Unveiling the Antiviral Capabilities of Targeting Human Dihydroorotate Dehydrogenase against SARS-CoV-2.

The urgent need for effective treatments against emerging viral diseases, driven by drug-resistant strains and new viral variants, remains critical. We focus on inhibiting the human dihydroorotate dehydrogenase (HsDHODH), one of the main enzymes responsible for pyrimidine nucleotide synthesis. This strategy could impede viral replication without provoking resistance. We evaluated naphthoquinone fragments, discovering potent HsDHODH inhibition with IC50 ranging from 48 to 684 nM, and promising in vitro anti-SARS-CoV-2 activity with EC50 ranging from 1.2 to 2.3 µM. These compounds exhibited low toxicity, indicating potential for further development. Additionally, we employed computational tools such as molecular docking and quantitative structure-activity relationship (QSAR) models to analyze protein-ligand interactions, revealing that these naphthoquinones exhibit a protein binding pattern similar to brequinar, a potent HsDHODH inhibitor. These findings represent a significant step forward in the search for effective antiviral treatments and have great potential to impact the development of new broad-spectrum antiviral drugs.

Cryptic circulation of chikungunya virus in São Jose do Rio Preto, Brazil, 2015-2019.

BACKGROUND: Chikungunya virus (CHIKV) has spread across Brazil with varying incidence rates depending on the affected areas. Due to cocirculation of arboviruses and overlapping disease symptoms, CHIKV infection may be underdiagnosed. To understand the lack of CHIKV epidemics in São José do Rio Preto (SJdRP), São Paulo (SP), Brazil, we evaluated viral circulation by investigating anti-CHIKV IgG seroconversion in a prospective study of asymptomatic individuals and detecting anti-CHIKV IgM in individuals suspected of dengue infection, as well as CHIKV presence in Aedes mosquitoes. The opportunity to assess two different groups (symptomatic and asymptomatic) exposed at the same geographic region aimed to broaden the possibility of identifying the viral circulation, which had been previously considered absent. METHODOLOGY/PRINCIPAL FINDINGS: Based on a prospective population study model and demographic characteristics (sex and age), we analyzed the anti-CHIKV IgG seroconversion rate in 341 subjects by ELISA over four years. The seroprevalence increased from 0.35% in the first year to 2.3% after 3 years of follow-up. Additionally, we investigated 497 samples from a blood panel collected from dengue-suspected individuals during the 2019 dengue outbreak in SJdRP. In total, 4.4% were positive for anti-CHIKV IgM, and 8.6% were positive for IgG. To exclude alphavirus cross-reactivity, we evaluated the presence of anti-Mayaro virus (MAYV) IgG by ELISA, and the positivity rate was 0.3% in the population study and 0.8% in the blood panel samples. In CHIKV and MAYV plaque reduction neutralization tests (PRNTs), the positivity rate for CHIKV-neutralizing antibodies in these ELISA-positive samples was 46.7%, while no MAYV-neutralizing antibodies were detected. Genomic sequencing and phylogenetic analysis revealed CHIKV genotype ECSA in São José do Rio Preto, SP. Finally, mosquitoes collected to complement human surveillance revealed CHIKV positivity of 2.76% of A. aegypti and 9.09% of A. albopictus (although it was far less abundant than A. aegypti) by RT-qPCR. CONCLUSIONS/SIGNIFICANCE: Our data suggest cryptic CHIKV circulation in SJdRP detected by continual active surveillance. These low levels, but increasing, of viral circulation highlight the possibility of CHIKV outbreaks, as there is a large naïve population. Improved knowledge of the epidemiological situation might aid in outbreaks prevention.

Severe influenza infection is associated with inflammatory programmed cell death in infected macrophages.

Introduction: Influenza A virus (IAV) is one of the leading causes of respiratory tract infections in humans, representing a major public health concern. The various types of cell death have a crucial role in IAV pathogenesis because this virus may trigger both apoptosis and necroptosis in airway epithelial cells in parallel. Macrophages play an important role in the clearance of virus particles, priming the adaptive immune response in influenza. However, the contribution of macrophage death to pathogenesis of IAV infection remains unclear. Methods: In this work, we investigated IAV-induced macrophage death, along with potential therapeutic intervention. We conducted in vitro and in vivo experiments to evaluate the mechanism and the contribution of macrophages death to the inflammatory response induced by IAV infection. Results: We found that IAV or its surface glycoprotein hemagglutinin (HA) triggers inflammatory programmed cell death in human and murine macrophages in a Toll-like receptor-4 (TLR4)- and TNF-dependent manner. Anti-TNF treatment in vivo with the clinically approved drug etanercept prevented the engagement of the necroptotic loop and mouse mortality. Etanercept impaired the IAV-induced proinflammatory cytokine storm and lung injury. Conclusion: In summary, we demonstrated a positive feedback loop of events that led to necroptosis and exacerbated inflammation in IAV-infected macrophages. Our results highlight an additional mechanism involved in severe influenza that could be attenuated with clinically available therapies.

Metabolic reprogramming and lipid droplets are involved in Zika virus replication in neural cells.

Zika virus (ZIKV) infection is a global public health concern linked to adult neurological disorders and congenital diseases in newborns. Host lipid metabolism, including lipid droplet (LD) biogenesis, has been associated with viral replication and pathogenesis of different viruses. However, the mechanisms of LD formation and their roles in ZIKV infection in neural cells are still unclear. Here, we demonstrate that ZIKV regulates the expression of pathways associated with lipid metabolism, including the upregulation and activation of lipogenesis-associated transcription factors and decreased expression of lipolysis-associated proteins, leading to significant LD accumulation in human neuroblastoma SH-SY5Y cells and in neural stem cells (NSCs). Pharmacological inhibition of DGAT-1 decreased LD accumulation and ZIKV replication in vitro in human cells and in an in vivo mouse model of infection. In accordance with the role of LDs in the regulation of inflammation and innate immunity, we show that blocking LD formation has major roles in inflammatory cytokine production in the brain. Moreover, we observed that inhibition of DGAT-1 inhibited the weight loss and mortality induced by ZIKV infection in vivo. Our results reveal that LD biogenesis triggered by ZIKV infection is a crucial step for ZIKV replication and pathogenesis in neural cells. Therefore, targeting lipid metabolism and LD biogenesis may represent potential strategies for anti-ZIKV treatment development.

Enzymatic Cocktail Formulation for Xylan Hydrolysis into Xylose and Xylooligosaccharides.

In the context of a biorefinery, lignocellulosic materials represent an important source of raw material for the bioconversion of cellulose, hemicellulose, and lignin into value-added products, such as xylose for fermentation, oligosaccharides, and bioplastics for packaging. Among the most abundant lignocellulosic materials in Brazil, sugarcane bagasse biomass stands out, as it is rich in cellulose and hemicellulose. In this context, through an experimental design, this study developed a robust enzyme cocktail containing xylanases and accessory enzymes to complete the hydrolysis of xylan from sugarcane bagasse, obtaining a low xylose yield and concentration (9% and 1.8 g/L, respectively, observed in experiment number 16 from the complete hydrolysis of a xylan assay), a fermentable sugar that is important in the production of second-generation ethanol, and a high xylooligosaccharides (XOS) yield and concentration (93.1% and 19.6 g/L, respectively, obtained from a xylooligosaccharides production assay); in general, xylan has prebiotic activities that favor an improvement in intestinal functions, with immunological and antimicrobial actions and other benefits to human health. In addition to completely hydrolyzing the sugarcane bagasse xylan, this enzymatic cocktail has great potential to be applied in other sources of lignocellulosic biomass for the conversion of xylan into xylose and XOS due to its enzymes content, involving both main chain and pendant groups hydrolysis of hemicelluloses.

Agathisflavone, a natural biflavonoid that inhibits SARS-CoV-2 replication by targeting its proteases.

Despite the fast development of vaccines, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) still circulates through variants of concern (VoC) and escape the humoral immune response. SARS-CoV-2 has provoked over 200,000 deaths/months since its emergence and only a few antiviral drugs showed clinical benefit up to this moment. Thus, chemical structures endowed with anti-SARS-CoV-2 activity are important for continuous antiviral development and natural products represent a fruitful source of substances with biological activity. In the present study, agathisflavone (AGT), a biflavonoid from Anacardium occidentale was investigated as a candidate anti-SARS-CoV-2 compound. In silico and enzymatic analysis indicated that AGT may target mainly the viral main protease (Mpro) and not the papain-like protease (PLpro) in a non-competitive way. Cell-based assays in type II pneumocytes cell lineage (Calu-3) showed that SARS-CoV-2 is more susceptible to AGT than to apigenin (APG, monomer of AGT), in a dose-dependent manner, with an EC50 of 4.23 ± 0.21 µM and CC50 of 61.3 ± 0.1 µM and with a capacity to inhibit the level of pro-inflammatory mediator tumor necrosis factor-alpha (TNF-α). These results configure AGT as an interesting chemical scaffold for the development of novel semisynthetic antivirals against SARS-CoV-2.

Development of novel montmorillonite-based sustained release system for oral bromopride delivery.

The drug delivery systems are an important strategy of pharmaceutical technology to modulate undesirable properties, increasing efficacy, and reducing the side effects of active pharmaceutical ingredients (API). The sustained release is a type of controlled-release system that provides a suitable drug level in the blood through a slow release rate. An interesting alternative to achieve a controlled release is the application of carrier materials such as polymers, cyclodextrins, and clays. Sodium montmorillonite (Na-MMT) is a biocompatible natural clay that allows the insertion of organic compounds in interlamellar space, owing to its high cation exchange capacity and large internal surface area. Bromopride (BPD) is an aminated compound with antiemetic properties classified as class II (low solubility, high permeability) of the Biopharmaceutical Classification System (BCS). Herein, the aim of the study was the development and investigation of a drug delivery system formed by intercalation of BPD with Na-MMT. The results indicate the successful intercalation of this API with the lamellar silicate, meanwhile, there was no evidence of BPD intercalation in organic montmorillonite. The Na-MMT/BPD molecular complex exhibits a sustained release in performed assays. Molecular dynamics simulations suggested that BPD molecules interact with the montmorillonite layer through ion-dipole interactions and also between BPD molecules, forming hydrogen bonds web into montmorillonite interlayer space. The new drug delivery system showed an alternative to achieve the BPD sustained release, which may improve its pharmacological performance in therapeutic applications.

Reações contratransferenciais de psicoterapeutas frente a uma paciente borderline em psicoterapia: um estudo qualitativo / Countertransferential reactions of psychotherapists towards a borderline patient in psychotherapy: a qualitative study

A contratransferência (CT) é um elemento do relacionamento terapêutico que possui valor clínico, especialmente com pacientes com personalidade borderline (PB). Este estudo, qualitativo e exploratório, visou identificar os sentimentos despertados em psicoterapeutas frente a um caso de paciente com PB, buscando compreendê-los em relação às características da paciente ou da sua narrativa, bem como explorar de que forma a CT seria idealmente manejada. Oito psicoterapeutas assistiram ao vídeo de uma sessão real com paciente com PB e, após, responderam a uma entrevista. As transcrições das entrevistas foram analisadas com o método Consensual Qualitative Research (CQR). Os dados organizaram-se em relação às facetas da CT, características da paciente que mobilizam afetos, e manejo da CT. Os resultados sugerem que pacientes com PB tendem a suscitar sentimentos intensos, vinculados a sua história e seu funcionamento em situação observacional, apontando para a viabilidade do método para o estudo empírico da CT. Resultados de estudos como esse podem ser utilizados como guia para jovens terapeutas compreenderem o mundo interno dos seus pacientes. A validação empírica de hipóteses clínicas fortalece a teorização e enriquece a prática psicanalítica.(AU)

Countertransference (CT) is an element of the therapeutic relationship that has clinical value, especially with borderline personality patients (BP). This qualitative and exploratory study aimed to identify the feelings aroused in psychotherapists in the case of a BP patient, seeking to understand them in relation to the patient's characteristics or her narrative, as well as exploring how the CT would be ideally managed. Eight psychotherapists watched the video of a real session with a BP patient and responded to an interview. The interview transcripts were analyzed using the Consensual Qualitative Research (CQR) method. The data were organized in relation to the facets of the CT, characteristics of the patient that mobilize affections, and management of the CT. The results suggest that patients with BP tend to elicit intense feelings, linked to their history and functioning, in an observational situation, pointing to the feasibility of the method for the empirical study of CT. Results from studies like this one can be used as a guide for young therapists to understand the inner world of their patients. Empirical validation of clinical hypotheses strengthens theorization and enriches psychoanalytic practice.(AU)

La contratransferencia (CT) es un elemento de la relación terapéutica que tiene valor clínico, especialmente en pacientes con personalidad límite (BP). Este estudio cualitativo y exploratorio tuvo como objetivo identificar los sentimientos que despiertan los psicoterapeutas en el caso de un paciente con BP, buscando comprenderlos en relación con las características de la paciente o su narrativa, así como explorar cómo se manejaría idealmente la CT. Ocho psicoterapeutas vieron el video de una sesión real y luego respondieron a una entrevista. Las transcripciones de las entrevistas se analizaron utilizando el método de Investigación Cualitativa Consensual (CQR). Los datos se organizaron en relación a las facetas del CT, características del paciente que movilizan afectos y manejo de la CT. Los resultados sugieren que los pacientes con BP tienden a provocar sentimientos intensos, ligados a su historia y funcionamiento, en una situación de observación, lo que apunta a la viabilidad del método para el estudio empírico de la CT. Los resultados de estudios como este pueden usarse como una guía para que los terapeutas jóvenes comprendan el mundo interior de sus pacientes. La validación empírica de hipótesis clínicas fortalece la teorización y enriquece la práctica psicoanalítica.(AU)

VIP plasma levels associate with survival in severe COVID-19 patients, correlating with protective effects in SARS-CoV-2-infected cells.

Infection by SARS-CoV-2 may elicit uncontrolled and damaging inflammatory responses. Thus, it is critical to identify compounds able to inhibit virus replication and thwart the inflammatory reaction. Here, we show that the plasma levels of the immunoregulatory neuropeptide VIP are elevated in patients with severe COVID-19, correlating with reduced inflammatory mediators and with survival on those patients. In vitro, vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP), highly similar neuropeptides, decreased the SARS-CoV-2 RNA content in human monocytes and viral production in lung epithelial cells, also reducing cell death. Both neuropeptides inhibited the production of proinflammatory mediators in lung epithelial cells and in monocytes. VIP and PACAP prevented in monocytes the SARS-CoV-2-induced activation of NF-kB and SREBP1 and SREBP2, transcriptions factors involved in proinflammatory reactions and lipid metabolism, respectively. They also promoted CREB activation, a transcription factor with antiapoptotic activity and negative regulator of NF-kB. Specific inhibition of NF-kB and SREBP1/2 reproduced the anti-inflammatory, antiviral, and cell death protection effects of VIP and PACAP. Our results support further clinical investigations of these neuropeptides against COVID-19.

Simvastatin Downregulates the SARS-CoV-2-Induced Inflammatory Response and Impairs Viral Infection Through Disruption of Lipid Rafts.

Coronavirus disease 2019 (COVID-19) is currently a worldwide emergency caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). In observational clinical studies, statins have been identified as beneficial to hospitalized patients with COVID-19. However, experimental evidence of underlying statins protection against SARS-CoV-2 remains elusive. Here we reported for the first-time experimental evidence of the protective effects of simvastatin treatment both in vitro and in vivo. We found that treatment with simvastatin significantly reduced the viral replication and lung damage in vivo, delaying SARS-CoV-2-associated physiopathology and mortality in the K18-hACE2-transgenic mice model. Moreover, simvastatin also downregulated the inflammation triggered by SARS-CoV-2 infection in pulmonary tissue and in human neutrophils, peripheral blood monocytes, and lung epithelial Calu-3 cells in vitro, showing its potential to modulate the inflammatory response both at the site of infection and systemically. Additionally, we also observed that simvastatin affected the course of SARS-CoV-2 infection through displacing ACE2 on cell membrane lipid rafts. In conclusion, our results show that simvastatin exhibits early protective effects on SARS-CoV-2 infection by inhibiting virus cell entry and inflammatory cytokine production, through mechanisms at least in part dependent on lipid rafts disruption.

Low Food Consumption Interferes with the Nutritional Status of Surgical Patients with Neoplasia of the Gastrointestinal Tract.

Nutritional support strongly influence the nutritional status of the surgical neoplastic patients. This study aimed to evaluate the influence of food consumption on the perioperative nutritional status of hospitalized patients with neoplasia of the upper (UGIT) and lower (LGIT) gastrointestinal tract. METHOD: Observational, longitudinal, and prospective study. Data collected: food consumption, Subjective Global Assessment, anthropometry, laboratory tests. RESULTS: Eighty patients were followed up: 43 (54%) in the UGIT and 37 (46%) in the LGIT. The consumption in the perioperative period was lower than the usual consumption in the UGIT and LGIT groups, respectively, of energy (14.2 ± 6.5; 22.8 ± 11.2 Kcal/kg/d, p < 0.001; 13.6 ± 1.2; 19.0 ± 2.0 Kcal/kg/d; p = 0.014), protein (1.1 ± 0.7; 0.6 ± 0.3 g/kg/d, p < 0.001; 0.8 ± 0.1; 0.5 ± 0.1 g/kg/d; p = 0.058), selenium, zinc and copper. Most patients presented in the UGIT and LGIT groups, respectively, worsening malnutrition and muscle depletion according to the Subjective Global Assessment (61.9%; 51.4%) and hypoalbuminemia, mainly in the UGIT in the postoperative. CONCLUSION: Low food consumption during the perioperative period associated with prolongation of the postoperative fasting period worsens the nutritional status of patients undergoing surgery of the gastrointestinal tract for neoplasia, especially in the UGIT group.

Increased postoperative fasting time aggravates the nutritional status in patients with gastrointestinal tract neoplasia

ABSTRACT Surgical patients with gastrointestinal cancer often suffer from malnutrition. This study aimed to evaluate the influence of fasting time on the nutritional status of patients hospitalized with preoperative and postoperative gastrointestinal tract neoplasms. Observational, longitudinal, and prospective study conducted in the surgical unit at a public-school hospital. The patients were divided into groups: upper (UGIT) and lower (LGIT) gastrointestinal tract. Follow-up started within 72 h of hospitalization with reassessment 72 h after surgery. Data collected: sex, age, type and duration of surgery, preoperative (compared with 8 h) and postoperative (compared with 24 h) fasting time, food acceptance, Subjective Global Assessment, anthropometry, and laboratory tests. Analyses: Student t, Wilcoxon, and chi-square tests. Fifty-one patients were followed up, 29 (57%) UGIT and 22 (43%) LGIT. The preoperative fasting time was 8.2±2.8 h in UGIT and 8.1±2.2 h in LGIT groups, respectively; however, postoperative fasting times in UGIT (60.4±40.7 h) and LGIT groups (57.6±38.2 h) were longer than 24 h (P<0.001). Although eutrophic in the preoperative period, in the postoperative most patients in the UGIT and LGIT groups presented, respectively, malnutrition (71%; 59%; P<0.001), severe weight loss (79%; 80%), a significant correlation between triceps skinfold and postoperative fasting time (r= -0.306; P= 0.03), and hemoglobin and albumin values (r= 0.633; P<0.001), additionally low dietary acceptance, especially in the UGIT group. Prolonging postoperative fasting time worsened the nutritional status of surgical patients, especially in the UGIT group.

RESUMEN Los pacientes quirúrgicos con cáncer gastrointestinal a menudo sufren desnutrición. El objetivo de este estudio fue evaluar la influencia del tiempo de ayuno en estado nutricional de pacientes hospitalizados con neoplasias del tracto gastrointestinal preoperatorio y posoperatorio. Estudio observacional, longitudinal y prospectivo realizado en unidad quirúrgica de un hospital escolar público. Los pacientes fueron divididos en grupos: tracto gastrointestinal superior (UGIT) y inferior (LGIT). El seguimiento se inició dentro del 72 h de la hospitalización con reevaluación 72 h después de la cirugía. Datos recolectados: sexo, edad, tipo y duración de la cirugía, tiempo de ayuno preoperatorio (comparado con 8 h) y postoperatorio (comparado con 24 h), aceptación de dieta, Evaluación Subjetiva Global, antropometría y pruebas de laboratorio. Análisis: pruebas de T, Wilcoxon y chi-cuadrado. Se siguió 51 pacientes, 29 (57%) en UGIT y 22 (43%) en LGIT. El tiempo de ayuno preoperatorio fue 8,2±2,8 h (UGIT) y 8,1±2,2 h (LGIT); sin embargo, los tiempos de ayuno posoperatorio en UGIT (60,4±40,7 h) y LGIT (57,6±38,2 h) fueron superiores a 24 h (P<0,001). Aunque eutróficos en preoperatorio, en postoperatorio la mayoría de los pacientes (UGIT y LGIT, respectivamente) presentaron desnutrición (71%; 59%; P<0,001), pérdida de peso severa (79%; 80%), correlación significativa entre pliegue cutáneo del tríceps y tiempo de ayuno posoperatorio (r= -0.306; P= 0.03), valores de hemoglobina y albúmina (r= 0,633; P<0,001), y baja aceptación de dieta, especialmente del UGIT. La prolongación del ayuno postoperatorio empeoró el estado nutricional de los pacientes quirúrgicos, especialmente del UGIT.

In vitro antiviral activity of the anti-HCV drugs daclatasvir and sofosbuvir against SARS-CoV-2, the aetiological agent of COVID-19.

BACKGROUND: Current approaches of drug repurposing against COVID-19 have not proven overwhelmingly successful and the SARS-CoV-2 pandemic continues to cause major global mortality. SARS-CoV-2 nsp12, its RNA polymerase, shares homology in the nucleotide uptake channel with the HCV orthologue enzyme NS5B. Besides, HCV enzyme NS5A has pleiotropic activities, such as RNA binding, that are shared with various SARS-CoV-2 proteins. Thus, anti-HCV NS5B and NS5A inhibitors, like sofosbuvir and daclatasvir, respectively, could be endowed with anti-SARS-CoV-2 activity. METHODS: SARS-CoV-2-infected Vero cells, HuH-7 cells, Calu-3 cells, neural stem cells and monocytes were used to investigate the effects of daclatasvir and sofosbuvir. In silico and cell-free based assays were performed with SARS-CoV-2 RNA and nsp12 to better comprehend the mechanism of inhibition of the investigated compounds. A physiologically based pharmacokinetic model was generated to estimate daclatasvir's dose and schedule to maximize the probability of success for COVID-19. RESULTS: Daclatasvir inhibited SARS-CoV-2 replication in Vero, HuH-7 and Calu-3 cells, with potencies of 0.8, 0.6 and 1.1 µM, respectively. Although less potent than daclatasvir, sofosbuvir alone and combined with daclatasvir inhibited replication in Calu-3 cells. Sofosbuvir and daclatasvir prevented virus-induced neuronal apoptosis and release of cytokine storm-related inflammatory mediators, respectively. Sofosbuvir inhibited RNA synthesis by chain termination and daclatasvir targeted the folding of secondary RNA structures in the SARS-CoV-2 genome. Concentrations required for partial daclatasvir in vitro activity are achieved in plasma at Cmax after administration of the approved dose to humans. CONCLUSIONS: Daclatasvir, alone or in combination with sofosbuvir, at higher doses than used against HCV, may be further fostered as an anti-COVID-19 therapy.

SARS-CoV-2 engages inflammasome and pyroptosis in human primary monocytes.

Infection by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been associated with leukopenia and uncontrolled inflammatory response in critically ill patients. A better comprehension of SARS-CoV-2-induced monocyte death is essential for the identification of therapies capable to control the hyper-inflammation and reduce viral replication in patients with 2019 coronavirus disease (COVID-19). Here, we show that SARS-CoV-2 engages inflammasome and triggers pyroptosis in human monocytes, experimentally infected, and from patients under intensive care. Pyroptosis associated with caspase-1 activation, IL-1ß production, gasdermin D cleavage, and enhanced pro-inflammatory cytokine levels in human primary monocytes. At least in part, our results originally describe mechanisms by which monocytes, a central cellular component recruited from peripheral blood to respiratory tract, succumb to control severe COVID-19.

Correlação entre incontinência urinária, disfunção sexual e avaliação subjetiva da contração muscular perineal em idosas fisicamente ativas / Correlación entre incontinencia urinaria, disfunción sexual y evaluación subjetiva de la contracción del músculo perineal en mujeres ancianas físicamente activas / Urinary incontinence and female sexual dysfunction in physically active older women

RESUMO Incontinência urinária (IU) e disfunção sexual feminina (DSF) são disfunções que afetam a qualidade de vida. Com a mudança na pirâmide etária brasileira, a prevalência de tais disfunções tende a aumentar. O objetivo deste estudo foi estabelecer a relação entre a IU, a DSF e a contração muscular perineal em idosas ativas fisicamente. Trata-se de estudo transversal, do qual participaram 35 idosas ≥60 anos. Inicialmente foram aplicados a ficha diagnóstica e os questionários International Consultation on Incontinence Questionnaire - Short Form (ICIQ-UI-SF) e Female Sexual Function Index (FSFI). Em seguida foram avaliados os músculos do assoalho pélvico (MAP) por meio do toque vaginal, de acordo com o que propõe o esquema perfect. Entre as idosas, 20% apresentaram ambas as disfunções dos MAP. No total, 17 (48,6%) apresentavam IU e 16 (45,7%) DSF. Observou-se diferença significativa no número de partos vaginais (p=0,028) e no item rapidez do esquema perfect (p=0,033) entre as idosas com e sem DSF. Não houve diferença estatística entre os grupos com e sem IU. A análise apontou uma correlação inversamente proporcional entre a gravidade da IU e a função sexual nas idosas incontinentes (p=0,008; r=−0,622), de forma que, para cada aumento na gravidade da IU pelo ICIQ-SF, diminuiu-se 0,507 de função sexual avaliada pelo FSFI (p=0,034; r=0,516). Há uma associação entre IU e DSF em idosas fisicamente ativas. Algumas idosas apresentam ambas as disfunções dos MAP, apesar de serem ativas fisicamente. Dessa forma, é necessário reforçar a importância de avaliações minuciosas, da atividade física orientada e da atuação multiprofissional em saúde.

RESUMEN La incontinencia urinaria (IU) y la disfunción sexual femenina (DSF) son trastornos que afectan la calidad de vida de las personas. Con el cambio en la pirámide de edad brasileña se aumentará la prevalencia de tales disfunciones. El objetivo de este estudio fue establecer la relación entre la IU, la DSF y la contracción del músculo perineal en mujeres mayores físicamente activas. Este es un estudio transversal, en el que participaron 35 mujeres ≥60 años. Inicialmente se aplicó el formulario de diagnóstico y los cuestionarios International Consultation on Incontinence Questionnaire - Short Form (ICIQ-UI-SF) e Índice de Función Sexual Femenina (FSFI). Luego, se evaluó la musculatura del piso pélvico (MAP) mediante toque vaginal siguiendo lo que propone el esquema perfect. Entre las ancianas, el 20% tenía ambas disfunciones de MAP. En total, 17 (48,6%) tenía IU, y 16 (45,7%) DSF. Hubo una diferencia significativa en el número de partos vaginales (p=0,028) y en el ítem rapidez del esquema perfect (p=0,033) entre mujeres ancianas con y sin DSF. No hubo diferencia estadística entre los grupos con y sin IU. El análisis mostró una correlación inversamente proporcional entre la gravedad de la IU y la función sexual en mujeres ancianas con incontinencia (p=0,008; r=−0,622), de modo que por cada aumento en la gravedad de la IU por el ICIQ-SF se disminuyó 0,507 de función sexual evaluada por el FSFI (p=0,034; r=0,516). Existe una asociación entre la IU y la DSF en mujeres ancianas físicamente activas. Algunas ancianas tuvieron ambas disfunciones del MAP a pesar de ser físicamente activas. Por lo tanto, es necesario reforzar la importancia de las evaluaciones detalladas, la actividad física orientada y la acción multiprofesional en salud.

ABSTRACT Urinary incontinence (UI) and female sexual dysfunction (FSD) are disorders that affect quality of life. With the change in the Brazilian age pyramid, the prevalence of such disorders tends to increase. The aim of the present study was to correlate the relationship between UI, FSD and the functionality of the pelvic floor muscles (PFM) in physically active older women. This is a cross-sectional study, in which 35 older women ≥60 years old participated. Initially, the diagnostic form, the International Consultation on Incontinence Questionnaire - Short form (ICIQ-UI-SF) and Female Sexual Function Index (FSFI) were applied. Then, the PFM were evaluated by vaginal touch, following what is proposed in the perfect scheme. Among older women, 20% had both PFM dysfunctions. In total, 17 (48.6%) had UI and 16 (45.7%) FSD. There was a significant difference in the number of vaginal deliveries (p=0.028) and in the perfect schedule (p=0.033) between older women with and without FSD. There was no statistical difference between the groups with and without UI. The analysis showed an inversely proportional correlation between UI severity and sexual function in incontinent older women. (p=0.008; r=−0.622). Since, for each increase in UI severity by the ICIQ-SF, 0.577 of sexual function assessed by the FSFI is decreased (p=0.034; r=0.516). There is an association between UI and FSD in physically active older women. Some older women have both PFM dysfunctions despite being physically active. Thus, it is important to reinforce the importance of thorough evaluations, oriented physical activity and multiprofessional action in health.

Lipid droplets fuel SARS-CoV-2 replication and production of inflammatory mediators.

Viruses are obligate intracellular parasites that make use of the host metabolic machineries to meet their biosynthetic needs. Thus, identifying the host pathways essential for the virus replication may lead to potential targets for therapeutic intervention. The mechanisms and pathways explored by SARS-CoV-2 to support its replication within host cells are not fully known. Lipid droplets (LD) are organelles with major functions in lipid metabolism, energy homeostasis and intracellular transport, and have multiple roles in infections and inflammation. Here we described that monocytes from COVID-19 patients have an increased LD accumulation compared to SARS-CoV-2 negative donors. In vitro, SARS-CoV-2 infection were seen to modulate pathways of lipid synthesis and uptake as monitored by testing for CD36, SREBP-1, PPARγ, and DGAT-1 expression in monocytes and triggered LD formation in different human cell lines. LDs were found in close apposition with SARS-CoV-2 proteins and double-stranded (ds)-RNA in infected Vero cells. Electron microscopy (EM) analysis of SARS-CoV-2 infected Vero cells show viral particles colocalizing with LDs, suggestive that LDs might serve as an assembly platform. Pharmacological modulation of LD formation by inhibition of DGAT-1 with A922500 significantly inhibited SARS-CoV-2 replication as well as reduced production of mediators pro-inflammatory response. Taken together, we demonstrate the essential role of lipid metabolic reprograming and LD formation in SARS-CoV-2 replication and pathogenesis, opening new opportunities for therapeutic strategies to COVID-19.

Atazanavir, Alone or in Combination with Ritonavir, Inhibits SARS-CoV-2 Replication and Proinflammatory Cytokine Production.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is already responsible for far more deaths than previous pathogenic coronaviruses (CoVs) from 2002 and 2012. The identification of clinically approved drugs to be repurposed to combat 2019 CoV disease (COVID-19) would allow the rapid implementation of potentially life-saving procedures. The major protease (Mpro) of SARS-CoV-2 is considered a promising target, based on previous results from related CoVs with lopinavir (LPV), an HIV protease inhibitor. However, limited evidence exists for other clinically approved antiretroviral protease inhibitors. Extensive use of atazanavir (ATV) as antiretroviral and previous evidence suggesting its bioavailability within the respiratory tract prompted us to study this molecule against SARS-CoV-2. Our results show that ATV docks in the active site of SARS-CoV-2 Mpro with greater strength than LPV, blocking Mpro activity. We confirmed that ATV inhibits SARS-CoV-2 replication, alone or in combination with ritonavir (RTV) in Vero cells and a human pulmonary epithelial cell line. ATV/RTV also impaired virus-induced enhancement of interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α) levels. Together, our data strongly suggest that ATV and ATV/RTV should be considered among the candidate repurposed drugs undergoing clinical trials in the fight against COVID-19.

Neuraminidase from Influenza A and B Viruses is Susceptible to the Compound 4-(4-Phenyl-1H-1,2,3-Triazol-1-yl)-2,2,6,6-Tetramethylpiperidine-1- Oxyl.

BACKGROUND: Since the influenza virus is the main cause of acute seasonal respiratory infections and pandemic outbreaks, antiviral drugs are critical to mitigate infections and impair chain of transmission. Neuraminidase inhibitors (NAIs) are the main class of anti-influenza drugs in clinical use. Nevertheless, resistance to oseltamivir (OST), the most used NAI, has been detected in circulating strains of the influenza virus. Therefore, novel compounds with anti-influenza activity are necessary. OBJECTIVE: To verify whether the NA from influenza A and B virus is susceptible to the compound 4-(4- phenyl-1H-1,2,3-triazol-1-yl)-2,2,6,6-tetramethylpiperidine-1-oxyl (Tritempo). METHODS: Cell-free neuraminidase inhibition assays were performed with Tritempo, using wild-type (WT) and OST-resistant influenza strains. Cell-based assays in MDCKs were performed to confirm Tritempo`s antiviral activity and cytotoxicity. Multiple passages of the influenza virus in increasing concentrations of our compound, followed by the sequencing of NA gene and molecular docking, were used to identify our Tritempo's target. RESULTS AND DISCUSSION: Indeed, Tritempo inhibited the neuraminidase activity of WT and OSTresistant strains of influenza A and B, at the nanomolar range. Tritempo bound to WT and OST-resistant influenza NA isoforms at the sialic acid binding site with low free binding energies. Cell-free assays were confirmed using a prototypic influenza A infection assay in MDCK cells, in which we found an EC50 of 0.38 µM, along with very low cytotoxicity, CC50 > 2,000 µM. When we passaged the influenza A virus in the presence of Tritempo, a mutant virus with the G248P change in the NA was detected. This mutant was resistant to Tritempo but remained sensitive to OST, indicating no cross-resistance between the studied and reference drugs. CONCLUSION: Our results suggest that Tritempo's chemical structure is a promising one for the development of novel antivirals against influenza.

Agathisflavone, a Biflavonoid from Anacardium occidentale L., Inhibits Influenza Virus Neuraminidase.

BACKGROUND: Neuraminidase inhibitors (NAIs) are the only class of antivirals in clinical use against influenza virus approved worldwide. However, approximately 1-3% of circulating strains present resistance mutations to oseltamivir (OST), the most used NAI. Therefore, it is important to catalogue new molecules to inhibit influenza virus, especially OST-resistant strains. Natural products from tropical plants used for human consumption represent a worthy class of substances. Their use could be stimulated in resource-limited setting where the access to expensive antiviral therapies is restricted. METHODS: We evaluated the anti-influenza virus activity of agathisflavone derived from Anacardium occidentale L. RESULTS: The neuraminidase (NA) activity of wild-type and OST-resistant influenza virus was inhibited by agathisflavone, with IC50 values ranging from 20 to 2.0 µM, respectively. Agathisflavone inhibited influenza virus replication with EC50 of 1.3 µM. Sequential passages of the virus in the presence of agathisflavone revealed the emergence of mutation R249S, A250S and R253Q in the NA gene. These changes are outside the OST binding region, meaning that agathisflavone targets this viral enzyme at a region different than conventional NAIs. CONCLUSION: Altogether our data suggest that agathisflavone has a promising chemical structure for the development of anti-influenza drugs.