Gyroxin increases blood-brain barrier permeability to Evans blue dye in mice

Abstract

Gyroxin is a serine protease enzyme component of the South American rattlesnake (Crotalus durissus terrificus) venom. This toxin displays several activities, including the induction of blood coagulation (fibrinogenolytic activity), vasodilation and neurotoxicity, resulting in an effect called barrel rotation. The mechanisms involved in this neurotoxic activity are not well known. Because gyroxin is a member of a potentially therapeutic family of enzymes, including thrombin, ancrod, batroxobin, trypsin and kallicrein, the identification of the mechanism of gyroxin's action is extremely important. In this study, gyroxin was isolated from crude venom by affinity and molecular exclusion chromatography. Analysis of the isolated gyroxin via sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed a single protein band with a molecular weight of approximately 28 kDa, confirming the identity of the molecule. Furthermore, intravenous administration of purified gyroxin (0.25 μg/g of body weight) to mice resulted in symptoms compatible with barrel rotation syndrome, confirming the neurotoxic activity of the toxin. Mice treated with gyroxin showed an increase in the concentration of albumin-Evans blue in brain extracts, indicating an increase in the blood-brain barrier (BBB) permeability. This gyroxin-induced increase in BBB permeability was time-dependent, reaching a peak within 15 min after exposure, similar to the time span in which the neurotoxic syndrome (barrel rotation) occurs. This work provides the first evidence of gyroxin's capacity to temporarily alter the permeability of the BBB.

Evans Blue Dye: A Revisit of Its Applications in Biomedicine

“…In addition, Soubeyrand et al [ 70 ] and Pettersson et al [ 71 ] used EB dye to measure the permeability of the blood-spinal cord barrier and Xu et al [ 72 ] detected the blood-retinal barrier breakdown by quantification using EB. It is worth noting that the stain administration route by intraperitoneal and intravenous injection had no difference on the amount of EB dye stain accumulated but a strong time-dependent tendency towards increase in stain accumulation [ 73 ]. As a rapid and inexpensive method, EB dye is still the most commonly used marker of brain barrier integrity and vascular permeability; however, its associated problems of tissue discoloration and potential toxicity need to be considered.…”

Silencing of the miR-17∼92 Cluster Family Inhibits Medulloblastoma Progression

“…Evans blue is a dye with a high affinity for serum albumin not normally present in the brain (28, 29). Mice that received Evans blue were euthanized by cardiac perfusion with 4% paraformaldehyde, 24 hours later.…”

Effects of global warming on reproduction and potential dispersal of Mediterranean Cnidarians

“…Actually, the progressive warming of the Mediterranean shallow waters is influencing the delicate coexistence of boreal, temperate, sub-tropical and tropical species (Moschella 2008;Corriero et al 2015;Longobardi et al 2017). Many of these deep environmental changes and its repercussions on single species or whole ecosystem functioning have been highlighted, such as the more frequent occurrence of harmful algal blooms (Mangialajo et al 2010;Faimali et al 2012;Privitera et al 2012), mass mortalities of sponges and anthozoans in shallow waters (Cerrano et al 2000;Linares et al 2005;Garrabou et al 2009;Rivetti et al 2014;Parravicini et al 2015), and changes in biodiversity of Mediterranean communities (Puce et al 2009;Gatti et al 2015;Gravili et al 2015a;Betti et al 2017a, and references therein).…”