Correlation of tight junction morphology with the expression of tight junction proteins in blood-brain barrier endothelial cells

Endothelial cells of the blood-brain barrier form complex tight junctions, which are more frequently associated with the protoplasmic (P-face) than wi th the exocytoplasmic (E-face) membrane leaflet. The association of tight j unctional particles with either membrane leaflet is a result of the express ion of various claudins, which are transmembrane constituents of tight junc tion strands. Mammalian brain endothelial tight junctions exhibit an almost balanced distribution of particles and lose this morphology and barrier fu nction in vitro. Since it was shown that the brain endothelial fight juncti ons of submammalian species form P-face-associated tight junctions of the e pithelial type, the question of which molecular composition underlies the m orphological differences and how do these brain endothelial cells behave in vitro arose. Therefore, rat and chicken brain endothelial cells were inves tigated for the expression of junctional proteins in vivo and in vitro and for the morphology of the tight junctions. In order to visualize morphologi cal differences, the complexity and the P-face association of tight junctio ns were quantified. Rat and chicken brain endothelial cells form tight junc tions which are positive for claudin-1, claudin-5, occludin and ZO-1. In ag reement,vith the higher P-face association of tight junctions in vivo, chic ken brain endothelia exhibited a slightly stronger labeling for claudin-1 a t membrane contacts. Brain endothelial cells of both species showed a signi ficant alteration of tight junctions in vitro, indicating a loss of barrier function. Rat endothelial cells showed a characteristic switch of tight ju nction particles from the P-face to the E-face, accompanied by the loss of claudin-1 in immunofluorescence labeling. In contrast, chicken brain endoth elial cells did not show such a switch of particles, although they also los t claudin-1 in culture. These results demonstrate that the maintenance of r at and chicken endothelial barrier function depends on the brain microenvir onment. Interestingly, the alteration of tight junctions is different in ra t and chicken. This implies that the rat and chicken brain endothelial tigh t junctions are regulated differently.