Fluorescent redox-dependent labeling of lipid droplets in cultured cells by reduced phenazine methosulfate.

Natural and synthetic phenazines are widely used in biomedical sciences. In dehydrogenase histochemistry, phenazine methosulfate (PMS) is applied as a redox reagent for coupling reduced coenzymes to the reduction of tetrazolium salts into colored formazans. PMS is also currently used for cytotoxicity and viability assays of cell cultures using sulfonated tetrazoliums. Under UV (340 nm) excitation, aqueous solutions of the cationic PMS show green fluorescence (λem: 526 nm), whereas the reduced hydrophobic derivative (methyl-phenazine, MPH) shows blue fluorescence (λem: 465 nm). Under UV (365 nm) excitation, cultured cells (LM2, IGROV-1, BGC-1, and 3T3-L1 adipocytes) treated with PMS (5 µg/mL, 30 min) showed cytoplasmic granules with bright blue fluorescence, which correspond to lipid droplets labeled by the lipophilic methyl-phenazine. After formaldehyde fixation blue-fluorescing droplets could be stained with oil red O. Interestingly, PMS-treated 3T3-L1 adipocytes observed under UV excitation 24 h after labeling showed large lipid droplets with a weak green emission within a diffuse pale blue-fluorescing cytoplasm, whereas a strong green emission was observed in small lipid droplets. This fluorescence change from blue to green indicates that reoxidation of methyl-phenazine to PMS can occur. Regarding cell uptake and labeling mechanisms, QSAR models predict that the hydrophilic PMS is not significantly membrane-permeant, so most PMS reduction is expected to be extracellular and associated with a plasma membrane NAD(P)H reductase. Once formed, the lipophilic and blue-fluorescing methyl-phenazine enters live cells and mainly accumulates in lipid droplets. Overall, the results reported here indicate that PMS is an excellent fluorescent probe to investigate labeling and redox dynamics of lipid droplets in cultured cells.

Novel 2-arylazoimidazole derivatives as inhibitors of Trypanosoma cruzi proliferation: Synthesis and evaluation of their biological activity.

In this work, the synthesis of a series of 2-arylazoimidazole derivatives 6-20 has been achieved through the reaction of imidazole with aryldiazonium salts, followed by ultrasound-assisted alkylation. This approach has important advantages including higher yield, shorter reaction times and milder reaction conditions. The structures of the compounds obtained were determined by MS, IR; and 1H and 13C NMR. The anti-Trypanosoma cruzi activity of the 15 compounds obtained was evaluated. Two compounds with piperidino substituents in the carboxamide moiety proved to be effective inhibitors of epimastigote proliferation, obtaining inhibition values comparable to those achieved with the reference drug Benznidazole. Besides, these compounds displayed low cytotoxicity on mammalian cells. In vivo, both compounds protected mice against a challenge with a lethal Trypanosoma cruzi strain. These results allow us to propose 2-arylazoimidazoles as lead compounds for the design of novel drugs to treat Chagas' disease.

Varicela complicada en pacientes del Hospital “Juan Pablo II” durante el periodo 2008 – 2011 / Varicella in patients of the Hospital

Se analizó la epidemiología de la Varicela complicada en pacientes del Hospital Pediátrico “Juan Pablo II” de laciudad de Corrientes en el periodo 2008 – 2011, se describió la evolución de los pacientes internados por varicelacomplicada y se la relacionó con el estado nutricional y las internaciones previas de los niños. Estudio descriptivo, retrospectivo, observacional. Población conformada por niños de ambos sexos desde 1 mes a 16 años. Se revisaron las historias clínicas correspondientes. Registramos 1.832 consultas por Varicela (solo en guardia de emergencias) y 75 hospitalizaciones. Hallamos 87% de niños eutróficos, 9% con desnutrición leve, 1% con desnutrición moderada y 3% con desnutrición severa. Solo 18 pacientes con internaciones previas y ningúnpaciente de la serie con inmunización antivaricela. Complicaciones cutáneas: sobreinfección bacteriana, rash,celulitis, impétigo. Otras: neumonía bacteriana, neumonitis, ataxia y encefalitis. El 61% de los internados fueregistrado como primer caso entre los contactos. No hubo óbitos...

We analyzed the epidemiology of varicella in children at Pediatric Hospital “Juan Pablo II” of Corrientes in 2008-2011. We describe the evolution of patients with complicated chickenpox and related infectious complications withnutritional status and previous hospitalizations. Descriptive, retrospective, observational study. Population composed of children of both sexes from 1 month to 16 years old. We reviewed clinical records; 1.832 consultations by chickenpox (only in emergency service); 75 patients required hospitalization. We associate nutritional status and number of previous internments with complications of the disease, and found 87% of normal children, 9% with mild malnutrition, 1% with moderate malnourished and 3% with severe malnourished. Only 18 had previous hospitalization. None of children with varicella immunization. Cutaneous complications: bacterial superinfection, rash, cellulitis, impetigo. Others: pneumonitis, bacterial pneumonia, ataxia and encephalitis. 61% of cases were registered as the first case between contacts. None of them dead.

Experimental Lactococcus garvieae infection in zebrafish and first evidence of its ability to invade non-phagocytic cells.

Zebrafish has been used for studying infections and host-pathogen interactions in different bacterial fish pathogens. In the present study we evaluated the ability of Lactococcus garvieae to infect zebrafish when inoculated intraperitoneally with 2 × 10(7)UFC of this pathogen. L. garvieae can colonize and invade zebrafish at multiple anatomical sites causing a lethal acute septicemic infection with clinical signs and lesions consistent with those observed in lactococcosis outbreaks. Immunohistochemical studies showed the presence of L. garvieae into macrophages as well as into non-phagocytic zebrafish cells of liver (hepatocytes). The internalization capacity showed by L. garvieae in zebrafish cells was confirmed in the rainbow trout cell line RTG-2. Our results provide the first evidence that L. garvieae is able to invade non-phagocytic host cells.

Analysis of the genome content of Lactococcus garvieae by genomic interspecies microarray hybridization.

BACKGROUND: Lactococcus garvieae is a bacterial pathogen that affects different animal species in addition to humans. Despite the widespread distribution and emerging clinical significance of L. garvieae in both veterinary and human medicine, there is almost a complete lack of knowledge about the genetic content of this microorganism. In the present study, the genomic content of L. garvieae CECT 4531 was analysed using bioinformatics tools and microarray-based comparative genomic hybridization (CGH) experiments. Lactococcus lactis subsp. lactis IL1403 and Streptococcus pneumoniae TIGR4 were used as reference microorganisms. RESULTS: The combination and integration of in silico analyses and in vitro CGH experiments, performed in comparison with the reference microorganisms, allowed establishment of an inter-species hybridization framework with a detection threshold based on a sequence similarity of >or= 70%. With this threshold value, 267 genes were identified as having an analogue in L. garvieae, most of which (n = 258) have been documented for the first time in this pathogen. Most of the genes are related to ribosomal, sugar metabolism or energy conversion systems. Some of the identified genes, such as als and mycA, could be involved in the pathogenesis of L. garvieae infections. CONCLUSIONS: In this study, we identified 267 genes that were potentially present in L. garvieae CECT 4531. Some of the identified genes could be involved in the pathogenesis of L. garvieae infections. These results provide the first insight into the genome content of L. garvieae.