Antiangiogenic and antitumoral activity of LQFM126 prototype against B16F10 melanoma cells.

Inhibition of mouse double minute 2 homolog (MDM2)-p53 interaction and reactivation of p53 signaling have been explored as effective anticancer therapeutic strategy. The potent and specific antitumor activity shown by Nutlins, first class of MDM2-p53 inhibitors discovered, has made these compounds potential antitumor candidates. To this end, we synthesized Nutlin-1 and Nutlin-2 analogs through molecular simplification and selected the compound with the most efficient antitumoral activity. Cytotoxicity of Nutlin-2 analog LQFM126 on B16F10 melanoma cells induced intense cytoplasmic vacuolization, reduction of cell size, chromatin condensation, cytoplasmic degeneration and nuclear fragmentation. LQFM126 antiproliferative effects mediated cell cycle retention in G0/G1 phase and increased the levels of cell cycle regulatory proteins p21 and p27. This Nutlin analog increased mitochondrial membrane potential, activated caspase-8, -9 and -3/7 and reduced VEGF levels in B16F10 cells. Therefore, LQFM126 promoted alterations suggestive of apoptosis, G0/G1 cell cycle arrest and suppression of angiogenesis through modulation of VEGF expression in B16F10 cells. Additionally, LQFM126 was classified as UN GHS category 4 (LD50 > 300-2000 mg/kg), suggesting it has low acute systemic toxicity. LQFM126 can be a promising prototype for anticancer therapy.

Innovative strategy based on mechanisms to substitute animal testing for ocular toxicity assessment of agrochemical formulations market in Brazil.

Considering the successful employment of alternative methods for eye toxicity assessment of products for regulatory purposes, and the recent advances in Brazilian legislative scenario, which adopted the UN GHS classification system for agrochemical formulations toxicity assessment, there is an emerging demand for strategies that allow the evaluation of such products. Based on this, the present study aimed to address the applicability of a mechanistic-based defined approach for eye toxicity assessment of agrochemical formulations. It was investigated the opacity/permeability, depth and location of corneal injury in bovine cornea, and vascular events in chorioallantoic membrane induced for different Brazilian agrochemicals using a Sequential Testing Strategy (STS). Cytotoxicity induced by the agrochemical formulations was evaluated by Short Time exposure (STE) (OECD TG 491) assay (step 1), corneal injury was investigated by standard Bovine Corneal Opacity and Permeability (BCOP) (OECD TG 437) followed by histopathological evaluation (step 2), and Hen Chorionic-allantoic Membrane test (HET-CAM) was used to evaluate vascular injury (step 3). The results demonstrated that the proposed defined approach enabled a classification corresponding UN GHS classification of agrochemical formulations while minimizing the use of live animals. Therefore, this approach may be useful for categorization of agrochemicals in Brazil according to the new regulatory scenario.

Corneal histomorphometric analysis: The depth of damage induced in the bovine cornea correlates with the severity of the ocular toxicity.

The Bovine Corneal Opacity and Permeability (BCOP) assay is an alternative method used to ocular toxicity potential assessment of chemicals and mixtures. The standard BCOP test provides information about permeability and opacity, however, corneal histopathological analysis has been recommended as an additional parameter to better categorize eye irritants. Moreover, such analysis associated with depth of substance-induced corneal injury analysis may provide additional scientific measurement for the refinement of BCOP test. The aim of this study was to measure the depth of injury into the bovine cornea induced by eye irritants and associate it with the damage severity. For this purpose, BCOP assay was performed for 12 substances from different Globally Harmonized System of Classification and Labelling of Chemicals (UN GHS) categories and, additionally, corneal sections of 5 µm thickness were obtained and analyzed by fluorescence microscopy. The results showed that the fluorescein permeation depth was directly proportional to the substances irritation degree. Severe irritants promoted highest rates of permeation followed by moderate and mild irritants, while non-irritants showed similar permeation indexes to the negative control. The refinement of BCOP by the depth of injury analysis through epithelial permeation of fluorescein can be considered a useful quantitative parameter to better categorize eye irritants.

Eugenia dysenterica DC. (Myrtaceae) exerts chemopreventive effects against hexavalent chromium-induced damage in vitro and in vivo.

CONTEXT: Eugenia dysenterica DC. (Myrtaceae) has been widely used in the folk medicine and it presents phytochemicals constituents associated to antioxidant properties. OBJECTIVE: The objective of this study was to investigate the protective effects of E. dysenterica leaf hydroalcoholic extract (EDE) in vitro and in vivo using AMJ2-C11 cells and Swiss mice exposed to hexavalent chromium [Cr(VI)], respectively. MATERIALS AND METHODS: AMJ2-C11 cells were pretreated with EDE and exposed to Cr(VI) to evaluate cytotoxicity and the pathways involved in the chemopreventive effects of the extract. Mice were daily pretreated with EDE and then exposed to Cr(VI). Survival analysis, histopathological examination and determination of Cr levels in biological tissues were carried out. RESULTS: In vitro studies showed that pretreatment of the AMJ2-C11 cells with EDE protected against the cytotoxicity and oxidative stress induced by Cr(VI). Consequently, the pretreatment with EDE reduced reactive oxygen species and apoptosis triggered by Cr(VI), probably by a marked antioxidant and chelating activities demonstrated by EDE. Regarding in vivo studies, pretreatment for 10 days with EDE increased survival of the mice exposed to Cr(VI). In addition, EDE prevented liver and kidney pathological damages, in parallel with reduction in chromium levels found in these organs and plasma. EDE also showed a marked antioxidant potential associated with the presence of polyphenols, especially flavonoids and tannins, as confirmed by HPLC-PDA. CONCLUSION: The study showed that EDE protects against Cr(VI)-induced damage in vitro and in vivo supporting further studies for the development of therapeutic products applied to prevent the damage induced by toxic metals, especially Cr(VI).

Antifungal and cytotoxicity activities of the fresh xylem sap of Hymenaea courbaril L. and its major constituent fisetin.

BACKGROUND: The great potential of plants as Hymenaea courbaril L (jatoba) has not yet been throughly explored scientifically and therefore it is very important to investigate their pharmacological and toxicological activities to establish their real efficacy and safety. This study investigated the cytotoxicity of xylem sap of Hymenaea courbaril L and its bioactivity against the fungi Cryptococcus neoformans species complex and dermatophytes. METHODS: The fresh xylem sap of H. courbaril was filtered resulting in an insoluble brown color precipitate and was identified as fisetin. In the filtrate was identified the mixture of fisetinediol, fustin, 3-O-methyl-2,3-trans-fustin and taxifolin, which were evaluated by broth microdilution antifungal susceptibility testing against C. neoformans species complex and dermatophytes. The fresh xylem sap and fisetin were screened for cytotoxicity against the 3T3-A31 cells of Balb/c using neutral red uptake (NRU) assay. RESULTS: The fresh xylem sap and the fisetin showed higher in vitro activity than the filtrate. The xylem sap of H. courbaril inhibited the growth of dermatophytes and of C. neoformans with minimal inhibition concentration (MIC) < 256 µg/mL, while the fisetin showed MIC < 128 µg/mL for these fungi. Fisetin showed lower toxicity (IC50 = 158 µg/mL) than the fresh xylem sap (IC50 = 109 µg/mL). CONCLUSION: Naturally occurring fisetin can provide excellent starting points for clinical application and can certainly represent a therapeutic potential against fungal infections, because it showed in vitro antifungal activity and low toxicity on animal cells.

Quantification of mast cells in the uterine cervix of women infected with human immunodeficiency virus.

The objective of the study was to identify mastocytosis in the chorionic epithelium of the uterine cervix in HIV-infected and non-HIV-infected women in autopsy specimens using histochemistry and immunohistochemistry techniques. Sixteen cervical tissue specimens were collected, of which 10 (62.50%) were from HIV-infected women. Histochemical and immunohistochemical techniques were used to evaluate mast cell density using Giemsa stain and anti-mast cell tryptase and anti-mast cell chymase antibodies, respectively. The study of the sheets and counting of mast cells with blue (Giemsa) or brown staining (anti-mast cell tryptase or chymase antibodies) were performed by 3 examiners, and 10 consecutive fields were examined under a light microscope at 400× magnification. A significant difference was found in mast cell density in the chorionic epithelium of the cervix in HIV-infected compared with non-HIV-infected women. The present study may contribute to the characterization of genital mucosa abnormalities and help better understand the potential role of mast cells in HIV-infected women.