Histopathological Changes in the Placentas and Fetuses of Mice Infected with Trypanosoma cruzi Isolated from the Myotis nigricans nigricans Bat

Histopathological changes and placental transmission were studied in the late stages of pregnancy in mice infected with a strain of Trypanosoma cruzi, isolated from a Myolis nigricans nigricans bat. Large amastigote nests were observed in uterine muscles, as well as in decidual and endothelial placental cells. In addition, persistent coagulative and fibrotic Vascular degeneration was observed. Large amastigote burdens were found in giant cells, spongioblasts and endothelial cells within the labyrinthine layer. Transplacental transmission was confirmed in 30% of the fetuses examined, in which amastrigote nests were seen only in striated muscle. During tire acute phase, intrauterine development was impaired as the result of parasitic invasion of the placenta, and fetal mortality rose to 10%. (C) 2008 Elsevier Ltd. All rights reserved.

Molecular characterization of BjussuSP-I, a new thrombin-like enzyme with procoagulant and kallikrein-like activity isolated from Bothrops jararacussu snake venom

A thrombin-like enzyme, named BjussuSP-I, isolated from Bothrops jararacussu snake venom, is an acidic single-chain glycoprotein with M-r = 61,000, pI similar to 3.8 and 6% sugar. BjussuSP-I shows high proteolytic activity upon synthetic substrates, such as S-2238 and S-2288. It also shows procoagulant and kallikrein-like activity, but is unable to act on platelets and plasmin. These activities are inhibited by specific inhibitors of this class of enzymes. The complete cDNA sequence of BjussuSP-I with 696 bp encodes open reading frames of 232 amino acid residues, which conserve the common domains of thrombin-like serine proteases. BjussuSP-I shows a high structural homology with other thrombin-like enzymes from snake venoms where common amino acid residues are identified as those corresponding to the catalytic site and subsites S1, S2 and S3 already reported. In this study, we also demonstrated the importance of N-linked glycans, to improve thrombin-like activity of BjussuSP-I toxin. (c) 2007 Elsevier Masson SAS. All rights reserved.

Evaluation of the genotoxicity of Crotalus durissus terrificus snake venom and its isolated toxins on human lymphocytes

In the present study, experiments were carried out to evaluate the mutagenic potential and genotoxic effects of Crotalus durissus terrificus snake venom and its isolated toxins on human lymphocytes, using the micronucleus and comet assays. Significant damage to DNA was observed for crotoxin and crotapotin (CA). Basic phospholipase A(2) (CB) and crotamine did not present any mutagenic potential when evaluated by the micronucleus test. C. d. terrificus crude venom was able to induce the formation of micronuclei, similarly to the mutagenic drug used as a positive control. In the comet assay, all the toxins tested (crotamine, crotoxin, CB and CA) and C. d. terrificus venom presented genotoxic activity. Studies on the cytogenetic toxicology of animal venoms and their isolated proteins are still very scarce in the literature, which emphasizes the importance of the present work for the identification and characterization of potential therapeutic agents, as well as for the better understanding of the mechanisms of action of toxins on the human body. (C) 2011 Elsevier B.V. All rights reserved.

Biochemical and functional properties of a thrombin-like enzyme isolated from Bothrops pauloensis snake venom

In the present study, a thrombin-like enzyme named BpSP-I was isolated from Bothrops pauloensis snake venom and its biochemical, enzymatic and pharmacological characteristics were determined. BpSP-I is a glycoprotein that contains both N-linked carbohydrates and sialic acid in its structure, with M(r) = 34,000 under reducing conditions and pI similar to 6.4. The N-terminal sequence of the enzyme (VIGGDECDINEHPFL) showed high similarity with other thrombin-like enzymes from snake venoms. BpSP-I showed high clotting activity upon bovine and human plasma and was inhibited by PMSF, benzamidine and leupeptin. Moreover, this enzyme showed stability when examined at different temperatures (-70 to 37 degrees C), pH values (3-9) or in the presence of divalent metal ions (Ca(2+), Mg(2+), Zn(2+) and Mn(2+)). BpSP-I showed high catalytic activity upon substrates, such as fibrinogen, TAME, S-2238 and S-2288. It also showed kallikrein-like activity, but was unable to act upon factor Xa and plasmin substrates. Indeed, the enzyme did not induce hemorrhage, myotoxicity or edema. Taken together, our data showed that BpSP-I is in fact a thrombin-like enzyme isoform isolated from Bothrops pauloensis snake venom. (C) 2009 Elsevier Ltd. All rights reserved.

Effect of a Pool of Peptides Isolated from Crotalus durissus terrificus (South American Rattlesnake) Venom on Glucose Levels of Mice Fed on a High-Fat Diet

This study investigated the effect of a pool of peptides, isolated from venom of Crotalus durissus terrificus (South American rattlesnake) on glucose concentration in C57BL/6 mice fed on a high-fat diet for 6 weeks. The pool of peptides (molecular mass around of 10 kDa) was obtained using a MidJet apparatus with a cartridge of 10 KDa. The peptide pool was injected intraperitoneally in mice in a single dose (0.5 mg/animal) or multiple doses (0.2 mg/dose). After predetermined times (30, 60, 90 and 120 min) post injections, venous blood samples were collected for enzymatic measurement of serum glucose using a commercial glucose kit (glucose oxidase method). High-fat fed mice showed an increase in blood glucose concentration, in comparison with mice fed on the chow diet. Thirty minutes after a single dose of the peptide pool, high-fat fed animals showed a significant decrease (similar to 47%) in glycemia. However, the glucose level increased again at 60 and 120 min. Conversely, after multiple injections of the pool of peptides administered every 30 min, the blood glucose concentration in the high-fat mice was significantly decreased (similar to 37%) and remained at low levels until 120 min. These results suggest that the tested pool of peptides from Crotalus durissus terrificus contained a peptide (or peptides) with a beneficial role on glucose-lowering action of high-fat fed mice.

The antitumoral, trypanocidal and antileishmanial activities of extract and alkaloids isolated from Duguetia furfuracea

The alkaloid extract and five alkaloids isolated from subterranean stem bark of Duguetia furfuracea (Annonaceae) were investigated for the following activities: antitumoral, trypanocidal and leishmanicidal. Dicentrinone showed weak cytotoxicity, but it had the strongest leishmanicidal activity IC(50) 0.01 mu M). Duguetine and duguetine beta-N-oxide caused considerable antitumoral activity in every cell lines evaluated, although duguetine was more active against trypomastigote forms (IC(50) 9.32 mu M) than other alkaloids tested. (C) 2009 Elsevier GmbH. All rights reserved.

Clonal relationships determined by multilocus sequence typing among enteropathogenic Escherichia coli isolated in Brazil

Enteropathogenic Escherichia coli (EPEC) infections are a leading cause of infantile diarrhea in developing nations. Multilocus sequence typing (MLST) characterizes bacterial strains based on the sequences of internal fragments in housekeeping genes. Little is known about strains of EPEC analyzed by MLST from Brazil. In this study, a diverse collection of 29 EPEC strains isolated from patients with diarrhea, admitted to the University Hospital of Ribeirao Preto, was characterized by MLST. Strain analysis demonstrated 22 different sequence types (STs), of which almost half (48%) were new, indicating a high genotype diversity. The 22 STs were divided by eBURST into 12 clonal complexes. It was not possible to correlate typical and atypical EPEC with other strains in the MLST database. This is the first study that analyzed EPEC strains from South America that are included in the E. coli MLST database. Nine (31%) out of 29 strains are part of the CC10 clonal complex, the major clonal complex in the database, which comprises 174 strains and 86 different STs, suggesting that these strains might be the most important intestinal pathogenic E. coli worldwide. Genetic relationships between typical and atypical EPEC, enterohemorrhagic E. coli, and enteroaggregative E. coli strains were not established by MLST.

Structural and functional properties of Bp-LAAO, a new L-amino acid oxidase isolated from Bothrops pauloensis snake venom

An L-amino acid oxidase (Bp-LAAO) from Bothrops pauloensis snake venom was highly purified using sequential chromatography steps on CM-Sepharose, Phenyl-Sepharose CL4B, Benzamidine Sepharose and C18 reverse-phase HPLC. Purified Bp-LAAO showed to be a homodimeric acidic glycoprotein with molecular weight around 65 kDa under reducing conditions in SDS-PAGE. The best substrates for Bp-LAAO were L-Met, L-Leu, L-Phe and L-Ile and the enzyme showed a strong reduction of its catalytic activity upon L-Met and L-Phe substrates at extreme temperatures. Bp-LAAO showed leishmanicidal, antitumoral and bactericidal activities dose dependently. Bp-LAAO induced platelet aggregation in platelet-rich plasma and this activity was inhibited by catalase. Bp-LAAC-cDNA of 1548 bp codified a mature protein with 516 amino acid residues corresponding to a theoretical isoelectric point and molecular weight of 6.3 and 58 kDa, respectively. Additionally, structural and phylogenetic studies identified residues under positive selection and their probable location in Elp-LAAO and other snake venom LAAOs (svLAAOs). Structural and functional investigations of these enzymes can contribute to the advancement of toxinology and to the elaboration of novel therapeutic agents. (C) 2009 Elsevier Masson SAS. All rights reserved.

Antibacterial activity of 15-deoxygoyazensolide isolated from the stems of Minasia alpestris (Asteraceae) against oral pathogens

This work reports the isolation of the sesquiterpene lactone 15-deoxygoyazensolide from the stems of Minasia alpestris and the evaluation of its antimicrobial activity against the following oral pathogens: Enterococcus faecalis, Streptococcus salivarius, Streptococcus sobrinus, Streptococcus mutans, Streptococcus mitis, Streptococcus sanguinis, and Lactobacillus casei. Despite the cytotoxicity and genotoxicity of other sesquiterpene lactones of the furanoheliangolide-type, our results revealed that this compound exhibits low antibacterial activity against the evaluated oral pathogens; however, an interesting selectivity against E. faecalis (minimum inhibitory concentration [MIC] = 40 mu g mL(-1)) and S. sobrinus (MIC = 60 mu g mL(-1)) was observed.

Functional and structural characterization of isolated perfused stingray liver including effects of ischaemia/reperfusion

The morphological and functional characteristics of stingray liver were studied, including the effect of ischaemia/reperfusion. With an isolated perfused model, it was shown that the stingray liver was more resistant than the rat liver to ischaemia/reperfusion injury; this was consistent with the differing partial oxygen tensions usually present in the two species. This study confirmed that whereas stingray hepatocytes form tubules with central bile canaliculi as in other fish, the stingray liver has portal triads and a lobular architecture as in mammals. Apoptosis of hepatocytes, demonstrated in the normal liver, was only marginally enhanced by ischaemia/reperfusion. Resulting apoptotic bodies were phagocytized by macrophage-like cells in hepatocyte tubules. In contrast to rat liver, the stingray liver showed no necrosis after ischaemia-reperfusion. (C) 1998 W.B. Saunders Company Limited.

Structure of Caribbean ciguatoxin isolated from Caranx latus

Caribbean ciguatoxins (C-CTXs) are responsible for the widespread occurrence of ciguatera in the Caribbean Sea. The structure and configuration of C-CTX-1 (1), the major ciguatoxin isolated from the horse-eye jack (Caranx latus), has been determined from DQF-COSY, E-COSY, TOCSY, NOESY, POESY, ge-HSQC. and HMQC experiments performed at 750 MHz and 500 MHz on a 0.13 pmol sample. C-CTX-1 ([M + H](+) m/z 1141.6 Da, molecular formula C62H92O19) has a ciguatoxin/breveroxin ladder structure comprising 14 trans-fused, ether-linked rings (7/6/6/7/8/9/7/6/8/6/7/6/7/6) assembled fi um 6 protonated fragments. The relative stereochemistry and ring configuration of 1 was determined from an analysis of coupling constant and NOE data. Like ciguatoxins in the Pacific Ocean (P-CTX), C-CTX-1 possesses a flexible nine-membered ring which may be a conserved feature among ciguatoxins. However, C-CTX-1 has a longer contiguous carbon backbone (57 vs 55 carbons for P-CTX-1), one extra ring, and a hemiketal in ring N but no spiroketal as found in P-CTX. C-CTX-1 possesses a primary hydroxyl which may allow selective derivatization. A minor analogue, C-CTX-2, was also isolated from fish and assigned the structure 56 epi-C-CTX-1 (2). since it slowly rearranged to C-CTX-1 in solution. Given the structural similarities between Caribbean and Pacific ciguatoxins, we propose that C-CTX-1 and C-CTX-2 arise from a Caribbean strain of the benthic dinoflagellate, Gambierdiscus toxicus.

Topical penetration of commercial salicylate esters and salts using human isolated skin and clinical microdialysis studies

Aims The penetration of active ingredients from topically applied anti-inflammatory pharmaceutical products into tissues below the skin is the basis of their therapeutic efficacy. There is still controversy as to whether these agents are capable of direct penetration by diffusion through the tissues or whether redistribution in the systemic circulation is responsible for their tissue deposition below the application site. Methods The extent of direct penetration of salicylate from commercial ester and salt formulations into the dermal and subcutaneous tissue of human volunteers was determined using the technique of cutaneous microdialysis. We also examined differences in the extent of hydrolysis of the methylester of salicylate applied topically in human volunteers and in vitro skin diffusion cells using full-thickness skin and epidermal membranes. Results The present study showed that whilst significant levels of salicylate could be detected in the dermis and subcutaneous tissue of volunteers treated with the methylsalicylate formulation, negligible levels of salicylate were seen following application of the triethanolamine salicylate formulation. The tissue levels of salicylate from the methylsalicylate formulation were approx. 30-fold higher than the plasma concentrations. Conclusion The absorption and tissue concentration profiles for the commercial methylsalicylate formulation are indicative of direct tissue penetration and not solely redistribution by the systemic blood supply.