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A considerable amount of NaCl reabsorption in proximal tubules (PTs) occurs via the paracellular transport regulated by the tight junction proteins claudins (Cldns). However, the paracellular transport properties in mouse superficial PTs remain unclear. We characterized these properties in superficial PT S1–S3 segments from mice expressing [wild-type (WT, WTS1–WTS3)] or lacking [knockout (KO, KOS1–KOS3)] claudin-2. We isolated and perfused segments with symmetrical solutions in the presence of bath ouabain and measured the diffusion potential upon changing the salt composition of the lumen or bath. Based on the diffusion potential corrected for the liquid junction potential (dVT), we calculated the paracellular Na+ over Cl− permeability (PNa/PCl) ratio. The PNa/PCl values upon reducing luminal NaCl averaged 1.27, 1.04, and 0.85 in WTS1, WTS2, and WTS3 and 0.34, 0.55, and 0.80 in KOS1, KOS2, and KOS3, respectively. The dVT values exhibited a symmetrical response to bidirectional NaCl concentration gradients in WTS1–WTS3 and KOS1–KOS3. WTS1 and WTS3 were monovalent cation-selective, with WTS1 demonstrating stronger cation selectivity. The order of permeabilities relative to Cl− was K+ > Rb+ > Na+ > Li+, whereas both KOS1 and KOS3 exhibited monovalent cation selectivity loss and, consequently, enhanced anion selectivity, especially in KOS1. Protamine addition to the lumen and bath similarly decreased PNa/PCl values upon reduced luminal NaCl in the order of WTS1 > WTS3 > KOS3 > KOS1. Therefore, this study presents evidence of axial heterogeneity in paracellular transport across superficial PTs in mice. NEW & NOTEWORTHY: Research on isolated perfused S2 segments of proximal tubules in mice, both expressing and lacking claudin-2, indicates that claudin-2 forms leaky monovalent cation-selective paracellular channels within the tight junctions of proximal tubules. This study characterized the paracellular transport properties in isolated and perfused superficial proximal tubule S1–S3 segments in both groups of mice. The findings demonstrate, for the first time, functional heterogeneity in the paracellular pathway along the axis of the superficial proximal tubules. [ABSTRACT FROM AUTHOR]

Duck circovirus (DuCV) is widely recognized as a prominent virus in China's duck farming industry, known for its ability to cause persistent infections and significant immunosuppression, which can lead to an increased susceptibility to secondary infections, posing a significant threat to the duck industry. Moreover, clinical evidence also indicates the potential vertical transmission of the virus through duck embryos to subsequent generations of ducklings. However, the limited availability of suitable cell lines for in vitro cultivation of DuCV has hindered further investigation into the molecular mechanisms underlying its infection and pathogenicity. In this study, we observed that oral DuCV infection in female breeding ducks can lead to oviduct, ovarian, and follicular infections. Subsequently, the infection can be transmitted to the fertilized eggs, resulting in the emergence of virus-carrying ducklings upon hatching. In contrast, the reproductive organs of male breeding ducks were unaffected by the virus, thus confirming that vertical transmission of DuCV primarily occurs through infection in female breeding ducks. By analyzing transcriptome sequencing data from the oviduct, we focused on claudin-2, a gene encoding the tight junction protein CLDN2 located on the cell membrane, which showed significantly increased expression in DuCV-infected oviducts of female breeding ducks. Notably, CLDN2 was confirmed to interact with the unique structural protein of DuCV, namely capsid protein (Cap), through a series of experimental approaches including co-immunoprecipitation (co-IP), GST pull-down, immunofluorescence, and adhesion-blocking assays. Furthermore, we demonstrated that the Cap protein binds to the extracellular loop structural domains EL1 and EL2 of CLDN2. Subsequently, by constructing a series of truncated bodies of the CLDN2 promoter region, we identified the transcription factor SP5 for CLDN2. Moreover, we found that DuCV infection triggers the activation of the MAPK-ERK signaling pathway in DEF cells and ducks, leading to an upregulation of SP5 and CLDN2 expression. This process ultimately leads to the transportation of mature CLDN2 to the cell surface, thereby facilitating increased virus adherence to the target organs. In conclusion, we discovered that DuCV utilizes host CLDN2 proteins to enhance adhesion and infection in oviducts and other target organs. Furthermore, we elucidated the signaling pathways involved in the interaction between DuCV Cap proteins and CLDN2, which provides valuable insights into the molecular mechanism underlying DuCV's infection and vertical transmission. [ABSTRACT FROM AUTHOR]

Background: Considering the increasing worldwide prevalence of inflammatory bowel disease (IBD), the early diagnosis of this disease is extremely important. However, non-invasive diagnostic methods remain limited, while invasive techniques are the most commonly used in daily practice. Therefore, there is a serious need to find new non-invasive biomarkers of IBD. Methods: The serum profiles of occludin, claudin-2, and zonulin were assessed in IBD patients using the ELISA method. The levels of the analyzed biomarkers were measured before and after a year of anti-inflammatory treatment, which was a tumor necrosis factor α (TNF-α) inhibitor (adalimumab) in patients with ulcerative colitis (UC) and conventional therapy in patients with Crohn's disease (CD). Results: In IBD patients, the serum level of occludin (p < 0.001) decreased compared to healthy individuals, while the level of claudin-2 (p < 0.001) increased. Additionally, zonulin (p < 0.01) concentration increased in CD patients compared to the control group. The highest diagnostic ability was presented by occludin measurements with the area under the curve (AUC) of 0.959 (95% CI 0.907–1) in UC and 0.948 (95% CI 0.879–1) in CD. Claudin-2 also demonstrated very good ability in diagnosing UC and CD with AUC values of 0.864 (95% CI 0.776–0.952) and 0.896 (95% CI 0.792–0.999), respectively. The ability of zonulin to diagnose CD was estimated as good with an AUC of 0.74 (95% CI 0.598–0.881). Moreover, a significant correlation was identified between C-reactive protein (CRP), claudin-2 (r = −0.37; p < 0.05), and zonulin (r = −0.44; p < 0.05) in UC patients. Treatment with adalimumab improved the level of occludin, claudin-2, and zonulin in UC patients, while anti-inflammatory conventional therapy decreased the concentration of zonulin in CD. Conclusions: Occludin and claudin-2 measurements present significant utility in diagnosing both UC and CD, while zonulin assessments may be useful in CD diagnosis. Additionally, claudin-2 and zonulin measurements may be helpful in evaluating the intensity of the inflammatory process. Anti-TNF-α treatment improved the value of occludin, claudin-2, and zonulin, indicating its beneficial effect on the integrity of tight junctions in UC. [ABSTRACT FROM AUTHOR]

Laurus nobilis L. (LNL) belongs to the evergreen Lauraceae family. It is native to the Mediterranean and widely distributed in the southern United States, Europe, and the Middle East. LNL is rich in active ingredients of the sesquiterpene lactone series and has been reported to have antioxidant, anti-inflammatory, and anticancer effects. And parthenolide, known as a sesquiterpene lactone-based compound, inhibits the activation of lipopolysaccharide-binding protein (LBP), which is a major trigger for leaky gut syndrome. However, the effectiveness of LNL in improving the state of increased intestinal permeability has not yet been reported. Therefore, we demonstrated the efficacy of LNL, which is known to be rich in parthenolide, in improving intestinal permeability induced by IL-13. We investigated the improvement in permeability and analyzed major tight junction proteins (TJs), permeability-related mechanisms, weight and disease activity indices, and corresponding cytokine mechanisms. LNL maintained TJs homeostasis and clinical improvement by reducing increased claudin-2 through the inhibition of IL-13/STAT6 activation in TJ-damaged conditions. These results are expected to be effective in preventing leaky gut syndrome through the TJ balance and to further improve intestinal-related diseases, such as inflammatory bowel disease. [ABSTRACT FROM AUTHOR]

Introduction: Pre-diabetes, a high-risk metabolic state, is situated between normal glucose homeostasis and diabetes. Early identification of pre-diabetes offers opportunities for intervention and diabetes reversal, highlighting the crucial need to investigate reliable biomarkers for this condition. Methods: We conducted an in-depth bioinformatics analysis of clinical samples from non-diabetic (ND), impaired glucose tolerance (IGT), and type 2 diabetes mellitus (T2DM) categories within the GSE164416 dataset. Thereafter the HFD and STZ treated mice were used for validation. Results: This analysis identified several codifferentially expressed genes (Co-DEGs) for IGT and T2DM, including CFB, TSHR, VNN2, APOC1, CLDN2, SLPI, LCN2, CXCL17, FAIM2, and REG3A. Validation of these genes and the determination of ROC curves were performed using the GSE76895 dataset. Thereafter, CLDN2 was selected for further verification. Gene expression analysis and immunofluorescence analysis revealed a significant upregulation of CLDN2 expression in the pancreas islets of mice in the high-fat diet and T2DM groups compared to the control group. Similarly, serum level of CLDN2 in patients with IGT and T2DM were significantly higher than those in the healthy group. Discussion: These results suggest that CLDN2 can serve as a novel biomarker for pre-diabetes, providing a new direction for future research in the prevention of type 2 diabetes. [ABSTRACT FROM AUTHOR]

Tight junction (TJ) protein cingulin (CGN) and transcription factor forkhead box protein O1 (FOXO1) contribute to the development of various cancers. Histone deacetylase (HDAC) inhibitors have a potential therapeutic role for some cancers. HDAC inhibitors affect the expression of both CGN and FOXO1. However, the roles and regulatory mechanisms of CGN and FOXO1 are unknown in non-small cell lung cancer (NSCLC) and normal human lung epithelial (HLE) cells. In the present study, to investigate the effects of CGN and FOXO1 on the malignancy of NSCLC, we used A549 cells as human lung adenocarcinoma and primary human lung epithelial (HLE) cells as normal lung tissues and performed the knockdown of CGN and FOXO1 by siRNAs. Furthermore, to investigate the detailed mechanisms in the antitumor effects of HDAC inhibitors for NSCLC via CGN and FOXO1, A549 cells and HLE cells were treated with the HDAC inhibitors trichostatin A (TSA) and Quisinostat (JNJ-2648158). In A549 cells, the knockdown of CGN increased bicellular TJ protein claudin-2 (CLDN-2) via mitogen-activated protein kinase/adenosine monophosphate-activated protein kinase (MAPK/AMPK) pathways and induced cell migration, while the knockdown of FOXO1 increased claudin-4 (CLDN-4), decreased CGN, and induced cell proliferation. The knockdown of CGN and FOXO1 induced cell metabolism in A549 cells. TSA and Quisinostat increased CGN and tricellular TJ protein angulin-1/lipolysis-stimulated lipoprotein receptor (LSR) in A549. In normal HLE cells, the knockdown of CGN and FOXO1 increased CLDN-4, while HDAC inhibitors increased CGN and CLDN-4. In conclusion, the knockdown of CGN via FOXO1 contributes to the malignancy of NSCLC. Both HDAC inhibitors, TSA and Quisinostat, may have potential for use in therapy for lung adenocarcinoma via changes in the expression of CGN and FOXO1. [ABSTRACT FROM AUTHOR]

Congcong Guo,1– 4 Mingwen Jiao,5 Yuying Cui,1– 4 Pingjiang Li,6 Jinming Yao,1– 3 Jianjun Dong,7 Lin Liao1– 4 1Department of Endocrinology and Metabology, The First Affiliated Hospital of Shandong First Medical University & Shandong Provincial Qianfoshan Hospital, Jinan, Shandong, People’s Republic of China; 2Shandong Key Laboratory of Rheumatic Disease and Translational medicine, Shandong Institute of Nephrology, Jinan, Shandong, People’s Republic of China; 3Shandong Institute of Nephrology, the First Affiliated Hospital of Shandong First Medical University& Shandong Provincial Qianfoshan Hospital, Jinan, Shandong, People’s Republic of China; 4Shandong University of Traditional Chinese Medicine, Jinan, Shandong, People’s Republic of China; 5Department of General Surgery, The First Affiliated Hospital of Shandong First Medical University & Shandong Provincial Qianfoshan Hospital, Jinan, Shandong, People’s Republic of China; 6Shandong First Medical University and Shandong Academy of Medical Sciences, Jinan, Shandong, People’s Republic of China; 7Department of Endocrinology, Qilu Hospital, Cheeloo College of Medicine, Shandong University, Jinan, Shandong, People’s Republic of ChinaCorrespondence: Lin Liao, Department of Endocrinology and Metabology, The First Affiliated Hospital of Shandong First Medical University and Shandong Provincial Qianfoshan Hospital, Jinan, Shandong, People’s Republic of China, Tel +86 15168888260, Email liaolin@sdu.edu.cn Jianjun Dong, Department of Endocrinology, Qilu Hospital, Cheeloo College of Medicine, Shandong University, Jinan, Shandong, People’s Republic of China, Tel +86 13791122910, Email dongjianjun@sdu.edu.cnPurpose: Proximal tubular epithelial cell (PTEC) is vulnerable to injury in diabetic kidney disease (DKD) due to high energy expenditure. The injured PTECs-derived profibrotic factors are thought to be driving forces in tubulointerstitial fibrosis (TIF) as they activate surrounding fibroblasts. However, the mechanisms remain unclear.Methods: The diabetes with uninephrectomy (DKD) rats were used to evaluated renal histological changes and the expression of Claudin-2 by immunofluorescence staining. Then, Claudin-2 expression in PTECs were modulated and subsequently determined the proliferation and activation of fibroblasts by building a transwell co-culture system in normal glucose (NG)or high glucose (HG) condition.Results: Decreased expression of Claudin-2 in PTECs accompanied by tight junction disruption and increased interstitial fibrosis, were detected in DKD rats. In vitro, downregulated Claudin-2 in PTECs promoted proliferation and activation of fibroblasts, which coincided with elevated expression of profibrotic connective tissue growth factor (CTGF) in PTECs. Silenced CTGF inhibited the profibrotic of PTECs via Claudin-2 inhibition. Fibroblasts co-cultured with PTECs transitioned more to myofibroblasts and generated extracellular matrix (ECM) significantly in response to high glucose (HG) stimulation whereas overexpression of Claudin-2 in PTECs reversed the above results. Upregulating CTGF disrupted the beneficial anti-fibrosis effects obtained by overexpression of Claudin-2 in HG condition.Conclusion: Our study suggested that Claudin-2 in PTECs, a key mediator of paracellular cation and water transport, promotes the activation and proliferation of surrounding fibroblasts significantly via CTGF in a paracrine manner.Keywords: proximal tubular epithelial cell, fibroblast, crosstalk, diabetic kidney disease, Claudin-2

Background: Considering the increasing worldwide prevalence of inflammatory bowel disease (IBD), the early diagnosis of this disease is extremely important. However, non-invasive diagnostic methods remain limited, while invasive techniques are the most commonly used in daily practice. Therefore, there is a serious need to find new non-invasive biomarkers of IBD. Methods: The serum profiles of occludin, claudin-2, and zonulin were assessed in IBD patients using the ELISA method. The levels of the analyzed biomarkers were measured before and after a year of anti-inflammatory treatment, which was a tumor necrosis factor α (TNF-α) inhibitor (adalimumab) in patients with ulcerative colitis (UC) and conventional therapy in patients with Crohn’s disease (CD). Results: In IBD patients, the serum level of occludin (p < 0.001) decreased compared to healthy individuals, while the level of claudin-2 (p < 0.001) increased. Additionally, zonulin (p < 0.01) concentration increased in CD patients compared to the control group. The highest diagnostic ability was presented by occludin measurements with the area under the curve (AUC) of 0.959 (95% CI 0.907–1) in UC and 0.948 (95% CI 0.879–1) in CD. Claudin-2 also demonstrated very good ability in diagnosing UC and CD with AUC values of 0.864 (95% CI 0.776–0.952) and 0.896 (95% CI 0.792–0.999), respectively. The ability of zonulin to diagnose CD was estimated as good with an AUC of 0.74 (95% CI 0.598–0.881). Moreover, a significant correlation was identified between C-reactive protein (CRP), claudin-2 (r = −0.37; p < 0.05), and zonulin (r = −0.44; p < 0.05) in UC patients. Treatment with adalimumab improved the level of occludin, claudin-2, and zonulin in UC patients, while anti-inflammatory conventional therapy decreased the concentration of zonulin in CD. Conclusions: Occludin and claudin-2 measurements present significant utility in diagnosing both UC and CD, while zonulin assessments may be useful in CD diagnosis. Additionally, claudin-2 and zonulin measurements may be helpful in evaluating the intensity of the inflammatory process. Anti-TNF-α treatment improved the value of occludin, claudin-2, and zonulin, indicating its beneficial effect on the integrity of tight junctions in UC.

Claudin‐2 is a tight junction protein found in various tissues including the epidermis of the skin. Intracellular signalling via claudin‐2 may have an effect on cell proliferation and migration. While the role of claudin‐2 in the epidermis has not been established, here we show an increase in claudin‐2 expression in hyperproliferative archival skin samples. To further examine the role of claudin‐2 in cell migration we examined its expression in cultured keratinocytes and found it was increased in wound margins in an in vitro scratch test assay. We then used a claudin‐2 knockdown assay using small interfering ribonucleic acid (siRNA) with a 77% transfection efficiency and decrease in claudin‐2 protein via Western blot analysis to examine cell migration, which was inhibited following claudin‐2 knockdown over a 5‐day period. Cells transfected with claudin‐2 siRNA also showed a decreased size compared to controls and a more diffuse staining pattern. Lastly we examined claudin‐2 expression in migrating keratinocytes by Western blot analysis and found a significant decrease in protein staining in scratch‐test assay cultures after 4 h, followed by a significant increase in claudin‐2 protein after 24 h. Taken together these results indicate a role for claudin‐2 signalling in proliferation and cell migration in the epidermis of the skin. [ABSTRACT FROM AUTHOR]

Gut barrier disintegrity and endotoxin translocation to the liver and systemic circulation are serious clinical complications associated with the stoppage of intestinal bile flow. There is no precise pharmacological option to prevent increased intestinal permeability after bile duct ligation (BDL). Lubiprostone, a chloride channel-2 agonist, has been shown to accelerate restoration of epithelial barrier dysfunction caused by injury, but the exact mechanisms underlying the beneficial effects of lubiprostone on intestine barrier integrity remain unknown. Here, we assessed the beneficial effect of lubiprostone on cholestasis caused by BDL and relevant mechanisms. Male rats were subjected to BDL for 21 days. Seven days after BDL induction, lubiprostone was administered twice daily (10 µg/kg of body weight). Intestinal permeability was assessed through measurements of serum lipopolysaccharide (LPS) concentration. Real-time PCR was conducted to assess expression of intestinal claudin-1 occludin and FXR genes, which are important in preserving the intestinal epithelial barrier integrity, as well as claudin-2 being involved in a leaky gut barrier. Histopathological alterations were also monitored for liver injury. Lubiprostone significantly decreased BDL-induced systemic LPS elevation in rats. BDL induced a significant reduction in FXR, occludin, and claudin-1 genes expression, while increased claudin-2 expression in rat colon. Treatment with lubiprostone significantly restored expression of these genes to the control values. BDL also increased the level of hepatic enzymes ALT, ALP, AST, and total bilirubin, while lubiprostone could preserve the hepatic enzymes and total bilirubin in the treated BDL rats. Lubiprostone also caused a significant reduction in BDL-induced liver fibrosis and intestinal damage in rats. Our results suggest that lubiprostone favorably prevents BDL-induced alterations in intestinal epithelial barrier integrity possibly via modulating intestinal FXRs and tight junction gene expression. [ABSTRACT FROM AUTHOR]

Cingulin (CGN) and paracingulin (CGNL1) are cytoplasmic proteins of tight junctions (TJs), where they play a role in tethering ZO-1 to the actomyosin and microtubule cytoskeletons. The role of CGN and CGNL1 in the barrier function of epithelia is not completely understood. Here, we analyzed the effect of the knock out (KO) of either CGN or CGNL1 or both on the paracellular permeability of monolayers of kidney epithelial (MDCK) cells. KO cells displayed a modest but significant increase in the transepithelial resistance (TER) of monolayers both in the steady state and during junction assembly by the calcium switch, whereas the permeability of the monolayers to 3 kDa dextran was not affected. The permeability to sodium was slightly but significantly decreased in KO cells. This phenotype correlated with slightly increased mRNA levels of claudin-2, slightly decreased protein levels of claudin-2, and reduced junctional accumulation of claudin-2, which was rescued by CGN or CGNL1 but not by ZO-1 overexpression. These results confirm previous observations indicating that CGN and CGNL1 are dispensable for the barrier function of epithelia and suggest that the increase in the TER in clonal lines of MDCK cells KO for CGN, CGNL1, or both is due to reduced protein expression and junctional accumulation of the sodium pore-forming claudin, claudin-2. [ABSTRACT FROM AUTHOR]

Background & aimsThe mechanisms by which macrophages regulate intestinal epithelial cell (IEC) barrier properties are poorly understood. Protein tyrosine phosphatase non-receptor type 2 (PTPN2) protects the IEC barrier from inflammation-induced disruption and regulates macrophage functions. We investigated whether PTPN2 controls interactions between IECs and macrophages to maintain intestinal barrier function.MethodsHuman IEC (Caco-2BBe/HT-29.cl19a cells) and mouse enteroid monolayers were cocultured with human macrophages (THP-1, U937, primary monocyte-derived macrophages from patients with inflammatory bowel disease [IBD]) or mouse macrophages, respectively. We assessed barrier function (transepithelial electrical resistance [TEER] and permeability to 4-kDa fluorescently labeled dextran or 70-kDa rhodamine B-dextran) and macrophage polarization. We analyzed intestinal tissues from mice with myeloid cell-specific deletion of PTPN2 (Ptpn2-LysMCre mice) and mice without disruption of Ptpn2 (controls); some mice were given injections of a neutralizing antibody against interleukin (IL) 6. Proteins were knocked down in macrophages and/or IECs with small hairpin RNAs.ResultsKnockdown of PTPN2 in either macrophages and/or IECs increased the permeability of IEC monolayers, had a synergistic effect when knocked down from both cell types, and increased the development of inflammatory macrophages in macrophage-IEC cocultures. Colon lamina propria from Ptpn2-LysMCre mice had significant increases in inflammatory macrophages; these mice had increased in vivo and ex vivo colon permeability to 4-kDa fluorescently labeled dextran and reduced ex vivo colon TEER. Nanostring analysis showed significant increases in the expression of IL6 in colon macrophages from Ptpn2-LysMCre mice. An IL6-blocking antibody reversed the effects of PTPN2-deficient macrophages, reducing the permeability of IEC monolayers in culture and in Ptpn2-LysMCre mice. Macrophages from patients with IBD carrying a single-nucleotide polymorphism associated with the disease (PTPN2 rs1893217) had the same features of PTPN2-deficient macrophages from mice, including reduced TEER and increased permeability in cocultures with human IEC or mouse enteroid monolayers, which were restored by anti-IL6.ConclusionsPTPN2 is required for interactions between macrophages and IECs; loss of PTPN2 from either cell type results in intestinal barrier defects, and loss from both cell types has a synergistic effect. We provide a mechanism by which the PTPN2 gene variants compromise intestinal epithelial barrier function and increase the risk of inflammatory disorders such as IBD.

The tight junction protein claudin-2 is upregulated in inflammatory bowel disease, and yet its deficit worsens infectious and chemical colitis. In this issue of the JCI, Raju and Shashikanth et al. examined the contribution of claudin-2 to immune-mediated colitis. The authors used transgenic mouse models to show that claudin-2 deficiency attenuated colitis progression as well as a leak barrier defect, albeit at the risk of intestinal obstruction. Further, inhibition of claudin-2 by targeting casein kinase 2 (CK2) also ameliorated colitis. The findings reveal unsuspected links between the pore and leak pathways of intestinal permeability and immune responses leading to colitis. They additionally suggest potential targets for therapeutic intervention in inflammatory bowel disease.

BackgroundAlterations to epithelial tight junctions can compromise the ability of the epithelium to act as a barrier between luminal contents and the underlying tissues, thereby increasing intestinal permeability, an early critical event in inflammatory bowel disease (IBD). Tofacitinib (Xeljanz), an orally administered pan-Janus kinase (JAK) inhibitor, was recently approved for the treatment of moderate to severe ulcerative colitis. Nevertheless, the effects of tofacitinib on intestinal epithelial cell functions are largely unknown. The aim of this study was to determine if JAK inhibition by tofacitinib can rescue cytokine-induced barrier dysfunction in intestinal epithelial cells (IECs).MethodsT84 IECs were used to evaluate the effects of tofacitinib on JAK-signal transducer and activator of transcription (STAT) activation, barrier permeability, and expression and localization of tight junction proteins. The impact of tofacitinib on claudin-2 promoter activity was assessed in HT-29 IECs. Tofacitinib rescue of barrier function was also tested in human colonic stem cell-derived organoids.ResultsPretreatment with tofacitinib prevented IFN-γ-induced decreases in transepithelial electrical resistance (TER) and increases in 4 kDa FITC-dextran permeability (FD4), partly due to claudin-2 transcriptional regulation and restriction of ZO-1 rearrangement at tight junctions. Although tofacitinib administered after IFN-γ challenge only partially normalized TER and claudin-2 levels, FD4 permeability and ZO-1 localization were fully recovered. The IFN-γ-induced FD4 permeability in primary human colonoids was fully rescued by tofacitinib.ConclusionsThese data suggest differential therapeutic efficacy of tofacitinib in the rescue of pore vs leak-tight junction barrier defects and indicate a potential contribution of improved epithelial barrier function to the beneficial effects of tofacitinib in IBD patients.

Epigenetic changes, host–gut microbiota interactions, and environmental factors contribute to inflammatory bowel disease (IBD) onset and progression. A healthy lifestyle may help to slow down the chronic or remitting/relapsing intestinal tract inflammation characteristic of IBD. In this scenario, the employment of a nutritional strategy to prevent the onset or supplement disease therapies included functional food consumption. Its formulation consists of the addition of a phytoextract enriched in bioactive molecules. A good candidate as an ingredient is the Cinnamon verum aqueous extract. Indeed, this extract, subjected to a process of gastrointestinal digestion simulation (INFOGEST), exhibits beneficial antioxidant and anti-inflammatory properties in an in vitro model of the inflamed intestinal barrier. Here, we deepen the study of the mechanisms related to the effect of digested cinnamon extract pre-treatment, showing a correlation between transepithelial electrical resistance (TEER) decrement and alterations in claudin-2 expression under Tumor necrosis factor-α/Interleukin-1β (TNF-α/IL-1) β cytokine administration. Our results show that pre-treatment with cinnamon extract prevents TEER loss by claudin-2 protein level regulation, influencing both gene transcription and autophagy-mediated degradation. Hence, cinnamon polyphenols and their metabolites probably work as mediators in gene regulation and receptor/pathway activation, leading to an adaptive response against renewed insults. [ABSTRACT FROM AUTHOR]

AimsThe role of long non-coding RNA's (lncRNA) in the biology of ulcerative colitis (UC) is not well understood. We have previously detected changes in lncRNA's associated with UC. This study aims to characterize one specific lncRNA, CDKN2B-AS1 whose expression was downregulated in UC patients.Main methodsUC biopsies were used to determine the levels of linear and circular CDKN2B-AS1 relative to healthy controls. In situ hybridization was used to determine the localization of CKDN2B-AS1 in the colon. The intestinal epithelial cell line, Caco-2, was used to study the effects of shRNA mediated loss of CDKN2B-AS1. Transepithelial electrical resistance was used to measure barrier function. An RT-PCR array, immunoblots and immunohistochemistry were used to determine tight junction proteins that CDKN2B-AS1 regulates.Key findingsCDKN2B-AS1 is transcribed into not only linear transcripts but also as circular RNA through back-splicing and both forms are decreased in IBD. CDKN2B-AS1 is expressed mainly in colonic epithelial cells. Cells with down-regulated CDKN2B-AS1 exhibited increased proliferation and no alterations in apoptosis. Targeting both the linear and circular transcripts of CDKN2B-AS1 with short hairpin RNAs enhanced barrier function. We subsequently determined that Claudin-2, a "leaky Claudin" known to decrease barrier function, was decreased in CDKN2B-AS1 knockdown cells.SignificanceThis study identifies a novel lncRNA with both linear and circular transcripts affecting UC biology.

Abstract Background The chronic infection with Helicobacter pylori (H. pylori), especially cytotoxin-associated gene A-positive (CagA+) strains, has been associated with various extragastric disorders. Evaluating the potential impacts of virulence factor CagA on intestine may provide a better understanding of H. pylori pathogenesis such as colitis. The intestinal mucosal barrier is essential for maintaining its integrity and functions. However, how persistent CagA+ H. pylori colonization influences barrier disruption and thereby affects chronic colitis is not fully understood. Results Chronic colitis models of CagA+ H. pylori-colonized mice treated with 2% Dextran sulphate sodium (DSS) were established to assess the disease activity and pertinent expression of tight junction proteins closely related to mucosal integrity. The aggravating effect of CagA+ H. pylori infection on DSS-induced chronic colitis was confirmed in mouse models. In addition, augmented Claudin-2 expression was detected in CagA+ H. pylori infection conditions and selected for mechanistic analysis. Next, GES-1 human gastric epithelial cells were cultured with CagA+ H. pylori or a recombinant CagA protein, and exosomes isolated from conditioned media were then identified. We assessed the Claudin-2 levels after exposure to CagA+ exosomes, CagA− exosomes, and IFN-γ incubation, revealing that CagA+ H. pylori compromised the colonic mucosal barrier and facilitated IFN-γ-induced intestinal epithelial destruction through CagA-containing exosome-mediated mechanisms. Specifically, CagA upregulated Claudin-2 expression at the transcriptional level via a CDX2-dependent mechanism to slow the restoration of wounded mucosa in colitis in vitro. Conclusions These data suggest that exosomes containing CagA facilitate CDX2-dependent Claudin-2 maintenance. The exosome-dependent mechanisms of CagA+ H. pylori infection are indispensable for damaging the mucosal barrier integrity in chronic colitis, which may provide a new idea for inflammatory bowel disease (IBD) treatment.

Claudin-2 (CLDN2), a component of tight junctions, is abnormally expressed in human lung adenocarcinoma tissue. CLDN2 contributes to chemoresistance in human lung adenocarcinoma-derived A549 cells, and it may be a target for cancer therapy. Here, we found that coffee ingredients, namely caffeine and theobromine, decreased the protein level of CLDN2 in human lung adenocarcinoma-derived A549 cells. In contrast, other components, such as theophylline and chlorogenic acid, had no effect. These results indicate that the 7-methyl group in methylxanthines may play a key role in the reduction in CLDN2 expression. The caffeine-induced reduction in the CLDN2 protein was inhibited by chloroquine, a lysosome inhibitor. In a protein-stability assay using cycloheximide, CLDN2 protein levels decreased faster in caffeine-treated cells than in vehicle-treated cells. These results suggest that caffeine accelerates the lysosomal degradation of CLDN2. The accumulation and cytotoxicity of doxorubicin were dose-dependently increased, which was exaggerated by caffeine but not by theophylline in spheroids. Caffeine decreased nuclear factor-erythroid 2-related factor 2 (Nrf2) levels without affecting hypoxia-inducible factor-1α levels. Furthermore, caffeine decreased the expression of Nrf2-targeted genes. The effects of caffeine on CLDN2 expression and anticancer-drug-induced toxicity were also observed in lung adenocarcinoma RERF-LC-MS cells. We suggest that caffeine enhances doxorubicin-induced toxicity in A549 spheroids mediated by the reduction in CLDN2 and Nrf2 expression. [ABSTRACT FROM AUTHOR]

Cingulin (CGN) and paracingulin (CGNL1) are cytoplasmic proteins of tight junctions (TJs), where they play a role in tethering ZO-1 to the actomyosin and microtubule cytoskeletons. The role of CGN and CGNL1 in the barrier function of epithelia is not completely understood. Here, we analyzed the effect of the knock out (KO) of either CGN or CGNL1 or both on the paracellular permeability of monolayers of kidney epithelial (MDCK) cells. KO cells displayed a modest but significant increase in the transepithelial resistance (TER) of monolayers both in the steady state and during junction assembly by the calcium switch, whereas the permeability of the monolayers to 3 kDa dextran was not affected. The permeability to sodium was slightly but significantly decreased in KO cells. This phenotype correlated with slightly increased mRNA levels of claudin-2, slightly decreased protein levels of claudin-2, and reduced junctional accumulation of claudin-2, which was rescued by CGN or CGNL1 but not by ZO-1 overexpression. These results confirm previous observations indicating that CGN and CGNL1 are dispensable for the barrier function of epithelia and suggest that the increase in the TER in clonal lines of MDCK cells KO for CGN, CGNL1, or both is due to reduced protein expression and junctional accumulation of the sodium pore-forming claudin, claudin-2.

The endocannabinoid system of the gastrointestinal tract is involved in the control of intestinal barrier function. Whether the cannabinoid 1 (CB1) receptor is expressed on the intestinal epithelium and acutely regulates barrier function has not been determined. Here, we tested the hypothesis that ligands of the CB1 receptor acutely modulate small intestinal permeability and that this is associated with altered distribution of tight junction proteins. We examined the acute effects of CB1 receptor ligands on small intestinal permeability both in chow-fed and 2-wk high-fat diet (HFD)-fed mice using Ussing chambers. We assessed the distribution of CB1 receptor and tight junction proteins using immunofluorescence and the expression of CB1 receptor using PCR. A low level of CB1 expression was found on the intestinal epithelium. CB1 receptor was highly expressed on enteric nerves in the lamina propria. Neither the CB1/CB2 agonist CP55,940 nor the CB1 neutral antagonist AM6545 altered the flux of 4kDa FITC dextran (FD4) across the jejunum or ileum of chow-fed mice. Remarkably, both CP55,940 and AM6545 reduced FD4 flux across the jejunum and ileum in HFD-fed mice that have elevated baseline intestinal permeability. These effects were absent in CB1 knockout mice. CP55,940 reduced the expression of claudin-2, whereas AM6545 had little effect on claudin-2 expression. Neither ligand altered the expression of ZO-1. Our data suggest that CB1 receptor on the intestinal epithelium regulates tight junction protein expression and restores barrier function when it is increased following exposure to a HFD for 2 wk. [ABSTRACT FROM AUTHOR]

The uterine endometrium uniquely regenerates after menses, postpartum, or after breaks in the uterine layer integrity throughout women's lives. Direct cell-cell contacts ensured by tight and adherens junctions play an important role in endometrial integrity. Any changes in these junctions can alter the endometrial permeability of the uterus and have an impact on the regeneration of uterine layers. Interleukin 22 (IL-22) is a cytokine that is recognized for its role in epithelial regeneration. Moreover, it is crucial in controlling the inflammatory response in mucosal tissues. Here, we studied the role of IL-22 in endometrial recovery after inflammation-triggered abortion. Fecundity of mice was studied i n consecutive ma t i ngs of the same animals after lipopolysaccharide (LPS) (10 µg per mouse)-triggered abortion. The fecundity rate after the second mating was substantially different between IL-22 knockout (IL-22-/-) (9.1%) and wild-type (WT) (71.4%) mice (p < 0.05), while there was no difference between the groups in the initial mating, suggesting that IL-22 deficiency might be associated with secondary infertility. A considerable difference was observed between IL-22-/- and WT mice in the uterine clearance following LPStriggered abortion. Gross examination of the uteri of IL-22-/- mice revealed non-viable fetuses retained inside the horns (delayed clearance). In contrast, all WT mice had completed abortion with total clearance after LPS exposure. We also discovered that IL-22 deficiency is associated with a decreased expression of tight junctions (claudin-2 and claudin-10) and cell surface pathogen protectors (mucin-1). Moreover, IL-22 has a role in the remodeling of the uterine tissue in the inflammatory environment by regulating epithelial-mesenchymal transition markers called E- and N-cadherin. Therefore, IL-22 contributes to the proper regeneration of endometrial layers after inflammation-triggered abortion. Thus, it might have a practical significance to be utilized as a treatment option postpartum (enhanced regeneration function) and in secondary infertility caused by inflammation (enhanced barrier/protector function). [ABSTRACT FROM AUTHOR]

T cell protein tyrosine phosphatase (TCPTP) dephosphorylates a number of substrates, including JAK-STAT (signal transducer and activator of transcription) signaling proteins, which are activated by interferon (IFN)-γ, a major proinflammatory cytokine involved in conditions such as inflammatory bowel disease. A critical function of the intestinal epithelium is formation of a selective barrier to luminal contents. The structural units of the epithelium that regulate barrier function are the tight junctions (TJs), and the protein composition of the TJ determines the tightness of the barrier. Claudin-2 is a TJ protein that increases permeability to cations and reduces transepithelial electrical resistance (TER). We previously showed that transient knockdown (KD) of TCPTP permits increased expression of claudin-2 by IFN-γ. Here, we demonstrate that the decreased TER in TCPTP-deficient epithelial cells is alleviated by STAT1 KD. Moreover, increased claudin-2 in TCPTP-deficient cells requires enhanced STAT1 activation and STAT1 binding to the CLDN2 promoter. We also show that mutation of this STAT-binding site prevents elevated CLDN2 promoter activity in TCPTP-deficient epithelial cells. In summary, we demonstrate that TCPTP protects the intestinal epithelial barrier by restricting STAT-induced claudin-2 expression. This is a potential mechanism by which loss-of-function mutations in the gene encoding TCPTP may contribute to barrier defects in chronic intestinal inflammatory disease.

The uterine endometrium uniquely regenerates after menses, postpartum, or after breaks in the uterine layer integrity throughout women’s lives. Direct cell–cell contacts ensured by tight and adherens junctions play an important role in endometrial integrity. Any changes in these junctions can alter the endometrial permeability of the uterus and have an impact on the regeneration of uterine layers. Interleukin 22 (IL-22) is a cytokine that is recognized for its role in epithelial regeneration. Moreover, it is crucial in controlling the inflammatory response in mucosal tissues. Here, we studied the role of IL-22 in endometrial recovery after inflammation-triggered abortion. Fecundity of mice was studied in consecutive matings of the same animals after lipopolysaccharide (LPS) (10 µg per mouse)-triggered abortion. The fecundity rate after the second mating was substantially different between IL-22 knockout (IL-22−/−) (9.1%) and wild-type (WT) (71.4%) mice (p < 0.05), while there was no difference between the groups in the initial mating, suggesting that IL-22 deficiency might be associated with secondary infertility. A considerable difference was observed between IL-22−/− and WT mice in the uterine clearance following LPS-triggered abortion. Gross examination of the uteri of IL-22−/− mice revealed non-viable fetuses retained inside the horns (delayed clearance). In contrast, all WT mice had completed abortion with total clearance after LPS exposure. We also discovered that IL-22 deficiency is associated with a decreased expression of tight junctions (claudin-2 and claudin-10) and cell surface pathogen protectors (mucin-1). Moreover, IL-22 has a role in the remodeling of the uterine tissue in the inflammatory environment by regulating epithelial–mesenchymal transition markers called E- and N-cadherin. Therefore, IL-22 contributes to the proper regeneration of endometrial layers after inflammation-triggered abortion. Thus, it might have a practical significance to be utilized as a treatment option postpartum (enhanced regeneration function) and in secondary infertility caused by inflammation (enhanced barrier/protector function).

Claudin-2 (CLDN2), a component of tight junction, is involved in the reduction of anticancer drug-induced toxicity in spheroids of A549 cells derived from human lung adenocarcinoma. Fisetin, a dietary flavonoid, inhibits cancer cell growth, but its effect on chemosensitivity in spheroids is unknown. Here, we found that fisetin (20 μM) decreases the protein level of CLDN2 to 22.3%. Therefore, the expression mechanisms were investigated by real-time polymerase chain reaction and Western blotting. Spheroids were formed in round-bottom plates, and anticancer drug-induced toxicity was measured by ATP content. Fisetin decreased the phosphorylated-Akt level, and CLDN2 expression was decreased by a phosphatidylinositol 3-kinase (PI3K) inhibitor, suggesting the inhibition of PI3K/Akt signal is involved in the reduction of CLDN2 expression. Hypoxia level, one of the hallmarks of tumor microenvironment, was reduced by fisetin. Although fisetin did not change hypoxia inducible factor-1α level, it decreased the protein level of nuclear factor erythroid 2-related factor 2, a stress response factor, by 25.4% in the spheroids. The toxicity of doxorubicin (20 μM) was enhanced by fisetin from 62.8% to 40.9%, which was rescued by CLDN2 overexpression (51.7%). These results suggest that fisetin can enhance anticancer drug toxicity in A549 spheroids mediated by the reduction of CLDN2 expression. [ABSTRACT FROM AUTHOR]

Background & Aims: Tight junctions form a barrier to the paracellular passage of luminal antigens. Although most tight junction proteins reside within the apical tight junction complex, claudin-18 localizes mainly to the basolateral membrane where its contribution to paracellular ion transport is undefined. Claudin-18 loss in mice results in gastric neoplasia development and tumorigenesis that may or may not be due to tight junction dysfunction. The aim here was to investigate paracellular permeability defects in stomach mucosa from claudin-18 knockout (Cldn18-KO) mice. Methods: Stomach tissue from wild-type, heterozygous, or Cldn18-KO mice were stripped of the external muscle layer and mounted in Ussing chambers. Transepithelial resistance, dextran 4 kDa flux, and potential difference (PD) were calculated from the chambered tissues after identifying differences in tissue histopathology that were used to normalize these measurements. Marker expression for claudins and ion transporters were investigated by transcriptomic and immunostaining analysis. Results: No paracellular permeability defects were evident in stomach mucosa from Cldn18-KO mice. RNAseq identified changes in 4 claudins from Cldn18–KO mice, particularly the up-regulation of claudin-2. Although claudin-2 localized to tight junctions in cells at the base of gastric glands, its presence did not contribute overall to mucosal permeability. Stomach tissue from Cldn18–KO mice also had no PD versus a lumen-negative PD in tissues from wild-type mice. This difference resulted from changes in transcellular Cl– permeability with the down-regulation of Cl– loading and Cl– secreting anion transporters. Conclusions: Our findings suggest that Cldn18-KO has no effect on tight junction permeability in the stomach from adult mice but rather affects anion permeability. The phenotype in these mice may thus be secondary to transcellular anion transporter expression/function in the absence of claudin-18.

Background: The chronic infection with Helicobacter pylori (H. pylori), especially cytotoxin-associated gene A-positive (CagA+) strains, has been associated with various extragastric disorders. Evaluating the potential impacts of virulence factor CagA on intestine may provide a better understanding of H. pylori pathogenesis such as colitis. The intestinal mucosal barrier is essential for maintaining its integrity and functions. However, how persistent CagA+H. pylori colonization influences barrier disruption and thereby affects chronic colitis is not fully understood. Results: Chronic colitis models of CagA+H. pylori-colonized mice treated with 2% Dextran sulphate sodium (DSS) were established to assess the disease activity and pertinent expression of tight junction proteins closely related to mucosal integrity. The aggravating effect of CagA+H. pylori infection on DSS-induced chronic colitis was confirmed in mouse models. In addition, augmented Claudin-2 expression was detected in CagA+H. pylori infection conditions and selected for mechanistic analysis. Next, GES-1 human gastric epithelial cells were cultured with CagA+H. pylori or a recombinant CagA protein, and exosomes isolated from conditioned media were then identified. We assessed the Claudin-2 levels after exposure to CagA+ exosomes, CagA− exosomes, and IFN-γ incubation, revealing that CagA+H. pylori compromised the colonic mucosal barrier and facilitated IFN-γ-induced intestinal epithelial destruction through CagA-containing exosome-mediated mechanisms. Specifically, CagA upregulated Claudin-2 expression at the transcriptional level via a CDX2-dependent mechanism to slow the restoration of wounded mucosa in colitis in vitro. Conclusions: These data suggest that exosomes containing CagA facilitate CDX2-dependent Claudin-2 maintenance. The exosome-dependent mechanisms of CagA+H. pylori infection are indispensable for damaging the mucosal barrier integrity in chronic colitis, which may provide a new idea for inflammatory bowel disease (IBD) treatment. [ABSTRACT FROM AUTHOR]

Abstract Increased gut permeability facilitates the uptake of food allergens into the bloodstream and triggers allergenic reactions. The present study aimed to evaluate the effect of Lactobacillus pentosus S-PT84 (S-PT84) and Rubus suavissimus leaf extract (RSLE) against egg ovomucoid (OVM)-uptake in lipopolysaccharide (LPS)-induced increased gut-permeability mice model of food allergy. Six-eight weeks old, female C57BL6 mice were continuously fed with LPS (300 μg/kg BW), for 3 months to increase gut permeability. Reduction in the expression of sealing claudin-4, increase in the expression of pore-forming claudin-2, and increase in D-mannitol absorption into the blood plasma in the LPS treated groups suggested the increase in gut permeability after LPS treatment. The oral administration of major egg allergen, OVM, after LPS intervention, significantly increased the plasma mast cell protease-1 and OVM-specific IgE compared to the negative control group. These results indicated that continuous LPS intervention developed OVM-induced food allergy. However, both the treatment of S-PT84 and RSLE suppressed the claudin-2 expression and the gut permeability induced by LPS. Furthermore, S-PT84 and RSLE treatment also reduced the plasma mast cell protease-1 and OVM-specific IgE, indicating the potential beneficial effect against LPS intervention developed OVM-induced food allergy. These findings suggest that S-PT84 and RSLE ameliorated LPS induced gut permeability and food allergic reactions.

Duck circovirus (DuCV) is widely recognized as a prominent virus in China's duck farming industry, known for its ability to cause persistent infections and significant immunosuppression, which can lead to an increased susceptibility to secondary infections, posing a significant threat to the duck industry. Moreover, clinical evidence also indicates the potential vertical transmission of the virus through duck embryos to subsequent generations of ducklings. However, the limited availability of suitable cell lines for in vitro cultivation of DuCV has hindered further investigation into the molecular mechanisms underlying its infection and pathogenicity. In this study, we observed that oral DuCV infection in female breeding ducks can lead to oviduct, ovarian, and follicular infections. Subsequently, the infection can be transmitted to the fertilized eggs, resulting in the emergence of virus-carrying ducklings upon hatching. In contrast, the reproductive organs of male breeding ducks were unaffected by the virus, thus confirming that vertical transmission of DuCV primarily occurs through infection in female breeding ducks. By analyzing transcriptome sequencing data from the oviduct, we focused on claudin-2 , a gene encoding the tight junction protein CLDN2 located on the cell membrane, which showed significantly increased expression in DuCV-infected oviducts of female breeding ducks. Notably, CLDN2 was confirmed to interact with the unique structural protein of DuCV, namely capsid protein (Cap), through a series of experimental approaches including co-immunoprecipitation (co-IP), GST pull-down, immunofluorescence, and adhesion-blocking assays. Furthermore, we demonstrated that the Cap protein binds to the extracellular loop structural domains EL1 and EL2 of CLDN2. Subsequently, by constructing a series of truncated bodies of the CLDN2 promoter region, we identified the transcription factor SP5 for CLDN2. Moreover, we found that DuCV infection triggers the activation of the MAPK-ERK signaling pathway in DEF cells and ducks, leading to an upregulation of SP5 and CLDN2 expression. This process ultimately leads to the transportation of mature CLDN2 to the cell surface, thereby facilitating increased virus adherence to the target organs. In conclusion, we discovered that DuCV utilizes host CLDN2 proteins to enhance adhesion and infection in oviducts and other target organs. Furthermore, we elucidated the signaling pathways involved in the interaction between DuCV Cap proteins and CLDN2, which provides valuable insights into the molecular mechanism underlying DuCV's infection and vertical transmission., Importance: Although duck circovirus (DuCV) poses a widespread infection and a serious hazard to the duck industry, the molecular mechanisms underlying DuCV infection and transmission remain elusive. We initially demonstrated vertical transmission of DuCV through female breeding ducks by simulating natural infection. Furthermore, a differentially expressed membrane protein CLDN2 was identified on the DuCV-infected oviduct of female ducks, and its extracellular loop structural domains EL1 and EL2 were identified as the interaction sites of DuCV Cap proteins. Moreover, the binding of DuCV Cap to CLDN2 triggered the intracellular MAPK-ERK pathway and activated the downstream transcription factor SP5. Importantly, we demonstrated that intracellular Cap also interacts with SP5, leading to upregulation of CLDN2 transcription and facilitating enhanced adherence of DuCV to target tissue, thereby promoting viral infection and transmission. Our study sheds light on the molecular mechanisms underlying vertical transmission of DuCV, highlighting CLDN2 as a promising target for drug development against DuCV infection., Competing Interests: The authors declare no conflict of interest.

Background: Histological mucosal healing has become a paramount target goal to achieve in the treatment of inflammatory bowel diseases. However, there is still a lack of agreement on the best way to reach this goal, since numerous histological scores are available worldwide., Aims: We investigated whether claudin-2, a member of claudin family involved in the regulation of intestinal tight junctions, might be useful to assess the presence of active disease in patients with inflammatory bowel diseases., Methods: Biopsies from 123 patients with ulcerative colitis, Crohn's disease, infectious colitides and irritable bowel syndrome patients where tested with immunohistochemistry for claudin-2., Results: Claudin-2 appeared to be a very sensitive marker of disease activity in inflammatory bowel diseases, but was negative in the other kinds of patients. In addition, immunohistochemistry for claudin-2 showed good reproducibility by different pathologists., Conclusions: Should these findings be confirmed in more numerous cohorts of patients, and especially in those with minimal or focal residual disease activity, this simple assessment could be useful in the routine daily practice to facilitate the task of pathologists and clinicians in the diagnosis and management of patients with inflammatory bowel diseases., Competing Interests: Conflict of interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Tight junction proteins (TJPs) play an important role in gut-barrier dysfunction in cirrhosis and its complications such as acute variceal bleed (AVB). However, the dynamics of TJPs expression after AVB, its relation to bacterial translocation, and impact on clinical outcome is largely unknown. The aim of this study was to study the expression of TJPs in cirrhosis and assess its dynamic changes in AVB. In addition, the relation of TJP expression to endotoxemia and clinical outcomes was assessed. In this prospective pilot study, 17 patients of cirrhosis with AVB, 59 patients of cirrhosis without AVB (non-AVB cirrhosis), and 20 controls were assessed for claudin-2 and claudin-4 expression in the duodenal biopsy. In the AVB-cirrhosis group, additional biopsies were obtained after 3 weeks. Endotoxemia was assessed by measuring IgG anti-endotoxin antibody levels. Claudin expression was correlated with a 6-month survival. Claudin-2 expression was downregulated in patients with AVB and non-AVB cirrhosis in villi (P < 0.001 and 0.013) and crypts (P < 0.001 and 0.012), respectively, compared with the controls. Claudin-4 expression was similar in villi (P = 0.079), but lower in crypts (P = 0.007) in patients with cirrhosis. Claudin-2 expression was upregulated on serial biopsies in both villi and crypts (P = 0.003 and 0.001, respectively) in AVB-cirrhosis with postbleed expression comparable with those with non-AVB cirrhosis. IgG anti-endotoxin antibody levels were elevated in cirrhosis with no correlation with claudin-2/4 expression. Claudin-2 expression independently predicted survival at 6 months. Both claudin-2 and claudin-4 expression are downregulated in cirrhosis. AVB is associated with dynamic changes in TJPs expression. Gut-barrier dysfunction might predict outcomes independent of bacterial endotoxemia in cirrhosis. [ABSTRACT FROM AUTHOR]

Tight junction proteins play crucial roles in maintaining the integrity of intestinal mucosal barrier. MiRNA-182-5p is capable of targeting claudin-2 which is one of the vital tight junction proteins and the effect and mechanism of miRNA-182-5p was explored here in the DSS-induced colitis model. The pathological conditions were evaluated via hematoxylin and eosin staining. The gene expression level was assessed via PCR. Quantitative immunohistochemistry analysis was performed for the measurement of claudin-2. microRNA.org online tool was used for target gene prediction. Luciferase reporter assay and RNA pull-down assay were performed to detect the target of miRNA-182-5p. The inflammatory and oxidative stress level were measured using corresponding kits. MiRNA-182-5p was highly expressed in colitis model and miRNA-182-5p inhibitor exerted protective effects on colitis induced by DSS in mice. The protective effects includded improvement of pathological changes, increases in anti-inflammation and anti-oxidative genes, and up-regulation of TGF-β1. Claudin-2 mRNA was predicted as the target of miRNA-182-5p, which was validated via luciferase reporter assay and RNA pull-down assay. Claudin-2 overexpression was found in miRNA-182-5p inhibitor group. Consistent with the role of miRNA-182-5p, claudin-2 overexpression also exerted protective effects on DSS-induced colitis in mice. Inhibition of miRNA-182-5p exerted protective effects on colitis via targeting and upregulating claudin-2. The findings in study provide a new therapeutic strategy for colitis treatment and lay the foundation for future study. [ABSTRACT FROM AUTHOR]

Claudin-2 is a major component of tight junctions (TJs), which play an important role in reovirus entry into host cells. The Bombyx mori cytoplasmic polyhedosis virus (BmCPV) relates to the cypovirus strain of the reovirus family. So far, the role of claudin-2 in the process of BmCPV infection is not known. In the present study, it was observed that increasing expression of the claudin-2 gene (CLDN2) may concomitantly elevate BmCPV infection. Contrarily, knockdown of CLDN2 expression by siRNAs can reduce BmCPV infection. Similarly, antibody-based blockage of claudin-2 could also decrease BmCPV cell entry. These results suggest that claudin-2 can promote BmCPV infection in vitro. Moreover, immunofluorescence (IF) assays showed that claudin-2 can interact with BmCPV during viral infection. Specifically, co-immunoprecipitation experiments indicated that claudin-2 binds the BmCPV VP7 (instead of VP3 proteins). The interaction between VP7 and claudin-2 was further confirmed by bimolecular fluorescence complementation (BIFC). Altogether, our results suggest that BmCPV cell entry can be promoted upon interaction of VP7 with claudin-2. These findings provide new mechanistic insights related to BmCPV infection. Key points: •Claudin-2 could promote BmCPV infection of cells. •Claudin-2 interacted with BmCPV during BmCPV infection. •Claudin-2 could interact with BmCPV VP7 protein, but not with VP3 proteins. [ABSTRACT FROM AUTHOR]

Histone deacetylase (HDAC) inhibitors have a potential therapeutic role for non-small cell lung cancer (NSCLC). However, more preclinical studies of HDAC inhibitors in NSCLC and normal lung epithelial cells are required to evaluate their antitumor activities and mechanisms. The bicellular tight junction molecule claudin-2 (CLDN-2) is highly expressed in lung adenocarcinoma tissues and increase the proliferation of adenocarcinoma cells. Downregulation of the tricellular tight junction molecule angulin-1/LSR induces malignancy via EGF-dependent CLDN-2 and TGF-β-dependent cellular metabolism in human lung adenocarcinoma cells. In the present study, to investigate the detailed mechanisms of the antitumor activities of HDAC inhibitors in lung adenocarcinoma, human lung adenocarcinoma A549 cells and normal lung epithelial cells were treated with the HDAC inibitors Trichostatin A (TSA) and Quisinostat (JNJ-2648158) with or without TGF-β. Both HDAC inhibitors increased anguin-1/LSR, decrease CLDN-2, promoted G1 arrest and prevented the migration of A549 cells. Furthermore, TSA but not Quisinostat with or without TGF-β induced cellular metabolism indicated as the mitochondrial respiration measured using the oxygen consumption rate. In normal human lung epithelial cells, treatment with TSA and Quisinostat increased expression of LSR and CLDN-2 and decreased that of CLDN-1 with or without TGF-β in 2D culture. Quisinostat but not TSA with TGF-β increased CLDN-7 expression in 2D culture. Both HDAC inhibitors prevented disruption of the epithelial barrier measured as the permeability of FD-4 induced by TGF-β in 2.5D culture. TSA and Quisinostat have potential for use in therapy for lung adenocarcinoma via changes in the expression of angulin-1/LSR and CLDN-2. [ABSTRACT FROM AUTHOR]

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Strenuous exercise triggers deleterious effects on the intestinal epithelium, but their mechanisms are still uncertain. Here, we investigated whether a prolonged training and an additional exhaustive training protocol alter intestinal permeability and the putative effect of alanyl-glutamine (AG) pretreatment in this condition. Rats were allocated into 5 different groups: 1) sedentary; 2 and 3) trained (50 min per day, 5 days per week for 12 weeks) with or without 6 weeks oral (1.5 g/kg) AG supplementation; 4 and 5) trained and subjected to an additional exhaustive test protocol with or without oral AG supplementation. Venous blood samples were collected to determine gasometrical indices at the end of the 12-week protocol or after exhaustive test. Lactate and glucose levels were determined before, during, and after the exhaustive test. Ileum tissue collected after all experimental procedures was used for gene expression analysis of Zonula occludens 1 (ZO-1), occludin, claudin-2, and oligopeptide transporter 1 (PepT-1). Intestinal permeability was assessed by urinary lactulose/mannitol test collected after the 12-week protocol or the exhaustive test. The exhaustive test decreased pH and base excess and increased pCO2. Training sessions delayed exhaustion time and reduced the changes in blood glucose and lactate levels. Trained rats exhibited upregulation of PEPT-1, ZO-1, and occludin mRNA, which were partially protected by AG. Exhaustive exercise induced intestinal paracellular leakage associated with the upregulation of claudin-2, a phenomenon protected by AG treatment. Thus, AG partially prevented intestinal training adaptations but also blocked paracellular leakage during exhaustive exercise involving claudin-2 and occludin gene expression.

The distribution of complementary metabolic functions in hepatocytes along a portocentral axis is called liver zonation. Endothelial secreted Wnt ligands maintain metabolic zonation in the adult murine liver but whether those ligands are necessary to initiate zonation in the immature liver has been only partially explored. Also, numerous non-metabolic proteins display zonated expression in the adult liver but it is not entirely clear if their localization requires endothelial Wnts. Here we used a novel transgenic mouse model to compare the spatial distribution of zonated non-metabolic proteins with that of typical zonated metabolic enzymes during liver maturation and after acute injury induced by carbon tetrachloride (CCl4). We also investigated how preventing Wnt ligand secretion from endothelial cells affects zonation patterns under homeostasis and after acute injury. Our study demonstrates that metabolic and non-metabolic zonation are established non-synchronously during maturation and regeneration and require multiple endothelial Wnt sources.

Abstract Background In an earlier study, we investigated the expression of tight junction protein claudins (CLDNs) in human osteosarcoma (OS) cells, and the CLDN2 was found to be down-regulated in primary tumor cells compared with normal osteoblast cells. Here, we sought to explore the effects of CLDN2 on the malignant phenotype of OS and the underlying molecular mechanisms. Methods The expression patterns of CLDN2 and afadin in OS tissues and histologically non-neoplastic bone tissues were explored via immunohistochemistry and western blotting. CLDN2 expression levels in an OS cell line stably expressing CLDN2 and an osteoblast cell line with a CLDN2 knockout were confirmed by western blotting and immunofluorescence staining. The malignant phenotype of OS cells and osteoblast cells in vitro was assessed using a cell counting kit-8 assay, transwell assay and wound-healing experiment. Western blotting was utilized to detect the activation state of Ras/Raf/MEK/ERK pathway. Moreover, an RNA interference method were used to silence afadin in CLDN2-expressing OS cells. Results Our research group found that CLDN2 and afadin was underexpressed in OS tissues, and the overexpression of CLDN2 significantly inhibited the migration abilities of OS cells. Genetic silencing of afadin in CLDN2-overexpressing OS cells promoted U2OS cell motility and activation of the Ras/Raf/MEK/ERK pathway. Conclusions In this study, we confirmed that CLDN2 expression significantly inhibited the malignant phenotype of OS cells in vitro. Inhibition of the ERK pathway by afadin may be one of the mechanisms by which CLDN2 blocks the metastasis phenotype of OS cells.

Abstract Background Several studies have suggested that claudin proteins, which are the main components of tight junction structures, are related to the regulation of cell polarity and cell differentiation. Method To explore the expression profiles of the tight junction proteins claudin-2, − 5, − 8 and − 9 in nasopharyngeal carcinoma, IHC (immunohistochemical analysis), Western blot and real-time PCR were used to detect the expression profiles of these claudin proteins in nasopharyngeal carcinoma tissues and in non-neoplastic mucosal tissues. Results According to our study, the expression levels of claudin-2 and claudin-5 were reduced, while the expression of claudin-8 was increased in nasopharyngeal carcinoma tissues in comparison with non-neoplastic mucosal tissues. Correlations between claudin-2 and -5 expression and metastatic progression in nasopharyngeal carcinoma patients were also found. Conclusion In summary, our research reveals distinct expression profiles of claudin-2, − 5 and − 8 in non-neoplastic mucosal tissues and nasopharyngeal carcinoma tissues. In addition, the expression of these claudin proteins was highly correlated with metastatic progression and prognosis in patients with nasopharyngeal carcinoma and had predictive value for the metastasis and survival of nasopharyngeal carcinoma patients.

BACKGROUND: Previous studies have found that chronic cerebral hypoperfusion can cause changes in the intestinal mucosal barrier in rats, and Occludin and Claudin are important components of the intestinal mucosal barrier. OBJECTIVE: To explore the effect of chronic cerebral hypoperfusion on the ileum mucosal barrier. METHODS: Twenty male Sprague-Dawley rats were randomly divided into sham operation (sham) group and chronic cerebral hypoperfusion model group. Animal model of chronic cerebral hypoperfusion was built by permanent bilateral common carotid artery ligation. The bilateral common carotid arteries of the sham group were isolated without ligation. The animals were killed at 4 weeks after operation, and their ileums were isolated for morphological observation and pathological scoring using hematoxylin-eosin staining. Apoptosis in the ileum cells was detected using TUNEL. Western blot was used to detect the expression of Claudin-2 and Occludin. immunohistochemical staining was used to detect the expression of Occludin. The study protocol was approved by the Animal Experiment Ethics Committee of General Hospital of Western Theater Command. RESULTS AND CONCLUSION: Compared with the sham group, hematoxylin-eosin staining showed that the ileum in the model group did not develop obvious injury with no higher pathological scores (P>0.05). Westernblot results indicated that compared with the sham group, the expression of claudin-2 was increased in the ileum tissue of the rats with chronic cerebral hypoperfusion, while the expression of Occludin was decreased (P< 0.05). TUNEL fluorescence staining showed that the apoptotic rate in the model group was significantly increased compared with the sham group (P<0.05). Immunohistochemical results showed that the expression of Occludin in the ileum tissue of the chronic cerebral hypoperfusion group was significantly reduced (P<0.05). These findings indicate that chronic cerebral hypoperfusion may damage the intestinal mucosal barrier by downregulating Occludin expression and upregulating Claudin-2 expression. [ABSTRACT FROM AUTHOR]

Claudin-2 (CLDN-2) is a leaky-type tight junction protein, and its overexpression increases tumorigenesis of some types of cancer cells. In the present study, to examine the possibility of targeting CLDN-2 in the therapy for endometrioid endometrial adenocarcinoma, we investigated the regulation and role of CLDN-2 in endometriosis and endometrioid endometrial adenocarcinoma. In endometrioid endometrial adenocarcinoma tissues, marked upregulation of CLDN-2 was observed together with malignancy, while in endometriosis tissues, a change in the localization of CLDN-2 was observed. In cells of the endometrial adenocarcinoma cell line Sawano, which highly express CLDN-2, downregulation of CLDN-2 induced by the siRNA upregulated the epithelial barrier and inhibited cell migration. Furthermore, the downregulation of CLDN-2 affected the cell cycle and inhibited cell proliferation. In Sawano cells cultured with high-glucose medium, CLDN-2 expression was downregulated at the mRNA and protein levels. The high-glucose medium upregulated the epithelial barrier, cell proliferation, and migration, and inhibited cell invasion. The histone deacetylase (HDAC) inhibitor tricostatin A (TSA), which has antitumor effects, downregulated CLDN-2 expression, cell proliferation, invasion, and migration, and upregulated the epithelial barrier. The mitochondrial respiration level, an indicator of cancer metabolism, was downregulated by CLDN-2 knockdown and upregulated by the high-glucose condition. Taken together, these results indicated that overexpression of CLDN-2 closely contributed to the malignancy of endometrioid endometrial adenocarcinoma. Downregulation of CLDN-2 via the changes of the glucose concentration and treatment with HDAC inhibitors may be important in the therapy for endometrial cancer. [ABSTRACT FROM AUTHOR]

The present study aimed to determine the expression level of claudin-2 (CLDN2) in the peripheral blood of patients with sepsis, and to investigate its potential function and mechanism of action in vascular endothelial injury. A total of 25 patients with sepsis were included in the present study. Reverse transcription-quantitative PCR was used to determine CLDN2 levels in peripheral blood. HUVECs stably expressing CLDN2 were prepared and Cell Counting Kit-8, flow cytometry and Transwell assays were performed to study the proliferation, apoptosis and migration of HUVECs, respectively. Using bioinformatics, microRNA (miR) molecules that interact with CLDN2 were predicted. A dual luciferase reporter assay was used to test whether miR-331 regulated CLDN2. Western blotting was employed to determine CLDN2 protein expression. In addition, in vitro transfection of HUVECs with miR-331 mimics was performed to test the rescue effects of miR-331 on the cell function changes induced by CLDN2. The results indicated that elevated CLDN2 expression altered the proliferation and cell cycle of peripheral vascular endothelial cells. CLDN2 overexpression inhibited HUVEC proliferation via mechanisms not associated with the cell cycle. CLDN2 mRNA levels in the peripheral blood of patients with sepsis were significantly higher than those in healthy subjects. Upregulated CLDN2 expression promoted the apoptosis of HUVECs, but reduced their proliferation and migration. Notably, miR-331 was able to bind with CLDN2 mRNA and regulate its expression. Upregulation of miR-331 expression inhibited the expression of CLDN2 and restored nearly normal proliferation, apoptosis and migration to HUVECs. The present study demonstrated that CLDN2 expression is elevated in peripheral blood from patients with sepsis, and promotes the injury of vascular endothelial cells. In addition, miR-331 participates in the direct regulation of CLDN2, and upregulation of miR-331 expression inhibits the expression of CLDN2 and restores cellular functions to HUVECs. [ABSTRACT FROM AUTHOR]

A non-histone chromatin-associated protein, high mobility group box 1 (HMGB1), which impairs the airway epithelial barrier, is involved in the induction of airway inflammation in patients with allergy, asthma, chronic obstructive pulmonary disease (COPD), and idiopathic pulmonary fibrosis (IPF). Tricellular tight junctions (tTJs) form at the convergence of bicellular tight junctions (bTJs). Angulin-1/lipolysis-stimulated lipoprotein receptor (LSR) is a novel molecule present at tricellular contacts and contributes to the epithelial barrier and cellular metabolism. Adenosine monophosphate-activated protein kinase (AMPK) is a central metabolic regulator and has a reciprocal association with TJs. In the present study, to examine how HMGB1 contributes to airway epithelial barrier disruption and the cellular metabolism indicated as mitochondrial respiration, bronchial epithelial Calu-3 cells were transfected with siRNAs of angulin-1/LSR or treated with HMGB1 and the relationship between HMGB1 and angulin-1/LSR was investigated. Knockdown of angulin-1/LSR upregulated the expression of the tight junction molecule claudin-2, AMPK activity, and mitochondrial respiration, and downregulated the epithelial barrier. Treatment with HMGB1 downregulated angulin-1/LSR expression and the epithelial barrier, and upregulated claudin-2 expression, AMPK activity and mitochondrial respiration. Treatment with EW-7197, a transforming growth factor-β (TGF-β) type I receptor kinase inhibitor, prevented all the effects of HMGB1 in Calu-3 cells. HMGB1-downregulated angulin-1/LSR induced epithelial barrier disruption via claudin-2 and cellular metabolism via AMPK in airway epithelial Calu-3 cells. The effects of HMGB1 contribute to TGF-β signaling and EW-7197 shows potential for use in therapy for HMGB1-induced airway inflammation. •HMGB1-impaired airway epithelial barrier is involved in airway inflammation. •Angulin-1/LSR contributes to the airway epithelial barrier and cellular metabolism. •AMPK has a reciprocal association with cellular metabolism and tight junctions. • Downregulation of angulin-1/LSR induces claudin-2 and AMPK activity. •HMGB1-downregulated LSR induces barrier disruption and cellular metabolism. [ABSTRACT FROM AUTHOR]

Purpose: Claudins as the major components of tight junctions are important in maintaining cell-cell integrity and thus function as a barrier. Dysregulation of the claudins is often associated with loss of the epithelial phenotype, a process called epithelial-mesenchymal transition (EMT), which most often results in gain of migrative and invasive properties. However, the role of claudins in the endometrium or endometriosis has only rarely been examined.Methods: In this study, we investigated localization of claudin-2 and claudin-3 in the eutopic and ectopic endometrium with immunohistochemistry. A detailed quantification with HSCORE was performed for claudin-2 and claudin-3 in endometrium without endometriosis and in cases with endometriosis compared to the three endometriotic entities: peritoneal, ovarian, and deep-infiltrating endometriosis.Results: We found a preferential localization of both claudins in the glandular and the luminal epithelial cells in the endometrium with and without endometriosis. Quantification of localization of both claudins showed no differences in eutopic endometrium of control cases compared to cases with endometriosis. Furthermore, both claudins are localized highly similar in the ectopic compared to the eutopic endometrium, which is in clear contrast to previously published data for claudin-3.Conclusion: From our results, we conclude that localization of claudin-2 and claudin-3 is highly stable in eutopic and ectopic endometrium without any loss of the epithelial phenotype and thus do not contribute to the pathogenesis of endometriosis. [ABSTRACT FROM AUTHOR]

Background/Aims To determine whether the expression of tight junction proteins (TJPs) differs depending on the subtype of functional dyspepsia (FD) and sex. Methods Control (n = 95) and FD (n = 165) groups based on Rome III criteria were prospectively enrolled. Gastric mucosal mRNA expression levels of various TJPs (claudins [CLDN] 1, 2, and 4; zonula occludens-1; occludin [OCLN]) were assessed by reverse transcription polymerase chain reaction. Western blot was performed to determine the levels of various TJPs. Helicobacter pylori infection status was evaluated by histology, rapid urease test, and culture. Questionnaires were analyzed. Results In all groups irrespective of H. pylori, FD group showed significantly higher CLDN2 mRNA levels than control group (P = 0.048). The level of CLDN4 mRNA expression was significantly lower in female FD group than in male FD group (P = 0.018). In H. pylori uninfected subjects, the level of CLDN1 mRNA expression in female FD group was significantly lower than that of male FD group (P = 0.014). The level of CLDN2 mRNA expression was significantly higher in the male postprandial distress syndrome (P = 0.001) and male epigastric pain syndrome (P = 0.023) groups than in the male control group. In Western blot analysis, the expression of OCLN was significantly elevated 48 hour after the culture with H. pylori strain 43504. Conclusions H. pylori can affect a variety of TJPs, particularly claudin-4 and occludin. Claudin-2 is thought to be involved in FD irrespective of H. pylori status, especially in the pathophysiology of male FD. [ABSTRACT FROM AUTHOR]

Objective: To investigate the effects and possible mechanisms of action of Curcuma wenyujin Y. H. Chen et C. Ling n-Butyl alcohol extract (CWNAE) on repression of human gastric cancer (GC) AGS cell invasion induced by co-culturing with Helicobacter pylori (HP). Methods: AGS cells were cultured with HP of positive or negative cytotoxin-associated gene A (CagA) and vacuolating cytotoxin gene A (VacA) expression (CagA+/- or VacA+/-) and divided into 5 group. Group A was cultured without HP as a control, Group B with HPCagA+VacA+, Group C with HPCagA-VacA-, Group D with HPCagA+VacA+ and CWNAE, and Group E with HPCagA-VacA- and CWNAE. Methylthiazolyldiphenyl-tetrazolium bromide (MTT) and tumor invasion assays, examinations of morphology and ultramicroscopic structures, quantitative real-time polymerase chain reaction and Western blots were performed to measure the effects and uncover the mechanisms behind these effects of HPCagA+VacA+ and CWNAE on the epithelial-mesenchymal transition (EMT) of AGS cells. Results: The 10% inhibitory concentration of CWNAE against AGS cells after a 48 h incubation was 19.73±1.30 μg/mL. More AGS cells were elongated after co-culturing with HPCagA+VacA+ than after culturing with HPCagA-VacA-. In tumor invasion assays, HPCagA+VacA+ significantly enhanced the invasiveness of AGS cells compared to the other experimental groups (all P value <0.05), and this effect was inhibited by CWNAE. Treatment with CWNAE normalized tight junctions and reduced the number of pseudopodia of AGS cells co-cultured with HPCagA+VacA+. HPCagA+VacA+ up-regulated zincfinger ebox binding homeobox 1 (ZEB1) in AGS cells after co-culturing for 24 h. Expression of caudal type homeobox transcription factor (CDX-2) and claudin-2 was significantly increased by HPCagA+VacA+ (P<0.05), but not by HPCagA-VacA-. Conclusion: HPCagA+VacA+ promoted the invasiveness of AGS cells through up-regulation of ZEB1 transcription and claudin-2 and CDX-2 expression. CWNAE inhibited these effects of HPCagA+VacA+ on AGS cells by down-regulating ZEB1 transcription, and CDX-2 and claudin-2 expression. [ABSTRACT FROM AUTHOR]

Claudin-2-dependent pore function mediates paracellular cation permeability and can result in pathogenesis of many diseases. Although existing various types of claudins, including barrier-forming and pore-forming claudins, their heterodimeric interaction affecting barrier and pore functions has never been fully elucidated yet. Recently, Shashikanth and colleagues demonstrated that expression of claudin-4 was able to antagonize paracellular pore activity of claudin-2. This commentary will emphasize the mechanism underlying claudin-4-mediated claudin-2-dependent pore inhibition and discuss its potential therapeutic and prognostic applications. [ABSTRACT FROM AUTHOR]

Oxidative phosphorylation (OXPHOS) consists of four enzyme complexes and ATP synthase, and is crucial for maintaining physiological tissue and cell growth by supporting the main bioenergy pool. Cytochrome c oxidase (COX) has been implicated as a primary regulatory site of OXPHOS. Recently, COX subunit 5B (COX5B) emerged as a potential biomarker associated with unfavorable prognosis by modulating cell behaviors in specific cancer types. However, its molecular mechanism remains unclear, particularly in colorectal cancers (CRCs). To understand the role of COX5B in CRCs, the expression and postoperative outcome associations using independent in-house patient cohorts were evaluated. A higher COX5B tumor/nontumor expression ratio was associated with unfavorable clinical outcomes (p = 0.001 and 0.011 for overall and disease-free survival, respectively. In cell-based experiments, the silencing of COX5B repressed cell growth and enhanced the susceptibility of CRCs cells to anticancer drugs. Finally, downstream effectors identified by RNA sequencing followed by RT-qPCR and functional compensation experiments revealed that the tight junction protein Claudin-2 (CLDN2) acts downstream of COX5B-mediated bioenergetic alterations in controlling cell growth and the sensitivity to anticancer drugs in CRCs cells. In conclusion, it was found that COX5B promoted cell growth and attenuated anticancer drugs susceptibility in CRCs cells by orchestrating CLDN2 expression, which may contribute to unfavorable postoperative outcomes of patients with CRCs.