Giardiasis is a worldwide parasitic disease that affects mainly children and immunosuppressed people. Side effects and the emergence of resistance over current used drugs make imperative looking for new antiparasitics through discovering of new biological targets and designing of novel drugs. Recently, it has determined that gastric proton-pump inhibitors (PPI) have anti-giardiasic activity. The glycolytic enzyme, triosephosphate isomerase (GlTIM), is one of its potential targets. Therefore, we employed the scaffold of PPI to design new compounds aimed to increase their antigiardial capacity by inactivating GlTIM. Here we demonstrated that two novel PPI-derivatives (BHO2 and BHO3), have better anti-giardiasic activity than omeprazole in concentrations around 120–130 µM, without cytotoxic effect on mammal cell cultures. The derivatives inactivated GlTIM through the chemical modification of Cys222 promoting local structural changes in the enzyme. Furthermore, derivatives forms adducts linked to Cys residues through a C-S bond. We demonstrated that PPI can be used as scaffolds to design better antiparasitic molecules; we also are proposing a molecular mechanism of reaction for these novel derivatives.

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Giardiasis is a neglected parasitic disease that affects primarily children, in whom it delays physical and mental development. The pathophysiology of giardiasis in not well understood, and most reports have identified Giardia intestinalis trophozoites only in the lumen and on the brush border of the small intestine. We identified Giardia trophozoites within the epithelium of the small intestine of a lactose intolerance patient. The Giardia trophozoites were obtained and cultured in vitro. In addition, we demonstrated Giardia trophozoite invasion in an animal model. Giardia trophozoites invaded the intestinal mucosa and submucosa of infected gerbils. The invasive trophozoites were observed at 21, 30 and 60 days age, and the average numbers of invaded sites were 17 ± 5, 15 ± 4, and 9 ± 3, respectively. We found trophozoites between epithelial cells, at the base of empty goblet cells, in lacteal vessels and within the submucosa. The morphological integrity of the invasive trophozoites was demonstrated via electron microscopy. The analysis of the gerbils infected with the trophozoites of the WB reference strain did not show intraepithelial trophozoites. These results demonstrate another Giardia pathogenic mechanism, opening the door to numerous future studies.

Giardiasis, the infestation of the intestinal tract by Giardia lamblia, is one of the most prevalent parasitosis worldwide. Even though effective therapies exist for it, the problems associated with its use indicate that new therapeutic options are needed. It has been shown that disulfiram eradicates trophozoites in vitro and is effective in vivo in a murine model of giardiasis; disulfiram inactivation of carbamate kinase by chemical modification of an active site cysteine has been proposed as the drug mechanism of action. The triosephosphate isomerase from G. lamblia (GlTIM) has been proposed as a plausible target for the development of novel antigiardial pharmacotherapies, and chemical modification of its cysteine 222 (C222) by thiol-reactive compounds is evidenced to inactivate the enzyme. Since disulfiram is a cysteine modifying agent and GlTIM can be inactivated by modification of C222, in this work we tested the effect of disulfiram over the recombinant and trophozoite-endogenous GlTIM. The results show that disulfiram inactivates GlTIM by modification of its C222. The inactivation is species-specific since disulfiram does not affect the human homologue enzyme. Disulfiram inactivation induces only minor conformational changes in the enzyme, but substantially decreases its stability. Recombinant and endogenous GlTIM inactivates similarly, indicating that the recombinant protein resembles the natural enzyme. Disulfiram induces loss of trophozoites viability and inactivation of intracellular GlTIM at similar rates, suggesting that both processes may be related. It is plausible that the giardicidal effect of disulfiram involves the inactivation of more than a single enzyme, thus increasing its potential for repurposing it as an antigiardial drug.

Giardiosis is one of the major intestinal parasitic diseases of human beings as well as wild and domesticated animals. Several protective mechanisms against infection have been described. However, specific information about relationship between giardiosis and the increased proliferation of goblet cells (GC) in patients infected with Giardia intestinalis (Syn. G. duodenalis, G. lamblia) is scarce. In this work, we compare and quantify the number of GC, and have inferred their metabolic state in the small intestine of dogs parasitized with Giardia intestinalis compared to dogs without parasites. Small intestine segments were processed using routine methods for histology and electron microscopy; areas and cells were screened with an Axiovision Ver. 4.0 system. Data were analyzed by ANOVA and comparison of averages. Parasitized dogs showed higher GC numbers than nonparasitized ones. Averages were: 20+/-0.81 GC/25 microm(2) with independent mucin granules and 11+/-1.53 GC/25 microm(2) that were expelling mucus, compared to 11+/-0.94 GC/25 microm(2) and 1+/-0.27 GC/25 microm(2), respectively, in nonparasitized dogs (Tukey, p<0.001). The increases in GC number seem to be an unspecific defensive mechanism against Giardia trophozoites. However, we found some evidence supporting that GC hyperplasia could be a prejudicial to epithelial barrier that gives rise to gates allowing for Giardia-tissue invasion.

BackgroundCnidarian venoms and extracts have shown a broad variety of biological activities including cytotoxic, antibacterial and antitumoral effects. Most of these studied extracts were obtained from sea anemones or jellyfish. The present study aimed to determine the toxic activity and assess the antitumor and antiparasitic potential of Palythoa caribaeorum venom by evaluating its in vitro toxicity on several models including human tumor cell lines and against the parasite Giardia intestinalis.MethodsThe presence of cytolysins and vasoconstrictor activity of P. caribaeorum venom were determined by hemolysis, PLA2 and isolated rat aortic ring assays, respectively. The cytotoxic effect was tested on HCT-15 (human colorectal adenocarcinoma), MCF-7 (human mammary adenocarcinoma), K562 (human chronic myelogenous leukemia), U251 (human glyoblastoma), PC-3 (human prostatic adenocarcinoma) and SKLU-1 (human lung adenocarcinoma). An in vivo toxicity assay was performed with crickets and the antiparasitic assay was performed against G. intestinalis at 24 h of incubation.ResultsP. caribaeorum venom produced hemolytic and PLA2 activity and showed specific cytotoxicity against U251 and SKLU-1 cell lines, with approximately 50% growing inhibition. The venom was toxic to insects and showed activity against G. intestinalis in a dose-dependent manner by possibly altering its membrane osmotic equilibrium.ConclusionThese results suggest that P. caribaeorum venom contains compounds with potential therapeutic value against microorganisms and cancer.