S Nag1, J L Manias1, D J Stewart2. 1. Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, and. 2. Ottawa Health Research Institute, University of Ottawa, Ottawa, ON, Canada.
Abstract
AIMS: Increased endothelial caveolae leading to transcytosis of plasma proteins is associated with blood-brain barrier (BBB) breakdown and cerebral oedema in brain injury. Increased expression of caveolin-1alpha (Cav-1), an integral caveolar membrane protein, was reported in endothelium of arterioles and veins with BBB breakdown to fibronectin post injury. In this study the phosphorylation state of Cav-1 and its association with BBB breakdown was determined in the rat cortical cold injury model over a period of days 0.5-6 post lesion. METHODS: Expression of phosphorylated Cav-1 was determined by immunoblotting and dual labelling immunofluorescence for phosphorylated caveolin-1 and fibronectin, a marker of BBB breakdown. A phospho-specific monoclonal antibody that selectively recognizes only tyrosine 14-phosphorylated Cav-1 (PY14Cav-1) was used. RESULTS: Immunoblots showed constitutive expression of PY14Cav-1 in cortex of control rats and a significant increase in PY14Cav-1 expression at the lesion site up to day 4 post lesion. PY14Cav-1 immunostaining was observed in the endothelium of lesion vessels at days 0.5-4 post lesion, in neutrophils at days 0.5 and 2 and in macrophages at day 6 post lesion. Dual labelling showed that 100% of vessels with BBB breakdown to fibronectin showed endothelial PY14Cav-1 on day 0.5, the percentage decreasing to 62% on day 4. On day 6, none of the vessels showed endothelial phosphorylated Cav-1. CONCLUSIONS: The presence of phosphorylated Cav-1 in endothelium of vessels showing BBB breakdown suggests that phosphorylated Cav-1 signalling may be one of the factors associated with early BBB breakdown and brain oedema in brain injury.
AIMS: Increased endothelial caveolae leading to transcytosis of plasma proteins is associated with blood-brain barrier (BBB) breakdown and cerebral oedema in brain injury. Increased expression of caveolin-1alpha (Cav-1), an integral caveolar membrane protein, was reported in endothelium of arterioles and veins with BBB breakdown to fibronectin post injury. In this study the phosphorylation state of Cav-1 and its association with BBB breakdown was determined in the rat cortical cold injury model over a period of days 0.5-6 post lesion. METHODS: Expression of phosphorylated Cav-1 was determined by immunoblotting and dual labelling immunofluorescence for phosphorylated caveolin-1 and fibronectin, a marker of BBB breakdown. A phospho-specific monoclonal antibody that selectively recognizes only tyrosine 14-phosphorylated Cav-1 (PY14Cav-1) was used. RESULTS: Immunoblots showed constitutive expression of PY14Cav-1 in cortex of control rats and a significant increase in PY14Cav-1 expression at the lesion site up to day 4 post lesion. PY14Cav-1 immunostaining was observed in the endothelium of lesion vessels at days 0.5-4 post lesion, in neutrophils at days 0.5 and 2 and in macrophages at day 6 post lesion. Dual labelling showed that 100% of vessels with BBB breakdown to fibronectin showed endothelial PY14Cav-1 on day 0.5, the percentage decreasing to 62% on day 4. On day 6, none of the vessels showed endothelial phosphorylated Cav-1. CONCLUSIONS: The presence of phosphorylated Cav-1 in endothelium of vessels showing BBB breakdown suggests that phosphorylated Cav-1 signalling may be one of the factors associated with early BBB breakdown and brain oedema in brain injury.
Authors: Max Franzblau; Chiara Gonzales-Portillo; Gabriel S Gonzales-Portillo; Theo Diamandis; Mia C Borlongan; Naoki Tajiri; Cesar V Borlongan Journal: Med Hypotheses Date: 2013-09-17 Impact factor: 1.538