Received for publication November 26, 2007.
Revised May 1, 2008.
Accepted for publication May 2, 2008.

PKC enhances tight junction barrier function of human nasal epithelial cells in primary culture by transcriptional regulation

Abstract

The epithelium of upper respiratory tissues such as human nasal mucosa forms a continuous barrier via tight junctions, which is in part thought to be regulated through a protein kinase C (PKC) signaling pathway. To investigate the mechanisms of the regulation of PKC-mediated tight junction barrier function of human nasal epithelium in detail, primary human nasal epithelial cells were treated with the PKC activator 12-O-tetradecanoylophorbol-13-acetate (TPA). In primary human nasal epithelial cells, treatment with TPA led to not only activation of phosphorylation of PKC, myristoylated alanine-rich C kinase substrate (MARCKS) and mitogen-activated protein kinase (MAPK) but also expression of novel PKC-, PKC- and PKC-. Treatment with TPA increased transepithelial electrical resistance (TER), with tight junction barrier function more than four-fold that of the control, together with upregulation of tight junction proteins, occludin, ZO-1, ZO-2 and claudin-1 at the transcriptional level. Furthermore, it affected the subcellular localization of the tight junction proteins and the numbers of tight junction strands. The upregulation of barrier function and tight junction proteins was prevented by a panPKC inhibitor, and the inhibitors of PKC-, PKC- but not PKC-. In primary human nasal epithelial cells, transcriptional factors GATA-3 and -6 were detected by RT-PCR. The knockdown of GATA-3 using RNAi resulted in inhibition of upregulation of ZO-1 and ZO-2 by treatment with TPA. These results suggest that TPA induced PKC signaling enhances the barrier function of human nasal epithelial cells via transcriptional upregulation of tight junction proteins and the mechanisms may contribute to a drug delivery system.

Key words: Protein Kinase C, MAP Kinase, P38 MAP Kinase, Immunocytochemistry, Regulation - transcriptional, RNA/siRNA