Claudin-7 Modulates Cl− and Na+ Homeostasis and WNK4 Expression in Renal Collecting Duct Cells

Claudin-7 knockout (CLDN7−/−) mice display renal salt wasting and dehydration phenotypes. To address the role of CLDN7 in kidneys, we established collecting duct (CD) cell lines from CLDN7+/+ and CLDN7−/− mouse kidneys. We found that deletion of CLDN7 increased the transepithelial resistance (TER) and decreased the paracellular permeability for Cl− and Na+ in CLDN7−/− CD cells. Inhibition of transcellular Cl− and Na+ channels has no significant effect on TER or dilution potentials. Current-voltage curves were linear in both CLDN7+/+ and CLDN7−/− CD cells, indicating that the ion flux was through the paracellular pathway. The impairment of Cl− and Na+ permeability phenotype can be rescued by CLDN7 re-expression. We also found that WNK4 (its mutations lead to hypertension) expression, but not WNK1, was significantly increased in CLDN7−/− CD cell lines as well as in primary CLDN7−/− CD cells, suggesting that the expression of WNK4 was modulated by CLDN7. In addition, deletion of CLDN7 upregulated the expression level of the apical epithelial sodium channel (ENaC), indicating a potential cross-talk between paracellular and transcellular transport systems. This study demonstrates that CLDN7 plays an important role in salt balance in renal CD cells and modulating WNK4 and ENaC expression levels that are vital in controlling salt-sensitive hypertension

Necrotizing enterocolitis leads to disruption of tight junctions and increase in gut permeability in a mouse model

Background: Necrotizing enterocolitis (NEC) is a leading cause of death in preterm infants. Neonates weighing <1500 grams are at the highest risk for acquiring NEC, with a prevalence of nearly 7-10%, mortality up to 30%, and several long-term complications among survivors. Despite advancements in neonatal medicine, this disease remains a challenge to treat. The aim of this study is to investigate the effect of NEC on gut epithelial tight junctions and its barrier function using a NEC mouse model. Methods: Three-day old C57BL/6 mouse pups were fed with Esbilac formula every 3 hours and then subjected to hypoxia twice daily followed by cold stress. Dam fed pups from the same litters served as controls. Pups were observed and sacrificed 96 hours after the treatments and intestines were removed for experiments. The successful induction of NEC was confirmed by histopathology. Changes in tight junction proteins in NEC intestines were studied by western blotting and immunofluorescent microscopy using specific protein markers. The gut leakage in NEC was visualized using biotin tracer molecules. Results: Our study results demonstrate that we induced NEC in >50% of experimental pups, pups lost nearly 40% of weight and their intestines showed gross changes and microscopic changes associated with NEC. There were inflammatory changes with loss of tight junction barrier function and disruption of tight junction claudin proteins in the intestines of NEC mouse model. We have demonstrated for the first time that NEC intestines develop increased leakiness as visualized by biotin tracer leakage. Conclusions: NEC leads to breakdown of epithelial barrier due to changes in tight junction proteins with increased leakiness which may explain the transmigration of microbes and microbial products from the gut lumen into the blood stream leading to sepsis like signs clinically witnessed

WNK4 phosphorylates ser206 of claudin-7 and promotes paracellular Cl− permeability

AbstractMutations in WNK4 have been linked to hypertension in PHAII. Paracellular ion transport has been reported to be involved in this disease process; however, the specific molecular target has not been identified. In this study, we found that TJ protein claudin-7 and WNK4 were partially co-localized in renal tubules of rat kidney and co-immunoprecipitated in kidney epithelial cells. The wild-type and PHAII-causing mutant, but not the kinase-dead mutant, phosphorylated claudin-7. We have identified ser206 in the COOH-terminus of claudin-7 as a putative phosphorylation site for WNK4. More importantly, disease-causing mutant enhanced claudin-7 phosphorylation and significantly increased paracellular permeability to Cl−

Claudin-7 Modulates Cl- and Na+ Homeostasis and WNK4 Expression in Renal Collecting Duct Cells

Claudin-7 knockout (CLDN7-/-) mice display renal salt wasting and dehydration\r\nphenotypes. To address the role of CLDN7 in kidneys, we established collecting duct (CD) cell\r\nlines from CLDN7+/+ and CLDN7-/- mouse kidneys. We found that deletion of CLDN7 increased\r\nthe transepithelial resistance (TER) and decreased the paracellular permeability for Cl- and Na+ in\r\nCLDN7-/- CD cells. Inhibition of transcellular Cl- and Na+ channels has no significant effect on TER\r\nor dilution potentials. Current-voltage curves were linear in both CLDN7+/+ and CLDN7-/- CD cells,\r\nindicating that the ion flux was through the paracellular pathway. The impairment of Cl- and Na+\r\npermeability phenotype can be rescued by CLDN7 re-expression. We also found that WNK4 (its\r\nmutations lead to hypertension) expression, but not WNK1, was significantly increased in CLDN7-/-\r\nCD cell lines as well as in primary CLDN7-/- CD cells, suggesting that the expression of WNK4\r\nwas modulated by CLDN7. In addition, deletion of CLDN7 upregulated the expression level of the\r\napical epithelial sodium channel (ENaC), indicating a potential cross-talk between paracellular and\r\ntranscellular transport systems. This study demonstrates that CLDN7 plays an important role in\r\nsalt balance in renal CD cells and modulating WNK4 and ENaC expression levels that are vital in\r\ncontrolling salt-sensitive hypertension

Claudin-7 Modulates Cl− and Na+ Homeostasis and WNK4 Expression in Renal Collecting Duct Cells

Claudin-7 knockout (CLDN7−/−) mice display renal salt wasting and dehydrationphenotypes. To address the role of CLDN7 in kidneys, we established collecting duct (CD) celllines from CLDN7+/+ and CLDN7−/− mouse kidneys. We found that deletion of CLDN7 increasedthe transepithelial resistance (TER) and decreased the paracellular permeability for Cl− and Na+ inCLDN7−/− CD cells. Inhibition of transcellular Cl− and Na+ channels has no significant effect on TERor dilution potentials. Current-voltage curves were linear in both CLDN7+/+ and CLDN7−/− CD cells,indicating that the ion flux was through the paracellular pathway. The impairment of Cl− and Na+permeability phenotype can be rescued by CLDN7 re-expression. We also found that WNK4 (itsmutations lead to hypertension) expression, but not WNK1, was significantly increased in CLDN7−/−CD cell lines as well as in primary CLDN7−/− CD cells, suggesting that the expression of WNK4was modulated by CLDN7. In addition, deletion of CLDN7 upregulated the expression level of theapical epithelial sodium channel (ENaC), indicating a potential cross-talk between paracellular andtranscellular transport systems. This study demonstrates that CLDN7 plays an important role insalt balance in renal CD cells and modulating WNK4 and ENaC expression levels that are vital incontrolling salt-sensitive hypertension

Inflammation and disruption of the mucosal architecture in claudin-7-deficient mice.

BACKGROUND & AIMS: Integrity of the intestinal epithelium is required for nutrition absorption and defense against pathogens. Claudins are cell adhesion molecules that localize at tight junctions (TJs); many are expressed in the intestinal tract, but little is known about their functions. Claudin-7 is unique in that it has a stronger basolateral membrane distribution than other claudins, which localize primarily to apical TJs in the intestinal epithelium. We investigated the basolateral functions of claudin-7 and assessed the effects of disruption of Cldn7 in intestines of mice. METHODS: We generated Cldn7(-/-) mice and examined their intestines by histology, molecular and cellular biology, and biochemistry approaches. We performed gene silencing experiments in epithelial cell lines using small interfering RNAs (siRNAs). RESULTS: The Cldn7(-/-) mice had severe intestinal defects that included mucosal ulcerations, epithelial cell sloughing, and inflammation. Intestines of Cldn7(-/-) mice produced significantly higher levels of cytokines, the nuclear factor κB p65 subunit, and cyclooxygenase 2; they also up-regulated expression of matrix metalloproteinases (MMPs)-3 and -7. siRNA in epithelial cell lines showed that the increased expression of MMP-3 resulted directly from claudin-7 depletion, whereas that of MMP-7 resulted from inflammation. Electron microscopy analysis showed that intestines of Cldn7(-/-) mice had intercellular gaps below TJs and cell matrix loosening. Deletion of Cldn7 reduced expression and altered localization of the integrin α2 subunit in addition to disrupting formation of complexes of claudin-7, integrin α2, and claudin-1 that normally form in epithelial basolateral compartments of intestines. CONCLUSIONS: In mice, claudin-7 has non-TJ functions, including maintenance of epithelial cell-matrix interactions and intestinal homeostasis

Necrotizing enterocolitis leads to disruption of tight junctions and increase in gut permeability in a mouse model

Background: Necrotizing enterocolitis (NEC) is a leading cause of death in preterm infants. Neonates weighing 50% of experimental pups, pups lost nearly 40% of weight and their intestines showed gross changes and microscopic changes associated with NEC. There were inflammatory changes with loss of tight junction barrier function and disruption of tight junction claudin proteins in the intestines of NEC mouse model. We have demonstrated for the first time that NEC intestines develop increased leakiness as visualized by biotin tracer leakage.Conclusions: NEC leads to breakdown of epithelial barrier due to changes in tight junction proteins with increased leakiness which may explain the transmigration of microbes and microbial products from the gut lumen into the blood stream leading to sepsis like signs clinically witnessed

Necrotizing enterocolitis leads to disruption of tight junctions and increase in gut permeability in a mouse model

Abstract Background Necrotizing enterocolitis (NEC) is a leading cause of death in preterm infants. Neonates weighing 50% of experimental pups, pups lost nearly 40% of weight and their intestines showed gross changes and microscopic changes associated with NEC. There were inflammatory changes with loss of tight junction barrier function and disruption of tight junction claudin proteins in the intestines of NEC mouse model. We have demonstrated for the first time that NEC intestines develop increased leakiness as visualized by biotin tracer leakage. Conclusions NEC leads to breakdown of epithelial barrier due to changes in tight junction proteins with increased leakiness which may explain the transmigration of microbes and microbial products from the gut lumen into the blood stream leading to sepsis like signs clinically witnessed

Presenilin-1 inhibits delta-catenin-induced cellular branching and promotes delta-catenin processing and turnover

Although delta-catenin/neural plakophilin-related armadillo protein (NPRAP) was reported to interact with presenilin-1 (PS-1), the effects of PS-1 on delta-catenin have not been established. In this study, we report that overexpression of PS-1 inhibits the delta-catenin-induced dendrite-like morphological changes in NIH 3T3 cells and promotes delta-catenin processing and turnover. The effects of PS-1 on endogenous delta-catenin processing were confirmed in hippocampal neurons overexpressing PS-1, as well as in the transgenic mice expressing the disease-causing mutant PS-1 (M146V). In addition, disease-causing mutant PS-1 (M146V and L286V) enhanced delta-catenin processing, whereas PS-1/gamma-secretase inhibitors could block the formation of processed forms of delta-catenin. Together, our findings suggest that PS-1 can affect delta-catenin-induced morphogenesis possibly through the regulation of its processing and stability. Originally published Biochem Biophys Res Commun, Vol. 351, No. 4, Dec 200

Presenilin-1 inhibits delta-catenin-induced cellular branching and promotes delta-catenin processing and turnover

Although delta-catenin/neural plakophilin-related armadillo protein (NPRAP) was reported to interact with presenilin-1 (PS-1) the effects of PS-1 on delta-catenin have not been established. In this study we report that overexpression of PS-1 inhibits the delta-catenin-induced dendrite-like morphological changes in NIH 3T3 cells and promotes delta-catenin processing and turnover. The effects of PS-1 on endogenous delta-catenin processing were confirmed in hippocampal neurons overexpressing PS-1 as well as in the transgenic mice expressing the disease-causing mutant PS-1 (M146V). In addition disease-causing mutant PS-1 (M146V and L286V) enhanced delta-catenin processing whereas PS-1/gamma-secretase inhibitors could block the formation of processed forms of delta-catenin. Together our findings suggest that PS-1 can affect delta-catenin-induced morphogenesis possibly through the regulation of its processing and stability. Originally published Biochem Biophys Res Commun Vol. 351 No. 4 Dec 200