Quantitative analysis of microvascular alterations in traumatic brain injury by endothelial barrier antigen immunohistochemistry

Endothelial barrier antigen (EBA) is a protein triplet located in the plasm a membrane of microvascular endothelium and selectively expressed in the no rmal nervous system. In this study, microvascular alterations following tra umatic brain injury were studied using EBA immunohistochemistry. Anesthetiz ed, physiologically regulated, normothermic Sprague-Dawley rats received mo derate (1.5-2.0 atm) parieto-occipital parasagittal fluid-percussion trauma tic brain injury (TBI). Control rats were subjected to similar anesthesia a nd physiological monitoring. Seven days after operative procedures, brains were perfusion-fixed, and coronal sections were reacted for EBA immunohisto chemistry using a monoclonal antibody to rat EBA. Selected sections were re acted for isolectin B-4 histochemistry. Computerized image analysis was use d to compute numbers of EBA-immunopositive vascular profiles and mean vascu lar profile areas. In control brains, virtually all brain microvessels were clearly and positively immunostained, acid antibody binding was specific f or blood vessels. In rats with TBI, EBA immunoreactivity was greatly reduce d in the zone of cortical contusion. Within the core contusion, fractional areas occupied by vascular profiles were markedly reduced (on average, by 5 7%), vascular profile counts were diminished, and lectin histochemistry rev ealed a robust inflammatory response with abundant macrophages. Taken toget her, these findings were thought to indicate frank microvascular destructio n. At adjacent peri-contusional sites, the intensity of EBA immunostaining was also diminished; and vascular profile counts were reduced at adjacent c ortical sites and homologous contralateral sites. The latter findings were interpreted as sublethal microvascular alterations possibly related to cere bral edema. The present results confirm that EBA is a specific immunohistoc hemical marker of normal central nervous system microvessels; that it is su itable for use in formaldehyde-fixed material; and that it is useful in qua ntitatively assessing microvascular alterations observed at contusional, pe ri-contusional and more remote sites following traumatic brain injury.