Antiviral activity of myricetin glycosylated compounds isolated from Marcetia taxifolia against chikungunya virus.

The chikungunya virus (CHIKV) has produced epidemic outbreaks of significant public health impact. The clinical symptoms of this disease are fever, polyarthralgia, and skin rash, generally self-limiting, although patients may develop a chronic disabling condition or suffer lethal complications. Unfortunately, there is no specific treatment or vaccine available. Thus, the search for effective therapies to control CHIKV infection is an urgent need. This study evaluated the antiviral activity of flavonoids isolated from Marcetia taxifolia by in vitro and in silico analysis. Cytotoxicity of compounds was determined by MTT assay and viral load was assessed in cell substrates supernatants by plaque-forming and RT-qPCR assays. Selected molecules were analyzed by molecular docking assays. Myricetin 3-rhamnoside (MR) and myricetin 3-(6-rhamnosylgalactoside) (MRG) were tested for antiviral assays and analyzed by the TCID50 method and RT-qPCR. MR exhibited dose-dependent antiviral activity, reducing viral titer at concentrations of 150-18.8 µg/mL by at least 1-log. Similarly, MRG showed a significant decrease in viral titer at concentrations of 37.5, 9.4, and 2.3 µg/mL. RT-qPCR analysis also displayed a substantial reduction of CHIKV RNA for both flavonoids. Furthermore, molecular docking of the selected flavonoids proposed the nsP3 macrodomain as a possible target of action. Our study reveals that MR and MRG could be considered promising anti-CHIKV therapeutic agents. Molecular modeling studies showed MR and MRG ligands with a high affinity for the N-terminal region of the nsP3 macrodomain, postulating them as a potential target of action for the CHIKV control.

Immunohistochemical subtype and its relationship with 5-year overall survival in breast cancer patients.

Background: Breast cancer (BC) is the malignant tumour that has been most frequently diagnosed, being the second most common cancer worldwide and the most frequent in women. Objective: To analyse the probability of 5-year overall survival according to age, stage of disease, immunohistochemical subtype, histological grade and histological type in patients with BC. Methodology: Operational research that used a cohort design of patients diagnosed with BC at the SOLCA Núcleo de Loja-Ecuador Hospital from 2009 to 2015 and with follow-up until December 2019. Survival was estimated with the actuarial method and Kaplan-Meier method, and, for multivariate analysis, the proportional hazards model or Cox regression was used to estimate the adjusted Hazard Ratios (HRs). Results: Two hundred and sixty-eight patients were studied. Mean overall survival was 4.35 years (95% confidence interval (95% CI): 40.20-4.51) and 66% survived to 5 years. The main predictors of survival were advanced stage of disease (III-IV) (HR = 7.03; 95% CI: 3.81-12.9); patients human epidermal growth factor receptor 2-neu (HER2-neu) overexpressed (HR = 2.26; 95% CI = 1.31-4.75) and triple negative (HR = 2.57; 95% CI = 1.39-4.75). The other variables were not significant. Conclusions: The results show a higher mortality associated with higher clinical stage, more aggressive histological grades and immunohistochemical subtype HER2-neu overexpressed and triple negative tumours.

Guía clínica multidisciplinaria para el uso de analgesia y sedación; prevención de delirium y abstinencia / Multidisciplinary clinical guide for analgesia and sedation; prevention of delirium and withdrawal

El ingreso a la UCIP somete al niño a una serie de estímulos y procedimientos estresantes y dolorosos. El uso de analgesia y sedación es un componente central en los cuidados del niño críticamente enfermo para asegurar su confort y seguridad. Es importante durante la prescripción de analgosedación la monitorización continua de sus efectos. Este proceso debe ser vigilado por todos los integrantes del equipo de salud y su familia. Esta guía se basa en los principios propuestos por la iniciativa "Libérame de la UCI", que consisten en un paquete (bundle) de medidas (ABCDEF) para optimizar la recuperación de los pacientes y sus desenlaces. Contiene tablas con la farmacología de los analgésicos y sedantes, con estrategias generales para el manejo de anal-gosedación y prevención del síndrome de abstinencia y delirium. Guía realizada por el equipo multidisciplinario para lograr una adherencia más completa.

ABSTRACT The admission of a child to the PICU subdue stressful and pain stimuli. The use of analgesia and sedation is the main component for the care of the critically ill children to guarantee comfort and security. During the prescription of analgesia and sedation, monitoring of their effects is important. This process must be supervised by the health team and supported by the family. This guide is based on the initiative "PICU liberation" where a bundle of measures (ABCDEF) is used to optimize the recovery and outcome. This guide contains tables of the pharmacology of analgesics and sedatives, some strategies on the use of analgosedation, prevention of withdrawal and delirium. A guide of the multidisciplinary team to achieve a complete adherence.

Purification of Trypanosoma cruzi metacyclic trypomastigotes by ion exchange chromatography in sepharose-DEAE, a novel methodology for host-pathogen interaction studies.

Metacyclic trypomastigotes are essential for the understanding of the biology of Trypanosoma cruzi, the agent of Chagas disease. However, obtaining these biological stages in axenic medium is difficult. Techniques based on charge and density of the parasite during different stages have been implemented, without showing a high efficiency in the purification of metacyclic trypomastigotes. So far, there is no protocol implemented where sepharose-DEAE is used as a resin. Therefore, herein we tested its ability to purify metacyclic trypomastigotes in Liver Infusion Triptose (LIT) medium cultures. A simple, easy-to-execute and effective protocol based on ion exchange chromatography on Sepharose-DEAE resin for the purification of T. cruzi trypomastigotes is described. T. cruzi strains from the Discrete Typing Units (DTUs) I and II were used. The strains were harvested in LIT medium at a concentration of 1×107epimastigotes/mL. We calculated the time of trypomastigotes increment (TTI). Based on the data obtained, Ion exchange chromatography was performed with DEAE-sepharose resin. To verify the purity and viability of the trypomastigotes, a culture was carried out in LIT medium with subsequent verification with giemsa staining. To evaluate if the technique affected the infectivity of trypomastigotes, in vitro assays were performed in Vero cells and in vivo in ICR-CD1 mice. The technique allowed the purification of metacyclic trypomastigotes of other stages of T. cruzi in a percentage of 100%, a greater recovery was observed in cultures of 12days. There were differences regarding the recovery of metacyclic trypomastigotes for both DTUs, being DTU TcI the one that recovered a greater amount of these forms. The technique did not affect parasite infectivity in vitro or/and in vivo.

Correction: Phytometabolite Dehydroleucodine Induces Cell Cycle Arrest, Apoptosis, and DNA Damage in Human Astrocytoma Cells through p73/p53 Regulation.

[This corrects the article DOI: 10.1371/journal.pone.0136527.].

Phytometabolite Dehydroleucodine Induces Cell Cycle Arrest, Apoptosis, and DNA Damage in Human Astrocytoma Cells through p73/p53 Regulation.

Accumulating evidence supports the idea that secondary metabolites obtained from medicinal plants (phytometabolites) may be important contributors in the development of new chemotherapeutic agents to reduce the occurrence or recurrence of cancer. Our study focused on Dehydroleucodine (DhL), a sesquiterpene found in the provinces of Loja and Zamora-Chinchipe. In this study, we showed that DhL displayed cytostatic and cytotoxic activities on the human cerebral astrocytoma D384 cell line. With lactone isolated from Gynoxys verrucosa Wedd, a medicinal plant from Ecuador, we found that DhL induced cell death in D384 cells by triggering cell cycle arrest and inducing apoptosis and DNA damage. We further found that the cell death resulted in the increased expression of CDKN1A and BAX proteins. A marked induction of the levels of total TP73 and phosphorylated TP53, TP73, and γ-H2AX proteins was observed in D384 cells exposed to DhL, but no increase in total TP53 levels was detected. Overall these studies demonstrated the marked effect of DhL on the diminished survival of human astrocytoma cells through the induced expression of TP73 and phosphorylation of TP73 and TP53, suggesting their key roles in the tumor cell response to DhL treatment.

Characterization of Plasmodium falciparum integral membrane protein Pf25-IMP and identification of its red blood cell binding sequences inhibiting merozoite invasion in vitro.

The identification of proteins present on the surface of Plasmodium falciparum-infected red blood cells as well as of free merozoites has been widely considered as one of the main areas of research in the development of an antimalarial vaccine due to their involvement in the parasite's pathogenesis and invasion mechanisms. Major advances had been accomplished in this area thanks to the analysis of the reported genomic sequence of P. falciparum, allowing for the identification of genes encoding for putative integral membrane proteins. This study reports for the first time the transcription of the MAL8P1.3 gene, which codifies for a 25-kDa integral membrane protein of P. falciparum (FCB-2 strain), namely, Pf25-IMP. Western blot and immunofluorescence assays using goat polyclonal sera indicate that this protein is expressed in erythrocytic asexual blood stages. A highly robust, sensible, and specific receptor-ligand interaction assay allowed identification of two high activity binding peptides (HABPs) derived from Pf25-IMP: 30577 ((41)YKTANENVKLASSLSDRLSR(60)) and 30583 ((161)LNKKTVVRKIAEGLGYTIVF(180)). Both HABPs bound with high affinity to human red blood cells (RBCs), and such binding was susceptible to enzyme treatment with trypsin. A common RBC surface receptor of apparently 48 kDa was found for both HABPs, plus an additional 31-kDa receptor for HABP 30577. HABP 30577 inhibited merozoite invasion in vitro by 73%, while HABP 30583 showed a 59% inhibition at 200 microM concentration. The data suggest a possible role of Pf25-IMP in merozoite invasion to RBCs and support its inclusion in further immunological studies for evaluating its potential as vaccine candidates.