Cardiotonic steroids are used to treat heart failure and arrhythmia and have promising anticancer effects. The prototypic cardiotonic steroid ouabain may also be a hormone that modulates epithelial cell adhesion. Cardiotonic steroids consist of a steroid nucleus and a lactone ring, and their biological effects depend on the binding to their receptor, Na,K-ATPase, through which, they inhibit Na+ and K+ ion transport and activate of several intracellular signaling pathways. In this study, we added a styrene group to the lactone ring of the cardiotonic steroid digoxin, to obtain 21-benzylidene digoxin (21-BD), and investigated the effects of this synthetic cardiotonic steroid in different cell models. Molecular modeling indicates that 21-BD binds to its target Na,K-ATPase with low affinity, adopting a different pharmacophoric conformation when bound to its receptor than digoxin. Accordingly, 21-DB, at relatively high µM amounts inhibits the activity of Na,K-ATPase α1, but not α2 and α3 isoforms. In addition, 21-BD targets other proteins outside the Na,K-ATPase, inhibiting the multidrug exporter Pdr5p. When used on whole cells at low µM concentrations, 21-BD produces several effects, including: 1) up-regulation of Na,K-ATPase expression and activity in HeLa and RKO cancer cells, which is not found for digoxin, 2) cell specific changes in cell viability, reducing it in HeLa and RKO cancer cells, but increasing it in normal epithelial MDCK cells, which is different from the response to digoxin, and 3) changes in cell-cell interaction, altering the molecular composition of tight junctions and elevating transepithelial electrical resistance of MDCK monolayers, an effect previously found for ouabain. These results indicate that modification of the lactone ring of digoxin provides new properties to the compound, and shows that the structural change introduced could be used for the design of cardiotonic steroid with novel functions.

The sensitivity of the larval stages of Schistosoma mansoni to chemotherapy with praziquantel and oxamniquine was tested in mice during primary and secondary infections and after different intervals from cercarial exposure. Worm recovery by perfusion of the porto-mesenteric system, followed by counting and a morphometric study of the parasite, allowed the conclusion that the relative resistance of the larval stages of S. mansoni to schistosomicide drugs, demonstrated in primary infections, also persists when the host is already infected. This indicates that a therapeutic failure may result when an infected host is treated some time after being re-infected, because of the presence of migrating, drug-resistant, immature forms of the parasite. Key-words: Schistosoma mansoni. Larval stages. Praziquantel. Oxamniquine.Resumo A susceptibilidade dos estágios larvais do Schistosoma mansoni aos esquistossomicidas praziquantel e oxamniquine foi testada em camundongos durante infecção primária ou secundária, e após diferentes intervalos de tempo após a exposição cercariana. A avaliação foi feita pela contagem dos vermes após recuperação destes por perfusão do sistema porto-mesentérico e pelo estudo morfométrico dos mesmos. O estudo revelou que a relativa resistência das formas larvais aos esquistossomicidas, já demonstrada em infecção primária, persiste no caso de hospedeiros já infectados. Este fato indica que uma falha terapêutica pode resultar quando o tratamento é feito em hospedeiros re-infectados recentemente, em virtude dos mesmos apresentarem formas migrantes e imaturas do parasita, as quais são particularmente resistentes aos esquistossomicidas. Palavras-chaves: Schistosoma mansoni. Estágio Larval. Praziquantel. Oxamniquine.

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Key words: modulation -granuloma -shistosomiasis -extracellular matrix Hepatic granulomas formed around mature eggs of Schistosoma mansoni undergo changes in size, appearance and cellular constitution with time. This has been interpreted as a mechanism of host protection, with the host reactions becoming more economical and efficient during chronic infection (Andrade & Warren 1964). The immunological counterpart has been thoroughly investigated, both in vivo (Colley 1981, Chensue et al. 1993, Bogen et al. 1995 and in vitro (Doughty & Phillips 1982, Parra et al. 1991, including its cytokine patterns , McKerrow 1997. However, there are data indicating that granulomas outside the liver may behave differently. Granulomas in the intestine were observed to be already modulated in the ileum from the very beginning, although modulation was said to occur in the colon and ileal Peyer's patch (Weinstock & Boros 1981, 1983 the same morphology regardless the time of infection (Vidal et al. 1992, Eltoum et al. 1995. Such findings suggest that the so-called "immunological modulation" of periovular granulomas in schistosomiasis is mainly influenced by local factors. If this is so, the current interpretation of the immunological modulation concept needs to be revised.To further investigate this possibility, a comparative study of size, appearance and matrix composition of granulomas formed in three different organs of mice (liver, intestine and lung), during three different periods of infection (early, intermediate and late), was undertaken. Methods of histopathology, immunofluorescence, biochemistry and morphometry were applied in an attempt to understand the real significance of the so-called "immunological modulation" of schistosomal granuloma. MATERIALS AND METHODSThrity-one albino Swiss mice of both sexes, weighing 18-23 g, maintained with free access to a commercial balanced diet and water, were used. Animals were submitted to transcutaneous infection with 50 recently eliminated S. mansoni cercariae each. Later, animals were randomly divided into three groups, according to time of infection: first group (acute phase), animals sacrificed eight

Antimicrobial peptides (AMPs) are currently being investigated as potential sources of novel therapeutics against an increasing number of microorganisms resistant to conventional antibiotics. The conjugation of an AMP to other bioactive compounds is an interesting approach for the development of new derivatives with increased antimicrobial efficiency and broader spectra of action. In this work, the synthesis of a new peptide−coumarin conjugate via copper(I)-catalyzed azide−alkyne cycloaddition is described. The conjugate was assayed for in vitro cytotoxicity and displayed antifungal activity against Cryptococcus gattii and Cryptococcus neoformans. Additionally, the conjugate exhibited increased antifungal efficacy when compared with the individual peptide, coumarin, or triazole moieties. Treatment of C. gattii with the peptide−coumarin conjugate enhanced the production of reactive oxygen species, suggesting that the oxidative burst plays an important role in the mechanism of action of the conjugate. KEYWORDS: Antimicrobial peptide, copper(I)-catalyzed azide−alkyne cycloaddition, coumarin, cryptococcosis, reactive oxygen species A ntimicrobial peptides (AMPs) are small molecular weight proteins with antimicrobial activity against bacteria, viruses, and fungi. 1 AMPs are currently being investigated as potential sources of novel therapeutic antimicrobial agents because of their broad-spectrum activity and low susceptibility for developing resistance. 2 Ubiquicidin (UBI) is an antimicrobial peptide, which was first isolated from murine macrophages and then found at low concentrations (as a first line of defense) inside human airway epithelial cells, activated macrophages, and in human colon mucosa. UBI is a cationic peptide consisting of 59 amino acid residues. It is not expected to be immunogenic to humans since it is of human origin. 3 UBI 1−59 and some of its fragments have been shown to be microbicidal against a broad spectrum of pathogens. Of particular interest is the fragment UBI 31−38 (Figure 1), which is relatively small, easily synthesized, and exhibits antibacterial activity toward methicillin resistant Staphylococcus aureus. 4 Peptides open up new perspectives in drug design by providing an entire range of highly selective and nontoxic pharmaceuticals. With growing applications of their synthesis and bioactivity, considerable attention has been focused on the research of peptide-based derivatives. 5 Coumarins form an important class of benzopyrones, which are found in nature. Many coumarins and their derivatives possess antimicrobial, anti-inflammatory, and anticancer properties. 6 Research on coumarin compounds as antifungal agents has shown that structural modification of the coumarin skeleton (i.e., the benzene ring, lactone ring, or both) results in derivatives, which possess potent antifungal activity when compared to clinically used antifungal drugs. 7 Therefore, numerous efforts have focused on the development of coumarins as potential drugs. 8 The copper(I)-catalyzed azide−alkyne cyclo...