Role of 11β-hydroxysteroid dehydrogenase 2 renal activity in potassium homeostasis in rats with chronic renal failure

Aldosterone concentrations vary in advanced chronic renal failure (CRF). The isozyme 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2), which confers aldosterone specificity for mineralocorticoid receptors in distal tubules and collecting ducts, has been reported to be decreased or normal in patients with renal diseases. Our objective was to determine the role of aldosterone and 11β-HSD2 renal microsome activity, normalized for glomerular filtration rate (GFR), in maintaining K+ homeostasis in 5/6 nephrectomized rats. Male Wistar rats weighing 180-220 g at the beginning of the study were used. Rats with experimental CRF obtained by 5/6 nephrectomy (N = 9) and sham rats (N = 10) were maintained for 4 months. Systolic blood pressure and plasma creatinine (Pcr) concentration were measured at the end of the experiment. Sodium and potassium excretion and GFR were evaluated before and after spironolactone administration (10 mg·kg-1·day-1 for 7 days) and 11β-HSD2 activity on renal microsomes was determined. Systolic blood pressure (means ± SEM; Sham = 105 ± 8 and CRF = 149 ± 10 mmHg) and Pcr (Sham = 0.42 ± 0.03 and CRF = 2.53 ± 0.26 mg/dL) were higher (P < 0.05) while GFR (Sham = 1.46 ± 0.26 and CRF = 0.61 ± 0.06 mL/min) was lower (P < 0.05) in CRF, and plasma aldosterone (Pald) was the same in the two groups. Urinary sodium and potassium excretion was similar in the two groups under basal conditions but, after spironolactone treatment, only potassium excretion was decreased in CRF rats (sham = 0.95 ± 0.090 (before) vs 0.89 ± 0.09 µEq/min (after) and CRF = 1.05 ± 0.05 (before) vs 0.37 ± 0.07 µEq/min (after); P < 0.05). 11β-HSD2 activity on renal microsomes was lower in CRF rats (sham = 0.807 ± 0.09 and CRF = 0.217 ± 0.07 nmol·min-1·mg protein-1; P < 0.05), although when normalized for mL GFR it was similar in both groups. We conclude that K+ homeostasis is maintained during CRF development despite normal Pald levels. This adaptation may be mediated by renal 11β-HSD2 activity, which, when normalized for GFR, became similar to that of control rats, suggesting that mineralocorticoid receptors maintain their aldosterone selectivity.

Homocysteine induces glyceraldehyde-3-phosphate dehydrogenase acetylation and apoptosis in the neuroblastoma cell line Neuro2a

High plasma levels of homocysteine (Hcy) promote the progression of neurodegenerative diseases. However, the mechanism by which Hcy mediates neurotoxicity has not been elucidated. We observed that upon incubation with Hcy, the viability of a neuroblastoma cell line Neuro2a declined in a dose-dependent manner, and apoptosis was induced within 48 h. The median effective concentration (EC50) of Hcy was approximately 5 mM. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) nuclear translocation and acylation has been implicated in the regulation of apoptosis. We found that nuclear translocation and acetylation of GAPDH increased in the presence of 5 mM Hcy and that higher levels of acetyltransferase p300/CBP were detected in Neuro2a cells. These findings implicate the involvement of GAPDH in the mechanism whereby Hcy induces apoptosis in neurons. This study highlights a potentially important pathway in neurodegenerative disorders, and a novel target pathway for neuroprotective therapy.

Determination of structural and mechanical properties, diffractometry, and thermal analysis of chitosan and hydroxypropylmethylcellulose (HPMC) films plasticized with sorbitol

In this work, the structural, mechanical, diffractometric, and thermal parameters of chitosan-hydroxypropylmethylcellulose (HPMC) films plasticized with sorbitol were studied. Solutions of HPMC (2% w/v) in water and chitosan (2% w/v) in 2% acetic acid solution were prepared. The concentration of sorbitol used was 10% (w/w) to both polymers. This solutions were mixed at different proportions (100/0; 70/30; 50/50; 30/70, and 0/100) of chitosan and HPMC, respectively, and 20 mL was cast in Petri dishes for further analysis of dried films. The miscibility of polymers was assessed by X-ray diffraction, scanning electronic microscopy (SEM), differential scanning calorimetry (DSC), and thermal gravimetric analysis (TGA). The results obtained indicate that the films are not fully miscible at a dry state despite the weak hydrogen bonding between the polymer functional groups.

Excipientes de medicamentos e as informações da bula

OBJETIVO: Avaliar a presença de conservantes, corantes, adoçantes e aromatizantes em 73 apresentações farmacêuticas de 35 medicamentos para uso oral, e as informações da bula sobre excipientes. MÉTODOS: Selecionamos 35 medicamentos, de venda livre ou sob prescrição médica, comercializados no Brasil. A amostra incluiu: analgésicos/antitérmicos, antimicrobianos, mucolíticos, antitussígenos, descongestionantes, anti-histamínicos, broncodilatadores, corticosteróides, antiinflamatórios e suplementos vitamínicos. Foram analisadas 73 apresentações desses fármacos, anotando-se as informações da bula sobre conservantes, corantes, adoçantes e aromatizantes. RESULTADOS: A bula de um medicamento (1,3%) não mencionava os ingredientes inativos. Os conservantes mais encontrados nos medicamentos foram metilparabeno e propilparabeno (43% e 35,6% respectivamente). Os adoçantes mais usados foram: sacarose (açúcar) (53,4%), sacarina sódica (38,3%) e sorbitol (36,9%). Vinte e um produtos (28,7%) continham dois adoçantes. Predominaram os medicamentos sem corante (43,8%), seguidos pelos coloridos por amarelo crepúsculo (amarelo FD&C no. 6) (15%). Cinco produtos (6,8%) continham mais de um corante. A tartrazina (amarelo FD&C no. 5) foi encontrada em sete formulações (9,5%). Os aromatizantes mais usados foram os de frutas (83%). Constatamos a freqüente omissão das bulas sobre o teor exato de açúcar dos produtos (77%). Duas das quatro bulas de medicamentos contendo aspartame não mencionavam as precauções no uso por fenilcetonúricos. CONCLUSÕES: A omissão e a imprecisão das informações da bula sobre os excipientes farmacêuticos expõem os indivíduos suscetíveis ao risco de reações adversas dos conservantes e corantes. Também podem ocorrer complicações do uso inadvertido de medicamentos contendo açúcar pelos pacientes diabéticos, ou de fármacos adoçados com aspartame pelos fenilcetonúricos.

Análise físico-química e sensorial de doce de leite produzido sem adição de sacarose

O objetivo foi avaliar doces de leite produzidos sem adição de açúcar. Realizaram-se três formulações: doce com adição de açúcar (A), com edulcorante sucralose (B) e doce com ciclamato, sacarina e sorbitol (C). Nos doces B e C utilizou-se a carragena. Realizaram-se análises físico-quimicas, microbiológicas, exigidas pela legislação vigente, e análise sensorial. Nas análises de acidez não houve diferença significativa entre as amostras. O teor de gordura, a atividade de água e textura das amostras não diferiram entre os doces B e C, e esses diferiram estatisticamente do A. Nas análises de pH houve diferença significativa entre os doces. O teor de sólidos solúveis do doce A apresentou 63 °Brix, e os doces B e C, 26 °Brix. Na aceitação, observou-se diferença significativa para todos os atributos avaliados. Para sabor, o doce A foi mais aceito que os doces B e C, e o doce B mais que o C. Para atributo de textura sensorial e impressão global, o doce A foi mais aceito que os doces B e C, e esses apresentaram a mesma aceitação. Na análise de textura instrumental, os doces B e C não obtiveram a mesma força de resistência à penetração que o doce A. Todas as análises microbiológicas dos doces A, B e C apresentaram-se dentro dos padrões exigidos pela legislação vigente. Doce de leite com sucralose (B) mostrou-se uma boa alternativa para substituição do doce de leite com sacarose (A), uma vez que obteve boa aceitação para o atributo sabor.

Methemoglobinemia associated with loxoscelism

In twenty five patients who presented the cutaneous form of loxoscelism, serum haptoglobin and lactic dehydrogenase, erythrocyte glucose-6-phosphate dehydrogenase, glutathione reductase, glutathione peroxidase, methemoglobin, bilirubin and reticulocytes were investigated after bite. No hemolysis was detected but an increase in methemoglobin was found in 54% of the cases; in 7% it was between 1.1% and 2%, in 27% it ranged from 2.1% to 4%, and in 20% from 4.1% to 8%. Blood samples of a normal, blood group 0 individual and of a patient who exhibited methemoglobinemia after Loxosceles bite were incubated separately with antisera against Loxosceles gaucho, Crotalus terrificus, Bothrops jararaca, with Loxosceles gaucho venom and 0.3% phenol. No methemoglobin was found after 1, 4,8 and 15 days in both sets of samples. At the 25th day all the samples, including the controls, exhibited similar methemoglobin reductase decrease. The data suggest that the methemoglobinemia which occurs in 50% of the patients probably arises from in vivo venom metabolism, inasmuch as the crude venom does not induce methemoglobinemia.

Comparative studies of Yersinia pestis outer membrane isolation techniques and their potential use in plague epidemiology

In the present study three techniques for obtaining outer membrane enriched fractions from Yersinia pestis were evaluated. The techniques analysed were: differential solubilization of the cytoplasmic membrane with Sarkosyl or Triton X-100, and centrifugation in sucrose density gradients. The sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) of outer membrane isolated by the different methods resulted in similar protein patterns. The measurement of NADH-dehydrogenase and succinate dehydrogenase (inner membrane enzymes) indicated that the outer membrane preparations obtained by the three methods were pure enough for analytical studies. In addition, preliminary evidences on the potential use of outer membrane proteins for the identification of geographic variants of Y. pestis wild isolates are presented.

Norfloxacin monodose use in patients with cholera in Salta, Argentina

The use ofmonodose (800 mg) per os of Norfloxacin was evaluated in 32 patients with cholera at Salvador Mazza's Hospital, Salta, Argentina. It was considered the celerity in negativization of stool culture (100% of cases: 12 hours post administration), its efficiency along time (24/24 controlled patients were negative at 10th day) and MIC of isolated strains (100% of strains were sensitive: range 0.008 to 0.016 mug/ml). It was included oral administration of sorbitol 70% in peanut oil in order to study patients at 10th day's control. This method could be an alternative one in the study of asymptomatic carrier. Norfloxacin monodose shows good performance in early negativization of stool culture and it was also effective along the whole observation period, suggesting it could prevent carriage.

Human cutaneous leishmaniasis caused by Leishmania (Viannia) braziliensis in Santiago del Estero, Argentina: identification of parasites by monoclonal antibodies and isoenzymes

Diagnostic and parasite characterization and identification studies were carried out in human patients with cutaneous leishmaniasis lesions in Santiago del Estero, Northern Province of Argentina. Diagnostic procedures were biopsies of lesions for smears and inoculations in hamster, needle aspirations of material from ulcers for "in vitro" cultures. Immunodiagnostic techniques applied were IFAT-IgG and Montenegro skin test. Primary isolation of eight stocks of leishmanial parasites was achieved from patients with active lesions. All stocks were biologically characterized by their behaviour in hamster, measurements of amastigote and promastigotes and growth "in vitro". Eight stocks were characterized and identified at species level by their reactivity to a cross-panel of sub-genus and specie-specific Monoclonal Antibodies through an Indirect Immunofluorescence technique and a Dot-ELISA. We conclude from the serodeme analysis of Argentina stocks that: stocks MHOM/AR/92/SE-1; SE-2; SE-4; SE-8; SE-8-I; SE-30; SE-34 and SE-36 are Leishmania (Viannia) braziliensis. Three Leishmania stocks (SE-1; SE-2 and SE-30) did not react with one highly specie-specific Monoclonal Antibody (Clone: B-18, Leishmania (Viannia) braziliensis marker) disclosing two serodeme group patterns. Five out of eight soluble extracts of leishmanial promastigotes were electrophoresed on thin-layer starch gels and examined for the enzyme MPI, Mannose Phosphate Isomerase; MDH, Malate Dehydrogenase; 6PGD, 6 Phosphogluconate Dehydrogenase; NH, Nucleoside Hydrolase, 2-deoxyinosinc as substrate; SOD, Superoxide Dismutase; GPI, Glucose Phosphate Isomerase and ES, Esterase. From the isoenzyme studies we concluded that stocks: MHOM/AR/92/SE-1; SE-2; SE-4; SE-8 and SE-8-I are isoenzymatically Leishmania (Viannia) braziliensis. We need to analyze more enzymes before assigning them to a braziliensis zymodeme.

Snakebites by Crotalus durissus ssp in children in Campinas, São Paulo, Brazil

From January, 1984 to March, 1999, 31 children under 15 y old (ages 1-14 y, median 8 y) were admitted after being bitten by rattlesnakes (Crotalus durissus ssp). One patient was classified as "dry-bite", 3 as mild envenoming, 9 as moderate envenoming and 18 as severe envenoming. Most patients had neuromuscular manifestations, such as palpebral ptosis (27/31), myalgia (23/31) and weakness (20/31). Laboratory tests suggesting rhabdomyolysis included an increase in total blood creatine kinase (CK, 28/29) and lactate dehydrogenase (LDH, 25/25) levels and myoglobinuria (14/15). The main local signs and symptoms were slight edema (20/31) and erythema (19/31). Before antivenom (AV) administration, blood coagulation disorders were observed in 20/25 children that received AV only at our hospital (incoagulable blood in 17/25). AV early reactions were observed in 20 of these 25 cases (9/9 patients not pretreated and 11/16 patients pretreated with hydrocortisone and histamine H1 and H2 antagonists). There were no significant differences in the frequency of patients with AV early reactions between the groups that were and were not pretreated (Fisher's exact test, p = 0.12). Patients admitted less than and more than 6 h after the bite showed the same risk of developing severe envenoming (Fisher's exact test, p = 1). No children of the first group (< 6 h) showed severe complications whereas 3/6 children admitted more than 6 h post-bite developed acute renal failure. Patients bitten in the legs had a higher risk of developing severe envenoming (Fisher's exact test, p = 0.04). There was a significant association between both total CK and LDH blood enzyme levels and severity (p < 0.001 for CK and p < 0.001 for LDH; Mann-Whitney U test). No deaths were recorded.

HYPOXIC STRESS, HEPATOCYTES AND CACO-2 VIABILITY AND SUSCEPTIBILITY TO Shigella flexneri INVASION

SUMMARY Inflammation due to Shigella flexneri can cause damage to the colonic mucosa and cell death by necrosis and apoptosis. This bacteria can reach the bloodstream in this way, and the liver through portal veins. Hypoxia is a condition present in many human diseases, and it may induce bacterial translocation from intestinal lumen. We studied the ability of S. flexneri to invade rat hepatocytes and Caco-2 cells both in normoxic and hypoxic microenvironments, as well as morphological and physiological alterations in these cells after infection under hypoxia. We used the primary culture of rat hepatocytes as a model of study. We analyzed the following parameters in normoxic and hypoxic conditions: morphology, cell viability, bacterial recovery and lactate dehydrogenase (LDH) released. The results showed that there were fewer bacteria within the Caco-2 cells than in hepatocytes in normoxic and hypoxic conditions. We observed that the higher the multiplicity of infection (MOI) the greater the bacterial recovery in hepatocytes. The hypoxic condition decreased the bacterial recovery in hepatocytes. The cytotoxicity evaluated by LDH released by cells was significantly higher in cells submitted to hypoxia than normoxia. Caco-2 cells in normoxia released 63% more LDH than hepatocytes. LDH increased 164% when hepatocytes were submitted to hypoxia and just 21% when Caco-2 cells were in the same condition. The apoptosis evaluated by Tunel was significantly higher in cells submitted to hypoxia than normoxia. When comparing hypoxic cells, we obtained more apoptotic hepatocytes than apoptotic Caco-2 cells. Concluding our results contribute to a better knowledge of interactions between studied cells and Shigella flexneri. These data may be useful in the future to define strategies to combat this virulent pathogen.

FATAL DISSEMINATED CRYPTOCOCCOSIS WITH RENAL INVOLVEMENT IN AN HIV-INFECTED PATIENT

SUMMARY Introduction: We present a fatal case of disseminated cryptococcosis in a young man whose diagnosis of HIV infection was made at the time of admission to the emergency room. Case report: The patient was a twenty-three-year-old man, with a history of daily fever during one month associated with diarrhea, weight loss, headache, vomiting and generalized seizures. He also had a history of diabetes mellitus, alcoholism and drug addiction. Upon physical examination the patient was pale, disoriented and had periods of agitation. White blood cells count was 3,440/mm3 (5% lymphocytes), hemoglobin was 10g/dL, platelets were 83,000/ mm3. Creatinine was 0.7 mg/dL; urea 19 mg/dL; Na, K, and liver enzymes were within normal limits. Lactic dehydrogenase was 494 IU/L. Cerebrospinal fluid (CSF) analysis revealed 10 white blood cells/mm3 (58% neutrophils, 31% lymphocytes, 11% monocytes) and 2 red blood cells/mm3. India ink test revealed six Cryptococcus yeasts/mm3. CSF glucose was 122 mg/dL and protein was 36 mg/ dL. VDRL test was negative and anti-HIV test was positive. Intravenous hydration, insulin, phenytoin, fluconazole, pyrimethamine, sulfadiazine, folinic acid, and amphotericin B were started. The patient did not improve and became obtunded and hypotensive. He was intubated and put on mechanical respiration. He received vasoactive drugs and died less than 24 hours after admission. A postmortem examination was performed and revealed disseminated cryptococcosis, with severe involvement of the kidneys. Conclusion: Cryptococcosis, as a rule, is a systemic disease that affects mostly immunocompromised individuals, especially patients with AIDS. When diagnosed late in its course it has a very high mortality.

Echinococcus granulosus sensu lato GENOTYPES IN DOMESTIC LIVESTOCK AND HUMANS IN GOLESTAN PROVINCE, IRAN

Cystic echinococcosis (CE) is a globally parasitic zoonosis caused by larval stages of Echinococcus granulosus. This study investigated E. granulosus genotypes isolated from livestock and humans in the Golestan province, northern Iran, southeast of the Caspian sea, using partial sequencing data of the cytochrome c oxidase subunit 1 (cox1) and NADH dehydrogenase 1 (nad1) mitochondrial genes. Seventy E. granulosus isolates were collected from animals in slaughterhouses: 18 isolates from sheep, 40 from cattle, nine from camels, two from buffaloes and one from a goat, along with four human isolates (formalin-fixed, paraffin-embedded tissues) from CE patients of provincial hospitals. All isolates were successfully analysed by PCR amplification and sequencing. The sequence analysis found four E. granulosus genotypes among the 74 CE isolates: G1 (78.3%), G2 (2.7%), G3 (15%) and G6 (4%). The G1-G3 complex genotype was found in all of the sheep, goat, cattle and buffalo isolates. Among the nine camel isolates, the frequency of G1-G3 and G6 genotypes were 66.7% and 33.3%, respectively. All four human CE isolates belonged to E. granulosus sensu stricto. This study reports the first occurrence of the G2 genotype in cattle from Iran and confirms the previously reported G3 genotype in camels in the same country.

Africanized honeybee stings: how to treat them

INTRODUCTION: In 1956, Africanized honeybees (AHB) migrated from Brazil to other regions of the Western Hemisphere, including South, Central, and North America, except for Canada. Despite being productive, they are highly aggressive and cause fatal accidents. This study aimed to evaluate patients at the Clinical Hospital of Botucatu Medical School (HC-FMB) and to propose treatment guidelines. METHODS: From 2005 to 2006, the clinical and laboratorial aspects of 11 patients (7 male and 4 female) and the anatomopathological aspects of one patient who had died in 2003 were analyzed. RESULTS: The age of the surviving patients varied from 5 to 87 years, with a mean of 42.5 years. The majority of accidents occurred in the afternoon, and the number of stings ranged from 20 to 500. The principal signs and symptoms were pain and local inflammatory signs, nausea, tachycardia, and vomiting. Biochemical findings presented increased levels of creatine phosphokinase, lactate dehydrogenase, and aspartate/alanine aminotransferase. An 11-year-old male patient died upon entering the attic of a two-storey building where he was attacked by a swarm, receiving more than 1,000 stings. He was sent to HC-FMB where he was treated, but he died 24h later. Observed at the autopsy were erythematous-purpuric skin lesions besides necrosis at the sting locations, rhabdomyolysis, focal myocardial necrosis, tubular hydropic degeneration and focal tubular acute necrosis of the kidneys, myoglobinuria, and centrolobular necrosis in the liver. CONCLUSIONS: Accidents caused by multiple AHB stings always constitute a medical emergency. As there is no specific antivenom, we have developed guidelines, including first aid, drugs, and the proper removal of stingers.

The influence of carbohydrates in the interaction of Paracoccidioides brasiliensis with CCL-6 cells in vitro

INTRODUCTION: Little is known about the early events in the interaction between Paracoccidioides brasiliensis and its host. To understand the effect of carbohydrates in the interaction between the fungus and epithelial cell in culture, we analyzed the influence of different carbohydrate solutions on the adhesion of P. brasiliensis yeast cells to CCL-6 cells in culture. METHODS: Fungal cells were cultivated with the epithelial cell line, and different concentrations of D-fucose, N-acetyl-glucosamine, D-mannose, D-glucosamine, D-galactosamine, sorbitol and fructose were added at the beginning of the experiment. Six hours after the treatment, the cells were fixed and observed by light microscopy. The number of P. brasiliensis cells that were adhered to the CCL-6 monolayer was estimated. RESULTS: The number of adhesion events was diminished following treatments with D-fucose, N-acetyl-glucosamine, D-mannose, D-glucosamine and D-galactosamine as compared to the untreated controls. Sorbitol and fructose-treated cells had the same adhesion behavior as the observed in the control. P. brasiliensis propagules were treated with fluorescent lectins. The FITC-labeled lectins WGA and Con-A bound to P. brasiliensis yeast cells, while SBA and PNA did not. CONCLUSIONS: The perceptual of adhesion between P. brasiliensis and CCL-6 cells decreased with the use of D-mannose, N-acetyl-glucosamine and D-glucosamine. The assay using FITC-labeled lectins suggests the presence of N-acetyl-glucosamine, α-mannose and α-glucose on the P. brasiliensis cell surface. An enhanced knowledge of the mediators of adhesion on P. brasiliensis could be useful in the future for the development of more efficient and less harmful methods for disease treatment and control.