Structure-Function Relationships and Pathophysiological Significance of Tight Junction Claudin Proteins

Renal epithelial cells are actively involved in transporting water and solutes, as well as in homeostatic regulation. Trans-epithelial transport of substances is by either transcellular or paracellular pathways. Paracellular permeability is thought to be tightly associated with tight junctions (TJs). TJs are found in the most apical portion of the lateral membrane of epithelial cells and are made up of transmembrane proteins, cytoplasmic scaffold proteins, and signaling proteins. Transmembrane proteins, through their extracellular domains which extend into the intercellular space, help to regulate the movement of solutes via the paracellular route and are thought to be the primary determinants of paracellular permeability. Claudins are the most abundant transmembrane proteins in TJs.

Claudin family proteins have four transmembrane domains, and both N- and C-termini are on the intracellular side. This results in two extracellular loops of unequal size and a short intracellular loop. Claudins can be roughly divided into two major functional groups on the basis of their regulatory effect on permeability. The first group, best described as a "barrier", seals the paracellular pathway and limits free diffusion of molecules. Members of this group include Claudin-1, -3 and -5. The second group acts as selective "channels" through which some ions (usually cations or anions) can pass. Examples include Claudin-2, -4, -10, and -16.

Distribution of Claudins in the Kidney

Members of the Claudin family have characteristic expression and distribution patterns in different tissues and cells, as well as in different areas of the same tissue. Claudin genes are expressed in a segmental pattern in the kidney in specific nephron segments with each nephron segment expressing one or more Claudin proteins.

For instance, Claudin-2 is highly expressed in the proximal tubule. It is located mostly in the late part of proximal tubule and the early part of the descending thin limb of the loop of Henle. Claudin-2 forms a cation channel and mediates reabsorption of cations such as Na⁺ and K⁺. Claudin-4 and Claudin-8 are expressed in the distal tubule. Claudin-4 and Claudin-8 form a cation barrier that prevents the diffusion of Na⁺, K⁺ and H+ between the cells in the renal tubules. Claudin-4 and Claudin-8 thus help to maintain the cation concentration gradients needed for the transport of substances across the renal tubules (transcellular transport). In addition, Claudins can be alternatively spliced to produce Claudin-10 isoforms that have different extracellular domains and functional characteristics. Claudin-10a is anion selective and is expressed in the renal proximal tubule. Claudin-10b is very cation-selective, and is expressed in the medulla. Claudin-16 and -19 are found in the thick ascending limb of the loop of Henle where they mediate paracellular reabsorption of divalent cations and contribute to the positive luminal potential.

Claudins are not restricted to epithelial cells of the renal tubules; they have also been found in other parts of the kidney. For example, Claudin-7 is present in the connecting tubule and collecting duct. Claudin-9 is exclusively located in the proximal tubule, while Claudin-5 and Claudin-15 are primarily distributed in the renal vascular endothelium. Claudin-1 is found in renal glomerular epithelial cells, among others.

Figure 1. A schematic diagram of the distribution of certain Claudins in rodent nephrons
(Source: Milatz S, et al. 2017)

Claudin and Kidney Disease

The human Claudin-10 gene is located on chromosome 13q31-q34 and contains 5 exons. Within the first exon, there are 2 Claudin-10 splice variants that encode 2 major Claudin-10 isoforms: Claudin-10a and Claudin-10b. Research teams have described the clinical manifestations of biallelic mutations in the Claudin-10 gene, with the acronym HELIX (Hypohidrosis, Electrolyte disturbances, hypoLacrimia, Ichthyosis, Xerostomia) used to name this syndrome. All HELIX syndrome patients have functional defects in sweat glands, salivary glands, and lacrimal glands, with most patients exhibiting blood electrolyte abnormalities, hypermagnesemia and/or hypokalemia. Gene knockout experiments have shown that Claudin-10b specifically increases paracellular permeability to Na⁺. Indeed, the reduced paracellular permeability to Na⁺ in the thick ascending limb (TAL), sweat glands, and salivary glands may be the primary clinical consequence of loss-of-function mutations in Claudin-10b. In the renal medullary TAL, 50% of Na⁺ is passively reabsorbed through the paracellular pathway, and decreased paracellular permeability to Na⁺ affects Na⁺ reabsorption, leading to renal Na⁺ loss, extracellular fluid depletion, secondary aldosteronism, and renal K⁺ loss.

FHHNC is an autosomal recessive genetic disease caused by mutations in the Claudin-16 and Claudin-19 genes, characterized by massive loss of calcium and magnesium in urine, resulting in hypomagnesemia and hypercalciuria. Claudin-16 and Claudin-19 are expressed exclusively in the TAL and play crucial roles in paracellular transport of Ca²⁺ and Mg²⁺ in the TAL. When Claudin-16 is independently expressed in low-resistance LLC-PK1 cells, it can significantly increase paracellular Na⁺ permeability and moderately increase Mg²⁺ permeability, thereby changing selectivity from anion selectivity to slight cation selectivity. Expression of Claudin-19 in this cell line also eliminates anion selectivity, but in this case by reducing Cl^- permeability. Co-expression of Claudin-16 and Claudin-19 results in more significant changes in Na⁺ and Cl^- permeability than their individual expression levels, indicating functional synergy between these two Claudins.

Genome-wide association studies have confirmed that Claudin-14 is a major risk gene for hypercalciuric nephrolithiasis. According to genome-wide standards, synonymous mutations at the Claudin-14 gene locus are associated with increased kidney stone risk and decreased bone density. Homozygous patients with the common synonymous variant rs219780 (C) have an estimated 1.64-fold increased risk of developing kidney stones compared to non-carriers. Overexpression of Claudin-14 in various cell lines can reduce the Na⁺/Cl^- permeability ratio as well as Ca²⁺ permeability. For example, in transfected LLC-PK1 cells, Claudin-14 can reduce the extracellular cation permeability of Claudin-16 through physical interaction.

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