Abstract

The epithelium of upper respiratory tissues such as human nasal mucosa forms a continuous barrier via tight junctions, which is thought to be regulated in part 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 not only to activation of phosphorylation of PKC, myristoylated alanine-rich C kinase substrate, and mitogen-activated protein kinase but also expression of novel PKC-delta, PKC-theta, and PKC-epsilon. Treatment with TPA increased transepithelial electrical resistance, with tight junction barrier function more than 4-fold that of the control, together with up-regulation of tight junction proteins, occludin, zona occludens (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 up-regulation of barrier function and tight junction proteins was prevented by a pan-PKC inhibitor, and the inhibitors of PKC-delta and PKC-theta but not PKC-epsilon. In primary human nasal epithelial cells, transcriptional factors GATA-3 and -6 were detected by reverse transcription-polymerase chain reaction. The knockdown of GATA-3 using RNA interference resulted in inhibition of up-regulation 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 up-regulation of tight junction proteins, and the mechanisms may contribute to a drug delivery system.