Low-salt diet induces claudin-3 expression and drives adaptive changes in collecting duct of claudin-3-deficient mice

Abstract Background Renal sodium reabsorption occurs via both transcellular and paracellular pathways. Tight junction proteins play a key role in mediating paracellular transport. The collecting duct (CD) is critical for the fine-tuning of Na + balance and is sensitive to changes in dietary salt intake. A low-sodium diet, which increases endogenous aldosterone secretion, stimulates transcellular sodium transport via epithelial Na + channels (ENaC) and Na,K-ATPase. We hypothesized that a low-sodium diet also modulates paracellular Na + permeability by regulating the expression or function of claudin-3, a major tight junction protein in the CD, in order to limit the back-leak of reabsorbed sodium and preserve sodium balance. Methods We used in vivo mouse models and cultured mouse CD principal cells (mCCD cl1 ) to assess aldosterone’s effects on tight junction proteins. In mCCD cl1 cells, aldosterone-induced changes in claudin-3 expression and localization were evaluated via Western blotting and immunofluorescence, and Ussing chamber assays were used to assess paracellular Na + and Cl − permeability after modulating claudin-3 expression. Wild-type and claudin-3 knockout mice were fed low (0.01%) or normal (0.18%) sodium diets for seven days. In subsets of low sodium diet mice, spironolactone (a mineralocorticoid receptor antagonist) was administered. Results In mice, a low-sodium diet upregulates renal claudin-3 expression. Concordantly, in vitro studies using mCCD cl1 cells showed that aldosterone treatment increased claudin-3 protein levels and promoted its localization to the lateral membrane. Functional analyses demonstrated that claudin-3 overexpression reduced paracellular permeability to both Na + and Cl − , while claudin-3 silencing increased it. Claudin-3 knockout mice subjected to a low-sodium diet exhibited compensatory upregulation of the α- and γ-subunits of ENaC, alongside increased expression of claudin-4, claudin-8, and claudin-10. This highlights an adaptive response that maintains sodium homeostasis in the absence of claudin-3. Importantly, this compensatory mechanism persists even under spironolactone treatment, suggesting that the adaptation of claudin-3-deficient mice occurs independently of mineralocorticoid receptor activation. Conclusions Our findings demonstrate that aldosterone enhances claudin-3 expression, reinforcing the paracellular barrier to Na + and complementing its classical role in transcellular Na + transport. Under low-sodium conditions, claudin-3-deficient mice adapt through complementary mechanisms aimed at increasing sodium reabsorption via ENaC activation and upregulation of claudin-4 and claudin-8, both barrier-forming claudins that restrict paracellular sodium leakage in the CD. This is associated with increased claudin-10 abundance in the thick ascending limb of Henle, a pore-forming claudin that facilitates paracellular sodium permeability. This study advances our understanding of the complex control of renal sodium handling, revealing adaptive mechanisms in response to low-salt diet and claudin-3 deficiency.