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Melanoma cells differentiation through inhibition of p-Mek activity following treatment with 5,7-dimethoxycoumarin
Get PDFIn questo lavoro è stata studiata l’attività antineoplastica della 5,7-dimetossicumarina, un metabolita secondario di origine vegetale, su cellule di melanoma murino (B16) e umano (A375). Il composto riduce significativamente la proliferazione cellulare in modo dose e tempo dipendente, bloccando nella fase G0/G1 il ciclo di entrambe le linee cellulari. In seguito a questo blocco, è stato inoltre osservato un processo di differenziamento nelle cellule trattate attraverso l’analisi di markers specifici quali lo sviluppo di proiezioni dendritiche dalla superficie cellulare, l’incremento nella sintesi di melanina e l’accumulo della protoporfirina IX (PpIX).
Considerando le attività antineoplastica e differenziativa della 5,7-dimetossicumarina, è stato esaminato il suo effetto sulla fosforilazione, e quindi sull’attività, delle protein chinasi attivate da mitogeni Ras/Raf/Mek/Erk (MAPK). Questa via chinasica regola la sopravvivenza, la proliferazione, e il differenziamento cellulari ed è costitutivamente attivata, in seguito a mutazioni, nel 70% dei casi di melanoma. Un’inibizione dell’attività chinasica di Mek 1/2 e una conseguente riduzione nell’attivazione di Erk 1/2 è stata osservata in entrambe le linee usate dopo incubazione delle cellule con la 5,7-dimetossicumarina. Inoltre, è stato investigato il processo di melanogenesi indotto dal trattamento andando a monitorare il grado di fosforilazione di CREB (proteina legante CRE), l’espressione di Mitf (fattore di trascrizione associato alla microftalmia) e l’attività della tirosinasi. Mitf è coinvolto nel differenziamento, nella proliferazione e nella sopravvivenza dei melanociti e delle cellule di melanoma, poiché regola la trascrizione di geni codificanti per proteine che controllano la progressione del ciclo cellulare o la melanogenesi, come la tirosinasi che è considerato l’enzima chiave nella sintesi di melanina. L’attività di Mitf è regolata da vari fattori, tra cui CREB che quando è fosforilato induce la trascrizione del gene corrispettivo. Noi abbiamo osservato che la fosforilazione di CREB e l’espressione di Mitf aumentano significativamente in seguito a trattamento con la 5,7-dimetossicumarina nelle cellule di melanoma; di conseguenza, un incremento nell’attività della tirosinasi è stato dimostrato.
Questi risulatati suggeriscono che la 5,7-dimetosicumarina esercita un’attività antineoplastica nelle cellule di melanoma murino e umano, causando un processo di differenziamento con induzione di melanogenesi. Sarà interessante effettuare studi in vitro, per investigare un’eventuale attività antimetastatica del composto in esame, ed in vivo al fine di dimostrare le proprietà antineoplastiche e antimetastatiche della molecola anche in ambito preclinico.In this work the antiproliferative activity of 5,7-dimethoxycoumarin, a plant secondary metabolite, on murine (B16) and human (A375) melanoma cells was analysed. The compound reduced significatively cell proliferation in time and dose-dependent manner blocking cell cycle in G0/G1 phase, in both cell lines. Also a differentiation process, following this block, was detected by monitoring some specific markers such as morphological changes with development of dendrite-like projections from cell surface, melanin synthesis and protoporphyrine IX (PpIX) accumulation. Taking into account the antiproliferative and differentiation activities of this compound, we examined Ras/Raf/Mek/Erk mitogen-activated protein kinase (MAPK) activity following treatment; this kinases pathway regulates cell survival, proliferation and differentiation processes and it is costitutively activated following mutations in 70 % of melanoma. Inhibition of Mek 1/2 kinase activity and subsequent reduction in Erk 1/2 activation were detected in both cell types. Furthermore, we analyzed melanogenesis process by monitoring the phosphorylation of cAMP-response element-binding protein (p-CREB) and the expression of microphthalmia-associated transcription factor (Mitf); also tyrosynase activity was investigated. Mitf is involved in differentiation, proliferation and survival of melanocyte and melanoma cells since it regulates transcription of genes encoding for proteins involved in cell cycle progression or in melanogenesis, such as the enzyme tyrosinase, the key enzyme in melanin synthesis. Transcriptional activity of Mitf is regulated also through the phosphorylation of CREB that upregulates its gene transcription. We detected that CREB phosphorylation and Mitf expression enhanced significantly following 5,7-dimethoxycoumarin treatment in melanoma cells; consequently, an increase of tyrosinase activity was observed. These findings suggest that 5,7-dimethoxycoumarin exerts an antineoplastic activity in murine and human melanoma cells, triggering a differentiation process with stimulation of melanin synthesis. It will be of note to carry out in vitro analyses to investigate the antimetastatic activity of the tested compound; and in vivo studies with the aim to demonstrate cancer antigrowth and antimetastatic properties of compound also in preclinical trials
Gas chromatography-mass spectrometry analysis of phenolic compounds from Carica papaya L. leaf
No full textPhenolic compounds are important components in vegetable foods, infusions and teas for their beneficial effects on human health. The presence of such compounds, evidenced for the first time in Carica papaya leaves, could partially explain the pharmacological properties of this plant and demonstrates its importance in alimentation and daily intake. C. papaya leaves were extracted with methanol in a Soxhlet apparatus and later with a liquid-liquid extraction (LLE) with the aim of identifying and quantifying secondary metabolites from this plant, using gas chromatography-mass spectrometry (GC-MS) in the selected ion-monitoring (SIM) mode. Derivatization procedure of the extract was necessary to analyze the polar compounds in GC-MS. 5,7-Dimethoxycoumarin and polar molecules such as protocatechuic acid, p-coumaric acid, caffeic acid, chlorogenic acid, kaempferol and quercetin were detected and identified in qualitative analysis. The quantitative analysis has shown the presence of phenolic acids as the main compound, while chlorogenic acid was found in trace amounts, compared to the flavonoids and coumarin compounds. The quantities detected were 0.25 mg/g (dry leaf) for caffeic acid, 0.33 mg/g for p-coumaric acid and 0.11 mg/g for protocatechuic acid. Kaetnpferol and quercetin were 0.03 and 0.04 mg/g, respectively, while that for 5,7-dimethoxycoumarin was 0.14 mg/g. (c) 2007 Elsevier Inc. All rights reserved
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Inhibition of Mek 1/2 kinase activity and stimulation of melanogenesis by 5,7-dimethoxycoumarin treatment of melanoma cells
No full textIn this study, the processes of differentiation and melanogenesis induced by 5.7-dimethoxycoumarin in murine (B16) and human (A375) melanoma cells were investigated. Taking into account the previously demonstrated anti-proliferative and differentiation activities of this compound, we examined Ras/Raf/Mek/Erk mitogen-activated protein kinase activity following treatment; inhibition of Mek 1/2 kinase activity and Subsequent reduction in Erk 1/2 activation were detected in both cell types. We observed melanogenesis induction associated to an increase in cAMP-response element-binding protein (CREB) and microphthalmia-associated transcription factor (Mitf) expression, both involved in its regulation. Mitf is fundamental for development, survival and differentiation of melanocyte and melanoma, since it regulates transcription of genes encoding for proteins involved in cell cycle progression or in melanogenesis, such as the enzyme tyrosinase. A significant increase of tyrosinase activity was revealed following treatment in B16 but not in A375 cells. although a strong synthesis of melanin was induced by 5,7-dimethoxycoumarin in both cell lines. The treatment induced protoporphyrine IX accumulation involved in melanogenesis since it promotes stability of cAMP. Finally, the Mek 1/2 inhibitor U0126 significantly potentiated growth inhibition of B16 cells triggered by 5,7-dimethoxycoumarin, suggesting that down-regulation of Raf/Mek/Erk pathway sensitizes melanoma cells to 5,7-dimethoxycoumarin treatment
Characterization of Phaseolus vulgaris L. landraces cultivated in Central Italy
No full textEight Phaseolus vulgaris L. landraces cultivated on farm in marginal areas of Central Italy (Lazio region) were investigated in order to evaluate chemical composition of storage proteins and secondary metabolites fractions. The total protein content showed some differences among landraces; the maximum value was next to 30 g for 100 g of dry weight. The seed storage proteins were screened by polyacrylamide gel electrophoresis (SDS/PAGE): seven landraces exhibited phaseolin patterns type S, one landrace showed a phaseolin pattern type T. A morphological analysis of cotyledon parenchyma performed by scanning electron microscopy (SEM) revealed differences in size of starch granules. Moreover the polyphenolic composition was investigated using HPLC-APCI; from the methanol extracts a flavonoid, kaempferol, and a coumarin, 5,7-dimethoxycoumarin, were identified. To our knowledge, this is the first tithe that 5,7-dimethoxycoumarin has been reported in P. vulgaris seeds
Variations on the Author
Get PDF“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Get PDFWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Get PDFWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
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