Cryo-electron microscopy (cryo-EM) maps usually show heterogeneous distributions of B-factors and electron density occupancies and are typically B-factor sharpened to improve their contrast and interpretability at high-resolutions. However, 'over-sharpening' due to the application of a single global B-factor can distort processed maps causing connected densities to appear broken and disconnected. This issue limits the interpretability of cryo-EM maps, i.e. ab initio modelling. In this work, we propose 1) approaches to enhance high-resolution features of cryo-EM maps, while preventing map distortions and 2) methods to obtain local B-factors and electron density occupancy maps. These algorithms have as common link the use of the spiral phase transformation and are called LocSpiral, LocBSharpen, LocBFactor and LocOccupancy. Our results, which include improved maps of recent SARS-CoV-2 structures, show that our methods can improve the interpretability and analysis of obtained reconstructions.

Alternative conformations and motions adopted by 30S ribosomal subunits visualized by cryo-electron microscopy.

It is only after recent advances in cryo-electron microscopy that it is now possible to describe at high-resolution structures of large macromolecules that do not crystalize. Purified 30S subunits interconvert between an "active" and "inactive" conformation. The active conformation was described by crystallography in the early 2000s, but the structure of the inactive form at high resolution remains unsolved. Here we used cryo-electron microscopy to obtain the structure of the inactive conformation of the 30S subunit to 3.6 Å resolution and study its motions. In the inactive conformation, an alternative base-pairing of three nucleotides causes the region of helix 44, forming the decoding center to adopt an unlatched conformation and the 3' end of the 16S rRNA positions similarly to the mRNA during translation. Incubation of inactive 30S subunits at 42°C reverts these structural changes. The air-water interface to which ribosome subunits are exposed during sample preparation also peel off some ribosomal proteins. Extended exposures to low magnesium concentrations make the ribosomal particles more susceptible to the air-water interface causing the unfolding of large rRNA structural domains. Overall, this study provides new insights about the conformational space explored by the 30S ribosomal subunit when the ribosomal particles are free in solution.

Acute administration of levetiracetam in tonic pain model modulates gene expression of 5HT1A and 5HT7 receptors in the thalamus of rats (Rattus norvergicus).

The nociceptive effect of Levetiracetam (LEV) on the expression of 5-HT1A and 5-HT7 receptors found in the thalamus was evaluated. Thirty-six male rats (Wistar) were randomized into six groups: in the Control group without treatment; LEV50 group LEV was administered in a single dose of 50 mg/kg i.g.; in the LEV300 group LEV dose of 300 mg/kg i.g.; in the FORMALIN group the formalin test was performed; in the LEV50/FORMALIN group LEV dose of 50 mg/kg i.g and the formalin test was performed; in the LEV300/FORMALIN group LEV dose of 300 mg/kg i.g and the formalin test was performed, subsequently the thalamus was dissected in all groups. In the formalin tests LEV exhibited an antinociceptive effect in the LEV300/FORMALIN group (p < 0.05) and a pronociceptive effect in the LEV50/FORMALIN group (p < 0.001). The results obtained by Real-time PCR confirmed the expression of the 5-HT1A and 5-HT7 receptors in the thalamus, 5-HT1A receptors increased significantly in the FORMALIN group and the LEV300/FORMALIN group (p < 0.05). 5-HT7 receptors are only over expressed at a dose of 300 mg/Kg of LEV with formalin (p < 0.05). This suggests that LEV modulates the sensation of pain by controlling the expression of 5-HT1A and 5-HT7 in a tonic pain model, and that changes in the expression of 5-HT1A and 5-HT7 receptors are associated with the sensation of pain, furthermore its possibility to be used in clinical treatments for pain.