Claudin-8 Expression in Madin-Darby Canine Kidney Cells Augments the Paracellular Barrier to Cation Permeation

Abstract

Claudins are a family of integral membrane proteins of the tight junction that are thought to participate in the permeation of solutes across epithelia via the paracellular pathway. Claudin-8 is expressed in the distal renal tubule, which has a characteristically low passive permeability to monovalent cations. To test the hypothesis that claudin-8 plays a role in forming a tight paracellular barrier to cations, stably transfected MadinDarby canine kidney II cell lines with inducible expression of claudin-8 were generated. Induction of claudin-8 expression was associated with down-regulation of endogenous claudin-2 protein. Other tight junction proteins were expressed and targeted normally, and the number of junctional strands was minimally altered. By Ussing chamber and radiotracer flux studies, claudin-8 expression was found to reduce paracellular permeability to monovalent inorganic and organic cations and to divalent cations but not to anions or neutral solutes. The size selectivity, charge dependence, and activation energy of paracellular cation permeation were all unchanged. These observations are consistent with a model in which claudin-2 encodes a highly cationpermeable channel, whereas claudin-8 acts primarily as a cation barrier. When exogenous claudin-8 is expressed, it replaces endogenous claudin-2, inserting in its place into existing tight junction strands, thereby reducing the apparent number of functional cation pores. Our findings suggest that claudin-8 plays an important role in the paracellular cation barrier of the distal renal tubule.

PPARγ‐regulated tight junction development during human urothelial cytodifferentiation

“…In MDCK cells, forced expression of claudin 2 is associated with decreased transepithelial resistance (Furuse et al, 2001), whereas overexpression of claudins 1, 4 and 8 resulted in an increased TER (McCarthy et al, 2000;Van Itallie et al, 2001;Yu et al, 2003). Furthermore, transfection of claudin 8 into MDCK cells was associated with a selective down-regulation of claudin 2 expression (Yu et al, 2003).…”

“…In MDCK cells, forced expression of claudin 2 is associated with decreased transepithelial resistance (Furuse et al, 2001), whereas overexpression of claudins 1, 4 and 8 resulted in an increased TER (McCarthy et al, 2000;Van Itallie et al, 2001;Yu et al, 2003). Furthermore, transfection of claudin 8 into MDCK cells was associated with a selective down-regulation of claudin 2 expression (Yu et al, 2003). It has been shown that claudin 8 acts as a cation barrier, whereas claudin 2 forms a cation-selective channel which is capable of transforming a barrier from 'tight' to 'leaky ' (Furuse et al, 2001;Amasheh et al, 2002).…”

Biology of claudins

“…For example, claudin-2, which forms high-conductance cation pores (4,34), is expressed in the proximal tubule and early thin descending limb (28,59) and is likely responsible for the substantial amount of paracellular Na ϩ reabsorption in these segments. In contrast, claudins-4 and -8 function primarily as cation barriers (102,116) and are expressed in the distal nephron (59,116), where they likely protect against the dissipation of transtubular Na ϩ , K ϩ , and H ϩ gradients established by transcellular active transport. In addition, claudins can undergo alternative splicing.…”

Claudin-4 Levels Are Associated with Intact Alveolar Fluid Clearance in Human Lungs

“…Although Intact tight junctions are required for AFC, differential expression of transmembrane tight junction proteins modulates epithelial paracellular permeability to ions and other solutes. [5][6][7][8][9][10][11] Foremost among these tight junction proteins is the claudin family. [12][13][14] In both types 1 and 2 pneumocytes, claudin-4 is expressed at high levels.…”