Anti-Inflammatory Activity and Chemical Analysis of Different Fractions from Solidago chilensis Inflorescence.

Solidago chilensis Meyen (Compositae) is a species native to South America (Brazil) popularly known as arnica. In Brazilian popular medicine, inflorescences and rhizomes of this plant have been used since the end of the 19th century to replace the exogenous and hepatotoxic Arnica montana L. in the treatment of edema and inflammatory pathologies. Although the anti-inflammatory activity of S. chilensis is evidenced in the literature, there is a lack of studies with enriched fractions or compounds isolated from it. The objective of the current study was to characterize phytochemically and to evaluate the pharmacological action in vivo and in vitro of the crude extract and the different fractions (hexane, dichloromethane, acetal, butanolic, and aqueous) isolated from the inflorescence of S. chilensis. The inflorescence crude extract (ScIE) and fractions were administered by intraperitoneal route to mice at different doses. In an LPS-induced pleurisy model, inhibition of leukocyte influx was observed for the ScIE and all fractions tested, as compared to controls. Dichloromethane (ScDicF), butanolic (ScButF), and aqueous (ScAquF) were selected for further analysis as they showed the best inhibitory effects in leukocyte migration and inflammatory cytokine and chemokine production: TNF-α, CXCL1/KC, CXCL2/MIP-2, and CCL11/eotaxin-1. In LPS-stimulated J774A.1 cell line, ScIE and the ScDicF exhibited an inhibitory effect on nitric oxide (NO) production and downmodulated the COX-2 expression; ScAquF failed to modulate NO production and COX-2 expression. In phytochemical analysis, HPLC-UV-DAD chromatograms of ScDicF and ScAquF showed the main peaks with UV spectrum characteristics of flavonoids; chlorogenic acid and isoquercetin were the most present phytochemicals identified in the ScAquF, and a high number of n-alkanes was found in ScHexF. Our study was the first to address biological effects and correlate them to phytochemically characterized fractions from inflorescences of S. chilensis.

Protective effect of Myrsine parvifolia plant extract against the inflammatory process induced by Bothrops jararaca snake venom.

Accidents involving snakes from the genus Bothrops sp. constitute the most important cause of snake envenomation in Brazil. The Myrsine genus has been reported to be used in folk medicine against snakebites. In this work, the phytochemical profiles and ability of extracts from Myrsine parvifolia leaves to reduce the inflammatory process (edema, vascular permeability increase and leukocyte migration) induced by Bothrops jararaca venom were investigated in vivo. Chemical compounds were identified by chromatographic and spectroscopy techniques. Total polyphenol, tannin, and flavonoid contents were determined by spectrophotometric methods. Swiss male mice received an oral administration of extracts (100 mg/kg) in different protocols. Paw edema, intraperitoneal vascular permeability and pleurisy models in mice were used to evaluate the antiophidic potential of the extracts. Paw edema was induced by subplantar injection of B. jararaca venom and quantified as the increase in paw volume. Changes in vascular permeability were assessed by measuring the amount of Evans blue dye extravasation. Leukocyte migration was assessed by total and differential counts in the pleural cavity washes. Myricetin, myricetin-3-O-ß-arabinopyranoside, quercetin and kaempferol were isolated from the ethyl acetate extract and identified as the primary compounds of the dichloromethane extract. Terpenes and fatty acids were identified in the hexane and dichloromethane extracts. The pretreated group with hydroethanolic and dichloromethane extract reduced total edema (40 and 52%, respectively), vascular permeability increase (32.4 and 32.2%, respectively) and leukocyte influx into the pleural cavity (42 and 39%, respectively), while the group treated with hexane extract showed only reduced edema (37%) induced by B. jararaca venom. The hydroethanolic extract showed better results in all of the tests performed and was also administered by the protocol of post-poisoning, showing maintenance of paw edema reduction and cell migration. These data indicate a potential anti-inflammatory activity of M. parvifolia in poisoning by B. jararaca, especially to reduce local poison effects.

Molecular detection, quantification and characterization of human polyomavirus JC from waste water in Rio De Janeiro, Brazil.

Polyomavirus JC (JCPyV) is largely excreted by the human population through the urinary route and has been recognized as a potential viral marker for human waste contamination. This study aims to investigate the dissemination of JCPyV in waste water from a sewage treatment plant (STP) located in Rio de Janeiro, Brazil, and to describe the prevalence of JCPyV subtypes currently present in this population. Raw and treated sewage samples were collected bimonthly during one year, and examined for the presence of JCPyV using nested polymerase chain reaction (nPCR) and quantitative real time PCR (qPCR). JCPyV was detected by nPCR in 96% and 43% of raw and treated sewage samples, respectively. The concentration of JCPyV present in the samples ranged from 1.2x10(3) to 3.2x10(5) and 2.6x10(2) to 6.2x10(3) genome copies per 2 ml of concentrated raw and treated sewage sample, respectively. The strains were characterized and the obtained nucleotide sequences indicated that the detected JCPyV strains clustered with subtypes of East African, West African and European origin. To our knowledge, this is the first study describing the incidence and diversity of JCPyV strains in raw and treated sewage in Brazil.

Adaptação da técnica de reação em cadeia de polimerase (PCR) quantitativa em tempo real para o diagnóstico da leucoencefalopatia multifocal progressiva (LMP) / Adapting the technique of polymerase chain reaction (PCR) quantitative real-time diagnosis of progressive multifocal leukoencephalopathy (PML)

A leucoencefalopatia multifocal progressiva (LMP), doença subaguda desmielinizante do sistema nervoso central (SNC), é causada pelo Poliomavirus Humano JC (JCPyV). Devido ao uso de terapias imunossupressoras e à SIDA a frequência de casos de LMP aumentou consideravelmente nas últimas décadas. O padrão ouro para o diagnóstico da LMP é a biópsia cerebral. Entretanto, apesar do elevado poder diagnóstico, este é um método invasivo, com uma taxa custo/benefício elevada. Assim, utiliza-se a tomografia computadorizada ou a ressonância magnética do crânio para o diagnóstico presuntivo de LMP. Métodos moleculares, em especial a reação em cadeia da polimerase (PCR), têm se mostrado eficiente no diagnóstico de várias doenças virais do SNC. A nested-PCR (nPCR) é uma técnica mais eficaz na detecção do DNA do JCPyV no liquidocefalorraqueano (LCR), proporcionando maior sensibilidade e especificidade quando comparada à PCR convencional. Entretanto, aproximadamente 15 por cento dos casos prováveis de LMP podem não se confirmar pela nPCR. Na era da terapia antiretroviral altamente potente (HAART), o percentual de casos não confirmados aumentou, chegando à aproximadamente 42 por cento. Publicações recentes nas quais se utilizou a PCR quantitativa (qPCR) em tempo real demonstram que esta técnica pode ser 10 vezes mais sensível quando comparada à PCR convencional na detecção e quantificação do JCPyV, podendo também ser utilizado como um marcador virológico em pacientes em uso de HAART com diagnóstico de LMP. Neste trabalho, elaboramos uma técnica que permite quantificar a carga viral do JCPyV no LCR de pacientes com LMP em razão de acompanhar a resposta do paciente ao tratamento com a HAART. O princípio da técnica qnPCR em tempo real estabelecida foi semelhante ao da nPCR, sendo constituído por dois passos de amplificação sequenciados. O primeiro passo trata-se de uma PCR convencional seguido de nova amplificação, agora utilizando a técnica de qPCR em tempo real e tendo como molde o produto da primeira amplificação. Foram analisadas amostras de LCR armazenadas pelo Laboratório de Pesquisa Clínica em Neuroinfecções. Destas, 16 eram de pacientes com confirmação diagnóstica pelo método de nPCR e 10 eram de casos compatíveis mas sem diagnóstico. Pela qnPCR em tempo real, nenhum dos casos compatíveis com LMP apresentou quantificação de carga viral e das 16 amostras positivas, apenas 12 apresentaram sinais de carga que ultrapassaram a linha de base apontando a presença do DNA do JCPyV. Em nossa casuística a nPCR teve sensibilidade de 69,6 por cento enquanto a sensibilidade da qnPCR foi de 52,2 por cento. Concluímos que a questão diagnóstica da LMP ainda persiste, pois não conseguimos aumentar a sensibilidade de detecção do JCPyV no LCR, entretanto, desenvolvemos um método quantitativo que poderá auxiliar como dado da resposta do paciente ao tratamento com a HAART.