Genotoxic evaluation of the antimalarial drugs artemisinin and artesunate in human HepG2 cells and effects on CASP3 and SOD1 gene expressions

The malaria treatment recommended by the World Health Organization involves medicines derived from artemisinin, an active compound extracted from the plant Artemisia annua, and some of its derivatives, such as artesunate. Considering the lack of data regarding the genotoxic effects of these compounds in human cells, the objective of this study was to evaluate the cytotoxicity and genotoxicity, and expressions of the CASP3 and SOD1 genes in a cultured human hepatocellular liver carcinoma cell line (HepG2 cells) treated with artemisinin and artesunate. We tested concentrations of 2.5, 5, 7.5, 10, and 20 ��g/mL of both substances with a resazurin cytotoxicity assay, and the concentrations used in the genotoxicity experiments (2.5, 5, and 10 ��g/mL) and gene expression analysis (5 mg/mL) were determined. The results of the comet assay in cells treated with artemisinin and artesunate showed a significant dosedependent increase (P < 0.001) in the number of cells with DNA damage at all concentrations tested. However, the gene expression analysis revealed no significant change in expression of CASP3 or SOD1. Our data showed that although artemisinin and artesunate exhibited genotoxic effects in cultured HepG2 cells, they did not significantly alter expression of the CASP3 and SOD1 genes at the doses tested. ��FUNPEC-RP.

Relevance of the Myeloid Differentiation Factor 88 (MyD88) on RANKL, OPG, and Nod Expressions Induced by TLR and IL-1R Signaling in Bone Marrow Stromal Cells

The myeloid differentiation factor 88 (MyD88) plays a pivotal role in Toll-like receptor (TLR)- and interleukin-1 receptor (IL-1R)-induced osteoclastogenesis. We examined the role of MyD88 on p38 mitogen-activated protein kinase (MAPK) and nuclear factor kappa-light-chain-enhancer of activated B cell (NF-��B) activation and nucleotide-binding oligomerization domain (Nod) induction by lipopolysaccharide (LPS) and IL-1 beta, and their effect on receptor activator of NF-��B ligand (RANKL) and osteoprotegerin (OPG) production in bone marrow stromal cell (BMSC). RANKL, Nod1, Nod2, NF-��B, and p38 protein levels were determined by Western blot. Nod2 was stimulated with muramyl dipeptide (MDP) prior to TLR4 stimulation with LPS. MyD88 deficiency markedly inhibited RANKL expression after LPS stimulation and increased OPG messenger RNA (mRNA) production. Also, MyD88 was necessary for NF-��B and p38 MAPK activation. MDP alone did not induce RANKL and OPG expressions; however, when combined with LPS, their expressions were significantly increased (p���<���0.05). Our results support that MyD88 signaling has a pivotal role in osteoclastogenesis thought NF-��B and p38 activation. Nod2 and especially Nod1 levels were influenced by MyD88.

Relevance of the Myeloid Differentiation Factor 88 (MyD88) on RANKL, OPG, and Nod Expressions Induced by TLR and IL-1R Signaling in Bone Marrow Stromal Cells

Coordena����o de Aperfei��oamento de Pessoal de N��vel Superior (CAPES)

Improvement of the physical performance is associated with activation of NO/PGC-l alpha/mtTFA signaling pathway and increased protein expressions of electron transport chain in gastrocnemius muscle from rats supplemented with L-arginine

Funda����o de Amparo �� Pesquisa do Estado de S��o Paulo (FAPESP)

Current Expressions of American Jewish Identity: An Analysis of 114 Teenagers

This chapter explores the characteristics of 114 American teenagers' Jewish identities using data from the National Study of Youth and Religion (NSYR). The NSYR includes a telephone survey of a nationally representative sample of 3,290 adolescents aged 13 to 17. Jewish teenagers were over-sampled, resulting in a total of 3,370 teenage participants. Of the NSYR teens surveyed, 141 have at least one Jewish parent and 114 of them identify as Jewish. The NSYR also includes in-depth face-to-face interviews with a total of 267 U.S. teens: 23 who have at least one Jewish parent and 18 who identify as Jewish. The following analysis draws upon quantitative data from the 114 teens who identified themselves as Jewish in the face-to-face interviews.

Pattern of humoral immune response to Plasmodium falciparum blood stages in individuals presenting different clinical expressions of malaria

Abstract Background The development of protective immunity against malaria is slow and to be maintained, it requires exposure to multiple antigenic variants of malaria parasites and age-associated maturation of the immune system. Evidence that the protective immunity is associated with different classes and subclasses of antibodies reveals the importance of considering the quality of the response. In this study, we have evaluated the humoral immune response against Plasmodium falciparum blood stages of individuals naturally exposed to malaria who live in endemic areas of Brazil in order to assess the prevalence of different specific isotypes and their association with different malaria clinical expressions. Methods Different isotypes against P. falciparum blood stages, IgG, IgG1, IgG2, IgG3, IgG4, IgM, IgE and IgA, were determined by ELISA. The results were based on the analysis of different clinical expressions of malaria (complicated, uncomplicated and asymptomatic) and factors related to prior malaria exposure such as age and the number of previous clinical malaria attacks. The occurrence of the H131 polymorphism of the Fc��IIA receptor was also investigated in part of the studied population. Results The highest levels of IgG, IgG1, IgG2 and IgG3 antibodies were observed in individuals with asymptomatic and uncomplicated malaria, while highest levels of IgG4, IgE and IgM antibodies were predominant among individuals with complicated malaria. Individuals reporting more than five previous clinical malaria attacks presented a predominance of IgG1, IgG2 and IgG3 antibodies, while IgM, IgA and IgE antibodies predominated among individuals reporting five or less previous clinical malaria attacks. Among individuals with uncomplicated and asymptomatic malaria, there was a predominance of high-avidity IgG, IgG1, IgG2 antibodies and low-avidity IgG3 antibodies. The H131 polymorphism was found in 44.4% of the individuals, and the highest IgG2 levels were observed among asymptomatic individuals with this allele, suggesting the protective role of IgG2 in this population. Conclusion Together, the results suggest a differential regulation in the anti-P. falciparum antibody pattern in different clinical expressions of malaria and showed that even in unstable transmission areas, protective immunity against malaria can be observed, when the appropriated antibodies are produced.

Expressions- und Funktionsanalysen von "Tetraspan Vesicle Membrane Proteins" in Caenorhabditis elegans

Tetraspan vesicle membrane proteins (TVPs) sind ubiquit��re Komponenten von Transportvesikeln. Bei den S��ugetieren unterscheidet man drei Familien, die Physine, Gyrine und SCAMPs (secretory carrier-associated membrane proteins). Ihre Funktion ist weitgehend unbekannt, es wird jedoch vermutet, dass sie eine Rolle bei der Vesikelbildung und der Vesikelrezirkulierung spielen. In Caenorhabditis elegans existiert von jeder Familie jeweils nur ein einziges Polypeptid: f��r die Physine Synaptophysin (SPH-1), f��r die Gyrine Synaptogyrin (SNG-1) und f��r die SCAMPs SCAMP (SCM-1). Ziel der Arbeit war es die Verteilung der C. elegans TVPs zu untersuchen und ihre Funktion unter besonderer Ber��cksichtigung der vesikelvermittelten synaptischen Kopplung zu bestimmen. Wenn die C. elegans TVPs in humanen Epithelzellen synthetisiert werden, lokalisieren sie in zytoplasmatischen Vesikeln. In Kotransfektionsexperimenten wurde gezeigt, dass sie gr����tenteils in den gleichen Strukturen enthalten sind. In C. elegans synthetisierte TVP-Reporterkonstrukte k��nnen in unterschiedlichen Geweben nachgewiesen werden. Dabei ist SNG-1 fast ausschlie��lich in Neuronen zu finden. SPH-1 und SCM-1 hingegen weisen komplexe und teilweise ��berlappende Verteilungsmuster auf. W��hrend f��r SPH-1 eine starke Fluoreszenz im Pharynx, auf der apikalen Seite der Darmzellen oberhalb des sog. terminal webs und in adluminalen Regionen von exkretorischen Geweben gefunden wurde, war SCM-1 stark in der Muskulatur und den Coelomozyten vertreten. Die Expression von SCM-1 in Pharynx und Darm war deutlich schw��cher. Die C. elegans TVPs werden fr��h in der Entwicklung ab der Gastrulation (SPH-1 und SCM-1) bzw. ab der Neurulation im sog. Komma-Stadium (SNG-1) produziert. Um die Funktion der TVPs in C. elegans zu untersuchen, wurden TVP-Mutanten analysiert. Durch Kombination aller drei TVP-Gen-Mutanten wurden TVP-Dreifachmutanten generiert. Diese wiesen keinen offensichtlichen Defekt im Bewegungsmuster auf, entwickelten sich normal und bildeten ein normales Nervensystem aus. Auch auf unterschiedliche chemische und physikalische Reize in sensorischen Tests reagierten die TVP-Dreifachmutanten in gleicher Weise wie Wildtyptiere. Ebenso zeigen die TVP-Dreifachmutanten elektrophysiologisch unter normalen Bedingungen keine anormalen Reaktionsmuster. In ultrastrukturellen Untersuchungen wurde lediglich eine signifikant erh��hte Anzahl Clathrin-ummantelter Vesikel in cholinergen Synapsen gefunden. Erst unter Stressbedingungen, hervorgerufen durch den GABA-Antagonisten Pentylentetrazol (PTZ), wiesen sowohl die TVP-Dreifach- als auch die TVP-Einzelmutanten eine deutlich erh��hte Krampfbereitschaft auf. Zusammengenommen zeigen die Analysen, dass TVPs zwar f��r grundlegende neuronale Prozesse nicht notwendig sind, dass sie aber auf der anderen Seite vermutlich an alternativen redundanten Wegen der Neurotransmitterfreisetzung beteiligt sind.

Expressions- und Funktionsanalysen von Neuroglobin und Cytoglobin

Im Rahmen der vorliegenden Dissertation wurden Untersuchungen zur Expression und Funktion der respiratorischen Proteine Neuroglobin (Ngb) und Cytoglobin (Cygb) in Vertebraten durchgef��hrt. Beide Globine wurden erst k��rzlich entdeckt, und ihre Funktionen konnten trotz vorliegender Daten zur Struktur und biochemischen Eigenschaften dieser Proteine bisher nicht eindeutig gekl��rt werden. Im ersten Abschnitt der vorliegenden Arbeit wurde die zellul��re und subzellul��re Lokalisation von Neuroglobin und Cytoglobin in murinen Gewebeschnitten untersucht. Die Expression von Ngb in neuronalen und endokrinen Geweben h��ngt offensichtlich mit den hohen metabolischen Aktivit��ten dieser Organe zusammen. Insbesondere im Gehirn konnten regionale Unterschiede in der Ngb-Expression beobachtet werden. Dabei korrelierte eine besonders starke Neuroglobin-Expression mit Gehirnbereichen, die bekannterma��en die h��chsten Grundaktivit��ten aufweisen. In Anbetracht dessen liegt die Funktion des Neuroglobins m��glicherweise im basalen O2-Metabolismus dieser Gewebe, wobei Ngb als O2-Lieferant und kurzfristiger O2-Speicher den vergleichsweise hohen Sauerstoffbedarf vor Ort sicherstellen k��nnte. Weitere Funktionen in der Entgiftung von ROS bzw. RNS oder die k��rzlich publizierte m��gliche Rolle des Ngb bei der Verhinderung der Mitochondrien-vermittelten Apoptose durch eine Reduktion des freigesetzten Cytochrom c w��ren dar��ber hinaus denkbar. Die Cygb-Expression im Gehirn beschr��nkte sich auf relativ wenige Neurone in verschiedenen Gehirnbereichen und zeigte dort vorwiegend eine Co-Lokalisation mit der neuronalen NO-Synthase. Dieser Befund legt eine Funktion des Cytoglobins im NO-Metabolismus nahe. Quantitative RT-PCR-Experimente zur mRNA-Expression von Ngb und Cygb in alternden S��ugern am Bsp. der Hamsterspezies Phodopus sungorus zeigten keine signifikanten ��nderungen der mRNA-Mengen beider Globine in alten im Vergleich zu jungen Tieren. Dies widerspricht publizierten Daten, in denen bei der Maus anhand von Western Blot-Analysen eine Abnahme der Neuroglobin-Menge im Alter gezeigt wurde. M��glicherweise handelt es sich hierbei um speziesspezifische Differenzen. Die im Rahmen dieser Arbeit durchgef��hrte vergleichende Sequenzanalyse der humanen und murinen NGB/Ngb-Genregion liefert zum einen Hinweise auf die m��gliche Regulation der Ngb-Expression und zum anderen eine wichtige Grundlage f��r die funktionellen Analysen dieses Gens. Es konnte ein minimaler Promotorbereich definiert werden, der zusammen mit einigen konservierten regulatorischen Elementen als Basis f��r experimentelle Untersuchungen der Promotoraktivit��t in Abh��ngigkeit von ��u��eren Einfl��ssen dienen wird. Bioinformatische Analysen f��hrten zur Identifizierung des sog. ���neuron restrictive silencer element��� (NRSE) im Ngb-Promotor, welches vermutlich f��r die vorwiegend neuronale Expression des Proteins verantwortlich ist. Die kontrovers diskutierte O2-abh��ngige Regulation der Ngb-Expression konnte hingegen anhand der durchgef��hrten komparativen Sequenzanalysen nicht best��tigt werden. Es wurden keine zwischen Mensch und Maus konservierten Bindestellen f��r den Transkriptionsfaktor HIF-1 identifiziert, der die Expression zahlreicher hypoxieregulierter Gene, z.B. Epo und VEGF, vermittelt. Zusammen mit den in vivo-Daten spricht dies eher gegen eine Regulation der Ngb-Expression bei verminderter Verf��gbarkeit von Sauerstoff. Die Komplexit��t der Funktionen von Ngb und Cygb im O2-Stoffwechsel der Vertebraten macht den Einsatz muriner Modellsysteme unerl��sslich, die eine sukzessive Aufkl��rung der Funktionen beider Proteine erlauben. Die vorliegende Arbeit liefert auch dazu einen wichtigen Beitrag. Die hergestellten ���gene-targeting���-Vektorkonstrukte liefern in Verbindung mit den etablierten Nachweisverfahren zur Genotypisierung von embryonalen Stammzellen die Grundlage zur erfolgreichen Generierung von Ngb-knock out sowie Ngb- und Cygb-��berexprimierenden transgenen Tieren. Diese werden f��r die endg��ltige Entschl��sselung funktionell relevanter Fragestellungen von enormer Bedeutung sein.

The Long Tail of the Rainbow Serpent: New Technologies and the Protection and Promotion of Traditional Cultural Expressions

New technologies, in particular those stemming from digitisation, allow amongst other things the production of perfect copies, instantaneous and ubiquitous distribution of and easy access to information with no real location restrictions. The effects of these technological advances have largely been perceived as negative for the protection of traditional cultural expressions (TCE), both because of the peculiarities of the digital networked environment and because of the lack of appropriate intellectual property protection models for TCE. The purpose of this article is, while accounting for the diversity and complexity of issues related to TCE, to reveal a more positive side of digital technologies. It shows the potential of these to be proactively applied and the further reaching possibilities for designing an efficient multi-level and multi-faceted toolbox for the protection and promotion of TCE in the digital ecology.

Intellectual Property and Traditional Cultural Expressions in a Digital Environment

In the face of increasing globalisation, and a collision between global communication systems and local traditions, this book offers innovative trans-disciplinary analyses of the value of traditional cultural expressions (TCE) and suggests appropriate protection mechanisms for them. It combines approaches from history, philosophy, anthropology, sociology and law, and charts previously untravelled paths for developing new policy tools and legal designs that go beyond conventional copyright models. Its authors extend their reflections to a consideration of the specific features of the digital environment, which, despite enhancing the risks of misappropriation of traditional knowledge and creativity, may equally offer new opportunities for revitalising indigenous peoples' values and provide for the sustainability of TCE.This book will appeal to scholars interested in multidisciplinary analyses of the fragmentation of international law in the field of intellectual property and traditional cultural expressions. It will also be valuable reading for those working on broader governance and human rights issues.

The Care and Shelter of Children and Adolescents in Brazil: Expressions of Social Issues

The future of Brazilian children who have the protection offered by familial bonds is threatened by social inequities that force them to seek shelter and grow up in shelters. According to the Institute of Applied Economic Research, an estimated 20,000 children and adolescents are served by institutions. The majority of these children are afro-descendent males between the ages of seven and fifteen years old. Of those researched, 87.6% have families (58.2% receive visits from their families, 22.7% are rarely visited by their families and 5.8 are legally prohibited from contacting or being by their families). The percentage of children and adolescents ���without families��� or with ���missing families��� is 11.3%. There is no information available for 2% of the children and adolescents residing in shelters. The principle factors that necessitate the placement of Brazilian children in institutions that provide care and shelter include poverty (including children forced to work, sell drugs or beg, for example); domestic violence; chemical dependence of parents or guardians; homelessness; death or parents or guardian; imprisonment of their parents; and sexual abuse committed by their parents or guardians. The issue of abandoned children and adolescents and their care and shelter in the Brazilian context expresses a perverse violation of Child and Adolescent Rights.

Sui Generis Rights for the Protection of Traditional Cultural Expressions Policy Implications

Within the international community there have been many calls for better protection of traditional cultural expressions (TCEs), for which classic instruments of intellectual property rights do not seem to fit. In response, at least five model laws have been advanced within the last 40 years. These are referred to as sui generis because, though they generally belong to the realm of intellectual property they structurally depart from classic copyright law to accommodate the needs of the holders of TCEs. The purpose of this paper is to provide a well-founded basis for national policy makers who wish to implement protection for TCEs within their country. This is achieved by systematically comparing and evaluating economic effects that can be expected to result from these regulatory alternatives and a related system or private ordering. Specifically, we compare if and how protection preferences of local communities are met as well as the social costs that are likely to arise from the different model laws.