The blood-brain barrier after stroke: Structural studies and the role of transcytotic vesicles

Blood-brain barrier breakdown worsens ischaemic damage, but it is unclear how molecules breach the blood-brain barrier in vivo. Using the obese ob/ob mouse as a model of enhanced blood-brain barrier breakdown, we investigated how stroke-induced structural changes to the microvasculature related to blood-brain barrier permeability. Ob/ob mice underwent middle cerebral artery occlusion, followed by 4 or 24 h reperfusion. Blood-brain barrier integrity was assessed using IgG and horseradish peroxidase staining, and blood-brain barrier structure by two-dimensional and three-dimensional electron microscopy. At 4 and 24 h post-stroke, ob/ob mice had increased ischaemic damage and blood-brain barrier breakdown compared to ob/- controls, and vessels from both genotypes showed astrocyte end-foot swelling and increased endothelial vesicles. Ob/ob mice had significantly more endothelial vesicles at 4 h in the striatum, where blood-brain barrier breakdown was most severe. Both stroke and genotype had no effect on tight junction structure visualised by electron microscopy, or protein expression in isolated microvessels. Astrocyte swelling severity did not correlate with tissue outcome, being unaffected by genotype or reperfusion times. However, the rare instances of vessel lumen collapse were always associated with severe astrocyte swelling in two-dimensional and three-dimensional electron microscopy. Endothelial vesicles were therefore the best spatial and temporal indicators of blood-brain barrier breakdown after cerebral ischaemia.