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Li H, Wang K, Hao M, Liu Y, Liang X, Yuan D, Ding L. Intestinal epithelial Cldn-7 regulates intestinal inflammation by altering the gut microbiota. Pathol Res Pract 2024; 260:155448. [PMID: 39004000 DOI: 10.1016/j.prp.2024.155448] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/01/2024] [Revised: 06/05/2024] [Accepted: 06/30/2024] [Indexed: 07/16/2024]
Abstract
BACKGROUND AND AIM Tight junctions maintain gut homeostasis by forming a physical barrier that protects the gut from invasion by microbiota. Cldn-7 is an important component involved in this protection, but the relationship between Cldn-7, intestinal inflammation, and gut microbiota has not been clarified. Here, we hypothesize that Cldn-7 depletion affects intestinal inflammation by altering the gut microbiota. METHODS Based on the induced intestinal condition of Cldn-7 knockout mice (Cldn7fl/fl;villin-CreaERT2), we established the intestinal flora depletion model and colitis model by antibiotic drinking and feeding with dextran sodium sulfate (DSS). The environment of Cldn-7 gene deletion mice was changed by co-housing experiment. AB-PAS staining and Muc2 were used to detect the effect of co-housing and Cldn-7 deficiency on the mucus layer after flora depletion. qRT-PCR was used to detect the expression of intestinal inflammatory factors and AMPs in mice. Feces were collected and proportions of microbiota were analyzed by 16 S rRNA amplicon sequencing. RESULTS Mice in the co-housing experiment had altered intestinal microbiota, including diversity, composition, and functional prediction, compared to controls. Intestinal inflammation was restored to some extent following altered intestinal microbiota. The intestinal inflammation caused by Cldn-7 deficiency and susceptibility to DSS could be reduced after antibiotic administration compared to controls, in terms of phenotype, pathological changes, inflammatory factors, mucus barrier, and expression of AMPs. CONCLUSIONS In analyses of intestinal tissues, colitis induction, and gut microbiota in mice with intestinal disruption of Cldn-7, we found this protein to prevent intestinal inflammation by regulating the gut microbiota. Cldn-7might therefore be an important mediator of host-microbiome interactions. Our research has revealed that Cldn-7 plays an indispensable role in maintaining intestinal homeostasis by regulating the gut microbiota and impacting intestinal inflammation. These findings provide new insights into the pathogenesis of ulcerative colitis.
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Affiliation(s)
- Huimin Li
- Department of Gastrointestinal Oncology Surgery, Beijing Shijitan Hospital, Capital Medical University, Beijing 100038, China
| | - Kun Wang
- Department of Gastrointestinal Oncology Surgery, Beijing Shijitan Hospital, Capital Medical University, Beijing 100038, China
| | - Mengdi Hao
- Department of Gastrointestinal Oncology Surgery, Beijing Shijitan Hospital, Capital Medical University, Beijing 100038, China
| | - Yin Liu
- Department of Gastrointestinal Oncology Surgery, Beijing Shijitan Hospital, Capital Medical University, Beijing 100038, China
| | - Xiaoqing Liang
- Department of Gastrointestinal Oncology Surgery, Beijing Shijitan Hospital, Capital Medical University, Beijing 100038, China
| | - Dajin Yuan
- Department of Gastrointestinal Oncology Surgery, Beijing Shijitan Hospital, Capital Medical University, Beijing 100038, China
| | - Lei Ding
- Department of Gastrointestinal Oncology Surgery, Beijing Shijitan Hospital, Capital Medical University, Beijing 100038, China.
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Gao YN, Wang ZW, Su CY, Wang JQ, Zheng N. Omics analysis revealed the intestinal toxicity induced by aflatoxin B1 and aflatoxin M1. ECOTOXICOLOGY AND ENVIRONMENTAL SAFETY 2024; 278:116336. [PMID: 38691883 DOI: 10.1016/j.ecoenv.2024.116336] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/09/2024] [Revised: 04/01/2024] [Accepted: 04/12/2024] [Indexed: 05/03/2024]
Abstract
Aflatoxin B1 (AFB1), a common mycotoxin, can occur in agricultural products. As a metabolite of AFB1, aflatoxin M1 (AFM1) mainly exist in dairy products. These two mycotoxins threaten human health, although it is unclear how they affect the function of the intestinal barrier. In this study, mice were exposed to AFB1 (0.3 mg/kg body b.w.) and AFM1(3.0 mg/kg b.w.) either individually or in combination for 28 days to explore the main differentially expressed proteins (DEPs) and the associated enriched pathways. These findings were preliminarily verified by the transcriptomic and proteomic analyses in differentiated Caco-2 cells. The results revealed that AFB1 and AFM1 exposure in mice disrupted the function of the intestinal barrier, and the combined toxicity was greater than that of each toxin alone. Further proteomic analysis in mice demonstrated that the mechanisms underlying these differences could be explained as follows: (i) lipid metabolism was enriched by AFB1-induced DEPs. (ii) protein export pathway was stimulated by AFM1-induced DEPs. (iii) cell metabolic ability was inhibited (as evidenced by changes in UDP-GT1, UDP-GT2, and Gatm6), apoptosis was induced (MAP4K3), and epithelial cell integrity was disrupted (Claudin7 and IQGAP2), resulting in more extensive intestinal damage after combined treatment. In conclusion, the hazardous impact of co-exposure to AFB1 and AFM1 from proteomic perspectives was demonstrated in the present study.
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Affiliation(s)
- Ya-Nan Gao
- Key Laboratory of Quality & Safety Control for Milk and Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China; Laboratory of Quality and Safety Risk Assessment for Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China; Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Zi-Wei Wang
- Key Laboratory of Quality & Safety Control for Milk and Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China; Laboratory of Quality and Safety Risk Assessment for Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Chuan-You Su
- College of Animal Science, Henan Agriculture University, Zhengzhou 450000, China
| | - Jia-Qi Wang
- Key Laboratory of Quality & Safety Control for Milk and Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China; Laboratory of Quality and Safety Risk Assessment for Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Nan Zheng
- Key Laboratory of Quality & Safety Control for Milk and Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China; Laboratory of Quality and Safety Risk Assessment for Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
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Wei W, Li W, Yang L, Weeramantry S, Ma L, Fu P, Zhao Y. Tight junctions and acute kidney injury. J Cell Physiol 2023; 238:727-741. [PMID: 36815285 DOI: 10.1002/jcp.30976] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2022] [Revised: 01/27/2023] [Accepted: 02/04/2023] [Indexed: 02/24/2023]
Abstract
Acute kidney injury (AKI) is characterized by a rapid reduction in kidney function caused by various etiologies. Tubular epithelial cell dysregulation plays a pivotal role in the pathogenesis of AKI. Tight junction (TJ) is the major molecular structure that connects adjacent epithelial cells and is critical in maintaining barrier function and determining the permeability of epithelia. TJ proteins are dysregulated in various types of AKI, and some reno-protective drugs can reverse TJ changes caused by insult. An in-depth understanding of TJ regulation and its causality with AKI will provide more insight to the disease pathogenesis and will shed light on the potential role of TJs to serve as novel therapeutic targets in AKI.
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Affiliation(s)
- Wei Wei
- Division of Nephrology and Kidney Research Institute, West China Hospital, Sichuan University, Chengdu, Sichuan, China
| | - Weiying Li
- Department of Internal Medicine, Florida Hospital/AdventHealth, Orlando, Florida, USA
| | - Letian Yang
- Division of Nephrology and Kidney Research Institute, West China Hospital, Sichuan University, Chengdu, Sichuan, China
| | - Savidya Weeramantry
- Department of Internal Medicine, West China School of Medicine, Sichuan University, Chengdu, Sichuan, China
| | - Liang Ma
- Division of Nephrology and Kidney Research Institute, West China Hospital, Sichuan University, Chengdu, Sichuan, China
| | - Ping Fu
- Division of Nephrology and Kidney Research Institute, West China Hospital, Sichuan University, Chengdu, Sichuan, China
| | - Yuliang Zhao
- Division of Nephrology and Kidney Research Institute, West China Hospital, Sichuan University, Chengdu, Sichuan, China
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Cetina-Palma A, Namorado-Tónix C, Rodríguez-Muñoz R, Vergara P, Reyes-Sánchez JL, Segovia J. Characterization of the pattern of expression of Gas1 in the kidney during postnatal development in the rat. PLoS One 2023; 18:e0284816. [PMID: 37093844 PMCID: PMC10124827 DOI: 10.1371/journal.pone.0284816] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2022] [Accepted: 04/09/2023] [Indexed: 04/25/2023] Open
Abstract
Growth Arrest-Specific 1 (Gas1) is a pleiotropic protein with different functions, in the adult kidney Gas1 acts as an endogenous inhibitor of cell proliferation but it is also necessary for the maintenance and proliferation of Renal Progenitor Cells (RPC) during early development, thus it fulfills important functions in the adult kidney. However, it is not known whether or not Gas1 is expressed during postnatal development, a critical stage for renal maturation. For this reason, the main objective of this work was to characterize the expression pattern of Gas1 in the different regions of the kidney by immunofluorescence and Western blot analysis during the postnatal development of the rat. We found that Gas1 is present and has a differential expression pattern in the various regions of the nephron during postnatal development. We observed that the highest levels of expression of Gas1 occur in the adult, however, Gas1 is also expressed in RPC and interestingly, the expression of RPC markers such as the Neural cell adhesion molecule (NCAM) and Cluster of differentiation 24 (CD24) were found to have an inverse pattern of expression to Gas1 (decreases as the kidney matures) during postnatal renal maturation, this indicates a role for Gas1 in the regulation of renal cell proliferation at this stage of development.
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Affiliation(s)
- Andrea Cetina-Palma
- Departamento de Farmacología, Centro de Investigación y de Estudios Avanzados del IPN, Mexico City, Mexico
| | - Carmen Namorado-Tónix
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados del IPN, Mexico City, Mexico
| | - Rafael Rodríguez-Muñoz
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados del IPN, Mexico City, Mexico
| | - Paula Vergara
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados del IPN, Mexico City, Mexico
| | - José Luis Reyes-Sánchez
- Departamento de Farmacología, Centro de Investigación y de Estudios Avanzados del IPN, Mexico City, Mexico
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados del IPN, Mexico City, Mexico
| | - José Segovia
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados del IPN, Mexico City, Mexico
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Li J. Context-Dependent Roles of Claudins in Tumorigenesis. Front Oncol 2021; 11:676781. [PMID: 34354941 PMCID: PMC8329526 DOI: 10.3389/fonc.2021.676781] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/06/2021] [Accepted: 07/05/2021] [Indexed: 12/16/2022] Open
Abstract
The barrier and fence functions of the claudin protein family are fundamental to tissue integrity and human health. Increasing evidence has linked claudins to signal transduction and tumorigenesis. The expression of claudins is frequently dysregulated in the context of neoplastic transformation. Studies have uncovered that claudins engage in nearly all aspects of tumor biology and steps of tumor development, suggesting their promise as targets for treatment or biomarkers for diagnosis and prognosis. However, claudins can be either tumor promoters or tumor suppressors depending on the context, which emphasizes the importance of taking various factors, including organ type, environmental context and genetic confounders, into account when studying the biological functions and targeting of claudins in cancer. This review discusses the complicated roles and intrinsic and extrinsic determinants of the context-specific effects of claudins in cancer.
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Affiliation(s)
- Jian Li
- Department of General Surgery, The Third Hospital of Mianyang, Sichuan Mental Health Center, Mianyang, China
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Xu C, Ding YH, Wang K, Hao M, Li H, Ding L. Claudin-7 deficiency promotes stemness properties in colorectal cancer through Sox9-mediated Wnt/β-catenin signalling. J Transl Med 2021; 19:311. [PMID: 34281572 PMCID: PMC8287764 DOI: 10.1186/s12967-021-02983-3] [Citation(s) in RCA: 19] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2021] [Accepted: 07/11/2021] [Indexed: 02/08/2023] Open
Abstract
Background Colorectal cancer (CRC) is a common malignant tumour of the digestive tract that is characterized by high patient morbidity and mortality rates. Claudin-7 (Cldn7), a tight junction protein, was recently reported to function as a candidate tumour suppressor gene in CRC. Our previous study demonstrated that the large intestine of C57/BL6 mice showed intestinal adenomas and abnormal Ki67 expression and distribution in the intestinal crypt when Cldn7 was knocked out. The aim of this study was to further investigate whether Cldn7 deficiency has non-tight junction functions, affects intestinal stemness properties, promotes CRC and to determine the specific mechanism. Methods Cell proliferation assays, migration assays, apoptosis assays, tumour sphere formation assays in vitro, and subcutaneous xenograft models in vivo were used to determine the effects of Cldn7 knockdown on the biological characteristics of CRC stem cells. Western blotting, qPCR and immunofluorescence staining were performed to identify the epithelial-mesenchymal transition and the activation of Wnt/β-catenin pathway in CRC stem cells. Cldn7 inducible conditional gene knockout mice and immunohistochemical staining further verified this hypothesis in vivo. The mechanism and target of Cldn7 were determined by performing a chromatin immunoprecipitation (ChIP) assay and coimmunoprecipitation (CoIP) assay. Results Cldn7 knock down in CRC stem cells promoted cell proliferation, migration, and globular growth in serum-free medium and the ability to form xenograft tumours; cell apoptosis was inhibited, while the cellular epithelial-mesenchymal transition was also observed. These changes in cell characteristics were achieved by activating the Wnt/β-catenin pathway and promoting the expression of downstream target genes after β-catenin entry into the nucleus, as observed in CRC cell lines and Cldn7 gene knockout mouse experiments. Using ChIP and CoIP experiments, we initially found that Cldn7 and Sox9 interacted at the protein level to activate the Wnt/β-catenin pathway. Conclusions Based on our research, Cldn7 deficiency confers stemness properties in CRC through Sox9-mediated Wnt/β-catenin signalling. This result clarifies that Cldn7 plays an inhibitory role in CRC and reveals a possible molecular mechanism, which is conducive to further research on Cldn7 and cancer stem cells.
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Affiliation(s)
- Chang Xu
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China.,Department of Hepato-Pancreato-Biliary Surgery, Key Laboratory of Carcinogenesis and Translational Research, Ministry of Education/Beijing , Peking University Cancer Hospital and Institute, Beijing, China
| | - Yu-Han Ding
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Kun Wang
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Mengdi Hao
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Huimin Li
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Lei Ding
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China.
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Kage H, Flodby P, Zhou B, Borok Z. Dichotomous roles of claudins as tumor promoters or suppressors: lessons from knockout mice. Cell Mol Life Sci 2019; 76:4663-4672. [PMID: 31332482 PMCID: PMC6858953 DOI: 10.1007/s00018-019-03238-7] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/27/2019] [Revised: 06/29/2019] [Accepted: 07/18/2019] [Indexed: 02/07/2023]
Abstract
Claudins are a family of integral tight junction proteins that regulate paracellular permeability in polarized epithelia. Overexpression or reduction of claudins can both promote and limit cancer progression, revealing complex dichotomous roles for claudins depending on cellular context. In contrast, recent studies demonstrating tumor formation in claudin knockout mouse models indicate a role for several claudin family members in suppressing tumor initiation. For example, intestine-specific claudin-7 knockout mice spontaneously develop atypical hyperplasia and intestinal adenomas, while claudin-18 knockout mice develop carcinomas in the lung and stomach. Claudin-4, -11, and -15 knockout mice show increased cell proliferation and/or hyperplasia in urothelium, Sertoli cells, and small intestinal crypts, respectively, possibly a precursor to cancer development. Pathways implicated in both cell proliferation and tumorigenesis include Yap/Taz and insulin-like growth factor-1 receptor (IGF-1R)/Akt pathways, among others. Consistent with the tumor suppressive role of claudins shown in mice, in humans, claudin-low breast cancer has been described as a distinct entity with a poor prognosis, and claudin-18-Rho GTPase activating protein 26 (CLDN18-ARHGAP26) fusion protein as a driver gene aberration in diffuse-type gastric cancer due to effects on RhoA. Paradoxically, claudins have also garnered interest as targets for therapy, as they are sometimes aberrantly expressed in cancer cells, which may or may not promote cancer progression. For example, a chimeric monoclonal antibody which targets cells expressing claudin-18.2 through antibody-dependent cell-mediated cytotoxicity has shown promise in multiple phase II studies. In this review, we focus on new findings supporting a tumor suppressive role for claudins during cancer initiation.
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Affiliation(s)
- Hidenori Kage
- Department of Respiratory Medicine, Graduate School of Medicine and Faculty of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8655, Japan
| | - Per Flodby
- Division of Pulmonary, Critical Care and Sleep Medicine and Hastings Center for Pulmonary Research, Keck School of Medicine, University of Southern California, IRD 620, M/C 9520, Los Angeles, CA, 90089-9520, USA
| | - Beiyun Zhou
- Division of Pulmonary, Critical Care and Sleep Medicine and Hastings Center for Pulmonary Research, Keck School of Medicine, University of Southern California, IRD 620, M/C 9520, Los Angeles, CA, 90089-9520, USA
| | - Zea Borok
- Division of Pulmonary, Critical Care and Sleep Medicine and Hastings Center for Pulmonary Research, Keck School of Medicine, University of Southern California, IRD 620, M/C 9520, Los Angeles, CA, 90089-9520, USA.
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Kim DH, Lu Q, Chen YH. Claudin-7 modulates cell-matrix adhesion that controls cell migration, invasion and attachment of human HCC827 lung cancer cells. Oncol Lett 2019; 17:2890-2896. [PMID: 30854065 PMCID: PMC6365970 DOI: 10.3892/ol.2019.9909] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2018] [Accepted: 12/31/2018] [Indexed: 11/12/2022] Open
Abstract
Claudins are a family of tight junction proteins, and serve important roles in epithelial barrier, selective ion transports and cancer metastasis. Although the exact role of claudin-7 in human lung cancer has not been completely elucidated, recent clinical studies have demonstrated that claudin-7 is associated with the survival of patients with lung cancer. Our previous studies have demonstrated that claudin-7 forms a protein complex with integrin β1 in human lung cancer cells. The knockdown (KD) of claudin-7 by short hairpin RNA (shRNA) reduced integrin β1 expression and increased the cell proliferative rate, whereas claudin-7 re-expression in the KD cells decreased the cell proliferation. It is unknown as to whether claudin-7 and integrin β1 regulate cell proliferation and invasion synergistically or independently. In the present study, it was observed that ectopic expression of integrin β1 in claudin-7 KD lung cancer cells did not reduce the cell proliferation. However, integrin β1-transfected cells migrated more effectively in wound healing and cell invasion assays and were more adhesive in a cell attachment assay when compared with those of claudin-7 KD cells. This indicates that claudin-7 controls cell proliferation, while cell attachment and motility were regulated partially through integrin β1. Additionally, claudin-7 overexpression in claudin-7 KD cells resulted in an improved ability to attach to the surface of cell culture plates and a higher expression of focal adhesion proteins when compared with claudin-7 non-KD control cells, which supports the role of claudin-7 in cell adhesion and motility. Taken together, these data suggest that claudin-7 regulates cell motility through integrin β1, providing additional insight into the roles of claudins in carcinogenesis and cancer cell metastasis.
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Affiliation(s)
- Do Hyung Kim
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC 27834, USA
| | - Qun Lu
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC 27834, USA.,Leo Jenkins Cancer Center, Brody School of Medicine, East Carolina University, Greenville, NC 27834, USA
| | - Yan-Hua Chen
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC 27834, USA.,Leo Jenkins Cancer Center, Brody School of Medicine, East Carolina University, Greenville, NC 27834, USA
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Wang K, Xu C, Li W, Ding L. Emerging clinical significance of claudin-7 in colorectal cancer: a review. Cancer Manag Res 2018; 10:3741-3752. [PMID: 30288105 PMCID: PMC6159786 DOI: 10.2147/cmar.s175383] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023] Open
Abstract
Tight junctions (TJs) play an important role in maintaining cell polarity and regulating cell permeability. In recent years, many studies have shown that TJ proteins, especially claudin-7, are closely related to inflammation and the development of various malignant tumors. Claudin-7 plays a significant role in maintaining the physiological functions and pathological conditions of the TJ barrier. The dysregulation of claudin-7 plays a tumor suppressor role or conversely has carcinogenic effects in different target tissues or cells, but the exact underlying mechanism is still unclear. In this review, we will summarize the expression pattern of claudin-7 in tumors, focusing on the expression and regulation of claudin-7 in colorectal cancer and discussing the correlation between claudin-7 and invasion, metastasis and epithelial–mesenchymal transition (EMT) in colorectal cancer. The construction of Cldn7−/− mice and conventional claudin-7 knockout mouse models has helped determine the mechanisms by which claudin-7 promotes tumorigenesis. Elucidation of the expression and subcellular localization of claudin-7 under pathological conditions will help develop claudin-7 as a useful biomarker for detecting and diagnosing cancer, and thus may help combat the occurrence, development, and invasion of cancers.
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Affiliation(s)
- Kun Wang
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, People's Republic of China,
| | - Chang Xu
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, People's Republic of China,
| | - Wenjing Li
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, People's Republic of China,
| | - Lei Ding
- Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, People's Republic of China,
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Li WJ, Xu C, Wang K, Li TY, Wang XN, Yang H, Xing T, Li WX, Chen YH, Gao H, Ding L. Severe Intestinal Inflammation in the Small Intestine of Mice Induced by Controllable Deletion of Claudin-7. Dig Dis Sci 2018; 63:1200-1209. [PMID: 29488037 PMCID: PMC5897149 DOI: 10.1007/s10620-018-4973-z] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/13/2017] [Accepted: 02/07/2018] [Indexed: 02/05/2023]
Abstract
BACKGROUND As a potential tumor suppressor gene, Claudin-7 (Cldn7), which is a component of tight junctions, may play an important role in colorectal cancer occurrence and development. AIMS To generate a knockout mouse model of inducible conditional Cldn7 in the intestine and analyze the phenotype of the mice after induction with tamoxifen. METHODS We constructed Cldn7-flox transgenic mice and crossed them with Villin-CreERT2 mice. The Cldn7 inducible conditional knockout mice appeared normal and were well developed at birth. We induced Cldn7 gene deletion by injecting different dosages of tamoxifen into the mice and then conducted a further phenotypic analysis. RESULTS After induction for 5 days in succession at a dose of 200 µl tamoxifen in sunflower oil at 10 mg/ml per mouse every time, the mice appeared dehydrated, had a lower temperature, and displayed inactivity or death. The results of hematoxylin-eosin staining showed that the intestines of the Cldn7 inducible conditional knockout mice had severe intestinal defects that included epithelial cell sloughing, necrosis, inflammation and hyperplasia. Owing to the death of ICKO mice, we adjusted the dose of tamoxifen to a dose of 100 µl in sunflower oil at 10 mg/ml per mouse (aged more than 8 weeks old) every 4 days. And we could induce atypical hyperplasia and adenoma in the intestine. Immunofluorescent staining indicated that the intestinal epithelial structure was destroyed. Electron microscopy experimental analysis indicated that the intercellular gap along the basolateral membrane of Cldn7 inducible conditional knockout mice in the intestine was increased and that contact between the cells and matrix was loosened. CONCLUSIONS We generated a model of intestinal Cldn7 inducible conditional knockout mice. Intestinal Cldn7 deletion induced by tamoxifen initiated inflammation and hyperplasia in mice.
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Affiliation(s)
- Wen-Jing Li
- The Cancer Center of Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Chang Xu
- The Cancer Center of Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Kun Wang
- The Cancer Center of Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Teng-Yan Li
- Liangxiang Hospital, Fangshan District, Beijing, 102401, China
| | - Xiao-Nan Wang
- Liangxiang Hospital, Fangshan District, Beijing, 102401, China
| | - Hui Yang
- The Cancer Center of Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Tiaosi Xing
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA
| | - Wen-Xia Li
- The Cancer Center of Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China
| | - Yan-Hua Chen
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
| | - Hong Gao
- The Cancer Center of Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China.
| | - Lei Ding
- The Cancer Center of Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China.
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Molina-Jijón E, Rodríguez-Muñoz R, González-Ramírez R, Namorado-Tónix C, Pedraza-Chaverri J, Reyes JL. Aldosterone signaling regulates the over-expression of claudin-4 and -8 at the distal nephron from type 1 diabetic rats. PLoS One 2017; 12:e0177362. [PMID: 28493961 PMCID: PMC5426686 DOI: 10.1371/journal.pone.0177362] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2016] [Accepted: 04/17/2017] [Indexed: 12/19/2022] Open
Abstract
Hyperglycemia in diabetes alters tight junction (TJ) proteins in the kidney. We evaluated the participation of aldosterone (ALD), and the effect of spironolactone (SPL), a mineralocorticoid receptor antagonist, on the expressions of claudin-2, -4, -5 and -8, and occludin in glomeruli, proximal and distal tubules isolated from diabetic rats. Type 1 diabetes was induced in female Wistar rats by a single tail vein injection of streptozotocin (STZ), and SPL was administrated daily by gavage, from days 3–21. Twenty-one days after STZ injection the rats were sacrificed. In diabetic rats, the serum ALD levels were increased, and SPL-treatment did not have effect on these levels or in hyperglycemia, however, proteinuria decreased in SPL-treated diabetic rats. Glomerular damage, evaluated by nephrin and Wilm’s tumor 1 (WT1) protein expressions, and proximal tubular damage, evaluated by kidney injury molecule 1 (Kim-1) and heat shock protein 72 kDa (Hsp72) expressions, were ameliorated by SPL. Also, SPL prevented decrement in claudin-5 in glomeruli, and claudin-2 and occludin in proximal tubules by decreasing oxidative stress, evaluated by superoxide anion (O2●―) production, and oxidative stress markers. In distal tubules, SPL ameliorated increase in mRNA, protein expression, and phosphorylation in threonine residues of claudin-4 and -8, through a serum and glucocorticoid-induced kinase 1 (SGK1), and with-no-lysine kinase 4 (WNK4) signaling pathway. In conclusion, this is the first study that demonstrates that ALD modulates the expression of renal TJ proteins in diabetes, and that the blockade of its actions with SPL, may be a promising therapeutic strategy to prevent alterations of TJ proteins in diabetic nephropathy.
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MESH Headings
- Aldosterone/metabolism
- Animals
- Claudin-4/metabolism
- Claudins/metabolism
- Diabetes Mellitus, Experimental/blood
- Diabetes Mellitus, Experimental/complications
- Diabetes Mellitus, Experimental/metabolism
- Diabetes Mellitus, Experimental/pathology
- Diabetes Mellitus, Type 1/blood
- Diabetes Mellitus, Type 1/complications
- Diabetes Mellitus, Type 1/metabolism
- Diabetes Mellitus, Type 1/pathology
- Female
- Hyperglycemia/blood
- Hyperglycemia/drug therapy
- Hyperglycemia/prevention & control
- Immediate-Early Proteins/metabolism
- Kidney Glomerulus/drug effects
- Kidney Glomerulus/pathology
- Kidney Tubules/drug effects
- Kidney Tubules/pathology
- Models, Biological
- Natriuresis/drug effects
- Nephrons/metabolism
- Oxidative Stress/drug effects
- Phosphorylation/drug effects
- Potassium/blood
- Protein Serine-Threonine Kinases/metabolism
- Proteinuria/blood
- Proteinuria/complications
- Proteinuria/drug therapy
- Proteinuria/prevention & control
- Rats, Wistar
- Signal Transduction/drug effects
- Spironolactone/pharmacology
- Spironolactone/therapeutic use
- Tight Junctions/drug effects
- Tight Junctions/metabolism
- Weight Loss/drug effects
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Affiliation(s)
- Eduardo Molina-Jijón
- Department of Physiology, Biophysics and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), Mexico City, México
- Departamento de Biociencias e Ingeniería, Centro Interdisciplinario de Investigaciones y Estudios sobre el Medio Ambiente y Desarrollo del Instituto Politécnico Nacional (CIIEMAD-IPN), Mexico City, México
| | - Rafael Rodríguez-Muñoz
- Department of Physiology, Biophysics and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), Mexico City, México
| | - Ricardo González-Ramírez
- Department of Molecular Biology and Histocompatibility, Dr. Manuel Gea González, General Hospital, Mexico City, México
| | - Carmen Namorado-Tónix
- Department of Physiology, Biophysics and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), Mexico City, México
| | - José Pedraza-Chaverri
- Department of Biology, Faculty of Chemistry, National Autonomous University of Mexico (UNAM), Mexico City, México
| | - Jose L. Reyes
- Department of Physiology, Biophysics and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), Mexico City, México
- * E-mail:
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Hagen SJ. Non-canonical functions of claudin proteins: Beyond the regulation of cell-cell adhesions. Tissue Barriers 2017; 5:e1327839. [PMID: 28548895 PMCID: PMC5501131 DOI: 10.1080/21688370.2017.1327839] [Citation(s) in RCA: 66] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2017] [Revised: 04/30/2017] [Accepted: 05/03/2017] [Indexed: 12/19/2022] Open
Abstract
Tight junctions form a barrier to the diffusion of apical and basolateral membrane proteins thus regulating membrane polarity. They also regulate the paracellular movement of ions and water across epithelial and endothelial cells so that functionally they constitute an important permselective barrier. Permselectivity at tight junctions is regulated by claudins, which confer anion or cation permeability, and tightness or leakiness, by forming several highly regulated pores within the apical tight junction complex. One interesting feature of claudins is that they are, more often than not, localized to the basolateral membrane, in intracellular cytoplasmic vesicles, or in the nucleus rather than to the apical tight junction complex. These intracellular pools of claudin molecules likely serve important functions in the epithelium. This review will address the widespread prevalence of claudins that are not associated with the apical tight junction complex and discuss the important and emerging non-traditional functions of these molecules in health and disease.
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Affiliation(s)
- Susan J. Hagen
- Department of Surgery/Division of General Surgery, Beth Israel Deaconess Medical Center, Boston, MA, USA
- Harvard Medical School, Boston, MA, USA
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13
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Luna-Antonio BI, Rodriguez-Muñoz R, Namorado-Tonix C, Vergara P, Segovia J, Reyes JL. Gas1 expression in parietal cells of Bowman’s capsule in experimental diabetic nephropathy. Histochem Cell Biol 2017; 148:33-47. [DOI: 10.1007/s00418-017-1550-z] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 02/15/2017] [Indexed: 12/25/2022]
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Martínez-Peña AA, Rivera-Baños J, Méndez-Carrillo LL, Ramírez-Solano MI, Galindo-Bustamante A, Páez-Franco JC, Morimoto S, González-Mariscal L, Cruz ME, Mendoza-Rodríguez CA. Perinatal administration of bisphenol A alters the expression of tight junction proteins in the uterus and reduces the implantation rate. Reprod Toxicol 2017; 69:106-120. [PMID: 28216266 DOI: 10.1016/j.reprotox.2017.02.009] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/21/2016] [Revised: 02/07/2017] [Accepted: 02/13/2017] [Indexed: 11/24/2022]
Abstract
We studied the effect of bisphenol-A (BPA) administration to rats, during the perinatal period, on the fertility of F1 generation and on the expression of tight junction (TJ) proteins in the uterus during early pregnancy. Pregnant Wistar dams (F0) received: BPA-L (0.05mg/kg/day), BPA-H (20mg/kg/day) or vehicle, from gestational day (GD) 6 to lactation day 21. F1 female pups were mated at 3 months of age and sacrificed at GD 1, 3, 6, and 7. Serum hormonal levels, ovulation rate, number of implantation sites and expression of TJ proteins in the uterus of F1 females were evaluated. BPA treatment induced no change in ovulation rate, but induced alterations in progesterone (P4) and estradiol (E2) serum levels, and in implantation rate. With regards to TJ proteins, BPA-H increased claudin-1 during all GDs; eliminated the peaks of claudins -3 and -4 at GD 3 and 6, respectively; and decreased claudin-7 at GD 6, ZO-1 from GD 1-6, and claudin-3 at GD 7 in stromal cells. BPA-L instead, eliminated claudin-3 peak at GD 3, increased claudin-4 and decreased claudin-7 from GD 1-6, decreased claudin-1 at GD 3 and 7 and claudin-4 at GD 7 in stromal cells. BPA-L also decreased ZO-1 at GDs 1 and 3 and increased ZO-1 at GD 6. Thus, BPA treatment during perinatal period perturbed, when the animals reached adulthood and became pregnant, the particular expression of TJ proteins in the uterine epithelium and reduced in consequence the number of implantation sites.
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Affiliation(s)
- Annia A Martínez-Peña
- Facultad de Química, Departamento de Biología, Universidad Nacional Autónoma de Mexico, Mexico D.F. 04510, Mexico
| | - Jorge Rivera-Baños
- Facultad de Química, Departamento de Biología, Universidad Nacional Autónoma de Mexico, Mexico D.F. 04510, Mexico
| | - Laura L Méndez-Carrillo
- Facultad de Química, Departamento de Biología, Universidad Nacional Autónoma de Mexico, Mexico D.F. 04510, Mexico
| | - Marcos I Ramírez-Solano
- Facultad de Química, Departamento de Biología, Universidad Nacional Autónoma de Mexico, Mexico D.F. 04510, Mexico
| | - Aarón Galindo-Bustamante
- Facultad de Química, Departamento de Biología, Universidad Nacional Autónoma de Mexico, Mexico D.F. 04510, Mexico
| | - J Carlos Páez-Franco
- Facultad de Química, Departamento de Biología, Universidad Nacional Autónoma de Mexico, Mexico D.F. 04510, Mexico
| | - Sumiko Morimoto
- Instituto Nacional de Ciencias Médicas y de la Nutrición Salvador Zubirán, Mexico, D.F. 14080, Mexico
| | - Lorenza González-Mariscal
- Centro de Investigación y Estudios Avanzados (CINVESTAV), Departamento de Fisiología, Biofísica y Neurociencias, Mexico, D.F. 07360, Mexico
| | - M Esther Cruz
- Facultad de Estudios Superiores Zaragoza, Laboratorio de Neuroendocrinología, Universidad Nacional Autónoma de Mexico, Mexico, D.F. 15000, Mexico
| | - C Adriana Mendoza-Rodríguez
- Facultad de Química, Departamento de Biología, Universidad Nacional Autónoma de Mexico, Mexico D.F. 04510, Mexico.
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Muto S. Physiological roles of claudins in kidney tubule paracellular transport. Am J Physiol Renal Physiol 2017; 312:F9-F24. [DOI: 10.1152/ajprenal.00204.2016] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2016] [Revised: 10/24/2016] [Accepted: 10/25/2016] [Indexed: 12/30/2022] Open
Abstract
The paracellular pathways in renal tubular epithelia such as the proximal tubules, which reabsorb the largest fraction of filtered solutes and water and are leaky epithelia, are important routes for transepithelial transport of solutes and water. Movement occurs passively via an extracellular route through the tight junction between cells. The characteristics of paracellular transport vary among different nephron segments with leaky or tighter epithelia. Claudins expressed at tight junctions form pores and barriers for paracellular transport. Claudins are from a multigene family, comprising at least 27 members in mammals. Multiple claudins are expressed at tight junctions of individual nephron segments in a nephron segment-specific manner. Over the last decade, there have been advances in our understanding of the structure and functions of claudins. This paper is a review of our current knowledge of claudins, with special emphasis on their physiological roles in proximal tubule paracellular solute and water transport.
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Affiliation(s)
- Shigeaki Muto
- Division of Nephrology, Department of Internal Medicine, Jichi Medical University, Shimotsuke, Tochigi, Japan
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16
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Heiler S, Wang Z, Zöller M. Pancreatic cancer stem cell markers and exosomes - the incentive push. World J Gastroenterol 2016; 22:5971-6007. [PMID: 27468191 PMCID: PMC4948278 DOI: 10.3748/wjg.v22.i26.5971] [Citation(s) in RCA: 65] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/19/2016] [Revised: 06/03/2016] [Accepted: 06/28/2016] [Indexed: 02/06/2023] Open
Abstract
Pancreatic cancer (PaCa) has the highest death rate and incidence is increasing. Poor prognosis is due to late diagnosis and early metastatic spread, which is ascribed to a minor population of so called cancer stem cells (CSC) within the mass of the primary tumor. CSC are defined by biological features, which they share with adult stem cells like longevity, rare cell division, the capacity for self renewal, differentiation, drug resistance and the requirement for a niche. CSC can also be identified by sets of markers, which for pancreatic CSC (Pa-CSC) include CD44v6, c-Met, Tspan8, alpha6beta4, CXCR4, CD133, EpCAM and claudin7. The functional relevance of CSC markers is still disputed. We hypothesize that Pa-CSC markers play a decisive role in tumor progression. This is fostered by the location in glycolipid-enriched membrane domains, which function as signaling platform and support connectivity of the individual Pa-CSC markers. Outside-in signaling supports apoptosis resistance, stem cell gene expression and tumor suppressor gene repression as well as miRNA transcription and silencing. Pa-CSC markers also contribute to motility and invasiveness. By ligand binding host cells are triggered towards creating a milieu supporting Pa-CSC maintenance. Furthermore, CSC markers contribute to the generation, loading and delivery of exosomes, whereby CSC gain the capacity for a cell-cell contact independent crosstalk with the host and neighboring non-CSC. This allows Pa-CSC exosomes (TEX) to reprogram neighboring non-CSC towards epithelial mesenchymal transition and to stimulate host cells towards preparing a niche for metastasizing tumor cells. Finally, TEX communicate with the matrix to support tumor cell motility, invasion and homing. We will discuss the possibility that CSC markers are the initial trigger for these processes and what is the special contribution of CSC-TEX.
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Thuma F, Heiler S, Schnölzer M, Zöller M. Palmitoylated claudin7 captured in glycolipid-enriched membrane microdomains promotes metastasis via associated transmembrane and cytosolic molecules. Oncotarget 2016; 7:30659-77. [PMID: 27120791 PMCID: PMC5058708 DOI: 10.18632/oncotarget.8928] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2015] [Accepted: 03/31/2016] [Indexed: 12/13/2022] Open
Abstract
In epithelial cells claudin7 (cld7) is a major component of tight junctions, but is also recovered from glycolipid-enriched membrane microdomains (GEM). In tumor cells, too, cld7 exists in two stages. Only GEM-located cld7, which is palmitoylated, promotes metastasis. Searching for the underlying mechanism(s) revealed the following.The metastatic capacity of the rat pancreatic adenocarcinoma cell line ASML is lost by a knockdown (kd) of cld7 and is not regained by rescuing cld7 with a mutated palmitoylation site (cld7mPalm). ASML-cld7kd and ASML-cld7mPalm cells show reduced motility and invasiveness. This is due to cld7, but not cld7mPalm associating with α6β4, ezrin, uPAR and MMP14, which jointly support motility and invasion. Palmitoylated cld7 also is engaged in drug resistance by repressing Pten, allowing activation of the antiapoptotic PI3K/Akt pathway. An association of cld7mPalm with the major Pten phosphorylating kinases does not restore apoptosis resistance as phosphorylated Pten is not guided towards GEM to compete with non-phosphorylated Pten. The pathway whereby palmitoylated cld7 supports expression of several EMT genes and nuclear translocation of EMT transcription factors remains to be unraveled. An association with Notch, reduced in ASML-cld7mPalm cells, might be the starting point. Finally, GEM-located, palmitoylated cld7 associates with several components of vesicle transport machineries engaged in exosome biogenesis.Taken together, prerequisites for cld7 acting as a cancer-initiating cell marker are GEM location and palmitoylation, which support a multitude of associations and integration into exosomes. The latter suggests palmitoylated cld7 contributing to message transfer via exosomes.
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Affiliation(s)
- Florian Thuma
- Department of Tumor Cell Biology, University Hospital of Surgery, Heidelberg, Germany
| | - Sarah Heiler
- Department of Tumor Cell Biology, University Hospital of Surgery, Heidelberg, Germany
| | - Martina Schnölzer
- Department of Functional Proteome Analysis, German Cancer Research Center, Heidelberg, Germany
| | - Margot Zöller
- Department of Tumor Cell Biology, University Hospital of Surgery, Heidelberg, Germany
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18
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Dong Y, Simske JS. Vertebrate Claudin/PMP22/EMP22/MP20 family protein TMEM47 regulates epithelial cell junction maturation and morphogenesis. Dev Dyn 2016; 245:653-66. [PMID: 26990309 DOI: 10.1002/dvdy.24404] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2015] [Revised: 02/18/2016] [Accepted: 02/24/2016] [Indexed: 01/22/2023] Open
Abstract
BACKGROUND TMEM47 is the vertebrate orthologue of C. elegans VAB-9, a tetraspan adherens junction protein in the PMP22/EMP/Claudin family of proteins. VAB-9 regulates cell morphology and adhesion in C. elegans and TMEM47 is expressed during kidney development and regulates the activity of Fyn. The conserved functions of VAB-9 and TMEM47 are not well understood. RESULTS expression of TMEM47 in C. elegans functionally rescues vab-9 mutations. Unlike Claudins, expression of TMEM47 in L fibroblasts does not generate tight junction strands; instead, membrane localization requires E-cadherin expression. Temporally, TMEM47 localizes at cell junctions first with E-cadherin before ZO-1 colocalization and in polarized epithelia, TMEM47 colocalizes with adherens junction proteins. By immunoprecipitation, TMEM47 associates with classical adherens junction proteins, but also with tight junction proteins Par6B and aPKCλ. Over-expression of TMEM47 in MDCK cells decreases apical surface area, increases activated myosin light chain at cell-cell contacts, disrupts cell polarity and morphology, delays cell junction reassembly following calcium switch, and selectively interferes with tight junction assembly. Reduced TMEM47 expression results in opposite phenotypes. CONCLUSIONS TMEM47 regulates the localization of a subset of tight junction proteins, associated actomyosin structures, cell morphology, and participates in developmental transitions from adherens to tight junctions. Developmental Dynamics 245:653-666, 2016. © 2016 Wiley Periodicals, Inc.
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Affiliation(s)
- Yi Dong
- Rammelkamp Center for Education and Research, MetroHealth Medical Center, Cleveland, Ohio
| | - Jeffrey S Simske
- Rammelkamp Center for Education and Research, MetroHealth Medical Center, Cleveland, Ohio
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Claudin-3, claudin-7, and claudin-10 show different distribution patterns during decidualization and trophoblast invasion in mouse and human. Histochem Cell Biol 2015; 144:571-85. [PMID: 26340953 DOI: 10.1007/s00418-015-1361-z] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 08/01/2015] [Indexed: 12/18/2022]
Abstract
Implantation of the mammalian embryo requires profound endometrial changes for successful pregnancy, including epithelial-mesenchymal transition of the luminal epithelium and stromal-epithelial transition of the stromal cells resulting in decidualization. Claudins (Cldn) determine the variability in tight junction paracellular permeability and may play a role during these epithelial and decidual changes. We here localized Cldn3, Cldn7 and Cldn10 proteins in the different compartments of murine endometrium up to day 8.5 of pregnancy (dpc) as well as in human endometrium and first trimester decidua. In murine estrous endometrium, luminal and glandular epithelium exhibited Cldn3 and Cldn7, whereas Cldn10 was only detectable in glandular epithelium. At 4.5 dpc, Cldn3 protein shifted to an apical localization, whereas Cldn7 vanished in the epithelium of the implantation chamber. At this stage, there was no stromal signal for Cldn3 and Cldn7, but a strong induction of Cldn10 in the primary decidual zone. Cldn3 proteins emerged at 5.5 dpc spreading considerably from 6.5 dpc onward in the endothelial cells of the decidual blood sinusoids and in the decidual cells of the compact antimesometrial region. In addition to Cldn3, Cldn10 was identified in human endometrial epithelia. Both proteins were not detected in human first trimester decidual cells. Cldn3 was shown in murine trophoblast giant cells as well as in human extravillous trophoblast cells and thus may have an impact on trophoblast invasion in both species. We here showed a specific claudin signature during early decidualization pointing to a role in decidual angiogenesis and regulation of trophoblast invasion.
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Lu Z, Kim DH, Fan J, Lu Q, Verbanac K, Ding L, Renegar R, Chen YH. A non-tight junction function of claudin-7-Interaction with integrin signaling in suppressing lung cancer cell proliferation and detachment. Mol Cancer 2015; 14:120. [PMID: 26081244 PMCID: PMC4470020 DOI: 10.1186/s12943-015-0387-0] [Citation(s) in RCA: 56] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2014] [Accepted: 05/18/2015] [Indexed: 02/08/2023] Open
Abstract
Background Claudins are a family of tight junction (TJ) membrane proteins involved in a broad spectrum of human diseases including cancer. Claudin-7 is a unique TJ membrane protein in that it has a strong basolateral membrane distribution in epithelial cells and in tissues. Therefore, this study aims to investigate the functional significance of this non-TJ localization of claudin-7 in human lung cancer cells. Methods Claudin-7 expression was suppressed or deleted by lentivirus shRNA or by targeted-gene deletion. Cell cycle analysis and antibody blocking methods were employed to assay cell proliferation and cell attachment, respectively. Electron microscopy and transepthelial electrical resistance measurement were performed to examine the TJ ultrastructure and barrier function. Co-immunolocalization and co-immunoprecipitation was used to study claudin-7 interaction with integrin β1. Tumor growth in vivo were analyzed using athymic nude mice. Results Claudin-7 co-localizes and forms a stable complex with integrin β1. Both suppressing claudin-7 expression by lentivirus shRNA in human lung cancer cells (KD cells) and deletion of claudin-7 in mouse lungs lead to the reduction in integrin β1 and phospho-FAK levels. Suppressing claudin-7 expression increases cell growth and cell cycle progression. More significantly, claudin-7 KD cells have severe defects in cell-matrix interactions and adhere poorly to culture plates with a remarkably reduced integrin β1 expression. When cultured on uncoated glass coverslips, claudin-7 KD cells grow on top of each other and form spheroids while the control cells adhere well and grow as a monolayer. Reintroducing claudin-7 reduces cell proliferation, upregulates integrin β1 expression and increases cell-matrix adhesion. Integrin β1 transfection partially rescues the cell attachment defect. When inoculated into nude mice, claudin-7 KD cells produced significantly larger tumors than control cells. Conclusion In this study, we identified a previously unrecognized function of claudin-7 in regulating cell proliferation and maintaining epithelial cell attachment through engaging integrin β1. Electronic supplementary material The online version of this article (doi:10.1186/s12943-015-0387-0) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Zhe Lu
- School of Medicine, Hangzhou Normal University, Hangzhou, 310036, China. .,Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
| | - Do Hyung Kim
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
| | - Junming Fan
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
| | - Qun Lu
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA. .,Leo Jenkins Cancer Center, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
| | - Kathryn Verbanac
- Leo Jenkins Cancer Center, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA. .,Department of Surgery, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
| | - Lei Ding
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
| | - Randall Renegar
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
| | - Yan-Hua Chen
- Department of Anatomy and Cell Biology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA. .,Leo Jenkins Cancer Center, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
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García-Hernández V, Flores-Maldonado C, Rincon-Heredia R, Verdejo-Torres O, Bonilla-Delgado J, Meneses-Morales I, Gariglio P, Contreras RG. EGF Regulates Claudin-2 and -4 Expression Through Src and STAT3 in MDCK Cells. J Cell Physiol 2014; 230:105-15. [DOI: 10.1002/jcp.24687] [Citation(s) in RCA: 36] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/20/2014] [Accepted: 05/22/2014] [Indexed: 12/20/2022]
Affiliation(s)
- Vicky García-Hernández
- Department of Physiology; Biophysics and Neurosciences; Center for Research and Advanced Studies (Cinvestav); México City México
| | - Catalina Flores-Maldonado
- Department of Physiology; Biophysics and Neurosciences; Center for Research and Advanced Studies (Cinvestav); México City México
| | - Ruth Rincon-Heredia
- Department of Physiology; Biophysics and Neurosciences; Center for Research and Advanced Studies (Cinvestav); México City México
- Department of Pharmacology; Center for Research and Advanced Studies (Cinvestav); México City México
| | - Odette Verdejo-Torres
- Department of Physiology; Biophysics and Neurosciences; Center for Research and Advanced Studies (Cinvestav); México City México
| | - José Bonilla-Delgado
- Laboratory of Genetics and Molecular Diagnosis; Research Unit; Hospital Juárez de México; México City México
| | - Ivan Meneses-Morales
- Breast Cancer investigation program; National Autonomous University of México (UNAM); México
- Department of Molecular Biology and Biotechnology; Biomedical Research Institute; National Autonomous University of México (UNAM); México
| | - Patricio Gariglio
- Department of Genetics and Molecular Biology; Center for Research and Advanced Studies (Cinvestav); México City México
| | - Rubén G. Contreras
- Department of Physiology; Biophysics and Neurosciences; Center for Research and Advanced Studies (Cinvestav); México City México
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Abstract
Alterations in water homeostasis can disturb cell size and function. Although most cells can internally regulate cell volume in response to osmolar stress, neurons are particularly at risk given a combination of complex cell function and space restriction within the calvarium. Thus, regulating water balance is fundamental to survival. Through specialized neuronal "osmoreceptors" that sense changes in plasma osmolality, vasopressin release and thirst are titrated in order to achieve water balance. Fine-tuning of water absorption occurs along the collecting duct, and depends on unique structural modifications of renal tubular epithelium that confer a wide range of water permeability. In this article, we review the mechanisms that ensure water homeostasis as well as the fundamentals of disorders of water balance.
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Affiliation(s)
- John Danziger
- Department of Medicine, Beth Israel Deaconess Medical Center, Boston, Massachusetts
| | - Mark L Zeidel
- Department of Medicine, Beth Israel Deaconess Medical Center, Boston, Massachusetts
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Molina-Jijón E, Rodríguez-Muñoz R, Namorado MDC, Pedraza-Chaverri J, Reyes JL. Oxidative stress induces claudin-2 nitration in experimental type 1 diabetic nephropathy. Free Radic Biol Med 2014; 72:162-75. [PMID: 24726862 DOI: 10.1016/j.freeradbiomed.2014.03.040] [Citation(s) in RCA: 45] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/02/2013] [Revised: 03/13/2014] [Accepted: 03/29/2014] [Indexed: 02/07/2023]
Abstract
Renal complications in diabetes are severe and may lead to renal insufficiency. Early alterations in tight junction (TJ) proteins in diabetic nephropathy (DN) have not been explored and the role of oxidative stress in their disassembly has been poorly characterized. We investigated the expression and distribution of TJ proteins: claudin-5 in glomeruli (GL), occludin and claudin-2 in proximal tubules (PTs), and ZO-1 and claudin-1, -4, and -8 in distal tubules (DTs) of rats 21 days after streptozotocin injection. Redox status along the nephron segments was evaluated. Diabetes increased kidney injury molecule-1 expression. Expression of sodium glucose cotransporters (SGLT1 and SGLT2) and facilitative glucose transporter (GLUT2) was induced. Increased oxidative stress was present in GL and PTs and to a lesser extent in DTs (measured by superoxide production and PKCβ2 expression), owing to NADPH oxidase activation and uncoupling of the endothelial nitric oxide synthase-dependent pathway. Claudin-5, occludin, and claudin-2 expression was decreased, whereas claudin-4 and -8 expression increased. ZO-1 was redistributed from membrane to cytosol. Increased nitration of tyrosine residues in claudin-2 was found, which might contribute to decrement of this protein in proximal tubule. In contrast, occludin was not nitrated. We suggest that loss of claudin-2 is associated with increased natriuresis and that loss of glomerular claudin-5 might explain early presence of proteinuria. These findings suggest that oxidative stress is related to alterations in TJ proteins in the kidney that are relevant to the pathogenesis and progression of DN and for altered sodium regulation in diabetes.
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Affiliation(s)
- Eduardo Molina-Jijón
- Department of Physiology, Biophysics, and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute, México, DF 07360, Mexico
| | - Rafael Rodríguez-Muñoz
- Department of Physiology, Biophysics, and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute, México, DF 07360, Mexico
| | - María del Carmen Namorado
- Department of Physiology, Biophysics, and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute, México, DF 07360, Mexico
| | - José Pedraza-Chaverri
- Department of Biology, Faculty of Chemistry, National Autonomous University of Mexico, 04510 University City, DF, Mexico
| | - José L Reyes
- Department of Physiology, Biophysics, and Neuroscience, Center for Research and Advanced Studies of the National Polytechnic Institute, México, DF 07360, Mexico.
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Retana C, Sanchez E, Perez-Lopez A, Cruz A, Lagunas J, Cruz C, Vital S, Reyes JL. Alterations of intercellular junctions in peritoneal mesothelial cells from patients undergoing dialysis: effect of retinoic Acid. Perit Dial Int 2014; 35:275-87. [PMID: 24584604 DOI: 10.3747/pdi.2012.00323] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2012] [Accepted: 10/01/2013] [Indexed: 11/15/2022] Open
Abstract
BACKGROUND Dialysis patients are classified according to their peritoneal permeability as low transporter (LT, low solute permeability) or high transporter (HT, high solute permeability). Tight junction (TJ) proteins are critical to maintain ions, molecules and water paracellular transport through peritoneum. Exposure to peritoneal dialysis solutions causes damage to TJ in human peritoneal mesothelial cells (HPMCs). We analyzed the quantity, distribution and function of TJ proteins: claudin-1, -2 and -8, ZO-1 and occludin, in HPMC cultures from LT and HT patients. Since all-trans retinoic acid (ATRA) might modify the expression of TJ proteins, we studied its effect on HPMCs. METHODS Control HPMCs were isolated from human omentum, while HT or LT cells were obtained from dialysis effluents. Cells were cultured in presence of ATRA 0, 50 or 100 nM. Transepithelial electrical resistance (TER) measurement, immunostaining and Western blot analyses were performed. RESULTS HT exhibited lower TER than control and LT monolayers. Immunofluorescence for TJ was weak and discontinuous along the cell contour, in LT and HT. Furthermore, claudin-1, occludin and ZO-1 expressions were decreased. In all groups, claudin-2 was localized at nuclei. We observed that ATRA improved TJ distribution and increased TJ expression in HT. This retinoid did not modify claudin-2 and -8 expressions. All-trans retinoic acid decreased TER in HT, but had no effect in LT. CONCLUSIONS Tight junctions were altered in HPMCs from dialyzed patients. The HT monolayer has lower TER than LT, which might be associated with the peritoneal permeability in these patients. ATRA might be a therapeutic alternative to maintain mesothelial integrity, since it improved TJ localization and expression.
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Affiliation(s)
- Carmen Retana
- Pharmacology Dept., Centre for Research and Advanced Studies National Polytechnic Institute, Mexico
| | - Elsa Sanchez
- Physiology, Biophysics and Neurosciences Dept., Centre for Research and Advanced Studies National Polytechnic Institute, Mexico
| | | | - Armando Cruz
- Unidad Medica Alta Especialidad, Hospital General la Raza, Instituto Mexicano del Seguro Social (IMSS), Mexico
| | - Jesus Lagunas
- Unidad Medica Alta Especialidad, Hospital General la Raza, Instituto Mexicano del Seguro Social (IMSS), Mexico
| | - Carmen Cruz
- Dept. of Nephrology, Hospital Regional No. 1 del IMSS, Mexico
| | - Socorro Vital
- Dept. of Nephrology, Hospital Juarez de Mexico, Mexico
| | - Jose L Reyes
- Physiology, Biophysics and Neurosciences Dept., Centre for Research and Advanced Studies National Polytechnic Institute, Mexico
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Szaszi K, Amoozadeh Y. New Insights into Functions, Regulation, and Pathological Roles of Tight Junctions in Kidney Tubular Epithelium. INTERNATIONAL REVIEW OF CELL AND MOLECULAR BIOLOGY 2014; 308:205-71. [DOI: 10.1016/b978-0-12-800097-7.00006-3] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
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Tight junction proteins and oxidative stress in heavy metals-induced nephrotoxicity. BIOMED RESEARCH INTERNATIONAL 2013; 2013:730789. [PMID: 23710457 PMCID: PMC3654622 DOI: 10.1155/2013/730789] [Citation(s) in RCA: 59] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/20/2012] [Accepted: 03/27/2013] [Indexed: 12/11/2022]
Abstract
Kidney is a target organ for heavy metals. They accumulate in several segments of the nephron and cause profound alterations in morphology and function. Acute intoxication frequently causes acute renal failure. The effects of chronic exposure have not been fully disclosed. In recent years increasing awareness of the consequences of their presence in the kidney has evolved. In this review we focus on the alterations induced by heavy metals on the intercellular junctions of the kidney. We describe that in addition to the proximal tubule, which has been recognized as the main site of accumulation and injury, other segments of the nephron, such as glomeruli, vessels, and distal nephron, show also deleterious effects. We also emphasize the participation of oxidative stress as a relevant component of the renal damage induced by heavy metals and the beneficial effect that some antioxidant drugs, such as vitamin A (all-trans-retinoic acid) and vitamin E (α-tocopherol), depict on the morphological and functional alterations induced by heavy metals.
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Regulation of Tight Junctions for Therapeutic Advantages. CANCER METASTASIS - BIOLOGY AND TREATMENT 2013. [DOI: 10.1007/978-94-007-6028-8_9] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/14/2023]
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Eadon MT, Hack BK, Xu C, Ko B, Toback FG, Cunningham PN. Endotoxemia alters tight junction gene and protein expression in the kidney. Am J Physiol Renal Physiol 2012; 303:F821-30. [PMID: 22791339 DOI: 10.1152/ajprenal.00023.2012] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023] Open
Abstract
Intact tight junctional (TJ) proteins are required for tubular ion transport and waste excretion. Disruption of TJs may contribute to a decreased glomerular filtration rate in acute kidney injury (AKI) via tubular backleak. The effect of LPS-mediated AKI on murine TJs has not been studied extensively. We hypothesized LPS endotoxin administration to mice would disrupt tubular TJ proteins including zonula occludens-1 (ZO-1), occludin, and claudins. ZO-1 and occludin immunofluorescence 24 h post-LPS revealed a marked change in localization from the usual circumferential fencework pattern to one with substantial fragmentation. Renal ZO-1 expression was significantly reduced 24 h after LPS (decrease of 56.1 ± 7.4%, P < 0.001), with subsequent recovery. ZO-1 mRNA expression was increased 24 h post-LPS (4.34 ± 0.87-fold, P = 0.0019), suggesting disruption of ZO-1 protein is not mediated by transcriptional regulation, but rather by degradation or changes in translation. Similarly, claudin-4 protein expression was decreased despite elevated mRNA. LPS administration resulted in dephosphorylation of occludin and fragmented tubular redistribution. Protein expression of claudin-1, and -3 was increased after LPS. ZO-1, occludin, and claudin-1, -3, and -4 gene expression were increased 48 h after LPS, suggesting a renal response to strengthen TJs following injury. Interestingly, reduced mRNA expression was found only for claudin-8. This study provides further support that LPS-induced AKI is associated with structural injury and is not merely due to hemodynamic changes.
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Affiliation(s)
- Michael T Eadon
- Section of Nephrology, The University of Chicago, Illinois, USA.
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Krug SM, Günzel D, Conrad MP, Lee IFM, Amasheh S, Fromm M, Yu ASL. Charge-selective claudin channels. Ann N Y Acad Sci 2012; 1257:20-8. [DOI: 10.1111/j.1749-6632.2012.06555.x] [Citation(s) in RCA: 85] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023]
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Ding L, Lu Z, Foreman O, Tatum R, Lu Q, Renegar R, Cao J, Chen YH. Inflammation and disruption of the mucosal architecture in claudin-7-deficient mice. Gastroenterology 2012; 142:305-15. [PMID: 22044670 PMCID: PMC3267838 DOI: 10.1053/j.gastro.2011.10.025] [Citation(s) in RCA: 175] [Impact Index Per Article: 13.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/16/2011] [Revised: 10/06/2011] [Accepted: 10/15/2011] [Indexed: 02/08/2023]
Abstract
BACKGROUND & AIMS Integrity of the intestinal epithelium is required for nutrition absorption and defense against pathogens. Claudins are cell adhesion molecules that localize at tight junctions (TJs); many are expressed in the intestinal tract, but little is known about their functions. Claudin-7 is unique in that it has a stronger basolateral membrane distribution than other claudins, which localize primarily to apical TJs in the intestinal epithelium. We investigated the basolateral functions of claudin-7 and assessed the effects of disruption of Cldn7 in intestines of mice. METHODS We generated Cldn7(-/-) mice and examined their intestines by histology, molecular and cellular biology, and biochemistry approaches. We performed gene silencing experiments in epithelial cell lines using small interfering RNAs (siRNAs). RESULTS The Cldn7(-/-) mice had severe intestinal defects that included mucosal ulcerations, epithelial cell sloughing, and inflammation. Intestines of Cldn7(-/-) mice produced significantly higher levels of cytokines, the nuclear factor κB p65 subunit, and cyclooxygenase 2; they also up-regulated expression of matrix metalloproteinases (MMPs)-3 and -7. siRNA in epithelial cell lines showed that the increased expression of MMP-3 resulted directly from claudin-7 depletion, whereas that of MMP-7 resulted from inflammation. Electron microscopy analysis showed that intestines of Cldn7(-/-) mice had intercellular gaps below TJs and cell matrix loosening. Deletion of Cldn7 reduced expression and altered localization of the integrin α2 subunit in addition to disrupting formation of complexes of claudin-7, integrin α2, and claudin-1 that normally form in epithelial basolateral compartments of intestines. CONCLUSIONS In mice, claudin-7 has non-TJ functions, including maintenance of epithelial cell-matrix interactions and intestinal homeostasis.
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Affiliation(s)
- Lei Ding
- Department of Anatomy and Cell Biology, East Carolina University Brody School of Medicine, Greenville, NC 27834,Department of Oncology, Beijing Shijitan Hospital, Capital Medical University, Beijing, China
| | - Zhe Lu
- Department of Anatomy and Cell Biology, East Carolina University Brody School of Medicine, Greenville, NC 27834
| | - Oded Foreman
- Department of Pathology, The Jackson Laboratory, Sacramento, CA 95838
| | - Rodney Tatum
- Department of Anatomy and Cell Biology, East Carolina University Brody School of Medicine, Greenville, NC 27834
| | - Qun Lu
- Department of Anatomy and Cell Biology, East Carolina University Brody School of Medicine, Greenville, NC 27834
| | - Randall Renegar
- Department of Anatomy and Cell Biology, East Carolina University Brody School of Medicine, Greenville, NC 27834
| | - Jian Cao
- Department of Medicine and Pathology, Stony Brook University, Stony Brook, NY 11794
| | - Yan-Hua Chen
- Department of Anatomy and Cell Biology, East Carolina University Brody School of Medicine, Greenville, NC 27834,Address correspondence to Yan-Hua Chen, Tel: (252) 744-1341; Fax: (252) 744-2850;
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Zöller M, Jung T. The Colorectal Cancer Initiating Cell: Markers and Their Role in Liver Metastasis. ACTA ACUST UNITED AC 2011. [DOI: 10.1007/978-94-007-0292-9_4] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
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González-Mariscal L, Garay E, Quirós M. Identification of claudins by western blot and immunofluorescence in different cell lines and tissues. Methods Mol Biol 2011; 762:213-231. [PMID: 21717359 DOI: 10.1007/978-1-61779-185-7_15] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 05/31/2023]
Abstract
Claudins are integral proteins of the TJ. Each epithelia in the organism expresses a unique set of claudins that determines the degree of sealing of the paracellular pathway and the ionic selectivity of the tissue. TJs are dynamic structures whose organization and composition change in response to alterations in the environment as well as under physiological and pathological conditions. Changes in claudin expression and subcellular distribution can be analyzed in western blot and immunofluorescence experiments, employing a wide array of available specific antibodies against claudins. In this chapter, we describe in detail protocols used for western blot and immunofluorescence detection of claudins in epithelial cell lines and in various tissue samples.
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Affiliation(s)
- Lorenza González-Mariscal
- Department of Physiology, Biophysics, and Neuroscience, Center for Research and Advanced Studies (CINVESTAV), México DF., Mexico.
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Collins JE, Kirk A, Campbell SK, Mason J, Wilson SJ. Enhanced immunohistochemical resolution of claudin proteins in glycolmethacrylate-embedded tissue biopsies. Methods Mol Biol 2011; 762:371-382. [PMID: 21717371 DOI: 10.1007/978-1-61779-185-7_27] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 05/31/2023]
Abstract
There are a number of disadvantages with conventional tissue immunohistochemistry for accurate -localisation of claudin proteins. Traditionally, tissue cryopreservation or formaldehyde fixation with wax embedding is utilised prior to sectioning and antibody localisation. Wax embedding gives better morphological preservation than frozen tissue, but the required use of chemical cross-linking fixatives renders many antigens inaccessible to antibody binding or results in subsequent disruption of antibody localisation patterns due to the use of harsh antigen retrieval methods. Use of frozen or wax-embedded tissue also requires the cutting of relatively thick>6-μm sections, making the interrogation of serial sections very limited. The use of glycolmethacrylate (GMA) tissue embedding with fixation in acetone is compatible with epitope preservation for many antibody reagents that are often destroyed by chemical cross-linking fixatives. GMA is a water-miscible embedding resin that maintains tissue hydration during processing, thus reducing tissue shrinkage, while embedding and cutting in the polymerised resin physically supports the tissue, thus improving morphology. This method also facilitates the cutting of 2-μm sequential sections for analysis of multiple antigens and maximises the information available from small tissue biopsies from human clinical sources.
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Affiliation(s)
- Jane E Collins
- Division of Infection, Inflammation, and Immunity, University of Southampton Medical School, Southampton, UK.
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Amasheh S, Fromm M, Günzel D. Claudins of intestine and nephron - a correlation of molecular tight junction structure and barrier function. Acta Physiol (Oxf) 2011; 201:133-40. [PMID: 20518752 DOI: 10.1111/j.1748-1716.2010.02148.x] [Citation(s) in RCA: 110] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
A prerequisite of epithelial transport is a paracellular barrier function, which seals the tissue against an uncontrolled leak flux. Moreover, selective paracellular permeability has been shown to be crucial for physiological epithelial transport function. Claudins are tetraspan tight junction proteins which play a major role in paracellular ion permeability across epithelia. The multigene family consists of 24 members and several splice variants which show distinct tissue-specific expression profiles. Moreover, in diseases associated with a loss of barrier function such as forms of inflammatory bowel disease, the expression of claudins is altered. Functional characterization of single claudins revealed specific contribution to barrier properties in epithelia. This review gives an overview on the exploration of molecular structure and barrier function along the intestine and nephron, which not only share mechanisms of selective restriction of the paracellular pathway but also exhibit distinct organ-specific characteristics.
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Affiliation(s)
- S Amasheh
- Institute of Clinical Physiology, Charité, Berlin, Germany.
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36
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Kirk A, Campbell S, Bass P, Mason J, Collins J. Differential expression of claudin tight junction proteins in the human cortical nephron. Nephrol Dial Transplant 2010; 25:2107-19. [PMID: 20124215 PMCID: PMC2891746 DOI: 10.1093/ndt/gfq006] [Citation(s) in RCA: 71] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023] Open
Abstract
Background. In renal tubules, paracellular permeability is tightly controlled to facilitate solute absorption and urinary concentration and is regulated by tight junctions, which incorporate claudin proteins. There is very limited information confirming the localization of these proteins in the human renal cortex. Most data is inferred from mouse, bovine and rabbit studies and differences exist between mouse and other species. Methods. A survey of claudin staining was performed on human kidney cortex embedded in glycolmethacrylate resin to enhance tissue morphology and facilitate the cutting of 2 µm serial sections. Results. Claudin-2, -10 and -11 antibodies labelled renal tubular epithelial cells, correlating with Lotus tetragonolobus and N-cadherin positive proximal tubules. Claudin-3, -10, -11 and -16 antibodies strongly stained a population of tubules that were positive for Tamm Horsfall protein on adjacent sections, confirming expression in the thick ascending limb of the Loop of Henle. Claudin-3, -4 and -8 antibodies reacted with tubules that correlated with the distal nephron markers, E-cadherin, epithelial membrane antigen and Dolichos biflorus and claudin-3, -4, -7 and -8 with the distal tubule marker, calbindin, and the collecting duct marker, aquaporin-2. Claudin-14 was localized in distal convoluted tubules, correlating positively with calbindin but negatively with aquaporin-2, whereas claudin-1 staining was identified in the parietal epithelium of Bowman's capsule, distal convoluted tubule and collecting duct. Cellular and tight junction localization of claudin staining in renal tubules was heterogeneous and is discussed. Conclusions. Complex variation in the expression of human claudins likely determines paracellular permeability in the kidney. Altered claudin expression may influence pathologies involving abnormalities of absorption.
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Affiliation(s)
- Adam Kirk
- 1Division of Infection, Inflammation and Immunity, Mailpoint 813, University of Southampton Medical School, Sir Henry Wellcome Laboratories, Southampton General Hospital, Tremona Road, Southampton SO16 6YD, UK
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Arreola-Mendoza L, Del Razo LM, Mendoza-Garrido ME, Martin D, Namorado MC, Calderon-Salinas JV, Reyes JL. The protective effect of alpha-tocopherol against dichromate-induced renal tight junction damage is mediated via ERK1/2. Toxicol Lett 2009; 191:279-88. [DOI: 10.1016/j.toxlet.2009.09.011] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2009] [Revised: 09/02/2009] [Accepted: 09/14/2009] [Indexed: 12/21/2022]
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Günzel D, Haisch L, Pfaffenbach S, Krug SM, Milatz S, Amasheh S, Hunziker W, Müller D. Claudin function in the thick ascending limb of Henle's loop. Ann N Y Acad Sci 2009; 1165:152-62. [PMID: 19538301 DOI: 10.1111/j.1749-6632.2009.04051.x] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/23/2023]
Abstract
During the past decade, claudins have been established as major determinants of paracellular permeablilty in epithelia. In the kidney, each nephron segment expresses a distinct pattern of claudins. Cells of the thick ascending limb of Henle's loop (TAL), which is characterized by high paracellular cation permeability, co-express an unusually large number of different claudins: claudin-10, -16, and -19 and, depending on the species, also claudin-3, -4, -8, and/or -11. The function of most of these claudins has been investigated in vitro. We present a summary of their function with special emphasis on claudin-16 and -19. Mutations in the corresponding human genes lead to severely impaired renal Ca(2+) and Mg(2+) handling. To date, 42 different claudin-16 mutations and three claudin-19 mutations have been reported. These mutations prevent the claudins from reaching the surface membrane, decrease membrane residence time, or render them functionless. In spite of the clear clinical symptoms such as hypomagnesemia, hypercalciuria, nephrocalcinosis, and renal insufficiency, mechanisms that link claudin-16 and -19 to these symptoms are still unknown. Depending on the cell type used in overexpression studies, claudin-16 appears to cause a mild increase in paracellular Mg(2+)-permeability or a pronounced increase in Na(+) permeability. Claudin-19 selectively decreases Cl(-) permeability, thus synergistically increasing relative cation permeability, or indiscriminately decreases paracellular permeability. In the light of these results it is hypothesized that the renal Mg(2+)/Ca(2+) waste may not be solely due to reduced resorption in the TAL but at least in part to paracellular back-leak of Mg(2+)/Ca(2+) into the tubular lumen of the distal convoluted tubule.
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Angelow S, Ahlstrom R, Yu ASL. Biology of claudins. Am J Physiol Renal Physiol 2008; 295:F867-76. [PMID: 18480174 PMCID: PMC2576152 DOI: 10.1152/ajprenal.90264.2008] [Citation(s) in RCA: 270] [Impact Index Per Article: 15.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2008] [Accepted: 05/13/2008] [Indexed: 12/13/2022] Open
Abstract
Claudins are a family of tight junction membrane proteins that regulate paracellular permeability of epithelia, likely by forming the lining of the paracellular pore. Claudins are expressed throughout the renal tubule, and mutations in two claudin genes are now known to cause familial hypercalciuric hypomagnesemia with nephrocalcinosis. In this review, we discuss recent advances in our understanding of the physiological role of various claudins in normal kidney function, and in understanding the fundamental biology of claudins, including the molecular basis for selectivity of permeation, claudin interactions in tight junction formation, and regulation of claudins by protein kinases and other intracellular signals.
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Affiliation(s)
- Susanne Angelow
- Department of Medicine, University of Southern California Keck School of Medicine, Division of Nephrology, 2025 Zonal Ave, RMR 406, Los Angeles, CA 90089, USA
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Günzel D, Yu ASL. Function and regulation of claudins in the thick ascending limb of Henle. Pflugers Arch 2008; 458:77-88. [PMID: 18795318 DOI: 10.1007/s00424-008-0589-z] [Citation(s) in RCA: 44] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2008] [Accepted: 09/04/2008] [Indexed: 01/25/2023]
Abstract
The thick ascending limb (TAL) of Henle mediates transcellular reabsorption of NaCl while generating a lumen-positive voltage that drives passive paracellular reabsorption of divalent cations. Disturbance of paracellular reabsorption leads to Ca(2+) and Mg(2+) wasting in patients with the rare inherited disorder of familial hypercalciuric hypomagnesemia with nephrocalcinosis (FHHNC). Recent work has shown that the claudin family of tight junction proteins form paracellular pores and determine the ion selectivity of paracellular permeability. Importantly, FHHNC has been found to be caused by mutations in two of these genes, claudin-16 and claudin-19, and mice with knockdown of claudin-16 reproduce many of the features of FHHNC. Here, we review the physiology of TAL ion transport, present the current view of the role and mechanism of claudins in determining paracellular permeability, and discuss the possible pathogenic mechanisms responsible for FHHNC.
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Affiliation(s)
- Dorothee Günzel
- Department of Clinical Physiology, Charité, Campus Benjamin Franklin, Berlin, Germany
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Angelow S, Schneeberger EE, Yu ASL. Claudin-8 expression in renal epithelial cells augments the paracellular barrier by replacing endogenous claudin-2. J Membr Biol 2007; 215:147-59. [PMID: 17516019 DOI: 10.1007/s00232-007-9014-3] [Citation(s) in RCA: 64] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/06/2006] [Accepted: 02/15/2007] [Indexed: 12/21/2022]
Abstract
Claudins are transmembrane proteins of the tight junction that determine and regulate paracellular ion permeability. We previously reported that claudin-8 reduces paracellular cation permeability when expressed in low-resistance Madin-Darby canine kidney (MDCK) II cells. Here, we address how the interaction of heterologously expressed claudin-8 with endogenous claudin isoforms impacts epithelial barrier properties. In MDCK II cells, barrier improvement by claudin-8 is accompanied by a reduction of endogenous claudin-2 protein at the tight junction. Here, we show that this is not because of relocalization of claudin-2 into the cytosolic pool but primarily due to a decrease in gene expression. Claudin-8 also affects the trafficking of claudin-2, which was displaced specifically from the junctions at which claudin-8 was inserted. To test whether replacement of cation-permeable claudin-2 mediates the effect of claudin-8 on the electrophysiological phenotype of the host cell line, we expressed claudin-8 in high-resistance MDCK I cells, which lack endogenous claudin-2. Unlike in MDCK II cells, induction of claudin-8 in MDCK I cells (which did not affect levels of endogenous claudins) did not alter paracellular ion permeability. Furthermore, when endogenous claudin-2 in MDCK II cells was downregulated by epidermal growth factor to create a cell model with low transepithelial resistance and low levels of claudin-2, the permeability effects of claudin-8 were also abolished. Our findings demonstrate that claudin overexpression studies measure the combined effect of alterations in both endogenous and exogenous claudins, thus explaining the dependence of the phenotype on the host cell line.
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Affiliation(s)
- Susanne Angelow
- Division of Nephrology, Department of Medicine, University of Southern California Keck School of Medicine, 2025 Zonal Avenue, Los Angeles, CA 90033, USA
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Levin M. Is the early left-right axis like a plant, a kidney, or a neuron? The integration of physiological signals in embryonic asymmetry. ACTA ACUST UNITED AC 2006; 78:191-223. [PMID: 17061264 DOI: 10.1002/bdrc.20078] [Citation(s) in RCA: 61] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023]
Abstract
Embryonic morphogenesis occurs along three orthogonal axes. While the patterning of the anterior-posterior and dorsal-ventral axes has been increasingly well-characterized, the left-right (LR) axis has only relatively recently begun to be understood at the molecular level. The mechanisms that ensure invariant LR asymmetry of the heart, viscera, and brain involve fundamental aspects of cell biology, biophysics, and evolutionary biology, and are important not only for basic science but also for the biomedicine of a wide range of birth defects and human genetic syndromes. The LR axis links biomolecular chirality to embryonic development and ultimately to behavior and cognition, revealing feedback loops and conserved functional modules occurring as widely as plants and mammals. This review focuses on the unique and fascinating physiological aspects of LR patterning in a number of vertebrate and invertebrate species, discusses several profound mechanistic analogies between biological regulation in diverse systems (specifically proposing a nonciliary parallel between kidney cells and the LR axis based on subcellular regulation of ion transporter targeting), highlights the possible importance of early, highly-conserved intracellular events that are magnified to embryo-wide scales, and lays out the most important open questions about the function, evolutionary origin, and conservation of mechanisms underlying embryonic asymmetry.
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Affiliation(s)
- Michael Levin
- Forsyth Center for Regenerative and Developmental Biology, The Forsyth Institute, and the Department of Developmental Biology, Harvard School of Dental Medicine, Boston, Massachusetts 02115, USA.
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