Contribution of Tight Junction Proteins to Ion, Macromolecule, and Water Barrier in Keratinocytes

Tight junctions (TJs) form a selective barrier for ions, water, and macromolecules in simple epithelia. In keratinocytes and epidermis, -Os were shown to be involved in individual barrier functions. The absence of the Ti protein claudin-1 (Cldn1) in mice results in a skin-barrier defect characterized by lethal water loss. However, detailed molecular analyses of the various Ti barriers in keratinocytes and the contribution of distinct Ti proteins are missing. Herein, we discriminate Ti-dependent paracellular resistance from transcellular resistance in cultured keratinocytes using the two-path impedance spectroscopy. We demonstrate that keratinocyte Tis form a barrier for Na+ Cl- and Ca2+ and contribute to barrier function for water and larger molecules of different size. In addition, knockdown of Cldn1, Cldn4, occludin, and zonula occludens-1 increased paracellular permeabilities for ions and larger molecules, demonstrating that all of these Ti proteins contribute to barrier formation. Remarkably, Cldn1 and Cldn4 are not critical for Ti barrier function for water in submerged keratinocyte cultures. However, Cldn1 influences stratum corneum (SC) proteins important for SC water barrier function, and is crucial for Ti barrier formation for allergen-sized macromolecules.