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12. Chapter 8 - In vitro fucoxanthin production by the Phaeodactylum tricornutum diatom

Author

Bauer, C. M., Schmitz, C., Corrêa, R. G., Herrera, C. M., Ramlov, F., Oliveira, E. R., Pizzato, A., Varela, L. A. C., Cabral, D. Q., Yunes, R. A., Lopes, R. G., Cella, H., Rocha, Miguel, Rorig, L. R., Derner, R. B., Abreu, P. C., Maraschin, M., and Universidade do Minho

Subjects
Carotenoid, In vitro cell cultures, Science & Technology, fungi, Elicitor, Metabolic engineering
Abstract

Diatoms, along with brown seaweeds, are the main producers of fucoxanthin, an oxycarotenoid with important biological functions such as antioxidant, anti-obesity, antitumor, antidiabetic, and anti-inflammatory activities. In this scenario, the marine diatom Phaeodactylum tricornutum is considered an important source of this carotenoid. Fucoxanthin production by this diatom has been studied through different approaches, e.g., in vitro and in vivo investigation of the effects of environmental factors and strains, as well as through metabolic engineering analysis. The optimization of environmental factors such as light exposure, pH, temperature and aeration has been reported to modulate fucoxanthin production by P. tricornutum cell cultures. In parallel, the identification and selection of strains with an elevated fucoxanthin biosynthesis and accumulation have also contributed to improve the biotechnological performance of that microalga. Indeed, much can be done from the screening of the genomic sequence of P. tricornutum, which has been completed in 2008. Candidate genes for specific reactions in carotenoid biosynthesis pathways can be cloned, expressed, and identified by combining genetic and analytical strategies. Furthermore, metabolic reactions can be used to perform in vitro simulations, mitigating laboratory costs. This chapter aims to present an overview of fucoxanthin biosynthesis in P. tricornutum as well as the different ways to enhance its production., CNPq -Conselho Nacional de Desenvolvimento Científico e Tecnológico(307099/2015-6), info:eu-repo/semantics/publishedVersion

Published
2019

18. Calcium-triggered acrosomal exocytosis in human spermatozoa requires the coordinated activation of Rab3A and N-ethylmaleimide-sensitive factor

Author

Michaut, M., Tomes, C. N., De Blas, G., Yunes, R., and Mayorga, L. S.

Subjects
Exocytosis -- Research, Spermatozoa -- Research, Cell membranes -- Research, Membrane fusion -- Research, Science and technology
Abstract

The acrosome reaction of spermatozoa is a complex, calcium-dependent, regulated exocytosis. Fusion at multiple sites between the outer acrosomal membrane and the cell membrane causes the release of the acrosomal contents and the loss of the membranes surrounding the acrosome. However, very little is known about the molecules that mediate and regulate this unique fusion process. Here, we show that N-ethylmaleimide-sensitive factor (NSF), a protein essential for most fusion events, is present in the acrosome of several mammalian spermatozoa. Moreover, we demonstrate that calcium-dependent exocytosis of permeabilized sperm requires active NSF. Previously, we have shown that the addition of the active (GTP-bound) form of the small GTPase Rab3A triggers exocytosis in permeabilized spermatozoa. In the present report we show that Rab3A is necessary for calcium-dependent exocytosis. The activation of Rab3A protects NSF from N-ethylmaleimide inhibition and precludes the exchange of the endogenous protein with recombinant dominant negative mutants of NSF. Furthermore, Rab3A activation of acrosomal exocytosis requires active NSF. Our results suggest that, upon calcium stimulation, Rab3A switches to its active GTP-bound form, triggering the formation of a protein complex in which NSF is protected. This process is suggested to be an essential part of the molecular mechanism of membrane fusion leading to the release of the acrosomal contents.

Published
2000

20. Microscopic diagnosis of the leaf and stem of Piper solmsianum C.DC.

Author

Bertocco, A. R. P., Migacz, I. P., Santos, V. L. P., Franco, C. R. C., Silva, R. Z., Yunes, R. A., Cechinel‐Filho, V., and Budel, J. M.

Abstract

Piper solmsianum C.DC., which is popularly known as pariparoba, is a shrub that measures 1-3 m in height and it inhabits areas with wet tropical soils. The objective of this study was to analyze the leaf and stem anatomy using light microscopy, scanning electron micrographs, and energy-dispersive X-ray spectroscopy in order to provide information for species identification. The anatomical profile showed the following main microscopic markers: hypostomatic leaf; hypodermis layer on both sides; pearl glands; biconvex midrib shape; five collateral vascular bundles in open arc with the central bundle larger than the others; circular stem shape; collateral vascular bundles arranged in two rings; sinuous sclerenchymatic sheath in the pith; secretory idioblasts; and starch grains in the mesophyll, in the ground parenchyma of the midrib, petiole, and in the stem; and six morphotypes of calcium oxalate crystals (styloids, cuneiform, tabular crystal rosettes, cuneiform crystal rosettes, elongated square dipyramids, as well as very elongated square dipyramids). [ABSTRACT FROM AUTHOR]

Published
2017
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23. Molecular Cytotoxic Mechanisms of 1-(3,4,5-Trihydroxyphenyl)-dodecylbenzoate in Human Leukemia Cell Lines.

Author

MAIORAL, M. F., BUBNIAK, L. D. S., MARZAROTTO, M. A. L., DE MORAES, A. C. R., LEAL, P., NUNES, R., YUNES, R. A., and SANTOS-SILVA, M. C.

Subjects
LEUKEMIA treatment, CELL lines, DRUG toxicity, CELL survival, GALLIC acid, APOPTOSIS
Abstract

Recent studies have shown that gallic acid and its alkylesters induce apoptosis in different cell lines. Since new compounds with biological activity and less cytotoxicity to normal cells are necessary for cancer therapy, the aim of this study was to evaluate the cytotoxic effect of 1-(3,4,5-trihydroxyphenyl)-dodecylbenzoate on human acute myeloid leukemia K562 cells and on human acute lymphoblastic leukemia Jurkat cells. The cell viability was determined by MTT method. The apoptosis induction was assessed by bromide and acridine orange staining and by Annexin V-FITC Apoptosis Detection kit. The cell cycle analysis was carried out by flow cytometry using propidium iodide. Cytometric analysis was also performed to evaluate the expression of the following proteins: AIF, p53, Bcl-2 and Bax. The mitochondrial potential was also assessed by flow cytometry using MitoView633 kit. The results showed that the compound significantly reduced the cell viability of K562 and Jurkat cells in a concentration and time dependent manner (IC50 of 30 μM). The compound induced cell cycle arrest in G0/G0 phase and significantly increased the proportion of cells in the sub-G0/G0 phase. Apoptosis was confirmed by the sight of morphological characteristics of apoptosis and by phosphatidylserine externalization (73.47±5.71% of cells expressing annexin). The results also showed that the compound promotes a modification in Bax:Bcl-2 ratio and increases p53 expression. Thus, it is possible to conclude that 1-(3,4,5-trihydroxyphenyl)-dodecylbenzoate induces apoptosis by inhibiting the antiapoptotic protein Bcl-2 and by increasing the release of AIF, Bax and p53. In addition, it blocks the cell cycle at G0/G0, stopping cell proliferation. So far, the results suggest that this compound may have a potential therapeutic effect against leukemia cells. [ABSTRACT FROM AUTHOR]

Published
2016
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