Effect of phenylmethylsulfonyl fluoride, a protease inhibitor, on enamel surface remineralization

Phenylmethylsulfonyl fluoride (PMSF) is a protease inhibitor widely used in research, but fluoride is released during its action and this knowledge has been neglected in dental research. Aim: to evaluate if fluoride released by salivary protease action on PMSF affects enamel remineralization and fluoride uptake. Methods: Groups of 10 enamel slabs, with caries-like lesions and known surface hardness (SH), were subjected to one of the following treatment groups: Stimulated human saliva (SHS), negative control; SHS containing 1.0 µg F/mL (NaF), positive control; and SHS containing 10, 50 or 100 µM PMSF. The slabs were subjected to a pH-cycling regimen consisting of 22 h/day in each treatment solution and 2 h/day in a demineralizing solution. After 12 days, SH was again measured to calculate the percentage of surface hardness recovery (%SHR), followed by enamel fluoride uptake determination. The time-related fluoride release from 100.0 µM PMSF by SHS action was also determined. Data were analyzed by ANOVA followed by Newman-Keuls test. Results: The release of fluoride from PMSF by SHS was rapid, reaching a maximum value after 10 min. Fluoride released from PMSF was more effective in enhancing %SHR and increasing fluoride uptake in enamel compared with SHS alone (p < 0.05); furthermore, it was equivalent to the positive control (p > 0.05). Conclusion: In conclusion, fluoride released by saliva from PMSF is available to react with enamel and needs to be taken into account in research using this protease inhibitor

A threefold approach including quantum chemical, molecular docking and molecular dynamic studies to explore the natural compounds from Centaurea jacea as the potential inhibitors for COVID-19 / Uma abordagem tríplice incluindo química quântica, docking molecular e estudos dinâmicos moleculares para explorar os compostos naturais de Centaurea jacea como inibidores potenciais para Covid-19

Abstract In the current report, we studied the possible inhibitors of COVID-19 from bioactive constituents of Centaurea jacea using a threefold approach consisting of quantum chemical, molecular docking and molecular dynamic techniques. Centaurea jacea is a perennial herb often used in folk medicines of dermatological complaints and fever. Moreover, anticancer, antioxidant, antibacterial and antiviral properties of its bioactive compounds are also reported. The Mpro (Main proteases) was docked with different compounds of Centaurea jacea through molecular docking. All the studied compounds including apigenin, axillarin, Centaureidin, Cirsiliol, Eupatorin and Isokaempferide, show suitable binding affinities to the binding site of SARS-CoV-2 main protease with their binding energies -6.7 kcal/mol, -7.4 kcal/mol, -7.0 kcal/mol, -5.8 kcal/mol, -6.2 kcal/mol and -6.8 kcal/mol, respectively. Among all studied compounds, axillarin was found to have maximum inhibitor efficiency followed by Centaureidin, Isokaempferide, Apigenin, Eupatorin and Cirsiliol. Our results suggested that axillarin binds with the most crucial catalytic residues CYS145 and HIS41 of the Mpro, moreover axillarin shows 5 hydrogen bond interactions and 5 hydrophobic interactions with various residues of Mpro. Furthermore, the molecular dynamic calculations over 60 ns (6×106 femtosecond) time scale also shown significant insights into the binding effects of axillarin with Mpro of SARS-CoV-2 by imitating protein like aqueous environment. From molecular dynamic calculations, the RMSD and RMSF computations indicate the stability and dynamics of the best docked complex in aqueous environment. The ADME properties and toxicity prediction analysis of axillarin also recommended it as safe drug candidate. Further, in vivo and in vitro investigations are essential to ensure the anti SARS-CoV-2 activity of all bioactive compounds particularly axillarin to encourage preventive use of Centaurea jacea against COVID-19 infections.

Resumo No presente relatório, estudamos os possíveis inibidores de Covid-19 de constituintes bioativos de Centaurea jacea usando uma abordagem tripla que consiste em técnicas de química quântica, docking molecular e dinâmica molecular. Centaurea jacea é uma erva perene frequentemente usada em remédios populares de doenças dermatológicas e febre. Além disso, as propriedades anticâncer, antioxidante, antibacteriana e antiviral de seus compostos bioativos também são relatadas. A Mpro (proteases principais) foi acoplada a diferentes compostos de Centaurea jacea por meio de docking molecular. Todos os compostos estudados, incluindo apigenina, axilarina, Centaureidina, Cirsiliol, Eupatorina e Isokaempferide, mostram afinidades de ligação adequadas ao sítio de ligação da protease principal SARS-CoV-2 com suas energias de ligação -6,7 kcal / mol, -7,4 kcal / mol, - 7,0 kcal / mol, -5,8 kcal / mol, -6,2 kcal / mol e -6,8 kcal / mol, respectivamente. Dentre todos os compostos estudados, a axilarina apresentou eficiência máxima de inibidor, seguida pela Centaureidina, Isokaempferida, Apigenina, Eupatorina e Cirsiliol. Nossos resultados sugeriram que a axilarina se liga aos resíduos catalíticos mais cruciais CYS145 e HIS41 do Mpro, além disso a axilarina mostra 5 interações de ligações de hidrogênio e 5 interações hidrofóbicas com vários resíduos de Mpro. Além disso, os cálculos de dinâmica molecular em uma escala de tempo de 60 ns (6 × 106 femtossegundos) também mostraram percepções significativas sobre os efeitos de ligação da axilarina com Mpro de SARS-CoV-2 por imitação de proteínas como o ambiente aquoso. A partir de cálculos de dinâmica molecular, os cálculos RMSD e RMSF indicam a estabilidade e dinâmica do melhor complexo ancorado em ambiente aquoso. As propriedades ADME e a análise de previsão de toxicidade da axilarina também a recomendaram como um candidato a medicamento seguro. Além disso, as investigações in vivo e in vitro são essenciais para garantir a atividade anti-SARS-CoV-2 de todos os compostos bioativos, particularmente a axilarina, para encorajar o uso preventivo de Centaurea jacea contra infecções por Covid-19.

High-throughput screening of SARS-CoV-2 main and papain-like protease inhibitors

The global COVID-19 coronavirus pandemic has infected over 109 million people, leading to over 2 million deaths up to date and still lacking of effective drugs for patient treatment. Here, we screened about 1.8 million small molecules against the main protease (Mpro) and papain like protease (PLpro), two major proteases in severe acute respiratory syndrome-coronavirus 2 genome, and identified 1851Mpro inhibitors and 205 PLpro inhibitors with low nmol/l activity of the best hits. Among these inhibitors, eight small molecules showed dual inhibition effects on both Mpro and PLpro, exhibiting potential as better candidates for COVID-19 treatment. The best inhibitors of each protease were tested in antiviral assay, with over 40% of Mpro inhibitors and over 20% of PLpro inhibitors showing high potency in viral inhibition with low cytotoxicity. The X-ray crystal structure of SARS-CoV-2 Mpro in complex with its potent inhibitor 4a was determined at 1.8 Å resolution. Together with docking assays, our results provide a comprehensive resource for future research on anti-SARS-CoV-2 drug development.

Genetic Subtypes and Pretreatment Drug Resistance in the Newly Reported Human Immunodeficiency Virus-Infected Men Aged≥50 Years Old in Guangxi / 中国医学科学院学报

Objective To analyze the genetic subtypes of human immunodeficiency virus (HIV) and the prevalence of pretreatment drug resistance in the newly reported HIV-infected men in Guangxi. Methods The stratified random sampling method was employed to select the newly reported HIV-infected men aged≥50 years old in 14 cities of Guangxi from January to June in 2020.The pol gene of HIV-1 was amplified by nested reverse transcription polymerase chain reaction and then sequenced.The mutation sites associated with drug resistance and the degree of drug resistance were then analyzed. Results A total of 615 HIV-infected men were included in the study.The genetic subtypes of CRF01_AE,CRF07_BC,and CRF08_BC accounted for 57.4% (353/615),17.1% (105/615),and 22.4% (138/615),respectively.The mutations associated with the resistance to nucleoside reverse transcriptase inhibitors (NRTI),non-nucleoside reverse transcriptase inhibitors (NNRTI),and protease inhibitors occurred in 8 (1.3%),18 (2.9%),and 0 patients,respectively.M184V (0.7%) and K103N (1.8%) were the mutations with the highest occurrence rates for the resistance to NRTIs and NNRTIs,respectively.Twenty-two (3.6%) patients were resistant to at least one type of inhibitors.Specifically,4 (0.7%),14 (2.3%),4 (0.7%),and 0 patients were resistant to NRTIs,NNRTIs,both NRTIs and NNRTIs,and protease inhibitors,respectively.The pretreatment resistance to NNRTIs had much higher frequency than that to NRTIs (2.9% vs.1.3%;χ2=3.929,P=0.047).The prevalence of pretreatment resistance to lamivudine,zidovudine,tenofovir,abacavir,rilpivirine,efavirenz,nevirapine,and lopinavir/ritonavir was 0.8%, 0.3%, 0.7%, 1.0%, 1.3%, 2.8%, 2.9%, and 0, respectively. Conclusions CRF01_AE,CRF07_BC,and CRF08_BC are the three major strains of HIV-infected men≥50 years old newly reported in Guangxi,2020,and the pretreatment drug resistance demonstrates low prevalence.

Nirmatrelvir-ritonavir (Paxlovid) para el tratamiento de pacientes con COVID-19: [2 de julio de 2022] / Nirmatrelvir-ritonavir (Paxlovid) for the treatment of patients with COVID-19: [July 2, 2022]

INTRODUCCIÓN: El nirmatrelvir es un inhibidor de la proteasa biodisponible por vía oral que es activo frente a MPRO, una proteasa viral que desempeña un papel esencial en la replicación viral al escindir las 2 poliproteínas virales.15 Ha demostrado actividad antiviral frente a todos los coronavirus que se sabe que infectan a los humanos.16 El nirmatrelvir es empaquetado con ritonavir (como Paxlovid), un inhibidor del citocromo P450 (CYP) 3A4 y agente potenciador farmacocinético que se ha utilizado para potenciar los inhibidores de la proteasa del VIH. Se requiere la coadministración de ritonavir para aumentar las concentraciones de nirmatrelvir al rango terapéutico objetivo. OBJETIVO: El objetivo del presente informe es evaluar parámetros de eficacia, seguridad, conveniencia y recomendaciones disponibles acerca del empleo de nirmatrelvir-ritonavir para el tratamiento de pacientes con COVID-19 en Argentina. MÉTODOS: Efectos en la Salud: Se desarrolló un protocolo sustentado en proyectos que resume activamente la evidencia científica a medida que la misma se hace disponible. Con este fin se utilizó la plataforma Love de Epistemonikos para identificar revisiones sistemáticas "vivas". Implementación: Este dominio contempla dos subdominios: la existencia de barreras y facilitadores en nuestro contexto para la implementación de la tecnología evaluada no consideradas en los otros dominios analizados, y los costos comparativos en relación con otras intervenciones similares. Con el objetivo de emitir un juicio de valor sobre la magnitud de dichos costos, en pacientes hospitalizados se utilizó como comparador al tratamiento con dexametasona, que ha demostrado ser una intervención accesible y de beneficios importantes en el contexto analizado. Recomendaciones: Para la identificación de recomendaciones sustentadas en evidencia y actualizadas, se utilizó la plataforma COVID recmap. RESULTADOS: Se identificaron dos sistemáticas que cumplieron con los criterios de inclusión del presente informe. Se identificó un ECA que incluyó 2085 pacientes en los que nirmatrelvir-ritonavir se comparó con el tratamiento estándar en pacientes con COVID-19. CONCLUSIONES: El cuerpo de la evidencia muestra que nirmatrelvir-ritonavir tiene un efecto incierto sobre la mortalidad en pacientes con COVID-19 leve o moderada de reciente comienzo, con factores de riesgo para progresar a enfermedad grave que no han sido vacunados. En esta población, probablemente disminuya la necesidad de hospitalización sin aumentar los eventos adversos severos. Esta tecnología no ha sido probada en personas vacunadas ni en otros escenarios. La tecnología no está autorizada para su comercialización por la Administración Nacional de Medicamentos, Alimentos y Tecnología Médica (ANMAT) de nuestro país. Aunque la forma de administración oral es simple, la experiencia en otros países muestra que el costo estimado es elevado siendo la población objetivo muy grande. Hay incertidumbre sobre si podrían existir además problemas de suministro que afecten la disponibilidad y la equidad en la distribución. Las guías de práctica clínica de alta calidad metodológica actualizadas incluidas tenderían a recomendar el tratamiento. Aquellas que recomiendan en forma condicional a favor se basan en el beneficio observado en personas de muy alto riesgo no vacunadas con enfermedad de reciente comienzo, y su facilidad de administración. Las que entregan recomendaciones en contra se basan en el costo comparativo muy elevado, para una población objetivo muy amplia y la baja certeza en su efecto sobre la mortalidad.

Nirmatrelvir-ritonavir (Paxlovid) para el tratamiento de pacientes con COVID-19: [17 de marzo de 2022] / Nirmatrelvir-ritonavir (Paxlovid) for the treatment of patients with COVID-19: [March 17, 2022]

INTRODUCCIÓN: El nirmatrelvir es un inhibidor de la proteasa que se administra por vía oral, que demostró ser activo frente a MPRO, una proteasa viral que desempeña un papel esencial en la replicación viral al escindir las 2 poliproteínas virales. 15 Además, ha demostrado actividad antiviral frente a todos los coronavirus que se sabe que infectan a los humanos. 16 El nirmatrelvir es empaquetado con ritonavir (como Paxlovid), un inhibidor del citocromo P450 (CYP) 3A4 y agente potenciador farmacocinético, que se ha utilizado para potenciar los inhibidores de la proteasa del virus de la inmunodeficiencia humana (VIH). La coadministración tiene como objetivo aumentar las concentraciones de nirmatrelvir al rango terapéutico objetivo. El 22 de diciembre de 2021, la Administración de Alimentos y Medicamentos (FDA, su siglas en inglés Food and Drug Administration) emitió una autorización de uso de emergencia para nirmatrelvir potenciado con ritonavir para el tratamiento de pacientes con COVID-19 leve a moderado de ≥12 años y un peso de ≥ 40 kg que están dentro de los 5 días de inicio de los síntomas y con alto riesgo de progresar a enfermedad grave. 17,18 Administración Nacional de Medicamentos, Alimentos y Tecnología Médica (ANMAT) aún no ha autorizado la comercialización del medicamento para este fin. OBJETIVO: El objetivo del presente informe es evaluar parámetros de eficacia, seguridad, conveniencia y recomendaciones disponibles acerca del empleo de nirmatrelvir-ritonavir para el tratamiento de pacientes con COVID-19 en Argentina. MÉTODOS: Teniendo en cuenta la velocidad con la que la información relacionada con la pandemia aparece y se modifica (link), se desarrolló un protocolo sustentado en proyectos que resume activamente la evidencia científica a medida que la misma se hace disponible. Con este fin se utilizó la plataforma L- ove de Epistemonikos para identificar revisiones sistemáticas "vivas". Se seleccionaron aquellas con una calidad metodológica apropiada evaluada a través de la herramienta AMSTAR-2, y que a su vez llevaran un proceso de actualización frecuente.19 De cada una de las revisiones sistemáticas identificadas se extractaron los efectos de la intervención sobre los desenlaces priorizados como importantes o críticos y la certeza en dichos efectos. Para la priorización de los desenlaces se adoptó una perspectiva desde el paciente considerando sus potenciales preferencias. La selección se realizó por consenso entre los autores y supervisores del informe considerando los resultados de múltiples ejercicios de priorización publicados, realizados en el marco del desarrollo de distintas guías de práctica clínica.20­23 Se seleccionaron "mortalidad", "ingreso en asistencia ventilatoria mecánica", "tiempo hasta resolución de síntomas", "hospitalización", "eventos adversos graves" como desenlaces críticos. Adicionalmente, se extractaron datos relacionados con efectos de subgrupo potencialmente relevantes para la toma de decisión, con especial énfasis en el tiempo de evolución, la severidad de la enfermedad y el estado de vacunación. En los casos en que no fue reportado por las revisiones sistemáticas incluidas, se calculó el efecto absoluto de las intervenciones en pacientes vacunados, tomando el riesgo basal reportado para pacientes no vacunados multiplicado por un riesgo relativo de 0,1 según el efecto de la vacunación observado en distintos estudios y sistemas de vigilancia.24­26 Para confeccionar las conclusiones en el efecto de las intervenciones evaluadas sobre los desenlaces priorizados, utilizamos lineamientos publicados, específicamente desarrollados a tal fin. RESULTADOS: Se identificaron dos sistemáticas que cumplieron con los criterios de inclusión del presente informe y reportaron resultados: Se identificó un ECA que incluyó 2.085 pacientes en los que nirmatrelvir-ritonavir se comparó con el tratamiento estándar en pacientes con COVID-19. Se describen los efectos absolutos y la certeza en dichos efectos de nirmatrelvir-ritonavir para pacientes con COVID-19. Los pacientes incluidos en el estudio no habían recibido un esquema completo de vacunación, por lo que no se realizó un análisis de subgrupos según el estado de vacunación. Sin embargo, un análisis de subgrupo según la presencia o no de anticuerpos anti SARS-CoV-2 en suero al momento de ingresar al estudio sugiere que los beneficios observados podrían ser mayores, o incluso limitarse, para aquellos pacientes que no posean inmunidad previa. Luego de la aprobación de emergencia por la FDA de nirmatrelvir-ritonavir (Paxlovir), el Departamento de Salud de los Estados Unidos estimó que tendrá distribuidos para el mes de marzo de 2022, a través del Organismo de Emergencia Pública en Salud, un total de 513.830 dosis a un costo unitario de 520 USD por 5 días de tratamiento; llevando a un gasto total de más de 266 millones de dólares (266.760 000 USD).32,33 La tecnología no está autorizada aún para su comercialización en Argentina para su uso en personas con COVID-19. La vía de administración oral podría asociarse con mayor facilidad de uso; sin embargo, no se conoce la disponibilidad del laboratorio para responder a la demanda sin afectar la equidad en la distribución en nuestro país. CONCLUSIONES: El cuerpo de la evidencia muestra que nirmatrelvir-ritonavir tiene un efecto incierto sobre la mortalidad en pacientes con COVID-19 leve o moderada de reciente comienzo, con factores de riesgo para progresar a enfermedad grave que no han sido vacunados. En esta población, probablemente disminuya la necesidad de hospitalización sin aumentar los eventos adversos severos. Esta tecnología no ha sido probada en personas vacunadas ni en otros escenarios. La tecnología no está autorizada para su comercialización por la Administración Nacional de Medicamentos, Alimentos y Tecnología Médica (ANMAT) de nuestro país. Aunque la forma de administración oral es simple, la experiencia en otros países muestra que el costo comparativo estimado es elevado, la población objetivo es muy alta y podrían existir además problemas de suministro y distribución que afecten la disponibilidad y la equidad en la distribución. Las guías de práctica clínica de alta calidad metodológica actualizadas entregan recomendaciones discordantes en cuanto a su uso. Aquellas que recomiendan en forma condicional a favor se basan en el beneficio observado en personas de muy alto riesgo, no vacunadas, con enfermedad de reciente comienzo, y su facilidad de administración. Las que entregan recomendaciones en contra se basan en el costo comparativo muy elevado, para una población objetivo muy amplia y la baja certeza en su efecto sobre la mortalidad.

Disfunción eréctil en personas con VIH ­ Revisión de alcance / Erectile Dysfunction in People with HIV ­ A Scoping Review

Objetivo Exponer los factores que han sido asociados al desarrollo de disfunción eréctil (DE) en las personas con virus de la inmunodeficiencia humana (VIH) y cuál es la prevalencia de la disfunción eréctil en esta población. Métodos Se realizó una revisión de alcance utilizando las siguientes bases de datos: MEDLINE, CENTRAL, Embase, Scopus, Lilacs, y Psycinfo. Los artículos incluidos debían tener cualquier información relacionada con la DE en personas con VIH, su prevalencia, y posibles factores asociados. Se filtraron un total de 2.726 artículos por título y resumen, y, de estos, se seleccionaron 22 referencias para revisión de texto completo y análisis. Resultados Se encontró que las personas con VIH tienen una mayor probabilidad de presentar DE en comparación con las personas sin VIH. Algunos estudios han establecido una asociación entre la DE y trastornos psicológicos o factores orgánicos, como la lipodistrofia, la hipertensión arterial, la hipercolesterolemia, la diabetes mellitus, la depresión, la ansiedad, y el hipogonadismo. La ingesta de terapia antirretroviral (TARV), más específicamente los inhibidores de la proteasa, sigue siendo cuestionada como causante de DE en pacientes con VIH. Conclusiones Ante un paciente con diagnóstico de VIH, se debe ahondar sobre DE con el fin de recomendar e iniciar conductas terapéuticas que aseguren un mejoramiento en su calidad de vida.

Aim To present factors that have been associated with the development of erectile dysfunction (ED) in people with human immunodeficiency virus (HIV) and the frequency of erectile dysfunction among this population. Methods We performed a scoping review on the following databases: MEDLINE, CENTRAL, Embase, Scopus, Lilacs, and Psycinfo. The articles included information related to ED in people with HIV, its frequency, and possible associated factors. A total of 2,726 articles were filtered by title and abstract, and, from these, 22 references were selected for full-text review and analysis. Results People with HIV were found to be more likely to have ED compared to people without HIV. Some studies have established an association between ED and psychological disorders or organic factors, such as lipodystrophy, high blood pressure, hypercholesterolemia, diabetes mellitus, depression, anxiety, and hypogonadism. The intake of antiretroviral therapy (ART), more specifically protease inhibitors, continues to be questioned as a cause of ED in patients with HIV. Conclusions In HIV-positive men, we recommend to actively seek ED in order to initiate therapeutic conducts that can ensure an improvement in their quality of life.

Sodium trimetaphosphate combined to calcium as a strategy to improve dentin-bonding interface / Trimetafosfato de sódio associado ao cálcio como uma estratégia para melhorar a interface adesiva da dentina

Objective: The aim of this study was to evaluate the effect of STMP as biomimetic analog of dentin matrix on the dentin bond strength submitted to artificial cariogenic challenge over time. Material and Methods: The total number of teeth used in the experiment was 60 teeth, which were divided into 6 groups (n = 10). Of these total amount, 10 teeth were not submitted to the artificial cariogenic challenge (ACC), serving as control group (Sound Dentin - SD) while the other 50 were submitted to an ACC (7d/37ºC), being treated with treatment solutions according to each group: SD- deionized water/sound dentin, CD- deionized water/ artificial caries dentin, GIII- STMP, GIV- STMP + Ca(OH)2, GV- STMP + NaF, and GVI- NaF. After treatments (24h), the specimens were restored (Adper Single Bond Universal + Filtek Z250), to obtain resin­dentin sticks with a cross sectional area of 0.8mm2, approximately. Two-third of these sticks were stored in artificial saliva (37°C) for analyzes after 6 and 12 months. The 1/3 remains were subjected to µTBS test (baseline). Data were analyzed by two-way ANOVA and Tukey tests (p<0.05). Results: In general, the highest µTBS values were obtained in sound condition (SD), while the artificial caries condition (CD) determined minimum values. Groups treated with NaF (with or without STMP- GV and GVI) were not able to improve adhesion over time. Only the use of STMP + Ca(OH)2(GIV) improved the µTBS compared to the others caries-challenged dentin after 1 year. The adhesive failure pattern was predominant in all time. Conclusion: The use of the STMP associated with Ca(OH)2 seems to be a viable therapeutic strategy conciliating the biomimetizing capacity to the adhesive process satisfactorily even its performance is not superior to initial condition (AU)

Objetivo: O objetivo deste estudo foi avaliar o efeito do STMP como análogo biomimético da matriz dentinária na resistência de união à dentina submetida a desafio cariogênico artificial ao longo do tempo. Material e Métodos:foram utilizados um total de 60 dentes neste experimento, os quais foram divididos em 6 grupos (n = 10). Desse total, 10 dentes não foram submetidos ao desafio cariogênico artificial (DCA), servindo como grupo controle (Dentina Hígida - DH) enquanto os outros 50 foram submetidos ao DCA (7d / 37ºC), sendo tratados com soluções de tratamento específicas para cada grupo: DH- água deionizada / dentina hígida, DC- água deionizada / dentina submetida ao DCA, GIII- STMP, GIV- STMP + Ca(OH)2, GV- STMP + NaF e GVI- NaF. Após os tratamentos (24h), os corpos-de-prova foram restaurados (Adper Single Bond Universal + Filtek Z250), para obtenção de palitos de resina-dentina com área transversal de aproximadamente 0,8mm2. Dois terços desses palitos foram armazenados em saliva artificial (37°C) para análises após 6 e 12 meses. Os outros 1/3 foram submetidos ao teste µTBS (baseline). Os dados foram analisados por ANOVA a dois fatores e testes de Tukey (p <0,05). Resultados:Em geral, os maiores valores de µTBS foram obtidos em condição hígidas (DH), enquanto a condição subtmetidas ao DCA determinou os menores valores. Os grupos tratados com NaF (com ou sem STMP associado -GV e GVI) não foram capazes de melhorar a resistência de união, ao longo do tempo. Somente o uso de STMP + Ca (OH)2(GIV) melhorou o µTBS em comparação com as outras condições desafiadas por cárie após 1 ano. O padrão de falha adesiva foi predominante em todos os tempos. Conclusão: O uso do STMP associado ao Ca (OH)2 parece ser uma estratégia terapêutica viável conciliando a capacidade biomimetizante ao processo adesivo de forma satisfatória mesmo que seu desempenho não seja superior à condição inicial.(AU)

Ulinastatin alleviates early brain injury after intracerebral hemorrhage by inhibiting oxidative stress and neuroinflammation via ROS/MAPK/Nrf2 signaling pathway

Purpose: Spontaneous intracerebral hemorrhage (ICH) is still a major public health problem, with high mortality and disability. Ulinastatin (UTI) was purified from human urine and has been reported to be anti-inflammatory, organ protective, and antioxidative stress. However, the neuroprotection of UTI in ICH has not been confirmed, and the potential mechanism is unclear. In the present study, we aimed to investigate the neuroprotection and potential molecular mechanisms of UTI in ICH-induced early brain injury in a C57BL/6 mouse model. Methods: The neurological score, brain water content, neuroinflammatory cytokine levels, oxidative stress levels, and neuronal damage were evaluated. Results: UTI treatment markedly increased the neurological score, alleviated brain edema, decreased the levels of the inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), IL-6, and NF-κB, decreased the levels of reactive oxygen species (ROS) and malondialdehyde (MDA), and upregulated the levels of glutathione (GSH), superoxide dismutase (SOD), and Nrf2. This finding indicated that UTI-mediated inhibition of neuroinflammation and oxidative stress alleviated neuronal damage after ICH. The neuroprotective capacity of UTI is partly dependent on the ROS/MAPK/Nrf2 signaling pathway. Conclusions: UTI improves neurological outcomes in mice and reduces neuronal death by protecting against neural neuroinflammation and oxidative stress.

Tratamiento Covid-19 / Covid-19 treatment

The main treatment for patients with severe Covid-19 is to maintain adequate ventilatory support and monitor the possible progression of the disease. Therapeutic strategies such as High Flow Nasal Cannula, awake prone position, antithrombotic prophylaxis and the use of dexamethasone. There have improved the probability of not presenting complications and not requiring invasive mechanical ventilation. The use of convalescent plasma is still under investigation and is currently only recommended in clinical trials. Finding antivirals that allow treating the infection and clinically improving patients has led to mounting studies with different methodologies, and currently there are specific and limited indications for their routine use, as in the case of Remdesivir, which has been approved by the FDA as emergency treatment in severe cases. Immunomodulatory treatments are still under study. An example of this is Tocilizumab and Anakinra, which have shown promising results for the management of seriously ill patients. It should be noted that there are many therapies that are being tested and that every day the information about the results obtained is changing. (AU)

Identification and characterization of a novel elastase inhibitor from Hirudinaria manillensis / 中国天然药物

A large number of protease inhibitors have been found from leeches, which are essential in various physiological and biological processes. In the curret study, a novel elastase inhibitor was purified and characterized from the leech of Hirudinaria manillensis, which was named HMEI-A. Primary structure analysis showed that HMEI-A belonged to a new family of proteins. HMEI-A exerted inhibitory effects on elastase and showed potent abilities to inhibit elastase with an inhibition constant (K

Key active sites of proteases and protease inhibitors: a review / 生物工程学报

Proteases are widely found in organisms participating in the decomposition of proteins to maintain the organisms' normal life activities. Protease inhibitors regulate the activities of target proteases by binding to their active sites, thereby affecting protein metabolism. The key amino acid mutations in proteases and protease inhibitors can affect their physiological functions, stability, catalytic activity, and inhibition specificity. More active, stable, specific, environmentally friendly and cheap proteases and protease inhibitors might be obtained by excavating various natural mutants of proteases and protease inhibitors, analyzing their key active sites by using protein engineering methods. Here, we review the studies on proteases' key active sites and protease inhibitors to deepen the understanding of the active mechanism of proteases and their inhibitors.

VIH y salud ósea / HIV and bone health

Con el advenimiento de la terapia antirretroviral, el pronóstico y la sobrevida de los pacientes infectados con el virus de la inmunodeficiencia humana (VIH) han cambiado de manera radical, por lo cual en la actualidad se evidencia un aumento en el riesgo de padecer enfermedades no relacionadas con el VIH como, por ejemplo, la osteoporosis. La disminución de la densidad mineral ósea (DMO) se observa en el 40-90% de las personas infectadas por el VIH, con una prevalencia de osteopenia y osteoporosis del 52 y 15%, respectivamente. Esta población de pacientes tiene un mayor riesgo de fracturas (60%) en comparación con personas no infectadas y un riesgo de fracturas vertebrales 2,3 veces mayor que en la población general. El tenofovir fumarato se asoció con un aumento de pérdida renal de fósforo e hiperparatiroidismo secundario. El efavirenz y los inhibidores de proteasas (IP) afectan el metabolismo de la vitamina D; actúan a nivel enzimático aumentando la expresión de la enzima CYP24 que lleva a producción de vitamina D inactiva. El FRAX es una herramienta sencilla y accesible, por lo que su uso está recomendado en pacientes con VIH. Además de las medidas higiénico-dietéticas, actividad física, calcio y vitamina D, el uso de bifosfonatos está indicado en el tratamiento de la osteoporosis en estos pacientes. (AU)

With the advent of antiretroviral therapy, the prognosis and survival of patients infected with the human immunodeficiency virus (HIV) have radically changed, which is why there is now evidence of an increased risk of suffering from diseases not related to HIV such as osteoporosis. The decrease in bone mineral density (BMD) is observed in 40-90% of people infected with HIV, with a prevalence of osteopenia and osteoporosis of 52 and 15%, respectively. This patient population has a 60% higher risk of fractures compared to uninfected people and a risk of vertebral fractures 2.3 times higher than in the general population. Tenofovir fumarate administration is associated with increased renal phosphorus loss and secondary hyperparathyroidism. Efavirenz and protease inhibitors (IP) affect the metabolism of vitamin D, they act at the enzymatic level by increasing the expression of the CYP24 enzyme that leads to the production of inactive vitamin D. The FRAX is a simple and accessible tool, so its use is recommended in patients with HIV and in addition to dietary hygiene measures, physical activity, calcium, and vitamin D, the use of bisphosphonates is indicated in the treatment of osteoporosis in these patients. (AU)

Advances of Kunitz-type serine protease inhibitors / 生物工程学报

Kunitz-type serine protease inhibitors are a class of ubiquitous protease inhibitors, which play important roles in various life activities. The structures of such inhibitors are generally stable, and are usually characterized by the presence of one or several Kunitz domains in tandem, which are able to bind to serine proteases in a manner similar to substrate binding, thereby inhibiting enzyme activity. In terms of function, Kunitz-type serine protease inhibitors are involved in processes such as blood coagulation and fibrinolysis, tumor immunity, inflammation regulation, and resistance to bacterial and fungal infections. This article summarizes the advances of Kunitz-type serine protease inhibitors and provides new ideas for the development of novel Kunitz-type serine protease inhibitors.

High-throughput screening identifies established drugs as SARS-CoV-2 PLpro inhibitors

A new coronavirus (SARS-CoV-2) has been identified as the etiologic agent for the COVID-19 outbreak. Currently, effective treatment options remain very limited for this disease; therefore, there is an urgent need to identify new anti-COVID-19 agents. In this study, we screened over 6,000 compounds that included approved drugs, drug candidates in clinical trials, and pharmacologically active compounds to identify leads that target the SARS-CoV-2 papain-like protease (PLpro). Together with main protease (M

Safety of protease inhibitors and Arbidol for SARS-CoV-2 pneumonia in Zhejiang Province, China / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

The aim of this study was to evaluate the safety of an antiviral regimen of protease inhibitors combined with Arbidol (umifenovir) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pneumonia patients. The genomic sequence of SARS-CoV-2 is highly homologous to that of SARS-CoV (Zhou et al., 2020). Previously published basic and clinical research on anti-SARS-CoV treatment found that lopinavir/ritonavir (LPV/r) could improve the prognosis of SARS patients (Chan et al., 2003; Chu et al., 2004). Darunavir (DRV) is another protease inhibitor that blocks the binding of SARS-CoV-2 to human angiotensin-converting enzyme 2 (Omotuyi et al., 2020). The broad-spectrum antiviral drug Arbidol (umifenovir) also shows in vitro anti-SARS-CoV activity (Khamitov et al., 2008).

Partial characterization of digestive proteases in juveniles of Microphis brachyurus (short-tailed pipefish) (Syngnathiformes: Syngnathidae)

Short-tailed pipe fish (Microphis brachyurus) is a freshwater organism with high economic potential for the aquarium hobby, so it is necessary to implement methods to promote its culture through studies of digestive physiology. General activities of acid and alkaline proteases were evaluated, as well as the effect of pH, temperature and inhibitors. The optimal pH of stomach proteases was 2, while the optimal pH of intestinal proteases was 10. Optimal temperature for the acidic proteases was 35 ºC, while for alkaline proteases it was 45 ºC. Thermal stability showed high resistance at 35 ºC for both acid and alkaline proteases (above 100% residual activity). Acid proteases are resistant at pH 2 (50% of residual activity), meanwhile alkaline proteases were highly resistant at pH 10 (90% of residual activity). Acid proteases were inhibited by 80% with pepstatin A and alkaline proteases were inhibited with TLCK and TPCK for trypsin (75%) and chymotrypsin (80%), respectively. Finally, metallo-proteases were 75% partially inhibited some serine proteases by 75% with EDTA. In conclusion, M. brachyurus has a good digestive capacity, since they can degrade a wide variety of proteins due to their greater proteolytic activity.(AU)

El pez pipa (Microphis brachyurus) es un organismo dulceacuícola con alto potencial económico para la acuarofilia; sin embargo, es necesario implementar su cultivo a través de estudios de fisiología digestiva. Se evaluó el efecto del pH, temperatura e inhibidores sobre las actividades enzimáticas de proteasas ácidas y alcalinas. El pH óptimo de proteasas estomacales es de 2, mientras que el de proteases intestinales es de 10. La temperatura óptima de proteasas ácidas es de 35 ºC y las alcalinas de 45 ºC. La estabilidad térmica para proteasas ácidas y alcalinas es a los 35 ºC (más de 100% de actividad residual). La estabilidad a los diferentes pH de las proteasas ácidas es en 2 (50 % de la actividad residual), mientras que para las proteasas alcalinas es en 10 (90 % de la actividad residual). Las proteasas ácidas fueron inhibidas en 80% con pepstatina A y las proteasas alcalinas fueron altamente inhibidas con TLCK para tripsina (75%) y TPCK quimitripsina (80%). Finalmente, las metaloproteasas fueron inactivadas con EDTA en 70%. En conclusión, M. brachyurus tiene una buena capacidad digestiva al degradar una amplia variedad de proteinas debido a su alta actividad proteolítica.(AU)

Coagulation modifiers targeting SARS-CoV-2 main protease Mpro for COVID-19 treatment: an in silico approach

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection depends on viral polyprotein processing, catalysed by the main proteinase (Mpro). The solution of the SARS-CoV-2 Mpro structure allowed the investigation of potential inhibitors. This work aims to provide first evidences of the applicability of commercially approved drugs to treat coronavirus disease-19 (COVID-19). We screened 4,334 compounds to found potential inhibitors of SARS-CoV-2 replication using an in silico approach. Our results evidenced the potential use of coagulation modifiers in COVID-19 treatment due to the structural similarity of SARS-CoV-2 Mpro and human coagulation factors thrombin and Factor Xa. Further in vitro and in vivo analysis are needed to corroborate these results.

Acquired antiretroviral drug resistance mutations upon treatment failure in HIV-1 infected pediatric patients in central Brazil

Antiretroviral drug-resistance mutations compromise the successful treatment of children and adolescents infected with human immunodeficiency virus type 1 (HIV-1). We describe the clinical, virological, and immunological follow-up of a cohort of children and adolescents perinatally infected with HIV-1 treated at Hospital Estadual de Doenças Tropicais Dr. Anuar Auad ­ HDT, in Central Brazil, after therapeutic failure related to drug resistance mutations. We analyzed the results of the genotypic test (protease codons 1­99 and reverse transcriptase codons 1­325) performed from 2003 to 2015. The ARV susceptibility profile was analyzed according to Stanford HIV drug resistance database. A total of 65 patients (median age of 10 years; range, 18 m­18 y) with therapeutic failure (after a median of 55 months of follow up; range, 9 m­13 y) and plasma levels of HIV-1 RNA greater than 1,000 copies/mL which were included and demonstrated mutations in: nucleoside reverse transcriptase inhibitors (NRTIs), 98.5%; non nucleoside reverse transcriptase inhibitors (NNRTIs), 75.4%; and protease inhibitors (PI), 44.6%. The most frequent NRTI mutations were found in codon T215 (83.1%) with a predominance of T215Y (56.9%), followed by M184V (69.3%). In the NNRTI class, mutations K103N (36.9%) and 190A (23.1%) were predominant, and, in the protease, mutations 54VL (35.4%) and 82ASTL (32.3%) were found in approximately the same proportion, with a predominance of the M54V mutation. These results demonstrate the high levels of resistance to different classes of antiretrovirals in HIV-infected children and adolescents and the importance of genotypic resistance tests in this population

Hora de analizar la evidencia de izquierda a derecha / It is time to analyze the evidence from left to right

En este artículo, el autor reflexiona sobre las expectativas de los profesionales de la salud acerca de la evidencia para recomendar tratamiento farmacológico a los pacientes con COVID-19. (AU)

In this article, the author reflects on the expectations of health professionals regarding the evidence to recommend pharmacological treatment to patients with COVID-19. (AU)