The evaluation of skin sensitization potential of the UVCB substance diisopentyl phthalate by in silico and in vitro methods.

Diisopentyl phthalate (DiPeP) is primarily used as a plasticizer or additive within the production of polyvinyl chloride (PVC), and has many additional industrial applications. Its metabolites were recently found in urinary samples of pregnant women; thus, this substance is of concern as relates to human exposure. Depending upon the nature of the alcohol used in its synthesis, DiPeP may exist either as a mixture consisting of several branched positional isomers, or as a single defined structure. This article investigates the skin sensitization potential and immunomodulatory effects of DiPeP CAS No. 84777-06-0, which is currently marketed and classified as a UVCB substance, by in silico and in vitro methods. Our findings showed an immunomodulatory effect for DiPeP in LPS-induced THP-1 activation assay (increased CD54 expression). In silico predictions using QSAR TOOLBOX 4.5, ToxTree, and VEGA did not identify DiPeP, in the form of a discrete compound, as a skin sensitizer. The keratinocyte activation (Key Event 2 (KE2) of the adverse outcome pathway (AOP) for skin sensitization) was evaluated by two different test methods (HaCaT assay and RHE assay), and results were discordant. While the HaCaT assay showed that DiPeP can activate keratinocytes (increased levels of IL-6, IL-8, IL-1α, and ILA gene expression), in the RHE assay, DiPeP slightly increased IL-6 release. Although inconclusive for KE2, the role of DiPeP in KE3 (dendritic cell activation) was demonstrated by the increased levels of CD54 and IL-8 and TNF-α in THP-1 cells (THP-1 activation assay). Altogether, findings were inconclusive regarding the skin sensitization potential of the UVCB DiPeP-disagreeing with the results of DiPeP in the form of discrete compound (skin sensitizer by the LLNA assay). Additional studies are needed to elucidate the differences between DiPeP isomer forms, and to better understand the applicability domains of non-animal methods in identifying skin sensitization hazards of UVCB substances.

An integrated in silico-in vitro investigation to assess the skin sensitization potential of 4-Octylphenol.

One of the major challenges in chemical toxicity testing is the possibility to protect human health against adverse effects with non-animal methods. In this paper, 4-Octylphenol (OP) was tested for skin sensitization and immunomodulatory effects using an integrated in silico-in vitro test approach. In silico tools (QSAR TOOLBOX 4.5, ToxTree and VEGA) were used together with several in vitro tests including HaCaT cells (quantification of IL-6; IL-8; IL-1α and IL-18 by ELISA and expression of genes TNF, IL1A, IL6 and IL8 by RT- qPCR), RHE model (quantification of IL-6; IL-8; IL-1α and IL-18 by ELISA) and THP-1 activation assay (CD86/CD54 expression and IL-8 release). Additionally, the immunomodulatory effect of OP was investigated using lncRNAs MALAT1 and NEAT1 expression and LPS-induced THP-1 activation (CD86/CD54 expression and IL-8 release). The in silico tools predicted OP as a sensitizer. In vitro tests are also concordant with the in silico prediction. OP increased IL-6 expression (HaCaT cells); IL-18 and IL-8 expressions (RHE model). An irritant potential was also shown by a great expression of IL-1α (RHE model); and increased expression of CD54 marker and IL-8 in THP-1 cells. Immunomodulatory effects of OP were demonstrated by the downregulation of NEAT1, MALAT1 (epigenetic markers), IL6 and IL8; and an increase in LPS-induced CD54 and IL-8 expressions. Overall, results indicate that OP is a skin sensitizer, being positive in three key events of the AOP for skin sensitization, also showing immunomodulatory effects.

The potential use of kraft lignins as natural ingredients for cosmetics: Evaluating their photoprotective activity and skin irritation potential.

The cosmetics market has recently undergone changes as consumers increasingly seek sustainable products. In this context, agro-derived lignins have the potential for cosmetics applications. Thus, this study investigated the photoprotective activity and skin irritation potential (OECD TG 439 with SkinVitro-RHE, an in-house reconstructed human epidermis (RHE) model) of a kraft lignin (LE) and two LE-derived lignins modified by enzymatic reactions to achieve higher molecular weight (Mw) (R1: intermediate Mw and E60: highest Mw). Results showed that LE and R1 lignins present adequate photoprotective activity with averages Sun Protection Factor (SPF) of 33.8 ± 0.02 and 22.7 ± 0.04, respectively. The E60 also present adequate SPF (22.4 ± 0.2); however, due to its poor solubility, this lignin has potential application as a physical filter. In terms of safety, these lignins did not cause skin irritation or cellular and structural damage to the epidermis. Additionally, using an analysis based on the autofluorescence feature of lignin, no sign of the tested lignins was found in exposed RHE models, indicating that these three lignins did not penetrate the skin. Altogether, the results indicate a promising application of kraft lignins for sunscreen products regarding safer alternatives and product sustainability. Also, the SkinVitro-RHE showed to be a good model for evaluating the skin irritation potential of substances, including natural cosmetic ingredients.

A Robust, Mechanistically Based In Silico Structural Profiler for Hepatic Cholestasis.

Owing to the primary role which it holds within metabolism of xenobiotics, the liver stands at heightened risk of exposure to, and injury from, potentially hazardous substances. A principal manifestation of liver dysfunction is cholestasis-the impairment of physiological bile circulation from its point of origin within the organ to the site of action in the small intestine. The capacity for early identification of compounds liable to exert cholestatic effects is of particular utility within the field of pharmaceutical development, where contribution toward candidate attrition is great. Shortcomings associated with the present in vitro methodologies forecasting cholestasis render their predictivity questionable, permitting scope for the adoption of computational toxicology techniques. As such, the intention of this study has been to construct an in silico profiler, founded upon clinical data, highlighting structural motifs most reliably associated with the end point. Drawing upon a list of >1500 small molecular drugs, compiled and annotated by Kotsampasakou, E. and Ecker, G. F. (J. Chem. Inf. Model. 2017, 57, 608-615), we have formulated a series of 15 structural alerts. These describe fragments intrinsic within distinct pharmaceutical classes including psychoactive tricyclics, ß-lactam antimicrobials, and estrogenic/androgenic steroids. Description of the coverage and selectivity of each are provided, alongside consideration of the underlying reactive mechanisms and relevant structure-activity concerns. Provision of mechanistic anchoring ensures that potential exists for framing within the adverse outcome pathway paradigm-the chemistry conveyed through the alert, in particular enabling rationalization at the level of the molecular initiating event.

Incorporating lines of evidence from New Approach Methodologies (NAMs) to reduce uncertainties in a category based read-across: A case study for repeated dose toxicity.

A group of triazole compounds was selected to investigate the confidence that may be associated with read-across of a complex data gap: repeated dose toxicity. The read-across was evaluated using Assessment Elements (AEs) from the European Chemicals Agency's (ECHA's) Read-Across Assessment Framework (RAAF), alongside appraisal of associated uncertainties. Following an initial read-across based on chemical structure and properties, uncertainties were reduced by the integration of data streams such as those from New Approach Methodologies (NAM) and other existing data. In addition, addressing the findings of the ECHA RAAF framework, complemented with specific questions concerning uncertainties, increased the confidence that can be placed in read-across. Although a data rich group of compounds with a strong mechanistic basis was analysed, it was clearly demonstrated that NAM data available from publicly available resources could be applied to support read-across. It is acknowledged that most read-across studies will not be so data rich or mechanistically robust, therefore some targeted experimentation may be required to fill the data gaps. In this sense, NAMs should constitute new experimental tests performed with the specific goal of reducing the uncertainties and demonstrating the read-across hypothesis.

Microscopia ótica de alta resoluçäo: adaptaçäo do método e sua aplicaçäo ao estudo da leptospirose experimental em cobaias / High resolution light microscopy: adaptation of the method and its use in the study of experimental leptospirosis in guinea pigs

As lesöes morfológicas em parênquina e vasos de fígado e rim foram estudadas em um lote de cobaias infectadas experimentalmente com Leptospira interrogans sorogrupos icterohaemorrhagiae, em comparaçäo com um grupo de cobaias normais näo infectadas. O material foi analisado a microscopia óptica convencional (MOC), através da inclusäo em parafina, e a de alta resoluçäo (MOAR), esta com cortes de um micrômetro de espessura e até-96 mm2 de superfície, em tecido incluido em glicol-metacrilato. A utilizaçäo pela primeira vez da microscopia óptica de alta resoluçäo para o estudo da leptospirose revelou novos aspectos da doença, na medida em que permitiu análise de detalhes celulares e visäo de conjunto do quadro histológico na mesma amostra. Assim foi possível constatar se tanto alteraçöes da coesäo intercelular como especializaçöes de membranas, incluindo borda em escova, e vesículas de pinocitose, além da distribuiçäo intracelular de organelas saculares, provavelmente mitocondrias. A introduçäo desta metodologia de baixo custo e fácil execuçäo poderátrazer contribuiçöes úteis a histotecnologia com objetivos diagnósticos e em pesquisa.

Expressäo antigênica e diferenciaçäo morfológica de neoplasias mamárias: um estudo comparativo / Antigenic characters and morphologic differences of the mammary neoplasms: a comparative study

Foram selecionados quatro carcinomas ductais invasivos de mama com graus de anaplasia progressivos e testados por métodos imuno-histoquímicos para marcadores de diferenciaçäo mamária com o objetivo de verificar a correlaçäo entre grau histológico e a presença desses marcadores. Foram utilizados anticorpos contra alfa-lactoalbumina, lactoferrina, caseína, componente secretor, antígeno epitelial de membrana e antígeno carcinoembriônico. A neoplasia mais diferenciada resultou positiva para todos os antígenos pesquisados, enquanto a neoplasia menos diferenciada resultou negativa, com exceçäo da alfa-lactoalbumina. As neoplasias com diferenciaçäo intermediária apresentaram coloraçäo focal nas áreas mais diferenciadas da lesäo. Uma correlaçäo entre a expressäo dos antígenos de diferenciaçäo celular mamária e uma boa diferenciaçäo histológica foi verificada para a maioria dos marcadores utilizados.

Utilizaçäo da imuno-histoquímica na detecçäo da sede primária em neoplasias metastáticas / Use of the immunohistochemistry in the detection of the primary site in neoplasm metastasis

O objetivo deste trabalho é demonstrar a utilizaçäo de marcadores teciduais, tendo como base amostra selecionada de tumores metastáticos em linfonodos de diversas origens para avaliar sua aplicabilidade, sensibilidade e especificidade na determinaçäo dos sítios primários. Para tanto foram selecionados 19 casos de metástases de neoplasias primárias dos seguintes órgäos: mama (6) , estômago (4), reto (1), cólon (1), tiróide (3), próstata (1), pâncreas (1), ovário (1) e melanoma cutâneo (1). Cortes histológicos desse material foram processados pelo método da imunoperoxidase utilizando-se dois anticorpos policlonais antitiroglobulina e antiantígeno específico de próstata e os anticorpos monoclonais: BRST-1, BRST-2, CAR-3, BD-5 E HMB-45, sendo os dois primeiros marcadores de mama, os outros dois de neoplasias gastrointestinais e o último marcador de melanoma. BRST-1 e BRST-2 coraram três das seis metástases de mama. BRST-1 foi também positivo para as metástases de melanoma e próstata. Os anticorpos monoclonais CAR-3 e BD-5 resultaram positivos em cinco das seis metástases gastrointestinais. Quatro das seis metástases de mama e as da tiróide foram positivas para CAR-3, assim como as metástases de próstata, ovário, pâncreas e melanoma. O anticorpo BD-5 corou também a metástase de próstata. Tiroglobulina e antígeno específico de próstata resultaram positivos nas metástases de tiróide e próstata, respectivamente. Em conclusäo, os anticorpos monoclonais e as técnicas imuno-histoquímicas têm se revelado ferramentas úteis na detecçäo da sede primária de neoplasias de origem desconhecida. Em alguns campos os resultados obtidos já säo satisfatórios, embora na maioria das áreas, estudos adicionais ainda sejam necessários

Patologia da leptospirose. Análise crítica dos aspectos morfológicos e imuno-histoquímicos relevantes para a compreensäo da patogenia / Pathology of leptospirosis. A critical analysis of morphological and immunochemical features relevant to pathogenesis of systemic lesion

Essa revisäo que se concentra nos aspectos morfológicos e imuno-histoquímicos da leptospirose humana e experimental analisa as principais possibilidades patogenéticas. O fígado pode apresentar hepatite colestática com destrabeculaçäo de hepatócitos nas fases de máxima gravidade. O acometimento renal varia desde nefrite intersticial até necrose tubular aguda. Miosites com predomínio de mononucleares säo descritas em vários músculos, especialmente na panturrilha. Miocardites e coronarites säo relacionadas aos casos de morte súbita. De fato, a vasculite pode ser considerada sistêmica e multifocal. Os pulmoes podem ter focos pneumônicos e até extensas hemorragias alveolares e pleurais. A detecç äo de antígenos de leptospira em biópsias e autópsias permite o diagnóstico etiológico, favorecendo também hipóteses patogenéticas que propoem que as principais lesoes decorram de açäo direta, sobre membranas de células parenquimatosas e endoteliais, inicialmente da leptospira íntegra e a seguir de produtos de sua degradaçäo por macrófagos