Tricellulin expression in brain endothelial and neural cells

Tricellulin is a tight junction (TJ) protein, which is not only concentrated at tricellular contacts but also present at bicellular contacts between epithelial tissues. We scrutinized the brain for tricellulin expression in endothelial and neural cells by using real-time polymerase chain reaction, Western blot and immunohistochemical and immunocytochemical analysis of cultured brain cells and paraffin sections of brain. Tricellulin mRNA was detected in primary cultures and in a cell line of human brain microvascular endothelial cells. Protein expression was confirmed by Western blot and immunofluorescence analysis, which further highlighted the localization of tricellulin in the cell membrane at tricellular and along bicellular contacts, and in the nucleus and perinuclear region. Compared with the well-studied TJ protein, zonula occludens-1, tricellulin expression was less marked at the cell membrane but more evident in the nuclear and perinuclear regions. The presence of tricellulin in cultured endothelial cells was corroborated by immunohistochemical and immunofluorescence staining in brain blood vessels, where it was colocalized with another TJ protein, claudin-5. Tricellulin mRNA was detected in neurons and astrocytes, whereas protein expression was observed in astrocytes but not in neurons, as shown by immunofluorescence analysis. This study reveals the presence and subcellular distribution of tricellulin in brain endothelial cells, both in vitro and in situ and its colocalization with other relevant TJ proteins. Moreover, it demonstrates the expression of the protein in astrocytes opening new avenues for future research to establish the biological significance of tricellulin expression in glial cells.