Avaliação dos efeitos renais da fração L-aminoácido Oxidase isolada do veneno da serpente Bothrops marajoensis

• Autor(a) principal: Dantas, Rodrigo Tavares
• Data de Publicação: 2010
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Repositório Institucional da Universidade Federal do Ceará (UFC)
• Texto Completo: http://www.repositorio.ufc.br/handle/riufc/2628
• topic: L-Aminoácido Oxidase; Bothrops

There are about 3.000 species of snakes worldwide, but only 10 to 14% are venomous. Among South American countries, Brazil is the one with the largest number of accidents, with approximately 20.000 snakebites each year. According to the Department of Health of Brazil, the genus Bothrops are the main involved in snakebites in the country. Acute renal failure (ARF) is a serious complication of snake poisoning. The fraction L-amino acid oxidase (LAAO) constitutes a main part of the total composition of snake venoms. In some cases this amount can reach 30% of total venom proteins. The renal effects of fraction L-amino acid oxidase isolated from the venom of Bothrops marajoensis (LAAOBM) was investigated in this study. Isolated perfused rat kidney and the cultured renal tubular cells line MDCK (Madin-Darby Canine Kidney) were used here. For the isolated perfused rat kidney method, we used Wistar rats weighing between 250 and 300. Their right kidneys were surgically excised and perfused with Krebs-Hanseleit containing 6% w/v bovine albumin previously dialyzed. The effects of LAAOBM (10 mg/mL, n=4) were analyzed on the Perfusion Pressure (PP), Renal Vascular Resistance (RVR), Urinary Flow (UF), Glomerular Filtration Rate (GFR) Percentage of Sodium Proximal Tubular Transport (%pTNa+), Percentage of Sodium (%TNa+), Potassium (%TK+) and Chloride (%TCl-) Tubular Transport. MDCK cells were cultured in RPMI 1640 medium supplemented with 10% v/v fetal bovine serum and incubated with LAAOBM at concentrations of 50, 25, 12.5, 6.25, 3.125 and 1.652 mg/mL. After 24 hours of incubation, assays were performed on cell proliferation and viability using the MTT method. The LAAOBM promoted a reduction of perfusion pressure at 90 minutes of the experiment and this reduction was even slightly higher at 120 minutes. It was also observed decrease in renal vascular resistance at 120 minutes. There was a sharp and sudden drop in urine flow at 90 minutes, despite the tendency of recovery observed at 120 minutes, still proved to be quite small when compared to the control group. The infusion of LAAOBM also promoted a reduction in glomerular filtration rate at 90 minutes compared to the control group and this parameter still remained at the same level of reduction at 120 minutes. The LAAOBM gradually reduced the percentage of sodium tubular transport at 90 and 120 minutes and the percentage of chloride tubular transport in the periods of 60, 90 and 120 minutes. Histological analysis of kidneys perfused with LAAOBM showed the presence of significant morphological changes such as accumulation of proteins in tubular and glomerular spaces. In the MDCK cell culture LAAOBM promoted a reduction in cell viability from concentrations of 3.25 mg / mL until 50μg/mL, with IC50 value of 2.43 mg/mL. Inverted light microscopy showed morphological changes of these cells, such as vacuolation, alteration of the state of confluence and detachment of the substrate culture. These results demonstrated that LAAOBM changed all the parameters evaluated in renal and vascular perfusion of isolated kidney and had cytotoxic activity on MDCK cells after 24 hours of incubation.