Análises proteômicas e toxicológicas do veneno de Bothrops itapetiningae

Abstract

Venomous organism’s ranged a wide distribution in all over animal kingdom. The snakes are not different, belonging to the suborder Ophidia and Squamata Order. Inside them, Bothrops genus is responsible for 90% of accidents in Latin America. The envenoming by snakes from Bothrops genus are commonly characterized by local pain, systemic bleeding, tachycardia, acute renal failure, as well as swelling and redness caused by proteolytic and other activities of venom. Among different species from Bothrops genus it was included Bothrops itapetiningae, which consists in the smallest pitviper known in Brazil. In order to clarify a little more about venom composition we start to elucidate the proteic composition of an adult individual of Bothrops itapetiningae from Cerrado of central Brazil. The venom was manually extracted by stimulation of the venom gland. The venom was lyophilized and stored at -20oC until using. Initially, unidirectional polyacrylamide gel (12.5%) (SDS-PAGE) analyses were conducted in order to evaluate protein content and protein pattern of B. itapetiningae venom such as the sample quality. Results showed several bands spread at unidirectional gel. The venom of Bothrops itapetiningae was subsequently subjected to proteomic analysis using two-dimensional electrophoresis (2DE) following analysis of the Bionumerics® software, where the two-dimensional gel of B. itapetiningae showed several spots distributed throughout all the gel, pIs and molecular mass of protein spots ranging between 6-180 kDa and 3-11 pI respectively. Subsequent analysis on Bionumerics® software showed a R2 of 0.90. Toxicological analysis were obtained in three groups, where, the first group received 100μl of NaCl 0.9%, the second one 0.7 μg.g-1 and the last one received 1.05 μg.g-1 of dried weight of B. itapetiningae venom. These analyses showed polymorphonuclear infiltration, hydropic degeneration and ischemic necrosis in the liver, kidney and skeletal muscle, watched in histological sections of the tissues in the microscope. Concentrations of alanine aminotransferase (ALT), aspartate aminotransferase, alkaline phosphatase and creatinin were measured in plasma indicating an increase of all enzymes after the venom injection.