Acute and short-term insulin-induced molecular adaptations of GLUT2 gene expression in the renal cortex of diabetic rats

Increased GLUT2 gene expression in the renal proximal tubule of diabetic rats is an adaptive condition, which may be important in the diabetic nephropathy development. We investigated the effects of insulin treatment upon the renal GLUT2 overexpression of diabetic rats. Acute treatment, surprisingly, induced a rapid further increase in GLUT2 mRNA content. Twelve hours after insulin injection, GLUT2 mRNA was twice the value of saline-injected rats (P < 0.001), when GLUT2 protein remained unchanged. In response to short-term treatment, both GLUT2 mRNA and protein were increased in 1-day treated rats (P < 0.05 versus saline-injected), decreasing after that, and reaching, within 6 days, values close to those of non-diabetic rats. Concluding, insulin treatment induced: initially, an additional upregulation of GLUT2 gene expression, involving posttranscriptional modulation; thereafter, downregulation of GLUT2 expression, which returns to non-diabetic levels. The former may be related to increased insulin concentration, the latter may be due to glycemic control. © 2005 Elsevier B.V. All rights reserved.

Effect of dietary supplementation with vitamin E and stocking density on macrophage recruitment and giant cell formation in the teleost fish, Piaractus mesopotamicus

The effect of dietary supplementation with 0, 100 and 450 mg of vitamin E (DL-α tocopheryl acetate)/kg of a dry diet on the kinetics of macrophage recruitment and giant cell formation in the pacu, maintained at different stocking densities (5 kg/m3 and 20 kg/m3), was investigated by insertion of round glass coverslips into the subcutaneous connective tissue. After a feeding period of 18 weeks, the coverslips were implanted and later removed for examination at 2, 7 and 15 days post-implantation. Fish fed diets supplemented with 450 mg of vitamin E showed an increase (P<0.05) in the accumulation of macrophages, foreign body giant cells and Langhans type cells. The kinetics of macrophage recruitment and giant cell formation on the glass coverslips appeared to be strongly influenced by vitamin E supplementation, since fish fed a basal diet and held at high stocking densities showed low numbers of adhering cells on the coverslips, and high concentrations of plasma corticosteroids. On the other hand, fish given a diet supplemented with 450 mg of vitamin E did not show a similar difference in plasma cortisol concentrations related to stocking density. The effect of cortisol concentrations on carbohydrate metabolism, analysed by assessment of plasma glycaemia, was not clear. Blood glucose concentrations did not vary substantially with the different treatments examined. These results suggest that vitamin E may contribute to the efficiency of the fish's inflammatory response by increasing macrophage recruitment and giant cell formation in the foreign body granulomatous reaction. Vitamin E appeared to act on the stress response of pacus by preventing a stress-related immunosuppression. © 2005 Elsevier Ltd. All rights reserved.

Low ethanol consumption induces enhancement of insulin sensitivity in liver of normal rats

Moderate amounts of alcohol intake have been reported to have a protective effect on the cardiovascular system and this may involve enhanced insulin sensitivity. We established an animal model of increased insulin sensitivity by low ethanol consumption and here we investigated metabolic parameters and molecular mechanisms potentially involved in this phenomenon. For that, Wistar rats have received drinking water either without (control) or with 3% ethanol for four weeks. The effect of ethanol intake on insulin sensitivity was analyzed by insulin resistance index (HOMA-IR), intravenous insulin tolerance test (IVITT) and lipid profile. The role of liver was investigated by the analysis of insulin signaling pathway, GLUT2 gene expression and tissue glycogen content. Rats consuming 3% ethanol showed lower values of HOMA-IR and plasma free fatty acids (FFA) levels and higher hepatic glycogen content and glucose disappearance constant during the IVITT. Neither the phosphorylation of insulin receptor (IR) and insulin receptor substrate-1 (IRS-1), nor its association with phosphatidylinositol-3-kinase (PI3-kinase), was affected by ethanol. However, ethanol consumption enhanced liver IRS-2 and protein kinase B (Akt) phosphorylation (3 times, P < 0.05), which can be involved in the 2-fold increased (P < 0.05) hepatic glycogen content. The GLUT2 protein content was unchanged. Our findings point out that liver plays a role in enhanced insulin sensitivity induced by low ethanol consumption. © 2005 Elsevier Inc. All rights reserved.

Efeito da cadência de pedalada sobre as respostas metabólica e cardiovascular durante o exercício incremental e de carga constante em indivíduos ativos

The main purpose of this study was to analyze the effect of the pedaling cadence (500 × 100 rpm) on the heart rate (HR) and the blood lactate response during incremental and constant workload exercises in active individuals. Nine active male individuals (20.9 ± 2.9 years old; 73.9 ± 6.5 kg; 1.79 ± 0.9 m) were submitted to two incremental tests, and to 6-8 constant workload tests to determine the intensity corresponding to the maximal steady state lactate (MLSSintens) in both cadences. The maximal power (Pmax) attained during the incremental test, and the MLSSintens were significantly lower at 100 rpm (240.9 ± 12.6 W; 148.1 ± 154.W) compared to 50 rpm (263.9 ± 18.6 W; 186.1 ± 21.2 W), respectively. The HRmax did not change between cadences (50 rpm = 191.1 ± 8.8 bpm; 100 rpm = 192.6 ± 9.9 bpm). Regardless the cadence, the HRmax percentage (70, 80, 90, and 100%) determined the same lactate concentrations during the incremental test. However, when the intensity was expressed in Pmax percentage or in absolute power, the lactate and the HR values were always higher at highest cadences. The HR corresponding to MLSSintens was similar between cadences (50 rpm = 162.5 ± 9.1 bpm; 100 rpm = 160.4 ± 9.2 bpm). Based on these results, it can be conclude that regardless the cadence employed (50 × 100 rpm), the use of the HR to individualize the exercise intensity indicates similar blood lactate responses, and this relationship is also kept in the exercise of constant intensity performed at MLSSintens. On the other hand, the use of the Pmax percentages depend on the cadence used, indicating different physiological responses to a same percentage.

Fermented soy product supplemented with isoflavones affected fat depots in juvenile rats

Objectives: This study investigated the effects of soy product fermented by Enterococcus faecium and Lactobacillus jugurti supplemented with isoflavones on adipose tissue, blood lipid, and glucose levels on juvenile rats. Methods: Rats were fed a cholesterol-enriched diet for 3 wk as a preliminary treatment to create hypercholesterolemia. They were then fed a chow diet (HC), a chow diet plus fermented soy product supplemented with isoflavones (HCFI), a chow diet plus placebo (HCP), or a chow diet plus placebo supplemented with isoflavones (HCPI), respectively, for an additional 3 wk. Results: The beneficial effects of fermented soy product supplemented with isoflavones on epididymal (EPI) and retroperitoneal (RET) fat pads was likely due to isoflavones because adipocyte circumference (micrometers) in the HC group was significantly larger (EPI: 105.66 ± 13.36; RET: 134.95 ± 25.40) than that in the HCFI group (EPI: 93.17 ± 12.80; RET: 108.62 ± 15.50) and HCPI group (EPI: 93.06 ± 15.10; RET: 112.34 ± 18.21). The probiotic micro-organism accentuated the antilipogenic effect of isoflavones on RET (HCFI: 108.62 ± 15.50 micrometers versus HCPI: 112.34 ± 18.21 micrometers). Moreover, the fermented product increased glucose concentration similar to that in the chow group but did not change blood lipids. Conclusion: This product may offer new approaches to obesity prevention. © 2005 Elsevier Inc. All rights reserved.

Maximal lactate steady state in running rats

The higher concentration during exercise at which lactate entry in blood equals its removal is known as maximal lactate steady state (MLSS) and is considered an important indicator of endurance exercise capacity. The aim of the present study was to determine MLSS in running rats. Adult male Wistar sedentary rats, which were selected and adapted to treadmill running for three weeks, were used. After becoming familiarized with treadmill running, the rats were submitted to five exercise tests at 15, 20, 25, 30 and 35 m/min velocities. The velocity sequence was distributed at random. Each test consisted of continuous running for 25 min at one velocity or until the exhaustion. Blood lactate was determined at rest and each 5 min of exercise to find the MLSS. The running rats presented MLSS at the 20 m/min velocity, with blood lactate of 3.9±1.1 mmol/L. At the 15 m/min velocity, the blood lactate also stabilized, but at a lower concentration (3.2±1.1 mmol/L). There was a progressive increase in blood lactate concentration at higher velocities, and some animals reached exhaustion between the 10 th and 25 th minute of exercise. These results indicate that the protocol of MLSS can be used for determination of the maximal aerobic intensity in running rats.

Liposomal daunorubicin and dexamethasone as a treatment for multiple myeloma - The DD protocol

Context and Objective: Lipasomial daunorubicin has been used to treat hematological malignancies, including multiple myelomo (MM). The goal was to evaluate efficacy, side-effects and toxicity of liposomal daunorubicin and dexamethasone (DD Protocol). Design and Setting: Prospective study of Sírio-Libonês, São Camilo, Brasil and Alemão Oswaldo Cruz hospitals. Methods: Twenty consecutive patients with active MM received four cycles of liposomal daunorubicin intravenously for two hours (25-30 mg/m 2/day) on three consecutive days per month, with oral dexamethasone, (10 mg every six hours) on four consecutive days three times a month. Results: The male/female ratio was 1:1 and median age 60. Nine patients were stage IIA, ten IIIA and one IIIB. The median from diagnosis to starting DD was 13 months. All patients received four cycles, except one. Fifteen had already received chemotherapy before DD. Responses of > 50% reduction in serum monoclonal paraprotein were observed in six patients after first cycle (30%), six after second (30%) and four after third (20%), while four (20%) did not obtain this. Initially, 17 patients (85%) had anemia: 12 (70%) achieved correction. Progressive disease was observed in three patients (15%), while one had minimal response, four (20%) partial and 12 (60%) complete. Hemotologlical toxicity was acceptable: three patients (15%) had neutrophils < 1,000/mm 3; none had thrombocyfopenia. Gastrointestinal toxicity was mild: nausea (10%), anorexio (15%) and no vomiting. Conclusions: This treatment has mild toxicity and good response rate. It may therefore be feasible before autologous bone marraw transplantation.

Diagnosis of 5α-reductase type 2 deficiency: Contribution of anti-Müllerian hormone evaluation

Aim: To evaluate anti-Müllerian hormone (AMH) levels in patients with clinical and molecular diagnosis of 5α-reductase 2 deficiency. Patients and methods: Data from 14 patients whose age ranged from 21 days to 29 years were analyzed according to age and pubertal stage. Sexual ambiguity was rated as Prader III in 11 patients. LH, FSH, testosterone (T), dihydrotestosterone (DHT) and AMH serum levels were measured in all but two patients, who had been previously submitted to gonadectomy; T and DHT were also measured in 20 age-matched controls. Results: Gonadotropin levels were normal in all but one patient who retained gonads (six of whom had reached puberty) and T/DHT ratio was elevated in all patients when compared to controls. All prepubertal patients had AMH levels < -1 SD for age, while most pubertal patients had AMH levels compatible with pubertal stage. Conclusions: Prepubertal patients with 5α-reductase 2 deficiency have AMH values in the lower part of the normal range. These data indicate that T does not need to be converted to DHT to inhibit AMH secretion by Sertoli cells. © Freund Publishing House Ltd., London.

Comparison of hemodynamic, clinicopathologic, and gastrointestinal motility effects and recovery characteristics of anesthesia with isoflurane and halothane in horses undergoing arthroscopic surgery

Objective - To compare hemodynamic, clinicopathologic, and gastrointestinal motility effects and recovery characteristics of halothane and isoflurane in horses undergoing arthroscopic surgery. Animals - 8 healthy adult horses. Procedure - Anesthesia was maintained with isoflurane or halothane (crossover study). At 6 intervals during anesthesia and surgery, cardiopulmonary variables and related derived values were recorded. Recovery from anesthesia was assessed; gastrointestinal tract motility was subjectively monitored for 72 hours after anesthesia. Horses were administered chromium, and fecal chromium concentration was used to assess intestinal transit time. Venous blood samples were collected for clinicopathologic analyses before and 2, 24, and 48 hours after anesthesia. Results - Compared with halothane-anesthetized horses, cardiac index, oxygen delivery, and heart rate were higher and systemic vascular resistance was lower in isoflurane-anesthetized horses. Mean arterial blood pressure and the dobutamine dose required to maintain blood pressure were similar for both treatments. Duration and quality of recovery from anesthesia did not differ between treatments, although the recovery periods were somewhat shorter with isoflurane. After isoflurane anesthesia, gastrointestinal motility normalized earlier and intestinal transit time of chromium was shorter than that detected after halothane anesthesia. Compared with isoflurane, halothane was associated with increases in serum aspartate transaminase and glutamate dehydrogenase activities, but there were no other important differences in clinicopathologic variables between treatments. Conclusions and clinical relevance - Compared with halothane, isoflurane appears to be associated with better hemodynamic stability during anesthesia, less hepatic and muscle damage, and more rapid return of normal intestinal motility after anesthesia in horses undergoing arthroscopic procedures.

Papel da eritropoetina recombinante humana sobre o metabolismo dos carboidratos, paratormônio, cálcio iônico, zinco, prolactina e níveis pressóricos em renais crônicos tratados com hemodiálise

Objective: To evaluate the influence of recombinant human erythropoietin (Epho) on carbohydrate metabolism, parathyroid hormone, calcium ionic, zinc, prolactin and blood pressure (BP) in chronic renal failure treated by hemodialysis. Methods: Ten patients in hemodialysis were followed during 24 weeks in two phases: 12 weeks pre-Epho (BP was measured pre and post hemodialysis sessions) and 12 weeks post-Epho (BP was measured as above and also the blood levels of glucose, insulin, parathyroid hormone, calcium ionic, prolactin, and zinc). Results: Patients were 39.8±8.5 y, 50% males. Hematocrit and hemoglobin presented a significant increase four weeks after Epho (22.3±2.3 to 28.1±2.6% and 7.4±0.8 to 9.4±0.9 g/dL, p<0.05). BP (mmHg) and weight pre-Epho: 158±99 and 59±13 (before hemodialysis), 147±96 and 55±13 (after hemo) and post-Epho: 161±100 and 59±13 (before hemo) 155±101 and 56±12 (after hemo) were all not statistically different in any moment. There are also no difference pre and post-Epho in fast glucose (91.8±6.5 and 90.8±6.1 mg/dL, p>0.05), parathyroid hormone (341.4±249.3 and 515.7±310 pg/ mL), calcium ionic (3,66±0.63 and 3.76±0.45 mmol/L), prolactin (males: 327±144.1 and 298.1 ±145.2 μg/mL; females: 666.2±426.6 and 659±395.3 μg/mL) and zinc (median of 0.73 and 0.71 μg\L). Basal insulin was lower after Epho (median of 9.1 to 3.8 μg/mL, p<0.05). Conclusion: These data suggest that recombinant human erythropoietin was effective to improve the anemia and the carbohydrate metabolism in patients with chronic renal failure treated by hemodialysis. © Copyright Moreira Jr. Editors. Todos os direitos reservados.

Clonagem e expressão da glicoproteína transmembrana do retrovírus HTLV-1 em células de mamíferos

The retrovirus HTLV-1 is the etiological agent of the adult T-cell leukemia and HTLV-1 associated myelopathy/tropical spastic paraparesis. The proviral genome has 9,032 base pairs, showing regulatory and structural genes. The env gene encodes for the transmembrane glycoprotein gp 21. The development of methodologies for heterologous protein expression, as well as the acquisition of a cellular line that constituently expresses the recombinant, were the main goals of this work. The DNA fragment that encodes for gp 21 was amplified by nested-PCR and cloned into a pCR2.1-TOPO vector. After which, a sub-cloning was realized using the expressing vector pcDNA3.1+. The transfection of mammalian cells HEK 293 was performed transitorily and permanently. Production of the recombinant gp 21 was confirmed by flux cytometry experiments and the cell line producing protein will be used in immunogenicity assays.

The number of podocyte and slit diaphragm is decreased in experimental diabetic nephropathy

Purpose: To determine the number of podocyte, slit diaphragms, slit diaphragm extensions and GBM thickness in diabetic nephropathy. Methods: Sixty Rattus Wistarof both sexes weighing 200-300g were divided in two experimental groups: normal group 10 animals, and alloxan diabetic rats - 50 animals. Alloxan was administered in a single IV dose of 42mg/kg body weight. Body weight, water and food intake, diuresis, and blood and urine glucose were determined in both groups before alloxan injection and two weeks, six and twelve months after alloxan injection. Proteinuria was measured at 12 months in both groups. After 12 months animals were sacrificed, and the right kidney processed for electron microscopy. Results: Clear clinical and laboratory signs of severe diabetes were seen, in all alloxan-diabetic rats at all follow-up times. Glomerular basement membrane (GBM) thickening, podocyte number, and slit diaphragm number and extension were determined. GBM of all diabetic rats was significantly thicker (median=0.29μm; semi-interquartile range=0.065μm) than in the normal rats (0.23μm; 0.035μm). Diabetic rat podocyte number (8; 1), slit diaphragm number (4; 1), and slit diaphragm extension (0.021μm; 0.00435μm) were significantly lower than in normal rats (11; 1) and (7; 1.5), and (0.031μm; 0.0058μm). Diabetic rat proteinuria (0.060mg/24h; 0.037mg/24h) was higher than in normal rats (0.00185mg/24h; 0.00055mg/24h). Conclusion: Experimental diabetes is associated with significant (p<0.05) changes in podocyte foot process, slit number, slit diaphragm extension, and GBM thickness.

Estudo multicêntrico Brasileiro - Adesivo contraceptivo transdérmico semanal: Preferência e satisfação das usuárias

Background: To assess satisfaction with the weekly transdermal contraceptive patch, compared to the previous contraceptive method. Adhesion, cycle control, safety and efficacy were secondary outcomes. Methods: A multicenter, open label, descriptive study. Subjects received the weekly transdermal contraceptive patch for 6 cycles. At the baseline and after the 1 st, 3 rd and 6 th cycles, satisfaction with the method was assessed. The contraceptive efficacy was assessed by the Pearl Index and by life table analysis. Results: An ITT analysis was performed with 494 subjects. At the endpoint, 95.3% of women were satisfied with the patch compared to the previous method; 59.5%, 58.0% and 63.2% of women reported improvement in the physical and emotional well-being, and pre-menstrual symptoms, respectively. Pre-menstrual symptoms frequency decreased from 85.0% (CI 95%: 80.8-87.4%) to 55.0% (CI 95%: 49.9-66.0%). There was a significant increase in hemoglobin levels, and also a significant decrease in total serum cholesterol and tryglicerides.. There were no significant adverse events. The adjusted Pearl Index was 1.1 pregnancies per 100 women-years. Conclusions: The weekly contraceptive transdermal patch is a safe and effective method, and there was a high rate of satisfaction with its use among brazilian women. The patch was associated to a good cycle control, with easiness of use, improvement in the physical and emotional well-being and improvement in the premenstrual symptoms and lipid profile. © Copyright Moreira Jr. Editora. Todos os direitos reservados.

Effects of acetylsalicylic acid and acetic acid solutions in VX2 carcinoma cells. In vitro analysis

Purpose: To analyze, in vitro, the effects of acetylsalicylic acid (aspirin) and acetic acid solutions on VX2 carcinoma cells in suspension and to examine the correlation between these effects and neoplastic cell death. Methods: The VX2 tumor cells (107 cells/ml) were incubated in solutions containing differing concentrations (2.5% and 5%) of either acetylsalicylic acid or acetic acid, or in saline solution (controls). Every five minutes, cell viability was tested (using the trypan blue test) and analyzed under light microscopy. Results: Tumor cell viability (in %) decreased progressively and, by 30 minutes, neoplastic cell death had occurred in all solutions. Conclusion: Based on this experimental model and the methodology employed, we conclude that these solutions cause neoplastic cell death in vitro.

Toxoplasma gondii: Comparison of a rhoptry-ELISA with IFAT and MAT for antibody detection in sera of experimentally infected pigs

Indirect ELISA and IFAT have been reported to be more sensitive and specific than agglutination tests. However, MAT is cheaper, easier than the others and does not need special equipment. The purpose of this study was to compare an enzyme linked immunosorbent assay using crude rhoptries of Toxoplasma gondii as coating wells (r-ELISA) with indirect fluorescence antibody test (IFAT) and modified agglutination test (MAT) to detect anti-T. gondii antibodies in sera of experimentally infected pigs. Ten mixed breed pigs between 6.5 and 7.5 weeks old were used. All pigs were negative for the presence of T. gondii antibodies by IFAT (titre < 16), r-ELISA (OD < 0.295) and MAT (titre < 16). Animals received 7 × 107 viable tachyzoites of the RH strain by intramuscular (IM) route at day 0. Serum samples were collected at days -6, 0, 7, 14, 21, 28, 35, 42, 50, and 57. IFAT detected anti-T. gondii antibodies earlier than r-ELISA and MAT. The average of antibody levels was higher at day 35 in IFAT (Log10 = 2.9) and in MAT (Log10 = 3.5), and at day 42 in r-ELISA (OD = 0.797). The antibody levels remained high through the 57th day after inoculation in MAT, and there was a decrease tendency in r-ELISA and IFAT. IFAT was used as gold standard and r-ELISA demonstrated a higher prevalence (73.3%), sensitivity (94.3%), negative predictive value (83.3%), and accuracy (95.6%) than MAT. Kappa agreements among tests were calculated, and the best results were shown by r-ELISA × IFAT (κ = 0.88, p < 0.001). Cross-reaction with Sarcocystis miescheriana was investigated in r-ELISA and OD mean was 0.163 ± 0.035 (n = 65). Additionally, none of the animals inoculated with Sarcocystis reacted positively in r-ELISA. Our results indicate that r-ELISA could be a good method for serological detection of T. gondii infection in pigs. © 2005 Elsevier Inc. All rights reserved.