Astrocytes and pericytes attenuate severely injured patient plasma mediated expression of tight junction proteins in endothelial cells.

Abstract

Blood Brain Barrier (BBB) breakdown is a secondary form of brain injury which has yet to be fully elucidated mechanistically. Existing research suggests that breakdown of tight junction proteins between endothelial cells is a primary driver of increased BBB permeability following injury, and intercellular signaling between primary cells of the neurovascular unit: endothelial cells, astrocytes, and pericytes; contribute to tight junction restoration. To expound upon this body of research, we analyzed the effects of severely injured patient plasma on each of the cell types in monoculture and together in a triculture model for the transcriptional and translational expression of the tight junction proteins Claudins 3 and 5, (CLDN3, CLDN5) and Zona Occludens 1 (ZO-1). Conditioned media transfer studies were performed to illuminate the cell type responsible for differential tight junction expression. Our data show that incubation with 5% human ex vivo severely injured patient plasma is sufficient to produce a differential response in endothelial cell tight junction mRNA and protein expression. Endothelial cells in monoculture produced a significant increase of CLDN3 and CLDN5 mRNA expression, (3.98 and 3.51 fold increase vs. control respectively, p<0.01) and CLDN5 protein expression, (2.58 fold change vs. control, p<0.01), whereas in triculture, this increase was attenuated. Our triculture model and conditioned media experiments suggest that conditioned media from astrocytes and pericytes and a triculture of astrocytes, pericytes and endothelial cells are sufficient in attenuating the transcriptional increases of tight junction proteins CLDN3 and CLDN5 observed in endothelial monocultures following incubation with severely injured trauma plasma. This data suggests that inhibitory molecular signals from astrocytes and pericytes contributes to prolonged BBB breakdown following injury via tight junction transcriptional and translational downregulation of CLDN5.