Effect of RNA interference of tight junction-related molecules on intercellular barrier function in cultured human keratinocytes.
Accumulating evidence shows that tight junctions (TJs) in the granular layer contribute to the epidermal barrier, suggesting that the regulation of TJ assembly in keratinocytes may provide a clue to understanding the role of barrier formation in epidermis. In this study, we investigated the behavior of TJ-related molecules in cultured human keratinocytes during keratinization induced by transfer to high-calcium medium, and the effect of RNA interference of TJ-related molecules on intercellular permeability and morphological features. The expression of TJ-related molecules and transepithelial electrical resistance were increased by transfer to high-calcium medium. In cells under the same conditions, we observed by freeze-fracture electron microscopy that TJ strands developed on the apposing cell membranes. In contrast, the transepithelial electrical resistance was clearly suppressed when the expression of claudin-1 or occludin was blocked by RNA interference. The morphological features of these knock-down cells were the same as those of MOCK cells, except for a marked decrease in the number of TJ strands. Furthermore, claudin-1 suppression inhibited occludin localization at the cell membrane, whereas suppression of occludin did not influence the localization of claudin-1. These results suggest that claudin-1 plays a crucial role in recruiting occludin to TJs, and that occludin is involved in intercellular barrier function.
Yamamoto
Takuya
T
Department of Dermatology, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15, Kuramoto, Tokushima, 770-8503, Japan.
Saeki
Yuko
Y
Kurasawa
Masumi
M
Kuroda
Shohei
S
Arase
Seiji
S
Sasaki
Hiroyuki
H
D002118:Calcium / Q000378:metabolism; D002450:Cell Communication / Q000502:physiology; D002463:Cell Membrane Permeability / Q000502:physiology*; D002478:Cells, Cultured; D062445:Claudin-1; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D062793:Occludin; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference / Q000502:physiology*; D019108:Tight Junctions / Q000378:metabolism*; D062826:Zonula Occludens-1 Protein
C568108:CLDN1 protein, human; D062445:Claudin-1; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; D010750:Phosphoproteins; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; D002118:Calcium