Nebulized enriched heparin improves respiratory parameters in patients with COVID-19: a phase I/II randomized and triple-blind clinical trial.

To evaluate the safety and the potential antiviral treatment of inhaled enriched heparin in patients with COVID-19. The specific objectives were to investigate the anticoagulation profile, antiviral and anti-inflammatory effects, and respiratory evolution of inhaled enriched heparin. We conducted a randomized, triple-blind, placebo-controlled Phase I/II clinical trial in hospitalized adults with COVID-19 receiving inhalation of enriched heparin or saline (placebo) every 4 h for 7 days. Among the 27 patients who completed the study, no changes in blood coagulation parameters were observed, indicating the safety of inhaled enriched heparin. The group receiving enriched heparin showed a significant reduction in the need for supplemental oxygen and improvement in respiratory parameters, such as the PaO2/FiO2 ratio. Inhalation of enriched heparin is shown to be safe and has also demonstrated potential therapeutic benefits for patients with COVID-19. These promising results justify the continuation of the study to the next phase, Phase II/III, to further evaluate the therapeutic efficacy of inhaled enriched heparin in the treatment of COVID-19-associated viral pneumonia.Trial registration: ClinicalTrials.gov. 08/02/2021. Identifier: NCT04743011.

Dichotomous outcomes vs. survival regression models for identification of predictors of mortality among patients with severe acute respiratory illness during COVID-19 pandemics.

Introduction: As the studies predicting mortality in severe acute respiratory illness (SARI) have inferred associations either from dichotomous outcomes or from time-event models, we identified some clinical-epidemiological characteristics and predictors of mortality by comparing and discussing two multivariate models. Methods: To identify factors associated with death among all SARI hospitalizations occurred in Botucatu (Brazil)/regardless of the infectious agent, and among the COVID-19 subgroup, from March 2020 to 2022, we used a multivariate Poisson regression model with binomial outcomes and Cox proportional hazards (time-event). The performance metrics of both models were also analyzed. Results: A total of 3,995 hospitalized subjects were included, of whom 1338 (33%) tested positive for SARS-CoV-2. We identified 866 deaths, of which 371 (43%) were due to the COVID-19. In the total number of SARI cases, using both Poisson and Cox models, the predictors of mortality were the presence of neurological diseases, immunosuppression, obesity, older age, and need for invasive ventilation support. However, the Poisson test also revealed that admission to an intensive care unit and the COVID-19 diagnosis were predictors of mortality, with the female gender having a protective effect against death. Likewise, Poisson proved to be more sensitive and specific, and indeed the most suitable model for analyzing risk factors for death in patients with SARI/COVID-19. Conclusion: Given these results and the acute course of SARI and COVID-19, to compare the associations and their different meanings is essential and, therefore, models with dichotomous outcomes are more appropriate than time-to-event/survival approaches.

Integrative Analysis of the Ethanol Tolerance of Saccharomyces cerevisiae.

Ethanol (EtOH) alters many cellular processes in yeast. An integrated view of different EtOH-tolerant phenotypes and their long noncoding RNAs (lncRNAs) is not yet available. Here, large-scale data integration showed the core EtOH-responsive pathways, lncRNAs, and triggers of higher (HT) and lower (LT) EtOH-tolerant phenotypes. LncRNAs act in a strain-specific manner in the EtOH stress response. Network and omics analyses revealed that cells prepare for stress relief by favoring activation of life-essential systems. Therefore, longevity, peroxisomal, energy, lipid, and RNA/protein metabolisms are the core processes that drive EtOH tolerance. By integrating omics, network analysis, and several other experiments, we showed how the HT and LT phenotypes may arise: (1) the divergence occurs after cell signaling reaches the longevity and peroxisomal pathways, with CTA1 and ROS playing key roles; (2) signals reaching essential ribosomal and RNA pathways via SUI2 enhance the divergence; (3) specific lipid metabolism pathways also act on phenotype-specific profiles; (4) HTs take greater advantage of degradation and membraneless structures to cope with EtOH stress; and (5) our EtOH stress-buffering model suggests that diauxic shift drives EtOH buffering through an energy burst, mainly in HTs. Finally, critical genes, pathways, and the first models including lncRNAs to describe nuances of EtOH tolerance are reported here.

Retrospective Insights of the COVID-19 Epidemic in the Major Latin American City, São Paulo, Southeastern Brazil.

São Paulo is the financial center of Brazil, with a population of over 12 million, that receives travelers from all over the world for business and tourism. It was the first city in Brazil to report a case of COVID-19 that rapidly spread across the city despite the implementation of the restriction measures. Despite many reports, much is still unknown regarding the genomic diversity and transmission dynamics of this virus in the city of São Paulo. Thus, in this study, we provide a retrospective overview of the COVID-19 epidemic in São Paulo City, Southeastern, Brazil, by generating a total of 9995 near-complete genome sequences from all the city's different macro-regions (North, West, Central, East, South, and Southeast). Our analysis revealed that multiple independent introduction events of different variants (mainly Gamma, Delta, and Omicron) occurred throughout time. Additionally, our estimates of viral movement within the different macro-regions further suggested that the East and the Southeast regions were the largest contributors to the Gamma and Delta viral exchanges to other regions. Meanwhile, the North region had a higher contribution to the dispersion of the Omicron variant. Together, our results reinforce the importance of increasing SARS-CoV-2 genomic monitoring within the city and the country to track the real-time evolution of the virus and to detect earlier any eventual emergency of new variants of concern that could undermine the fight against COVID-19 in Brazil and worldwide.

Degree of piRNA sharing and Piwi gene expression in the skeletal muscle of Piaractus mesopotamicus (pacu), Colossoma macropomum (tambaqui), and the hybrid tambacu.

PiRNAs are a class of small noncoding RNAs that, in their mature form, bind to Piwi proteins to repress transposable element activity. Besides their role in gametogenesis and genome integrity, recent evidence indicates their action in non-germinative tissues. We performed a global analysis of piRNA and Piwi gene expression in the skeletal muscle of juveniles pacu (Piaractus mesopotamicus), tambaqui (Colossoma macropomum), and the hybrid tambacu to evaluate the degree of piRNA sharing among these three genotypes. Total RNA was sequenced and analyzed using specific parameters of piRNAs by bioinformatics tools. piRNA and Piwi gene expression was analyzed by RT-qPCR. We detected 24 piRNA clusters common to the three genotypes, with eight shared between pacu and tambacu, three between pacu and tambaqui, and five between tambaqui and tambacu; seven, five, and four clusters were unique to pacu, tambacu, and tambaqui, respectively. Genomic localization and fold change values showed two clusters and 100 piRNAs shared among the three genotypes. The gene expression of four piRNAs was evaluated to validate our bioinformatics results. piRNAs from cluster 17 were higher in tambacu than pacu and piRNAs from cluster 18 were more highly expressed in tambacu than tambaqui and pacu. In addition, the expression of Piwis 1 and 2 was higher in tambacu and tambaqui than pacu. Our results open an important window to investigate whether these small noncoding RNAs benefit the hybrid in terms of faster growth and offer a new perspective on the function of piRNAs and Piwis in fish skeletal muscle.

Decreased miR-497-5p Suppresses IL-6 Induced Atrophy in Muscle Cells.

Interleukin-6 (IL-6) is a pro-inflammatory cytokine associated with skeletal muscle wasting in cancer cachexia. The control of gene expression by microRNAs (miRNAs) in muscle wasting involves the regulation of thousands of target transcripts. However, the miRNA-target networks associated with IL6-induced muscle atrophy remain to be characterized. Here, we show that IL-6 promotes the atrophy of C2C12 myotubes and changes the expression of 20 miRNAs (5 up-regulated and 15 down-regulated). Gene Ontology analysis of predicted miRNAs targets revealed post-transcriptional regulation of genes involved in cell differentiation, apoptosis, migration, and catabolic processes. Next, we performed a meta-analysis of miRNA-published data that identified miR-497-5p, a down-regulated miRNAs induced by IL-6, also down-regulated in other muscle-wasting conditions. We used miR-497-5p mimics and inhibitors to explore the function of miR-497-5p in C2C12 myoblasts and myotubes. We found that miR-497-5p can regulate the expression of the cell cycle genes CcnD2 and CcnE1 without affecting the rate of myoblast cellular proliferation. Notably, miR-497-5p mimics induced myotube atrophy and reduced Insr expression. Treatment with miR-497-5p inhibitors did not change the diameter of the myotubes but increased the expression of its target genes Insr and Igf1r. These genes are known to regulate skeletal muscle regeneration and hypertrophy via insulin-like growth factor pathway and were up-regulated in cachectic muscle samples. Our miRNA-regulated network analysis revealed a potential role for miR-497-5p during IL6-induced muscle cell atrophy and suggests that miR-497-5p is likely involved in a compensatory mechanism of muscle atrophy in response to IL-6.

Virucidal Activity of the Antiseptic Mouthwash and Dental Gel Containing Anionic Phthalocyanine Derivative: In vitro Study.

AIM: This research suggested an in vitro virucidal action of a dental gel and a mouthwash with phthalocyanine derivative. PURPOSE: The aim of this study was to report an in vitro study evaluating the virucidal capacity of mouthwash and dental gel containing anionic phthalocyanine derivate (APD). METHODS: The research followed the recommendations of the National Health Surveillance Agency (ANVISA) and adapted methodology, described in the standards EN14776: 2015; ASTM E1053-11 and the Robert Koch Institute - RKI, in addition to good laboratory practices (GLP). The determination of the percentage of inactivation of the SARS-CoV-2 virus particles was carried out by imposing the viral solution in contact with the respective tested products, with intervals of 30 seconds, 1 and 5 minutes, with subsequent submission of the aliquots, recovered in cell culture microplates following virus titration using the TCID50 (50% Median Tissue Culture Infectious Dose). RESULTS: The Mouthwash APD presented 90% of viral inactivation percentage, while the dental gel APD demonstrated 99.99% of viral inactivation. CONCLUSION: In vitro analyses showed that mouthwash and dental gel APD can reduce the viability of SARS-CoV-2 virus particles.

Comparison of microRNA Expression Profile in Chronic Myeloid Leukemia Patients Newly Diagnosed and Treated by Allogeneic Hematopoietic Stem Cell Transplantation.

Chronic myeloid leukemia (CML) results from a translocation between chromosomes 9 and 22, which generates the Philadelphia chromosome. This forms BCR/ABL1, an active tyrosine kinase protein that promotes cell growth and replication. Despite great progress in CML treatment in the form of tyrosine kinase inhibitors, allogeneic-hematopoietic stem cell transplantation (allo-HSCT) is currently used as an important treatment alternative for patients resistant to these inhibitors. Studies have shown that unregulated expression of microRNAs, which act as oncogenes or tumor suppressors, is associated with human cancers. This contributes to tumor formation and development by stimulating proliferation, angiogenesis, and invasion. Research has demonstrated the potential of microRNAs as biomarkers for cancer diagnosis, prognosis, and therapeutic targets. In the present study, we compared the circulating microRNA expression profiles of 14 newly diagnosed patients with chronic phase-CML and 14 Philadelphia chromosome-negative patients after allo-HSCT. For each patient, we tested 758 microRNAs by reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis. The global expression profile of microRNAs revealed 16 upregulated and 30 downregulated microRNAs. Target genes were analyzed, and key pathways were extracted and compared. Bioinformatics tools were used to analyze data. Among the downregulated miRNA target genes, some genes related to cell proliferation pathways were identified. These results reveal the comprehensive microRNA profile of CML patients and the main pathways related to the target genes of these miRNAs in cytogenetic remission after allo-HSCT. These results provide new resources for exploring stem cell transplantation-based CML treatment strategies.

MicroRNA-mRNA Co-sequencing Identifies Transcriptional and Post-transcriptional Regulatory Networks Underlying Muscle Wasting in Cancer Cachexia.

Cancer cachexia is a metabolic syndrome with alterations in gene regulatory networks that consequently lead to skeletal muscle wasting. Integrating microRNAs-mRNAs omics profiles offers an opportunity to understand transcriptional and post-transcriptional regulatory networks underlying muscle wasting. Here, we used RNA sequencing to simultaneously integrate and explore microRNAs and mRNAs expression profiles in the tibialis anterior (TA) muscles of the Lewis Lung Carcinoma (LLC) model of cancer cachexia. We found 1,008 mRNAs and 18 microRNAs differentially expressed in cachectic mice compared with controls. Although our transcriptomic analysis demonstrated a high heterogeneity in mRNA profiles of cachectic mice, we identified a reduced number of differentially expressed genes that were uniformly regulated within cachectic muscles. This set of uniformly regulated genes is associated with the extracellular matrix (ECM), proteolysis, and inflammatory response. We also used transcriptomic data to perform enrichment analysis of transcriptional factor binding sites in promoter sequences, which revealed activation of the atrophy-related transcription factors NF-κB, Stat3, AP-1, and FoxO. Furthermore, the integration of mRNA and microRNA expression profiles identified post-transcriptional regulation by microRNAs of genes involved in ECM organization, cell migration, transcription factors binding, ion transport, and the FoxO signaling pathway. Our integrative analysis of microRNA-mRNA co-profiles comprehensively characterized regulatory relationships of molecular pathways and revealed microRNAs targeting ECM-associated genes in cancer cachexia.

Defining suitable reference genes for RT-qPCR analysis on human sertoli cells after 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure.

Quantitative real-time RT-PCR (qPCR) has proven to be a valuable molecular technique to quantify gene expression. There are few studies in the literature that describe suitable reference genes to normalize gene expression data. Studies of transcriptionally disruptive toxins, like tetrachlorodibenzo-p-dioxin (TCDD), require careful consideration of reference genes. The present study was designed to validate potential reference genes in human Sertoli cells after exposure to TCDD. 32 candidate reference genes were analyzed to determine their applicability. geNorm and NormFinder softwares were used to obtain an estimation of the expression stability of the 32 genes and to identify the most suitable genes for qPCR data normalization.

Involvement of renal corpuscle microRNA expression on epithelial-to-mesenchymal transition in maternal low protein diet in adult programmed rats.

Prior study shows that maternal protein-restricted (LP) 16-wk-old offspring have pronounced reduction of nephron number and arterial hypertension associated with unchanged glomerular filtration rate, besides enhanced glomerular area, which may be related to glomerular hyperfiltration/overflow and which accounts for the glomerular filtration barrier breakdown and early glomerulosclerosis. In the current study, LP rats showed heavy proteinuria associated with podocyte simplification and foot process effacement. TGF-ß1 glomerular expression was significantly enhanced in LP. Isolated LP glomeruli show a reduced level of miR-200a, miR-141, miR-429 and ZEB2 mRNA and upregulated collagen 1α1/2 mRNA expression. By western blot analyzes of whole kidney tissue, we found significant reduction of both podocin and nephrin and enhanced expression of mesenchymal protein markers such as desmin, collagen type I and fibronectin. From our present knowledge, these are the first data showing renal miRNA modulation in the protein restriction model of fetal programming. The fetal-programmed adult offspring showed pronounced structural glomerular disorders with an accentuated and advanced stage of fibrosis, which led us to state that the glomerular miR-200 family would be downregulated by TGF-ß1 action inducing ZEB 2 expression that may subsequently cause glomeruli epithelial-to-mesenchymal transition.