CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / De acordo com o MinistÃrio da SaÃde do Brasil, as serpentes do gÃnero Bothrops sÃo as principais envolvidas nos acidentes ofÃdicos no paÃs e a insuficiÃncia renal aguda (IRA) à uma complicaÃÃo grave dos envenenamentos por estas serpentes. Neste trabalho, foram investigados os efeitos renais do veneno total de Bothropoides pauloensis (VBp) em sistema de rim isolado de rato e em cultura de cÃlulas tubulares renais da linhagem MDCK (Madin-Darby Canine Kidney). Para perfusÃo de rim isolado foram utilizados ratos Wistar pesando entre 250 e 300g, cujos rins foram excisados cirurgicamente e perfundidos com soluÃÃo de Krebs-Henseleit contendo 6%p/v de albumina bovina previamente dialisada. Foram investigados os efeitos do VBp (3Âg/mL e 10 Âg/mL; n=6). Verificamos queda na pressÃo de perfusÃo (PP) na concentraÃÃo de 10μg/mL aos 90 e 120 min. A resistÃncia vascular renal (RVR) tambÃm diminuiu na concentraÃÃo de 10μg/mL aos 90 e 120 minutos e na concentraÃÃo de 3 μg/mL aos 120 min. O fluxo urinÃrio (FU) diminuiu aos 90 e 120 minutos em ambas as concentraÃÃes, nÃo apresentando diferenÃa significativa entre as concentraÃÃes no tempo de 120min. O ritmo de filtraÃÃo glomerular (RFG) apresentou uma reduÃÃo aos 60 minutos em ambas as concentraÃÃes, mantendo-se diminuÃda atà o final do experimento, nÃo havendo diferenÃa significativa entre as concentraÃÃes no tempo de 60min. O percentual de transporte tubular de sÃdio (%TNA+) foi reduzido em ambas as concentraÃÃes nos tempos de 60,90 e 120min, nÃo apresentando diferenÃa significativa entre as concentraÃÃes no tempo de 60 min; O transporte tubular de cloreto(%TCl-) tambÃm foi reduzido aos 60, 90 e 120min. Em ambas as concentraÃÃes; O transporte tubular de potÃssio (%TK+) diminuÃram aos 60, 90 e 120 minutos na concentraÃÃo de 10μg/mL, na concentraÃÃo de 3μg/mL reduziu aos 60 min,retornando a valores prÃximos ao controle no tempo de 90 e 120min. NÃo havendo diferenÃa significativa entre as concentraÃÃes nos tempos de 60 e 90 min. A anÃlise mostrou a presenÃa de alteraÃÃes morfolÃgicas significativas, como acÃmulo de proteÃnas a nÃvel tubular. Na cultura de cÃlulas MDCK, o VBp foi capaz de diminuir a viabilidade celular (CI50= 7,5μg/mL). Citometria de fluxo com anexina V e iodeto de propÃdio mostrou que a morte celular ocorreu predominantemente por apoptose. Tratamento com VBp (7,5 μg / mL) levou a despolarizaÃÃo significativa do potencial de membrana mitocondrial. Verificamos a geraÃÃo de EspÃcies Reativas de OxigÃnio. A anÃlise morfolÃgica das cÃlulas mostrou caracterÃsticas indicativas de apoptose. Esses resultados demonstram que o veneno de B. pauloensis alterou todos os parÃmetros renais avaliados na perfusÃo de rim isolado e foi citotÃxico para cultura de cÃlulas MDCK. / According to the Ministry of Health of Brazil, Bothrops are the main involved in snakebites in the country and acute kidney injury is a serious complication of poisoning by these snakes. In this work, we investigated the renal effects of total venom Bothropoidespauloensis (VBP) in isolated rat kidney system and in cultured renal tubular cell line MDCK (Madin - Darby Canine Kidney). For perfusion of isolated kidney Wistar rats were used weighing 250- 300g, whose kidneys were surgically excised and perfused with Krebs- Henseleit solution containing 6% w/v pre-dialysed bovine serum albumin. The effects of VBp (n = 3μg/mL and 10 mg/mL; n=6) was investigated. Reduction was observedin perfusion pressure (PP) at a concentration of 10μg/mL at 90 and 120 min. Renal vascular resistance (RVR) also decreased the concentration of 10μg/mL at 90 and 120 minutes and at a concentration of 3 mg/mL at 120 min. The urinary flow (FU) decreased at 90 and 120 minutes in both concentrations, no significant difference between the concentrations at the time of 120min. The glomerular filtration rate (GFR) was reduced to 60 minutes in both concentrations and remained decreased until the end of the experiment, no significant difference between the concentrations in the time of 60min. The percentage of tubular sodium transport (%TNA+) was reduced by both concentrations in times of 60, 90 and 120 min, with no significant difference between the concentrations at time 60 min. The tubular transport of chloride (%TCl-) was also reduced to 60, 90 and 120 min. In both concentrations; the tubular potassium transport (%TK+) decreased at 60, 90 and 120 minutes at a concentration of 10μg/mL in a concentration of 3μg/mL reduced to 60 min, returning to near control at 90 and 120min values. No significant difference between the concentrations in the times of 60 and 90 min. The analysis showed the presence of significant morphological changes such as the accumulation of proteins plate level in the culture of MDCK cells, the VBP was able to reduce cell viability (IC50 = 7.5 g/ mL). Flow cytometry with Annexin V and propidium iodide showed that cell death occurred by apoptosis predominantly. VBP treatment ( 7.5 mg/ml) led to a significant depolarization of the mitochondrial membrane potential. We verified the generation of Reactive Oxygen Species. Morphological analysis of the cells showed features indicative of apoptosis. These results demonstrate that the venom of B. pauloensis altered all renal parameters evaluated in the kidney perfusion was cytotoxic to isolated and MDCK cell culture.
| Identifer | oai:union.ndltd.org:IBICT/oai:www.teses.ufc.br:8099 |
| Date | 17 December 2013 |
| Creators | Aline Diogo Marinho |
| Contributors | Helena Serra Azul Monteiro, Nylane Maria Nunes de Alencar, Roberto CÃsar Pereira Lima JÃnior |
| Publisher | Universidade Federal do CearÃ, Programa de PÃs-GraduaÃÃo em Farmacologia, UFC, BR |
| Source Sets | IBICT Brazilian ETDs |
| Language | Portuguese |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, info:eu-repo/semantics/masterThesis |
| Format | application/pdf |
| Source | reponame:Biblioteca Digital de Teses e Dissertações da UFC, instname:Universidade Federal do Ceará, instacron:UFC |
| Rights | info:eu-repo/semantics/openAccess |
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