Increased Claudin-2 Expression Research Articles

RhoA‐dependent tight junction regulation is associated with dysregulated Akt and ERK signaling and increased claudin‐2 expression

Previous studies have demonstrated that expression of either constitutively‐active (CA) or dominant negative (DN) RhoA reduced tight junction barrier function in cultured MDCK monolayers (Jou et al. 1998). However, the mechanisms by which RhoA disruption causes barrier loss have not been defined. The aims of this study were to determine whether RhoA regulates barrier function in mammary epithelia and to identify the molecular events responsible. V14 and N19 (CA and DN, respectively) RhoA were expressed in EpH4 mammary epithelial cells. RhoA (CA or DN) expression in confluent monolayers caused a dose‐dependent loss of transepithelial electrical resistance. Neither trafficking of ZO‐1 and occludin to the tight junction nor cell viability were affected by CA or DN RhoA expression, but western blots showed that ERK and Akt were phosphorylated (consistent with activation) after either CA or DN RhoA expression. Transcript content of several claudins was altered after CA or DN RhoA expression. Most notably, claudin‐2 mRNA and protein expression were increased following either CA or DN RhoA expression. These data demonstrate that RhoA regulates the barrier in mammary epithelia and suggest that increased claudin‐2 expression contributes to RhoA‐mediated tight junction regulation.Supported by Charite Universitaetsmedizin Berlin.

The Role of SAHA for Preventing IL-13-Induced Colon Epithelial Cell Dysfunction

IL-13 has been implicated as an effector cytokine in the pathophysiology of ulcerative colitis (UC) through increasing colon epithelial cell apoptosis and expression of the pore-forming tight junction protein claudin-2. Binding of IL-13 to its receptor leads to activation of STAT6. The STAT6 phosphorylation status in patients with UC, however, is unknown, as is the effect of STAT6 inhibition on colonic epithelium exposed to IL-13. Suberoylanilide hydroxamic acid (SAHA) is a FDA-approved histone deacetylase inhibitor that inhibits constitutive STAT6 activation in lymphoma cell lines. Therefore, the aim of this study was to determine if STAT6 phosphorylation is increased in patients with UC, and if inhibition of STAT6 signaling with SAHA attenuates IL-13-induced apoptosis and claudin-2 expression. To assess IL-13 signaling, immunohistochemical staining for phosphorylated (p) STAT6 was performed on colonic tissue from pediatric subjects with UC (early UC, n= 10) and Crohn's disease (CD, n= 10) at diagnosis, colectomy tissue from adults with UC (late UC, n= 5) and controls (n= 10). A pathologist blinded to the diagnosis scored histological sections on a 0-4 scale for pSTAT6. In cell culture, human colon HT-29 cells were pre-treated with SAHA (1 or 5 μM) or vehicle control prior to treatment with IL-13 (10 ng/ml). Western blot analysis was performed on cellular lysates for pSTAT6 and claudin-2 with results quantified by densitometry. Apoptosis was determined by flow cytometry of Annexin V-PE-stained cells and by immunodetection of cleaved caspase-3. Epithelial pSTAT6 staining was increased in UC with median score of 0 in control subjects, 2 in early UC (vs. control P= 0.004), 4 in late UC (P= 0.014), and 0 in CD. In HT-29 cell culture, SAHA inhibited IL-13-induced pSTAT6 in a dose-dependent fashion with a 57% reduction of pSTAT6 induction at 5 μM (P < 0.01). IL-13 induced a 2-fold increase in claudin-2 expression (p < 0.05), and SAHA reduced IL-13-induced claudin-2 expression by 70% to baseline levels (p<0.05). By Annexin V staining, IL-13 increased apoptosis 2.7-fold (3.7 ±1.6% vs. 10.0±1.4% Annexin V+, P<0.01). SAHA alone did not increase apoptosis and reduced IL-13-induced apoptosis in a dose-dependent manner with complete suppression to baseline at 5 μM (IL-13 + SAHA 1 μM 7.5±0.8%, + 5 μM 4.8±0.9, Ptrend< 0.001). Similarly, by immunodetection of cleaved caspase-3, SAHA reduced IL-13 induced apoptosis by 57% at 1 μM and 94% at 5 μM (P<0.001 for each compared to IL-13 alone). Increased STAT6 phosphorylation in the colonic epithelium of pediatric subjects at presentation with UC supports an effector role of IL-13 early in the disease. Treatment of colonic epithelial cells with SAHA, a drug that inhibits IL-13-induced STAT6 activation, reduces IL-13-induced apoptosis and claudin-2 expression, suggesting that STAT6 may be a novel therapeutic target for UC.

Enterocyte-specific epidermal growth factor prevents barrier dysfunction and improves mortality in murine peritonitis

Systemic administration of epidermal growth factor (EGF) decreases mortality in a murine model of septic peritonitis. Although EGF can have direct healing effects on the intestinal mucosa, it is unknown whether the benefits of systemic EGF in peritonitis are mediated through the intestine. Here, we demonstrate that enterocyte-specific overexpression of EGF is sufficient to prevent intestinal barrier dysfunction and improve survival in peritonitis. Transgenic FVB/N mice that overexpress EGF exclusively in enterocytes (IFABP-EGF) and wild-type (WT) mice were subjected to either sham laparotomy or cecal ligation and puncture (CLP). Intestinal permeability, expression of the tight junction proteins claudins-1, -2, -3, -4, -5, -7, and -8, occludin, and zonula occludens-1; villus length; intestinal epithelial proliferation; and epithelial apoptosis were evaluated. A separate cohort of mice was followed for survival. Peritonitis induced a threefold increase in intestinal permeability in WT mice. This was associated with increased claudin-2 expression and a change in subcellular localization. Permeability decreased to basal levels in IFABP-EGF septic mice, and claudin-2 expression and localization were similar to those of sham animals. Claudin-4 expression was decreased following CLP but was not different between WT septic mice and IFABP-EGF septic mice. Peritonitis-induced decreases in villus length and proliferation and increases in apoptosis seen in WT septic mice did not occur in IFABP-EGF septic mice. IFABP-EGF mice had improved 7-day mortality compared with WT septic mice (6% vs. 64%). Since enterocyte-specific overexpression of EGF is sufficient to prevent peritonitis-induced intestinal barrier dysfunction and confers a survival advantage, the protective effects of systemic EGF in septic peritonitis appear to be mediated in an intestine-specific fashion.

Claudin-4 augments alveolar epithelial barrier function and is induced in acute lung injury

Intact alveolar barrier function is associated with better outcomes in acute lung injury patients; however, the regulation of alveolar epithelial paracellular transport during lung injury has not been extensively investigated. This study was undertaken to determine whether changes in tight junction claudin expression affect alveolar epithelial barrier properties and to determine the mechanisms of altered expression. In anesthetized mice exposed to ventilator-induced lung injury, claudin-4 was specifically induced among tight junction structural proteins. Real-time PCR showed an eightfold increase in claudin-4 expression in the lung injury model. To examine the role of this protein in barrier regulation, claudin-4 function was inhibited with small interfering RNA (siRNA) and a blocking peptide derived from the binding domain of Clostridium perfringens enterotoxin (CPE(BD)). Inhibition of claudin-4 decreased transepithelial electrical resistance but did not alter macromolecule permeability in primary rat and human epithelial cells. In mice, CPE(BD) decreased air space fluid clearance >33% and resulted in pulmonary edema during moderate tidal volume ventilation that did not induce edema in control peptide-treated mice. In vitro phorbol ester induced a ninefold increase in claudin-4 expression that was dependent on PKC activation and the JNK MAPK pathway. These data establish that changes in alveolar epithelial claudin expression influence paracellular transport, alveolar fluid clearance rates, and susceptibility to pulmonary edema. We hypothesize that increased claudin-4 expression early in acute lung injury represents a mechanism to limit pulmonary edema and that the regulation of alveolar epithelial claudin expression may be a novel target for acute lung injury therapy.

Epidermal growth factor increases claudin-4 expression mediated by Sp1 elevation in MDCK cells

Epidermal growth factor (EGF) increases claudin-4 expression in Madin–Darby canine kidney (MDCK) cells. Here we examined what regulatory mechanisms are involved in the EGF-induced claudin-4 elevation. EGF transiently increased claudin-4 mRNA at 3 h and persistently increased its protein for 24 h without affecting claudin-1 expression. EGF increased p-ERK1/2 levels, which were inhibited by U0126, a MEK inhibitor. The exogenous expression of constitutively activated MEK increased claudin-4 expression. These results indicate that the activation of ERK1/2 is involved in the EGF-induced claudin-4 elevation. EGF increased Sp1 expression within 1 h, which was inhibited by U0126. In immunocytochemistry, Sp1 was distributed in nucleus in control and the EGF-treated cells. The EGF-induced claudin-4 elevation was inhibited by mithramycin, a Sp1 inhibitor, and Sp1 small interfering RNA. We suggest that EGF activates a MEK/ERK pathway and increases Sp1 expression, resulting in an elevation of claudin-4 expression.

Increased claudin‐4 expression is associated with poor prognosis and high tumour grade in breast cancer

The role of intercellular tight junctions in breast epithelial cells is traditionally thought to be in maintaining polarity and barrier function. However, claudin-4, a tight junction protein, is overexpressed in breast tumour cells compared to normal epithelial cells, which generally corresponds to a loss in polarity. The aim of this study was to investigate the distribution and potential clinical value of claudin-4 in breast cancer, and to evaluate its usefulness as a prognostic and predictive biomarker. Expression of claudin-4 was initially examined by Western blot analysis in a cohort of 88 breast tumours, and was found to correlate positively with tumour grade and negatively with ER. Claudin-4 expression was then evaluated by immunohistochemistry in a larger cohort of 299 tumours represented on a tissue microarray. Claudin-4 expression correlated positively with tumour grade and Her2, and negatively with ER. High claudin-4 expression was also associated with worse breast cancer-specific survival (p = 0.003), recurrence-free survival (p = 0.025) and overall survival (p = 0.034). Multivariate analysis revealed that claudin-4 independently predicted survival in the entire cohort (HR 1.95; 95%CI 1.01-3.79; p = 0.047) and in the ER positive subgroup treated with adjuvant tamoxifen (HR 4.34; 95%CI 1.14-16.53; p = 0.032). This relationship between increased claudin-4 expression and adverse outcome was validated at the mRNA level in a DNA microarray dataset of 295 breast tumours. We conclude that high levels of claudin-4 protein are associated with adverse outcome in breast cancer patients, including the subgroup of patients treated with adjuvant tamoxifen.

Hypertonic stress increases claudin-4 expression and tight junction integrity in association with MUPP1 in IMCD3 cells

We reported that the multiple PDZ protein 1 (MUPP1) is an osmotic response protein in kidney cells. This up-regulation was found to be necessary for the maintenance of tight epithelial properties in these cells. We investigated whether an interaction with one or more members of the claudin family is responsible for this observation. In response to hypertonicity, the up-regulation of claudin-4 (Cldn4) expression, and not other claudins, was initially identified in inner medullary collecting duct (IMCD3) cells by gene array and further verified by quantitative PCR and Western blotting. In kidney tissues, Cldn4 expression was substantial in the papilla and absent in the cortex. Furthermore, Cldn4 expression significantly increased in the papilla of mice after 36 h of thirsting. Cldn4 immunofluorescence in hypertonically stressed cells revealed colocalization with MUPP1 at the tight junctions. Interaction between Cldn4 and MUPP1 was also demonstrated by coimmunoprecipitation of both proteins from IMCD3 cells chronically adapted to hypertonicity. In IMCD3 cells stably silenced for MUPP1 expression under hypertonic conditions, a significant decrement in Cldn4 expression was observed that was restored after inhibition of lysosome activity. Immunofluorescence detection identified that in these MUPP1-silenced cells Cldn4 was mistargeted to the lysosomes. Functionally, silencing Cldn4 expression in IMCD3 cells resulted in a decrease in the transepithelial resistance to the same degree as observed when MUPP1 expression was silenced, suggesting that MUPP1 contributes to the maintenance of a tight epithelium in the medulla of the kidney under hypertonic stress by correctly localizing Cldn4 to the tight junctions.

Quercetin Enhances Epithelial Barrier Function and Increases Claudin-4 Expression in Caco-2 Cells 3

Quercetin is the most abundant flavonoid and is assumed to have positive effects on the gastrointestinal mucosa after dietary intake. The aim of the study was to analyze the influence of quercetin on intestinal barrier function using the human colonic epithelial cell line Caco-2. Transepithelial resistance (Rt), tracer fluxes of [3H]-mannitol, 22Na+, and 36Cl− as well as electrogenic ion transport were determined in Ussing chambers. Expression of tight junction (TJ) proteins and mRNA was analyzed in Western blots and quantitative RT-PCR, respectively. Regulation of transcription was analyzed by reporter gene assay. Cellular distribution of TJ proteins was examined by confocal laser scanning microscopy (LSM). Apoptotic rate was determined by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining. Quercetin induced a dose-dependent increase of Rt persisting for >2 d. Daily addition of quercetin was able to perpetuate the effect, which was seen whether quercetin was added apically or to the basolateral compartment. Parallel to the Rt increase, quercetin induced a strong increase of the TJ protein claudin-4 but not of other claudins. Confocal LSM showed a localization of claudin-4 in TJ. Apoptotic rate was not affected by quercetin. Consistent with these changes, fluxes of Na+ and Cl−, but not of mannitol, were reduced. Reporter gene assays revealed a stimulatory effect of quercetin on claudin-4 transcription. The flavonoid quercetin enhances barrier function via transcriptional expression regulation of the TJ protein claudin-4, which represents an important protective effect of this food component against barrier disturbance in intestinal inflammation.

Hepatitis C virus receptor expression in normal and diseased liver tissue

The principal site of hepatitis C virus (HCV) replication is the liver. HCV pseudoparticles infect human liver derived cell lines and this suggests that liver-specific receptors contribute to defining HCV hepatotropism. At least three host cell molecules have been reported to be important for HCV entry: the tetraspanin CD81, scavenger receptor class B member I (SR-BI), and the tight junction (TJ) protein Claudin 1 (CLDN1). Hepatocytes in liver tissue coexpress CD81, SR-BI, and CLDN1, consistent with their ability to support HCV entry. CLDN1 localized at the apical-canalicular TJ region and at basolateral-sinusoidal hepatocyte surfaces in normal tissue and colocalized with CD81 at both sites. In contrast, CLDN1 appeared to colocalize with SR-BI at the basolateral-sinusoidal surface. CLDN1 expression was increased on basolateral hepatocyte membranes in HCV-infected and other chronically inflamed liver tissue compared with normal liver. In contrast, CLDN4 hepatocellular staining was comparable in normal and diseased liver tissue. HCV infection of Huh-7.5 hepatoma cells in vitro significantly increased CLDN1 expression levels, consistent with a direct modulation of CLDN1 by virus infection. In HCV infected livers, immunohistochemical studies revealed focal patterns of CLDN1 staining, suggesting localized areas of increased CLDN1 expression in vivo which may potentiate local viral spread within the liver.

Smad4 Regulates Claudin-1 Expression in a Transforming Growth Factor-β–Independent Manner in Colon Cancer Cells

We have recently reported that the expression of a tight junction protein, claudin-1, is increased during colon carcinogenesis and particularly metastatic colorectal cancer. Manipulation of claudin-1 levels in colon cancer cells showed a positive correlation between claudin-1 expression and tumor growth and metastasis. However, the mechanisms underlying the increased claudin-1 expression in colorectal cancer remains unknown. The tumor suppressor Smad4 is a central intracellular signal transduction component of the transforming growth factor-beta (TGF-beta) family of cytokines. Loss of Smad4 protein expression is correlated with poor prognosis and is frequently observed in invasive and metastatic colorectal carcinoma. In the present study, we report an inverse relationship between Smad4 and claudin-1 expression in human colorectal carcinoma tumor samples and in human colon cancer cell lines. We found that the expression of Smad4 in Smad4-deficient but claudin-1-positive SW480 or HT29 colon cancer cell lines down-regulates claudin-1 expression through transcriptional repression by modulating beta-catenin/T-cell factor/lymphocyte enhancer factor activity. Furthermore, this Smad4-dependent inhibition of claudin-1 expression is independent of TGF-beta signaling because Smad4 expression alone is insufficient to restore TGF-beta signaling in the SW480 cells, and the selective TGF-beta receptor kinase inhibitor LY364947 did not prevent the Smad4 suppression of claudin-1 protein expression in either SW480 or HT29 cells. Taken together, these findings suggest a novel mechanism underlying Smad4 tumor-suppressive function through regulation of a potential metastatic modulator, claudin-1, in a TGF-beta-independent manner.

When the levee breaks: The etiologic role of epithelial barrier function defects in ileitis

When the levee breaks: The etiologic role of epithelial barrier function defects in ileitis

Claudin-4 differentiates biliary tract cancers from hepatocellular carcinomas

The recently identified claudins are dominant components of tight junctions, responsible for cell adhesion, polarity and paracellular permeability. Certain claudins have been shown to have relevance in tumor development, with some of them, especially claudin-4, even suggested as future therapeutic target. The aim of the present study was to analyze the expression of claudin-4 in the biliary tree, biliary tract cancers and hepatocellular carcinomas. A total of 107 cases were studied: 53 biliary tract cancers, 50 hepatocellular carcinomas, 10 normal liver and 10 normal extrahepatic biliary duct samples. Immunohistochemical analysis was performed on conventional specimens and on tissue microarrays as well. Claudin-4 was further investigated by Western blot analysis and real-time RT-PCR. Intense membranous immunolabeling was found for claudin-4 in all biliary tract cancers unrelated to the primary site of origin, namely intrahepatic, extrahepatic or gallbladder cancers. Normal biliary epithelium showed weak positivity for claudin-4. In contrast, normal hepatocytes and tumor cells of hepatocellular carcinomas did not express claudin-4. The results of Western immunoblot analysis and real-time RT-PCR were in correlation with the immunohistochemical findings. Cytokeratins, as CK7 (92%) and CK19 (83%) were mostly positive in biliary tract cancers, however, one-third of hepatocellular carcinomas also expressed CK7 (34%). HSA antibody (HepPar1) reacted with the majority of hepatocellular carcinomas (86%), while being positive in a low percentage of the biliary tract cancers (8%). In conclusion, this is the first report of a significantly increased claudin-4 expression in biliary tract cancers, which represents a novel feature of tumors of biliary tract origin. Claudin-4 expression seems to be a useful marker in differentiating biliary tract cancers from hepatocellular carcinomas and could well become a potential diagnostic tool.

Experimental obstructive jaundice alters claudin-4 expression in intestinal mucosa: Effect of bombesin and neurotensin

To investigate the influence of experimental obstructive jaundice and exogenous bombesin (BBS) and neurotensin (NT) administration on the expression of the tight junction (TJ)-protein claudin-4 in intestinal epithelium of rats. Forty male Wistar rats were randomly divided into five groups: I = controls, II = sham operated, III = bile duct ligation (BDL), IV = BDL+BBS (30 microg/kg per d), V = BDL+NT (300 microg/kg per d). At the end of the experiment on d 10, endotoxin was measured in portal and aortic blood. Tissue sections of the terminal ileum were examined histologically and immunohistochemically for evaluation of claudin-4 expression in intestinal epithelium. Obstructive jaundice led to intestinal barrier failure demonstrated by significant portal and aortic endotoxemia. Claudin-4 expression was significantly increased in the upper third of the villi in jaundiced rats and an upregulation of its lateral distribution was noted. Administration of BBS or NT restored claudin-4 expression to the control state and significantly reduced portal and aortic endotoxemia. Experimental obstructive jaundice increases claudin-4 expression in intestinal epithelium, which may be a key factor contributing to the disruption of the mucosal barrier. Gut regulatory peptides BBS and NT can prevent this alteration and reduce portal and systemic endotoxemia.

Interleukin-13 Is the Key Effector Th2 Cytokine in Ulcerative Colitis That Affects Epithelial Tight Junctions, Apoptosis, and Cell Restitution

Interleukin-13 Is the Key Effector Th2 Cytokine in Ulcerative Colitis That Affects Epithelial Tight Junctions, Apoptosis, and Cell Restitution

Interleukin-13 Is the Key Effector Th2 Cytokine in Ulcerative Colitis That Affects Epithelial Tight Junctions, Apoptosis, and Cell Restitution

Ulcerative colitis (UC) is characterized by a Th2 immune response with inflammation and epithelial barrier dysfunction. So far, Th2 cytokines have not been shown to directly influence epithelial barrier function. Lamina propria mononuclear cells (LPMCs) were stimulated and interleukin (IL)-13 was measured by enzyme-linked immunosorbent assay. Functional IL-13 and IL-4 effects were studied on HT-29/B6 colonic epithelial cells in Ussing chambers and by conductance scanning. Apoptosis was detected by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assays. IL-13/IL-4 receptors were analyzed by reverse-transcription polymerase chain reaction and immunofluorescence. Western blotting combined with immunofluorescence was used to detect tight junction proteins. Furthermore, restitution velocity was measured. Finally, mucosal biopsy specimens from patients with UC were compared with cultured cells for these features. LPMCs from patients with UC produced large amounts of IL-13 (985 +/- 73 pg/mL), much more than from controls or patients with Crohn's disease. IL-13Ralpha1 and IL-4Ralpha receptors were present in HT-29/B6 cells and colonic epithelial cells of control patients and patients with UC. IL-13 had a dose-dependent effect on transepithelial resistance of HT-29/B6 monolayers (reduction to 60% +/- 4%), whereas IL-4 had no effect. This was due to an increased number of apoptotic cells (5.6-fold +/- 0.9-fold) and an increased expression of the pore-forming tight junction protein claudin-2 to 295% +/- 37%, both of which contributed equally. Finally, epithelial restitution velocity decreased from 15.1 +/- 0.6 to 10.6 +/- 0.5 microm/h after treatment with IL-13. Parallel changes were observed in human samples, with an increase in claudin-2 expression to 956% +/- 252%. IL-13 was identified as an important effector cytokine in UC that impairs epithelial barrier function by affecting epithelial apoptosis, tight junctions, and restitution velocity.

Extracellular Signal-regulated Kinases 1/2 Control Claudin-2 Expression in Madin-Darby Canine Kidney Strain I and II Cells

The tight junction of the epithelial cell determines the characteristics of paracellular permeability across epithelium. Recent work points toward the claudin family of tight junction proteins as leading candidates for the molecular components that regulate paracellular permeability properties in epithelial tissues. Madin-Darby canine kidney (MDCK) strain I and II cells are models for the study of tight junctions and based on transepithelial electrical resistance (TER) contain "tight" and "leaky" tight junctions, respectively. Overexpression studies suggest that tight junction leakiness in these two strains of MDCK cells is conferred by expression of the tight junction protein claudin-2. Extracellular signal-regulated kinase (ERK) 1/2 activation by hepatocyte growth factor treatment of MDCK strain II cells inhibited claudin-2 expression and transiently increased TER. This process was blocked by the ERK 1/2 inhibitor U0126. Transfection of constitutively active mitogen-activated protein kinase/extracellular signal-regulated kinase kinase into MDCK strain II cells also inhibited claudin-2 expression and increased TER. MDCK strain I cells have higher levels of active ERK 1/2 than do MDCK strain II cells. U0126 treatment of MDCK strain I cells decreased active ERK 1/2 levels, induced expression of claudin-2 protein, and decreased TER by approximately 20-fold. U0126 treatment also induced claudin-2 expression and decreased TER in a high resistance mouse cortical collecting duct cell line (94D). These data show for the first time that the ERK 1/2 signaling pathway negatively controls claudin-2 expression in mammalian renal epithelial cells and provide evidence for regulation of tight junction paracellular transport by alterations in claudin composition within tight junction complexes.

EXPRESSION OF CLAUDIN-1 IN MOUSE TESTIS

In testis, tight junctions (TJs) between adjacent Sertoli cells are important for the formation of the blood-testis barrier (BTB) and crucial for spermatogenesis. The present study aimed to find postnatal changes in the expression of claudin-1, one of the TJ genes in mouse testis. By semiquantitative RT-PCR, it was found that claudin-1 expression in testis increased up to a peak at 10 days after birth and decreased thereafter. Western blot analysis showed abundant expression of 21-kDa protein in testis, lung, and brain from the adult mouse. The developmental change in the expression of claudin-1 protein in testis coincided with that from the RT-PCR. Testosterone treatment significantly increased claudin-1 expression in immature Sertoli cells, suggesting the possible regulation of claudin-1 expression by androgen in mouse Sertoli cells. Claudin-1 expression appears to be developmentally regulated in the mouse testis.

Expression of claudin-1 in mouse testis.

In testis, tight junctions (TJs) between adjacent Sertoli cells are important for the formation of the blood-testis barrier (BTB) and crucial for spermatogenesis. The present study aimed to find postnatal changes in the expression of claudin-1, one of the TJ genes in mouse testis. By semiquantitative RT-PCR, it was found that claudin-1 expression in testis increased up to a peak at 10 days after birth and decreased thereafter. Western blot analysis showed abundant expression of 21-kDa protein in testis, lung, and brain from the adult mouse. The developmental change in the expression of claudin-1 protein in testis coincided with that from the RT-PCR. Testosterone treatment significantly increased claudin-1 expression in immature Sertoli cells, suggesting the possible regulation of claudin-1 expression by androgen in mouse Sertoli cells. Claudin-1 expression appears to be developmentally regulated in the mouse testis.