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Rosenthal R, Waesch A, Masete KV, Massarani AS, Schulzke JD, Hering NA. The green tea component (-)-epigallocatechin-3-gallate protects against cytokine-induced epithelial barrier damage in intestinal epithelial cells. Front Pharmacol 2025; 16:1559812. [PMID: 40438590 PMCID: PMC12117334 DOI: 10.3389/fphar.2025.1559812] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2025] [Accepted: 04/28/2025] [Indexed: 06/01/2025] Open
Abstract
Background Green tea consumption is associated with health benefits, which are mainly attributed to its catechins, especially the main catechin (-)-epigallocatechin-3-gallate (EGCG). Among other beneficial effects, EGCG was shown to be protective in inflammatory bowel disease (IBD), a condition associated with barrier dysfunction. To elucidate the mechanisms behind this, the present study analyzed the impact of EGCG on barrier properties and inflammatory cytokine-induced barrier dysfunction in three different intestinal cell models. Methods T84 cells served as a colon model, while Caco-2 and 2D-organoids derived from human duodenum biopsies were used as small intestinal models. Epithelial monolayers grown on filter supports were challenged with EGCG and a combination of the two main IBD cytokines, tumor necrosis factor α (TNFα) and interferon γ (IFNγ). Barrier properties were monitored by measuring transepithelial resistance (TER), macromolecule permeability, apoptosis, and tight junction protein expression and localization. Results EGCG protected against barrier defects induced by TNFα and IFNγ. The cytokines decreased TER (T84: 11% ± 1% of initial value; Caco-2: 65% ± 2% of initial value; 2D-organoids: 57 ± 8 Ω*cm2 versus control 239 ± 29 Ω*cm2) which was prevented by 200 µM EGCG (T84: 89% ± 5%; Caco-2: 89% ± 3%; 2D-organoids: 343 ± 24 Ω*cm2; in all three models p < 0.001). In parallel, EGCG attenuated the cytokine-induced increase in macromolecular permeability by reducing apoptosis, as shown by reduced caspase-3 cleavage by >50% compared to cytokine-stimulated controls in all three models (p < 0.001). Furthermore, alterations in tight junction protein expression and localization contributed to barrier protection. In the small intestinal models, 200 µM EGCG stabilized barrier function, as demonstrated by an increase in TER (Caco-2: 105% ± 3% versus control 90% ± 3%; 2D-organoids: 182% ± 12% versus control 105% ± 2%, in both models p < 0.001), upregulation of claudin-4 (Caco-2: 140% ± 15%, p < 0.05; 2D-organoids: 115% ± 5%, p < 0.01) and reduced expression of claudin-2 (Caco-2: 75% ± 10%, p < 0.5; 2D-organoids: 66% ± 6%, p < 0.01) and claudin-7 (Caco-2: 64% ± 7%, p < 0.001; 2D-organoids: 65% ± 9%, p < 0.01). In the colon model, EGCG prevented the delocalization of claudin-1 and -5 that was induced by TNFα and IFNγ. Conclusion The green tea component EGCG stabilizes the intestinal barrier and protects against barrier dysfunction induced by pro-inflammatory cytokines. These findings highlight the potential of EGCG as a supportive treatment strategy for IBD.
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Affiliation(s)
- Rita Rosenthal
- Department of Gastroenterology, Rheumatology and Infectious Diseases, Clinical Physiology/Nutritional Medicine, Charité – Universitätsmedizin Berlin, Berlin, Germany
| | - Anne Waesch
- Department of Gastroenterology, Rheumatology and Infectious Diseases, Clinical Physiology/Nutritional Medicine, Charité – Universitätsmedizin Berlin, Berlin, Germany
| | - Kopano Valerie Masete
- Department of Gastroenterology, Rheumatology and Infectious Diseases, Clinical Physiology/Nutritional Medicine, Charité – Universitätsmedizin Berlin, Berlin, Germany
| | - Alain S. Massarani
- Department of Gastroenterology, Rheumatology and Infectious Diseases, Clinical Physiology/Nutritional Medicine, Charité – Universitätsmedizin Berlin, Berlin, Germany
| | - Jörg-Dieter Schulzke
- Department of Gastroenterology, Rheumatology and Infectious Diseases, Clinical Physiology/Nutritional Medicine, Charité – Universitätsmedizin Berlin, Berlin, Germany
| | - Nina A. Hering
- Department of General and Visceral Surgery, Charité – Universitätsmedizin Berlin, Berlin, Germany
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Lacoste B, Prat A, Freitas-Andrade M, Gu C. The Blood-Brain Barrier: Composition, Properties, and Roles in Brain Health. Cold Spring Harb Perspect Biol 2025; 17:a041422. [PMID: 38951020 PMCID: PMC12047665 DOI: 10.1101/cshperspect.a041422] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 07/03/2024]
Abstract
Blood vessels are critical to deliver oxygen and nutrients to tissues and organs throughout the body. The blood vessels that vascularize the central nervous system (CNS) possess unique properties, termed the blood-brain barrier (BBB), which allow these vessels to tightly regulate the movement of ions, molecules, and cells between the blood and the brain. This precise control of CNS homeostasis allows for proper neuronal function and protects the neural tissue from toxins and pathogens, and alterations of this barrier are important components of the pathogenesis and progression of various neurological diseases. The physiological barrier is coordinated by a series of physical, transport, and metabolic properties possessed by the brain endothelial cells (ECs) that form the walls of the blood vessels. These properties are regulated by interactions between different vascular, perivascular, immune, and neural cells. Understanding how these cell populations interact to regulate barrier properties is essential for understanding how the brain functions in both health and disease contexts.
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Affiliation(s)
- Baptiste Lacoste
- Ottawa Hospital Research Institute, Neuroscience Program, Ottawa, Ontario K1H 8M5, Canada
- Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario K1H 8M5, Canada
- University of Ottawa Brain and Mind Research Institute, Ottawa, Ontario K1H 8M5, Canada
| | - Alexandre Prat
- Department of Neuroscience, Université de Montréal, Montréal, Québec H2X 0A9, Canada
| | - Moises Freitas-Andrade
- Ottawa Hospital Research Institute, Neuroscience Program, Ottawa, Ontario K1H 8M5, Canada
| | - Chenghua Gu
- Department of Neurobiology, Howard Hughes Medical Institute, Harvard Medical School, Boston, Massachusetts 02115, USA
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Dey S, Gope TK, Ain R. Cellular prion: a novel regulator of decidual cell function at the maternal-fetal interface. Mol Hum Reprod 2025; 31:gaaf016. [PMID: 40323306 DOI: 10.1093/molehr/gaaf016] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2024] [Revised: 03/26/2025] [Indexed: 05/25/2025] Open
Abstract
Cellular prion (PRNP) is a GPI-anchored extrinsic membrane glycoprotein, which has been implicated in mouse decidualization. However, the molecular function of this protein in mouse decidua is not known. In this article, we have characterized and elucidated the possible role of PRNP in mouse decidua. We demonstrated that PRNP expression is evident on embryonic day (E) 6.5 to E9.5 across the primary decidual zone (PDZ) in the mouse implantation site. As gestation progressed, PRNP continued to be expressed in the receding decidua, up to E17.5. shRNA-mediated knockdown of PRNP on E5.5 through E7.5 led to decreased expression of tight junction proteins (TJPs) in PDZ in vivo. These included Cingulin, Afadin, Catenin-α1, Lethal (2) giant larvae protein homolog 1, Claudin-5, and ICAM1. Furthermore, PRNP-positive decidual cells showed augmented expression of autophagic machinery. PRNP knockdown curtailed expression of autophagy-related genes in decidua in vivo. Our results highlight hitherto unknown novel functions of PRNP: (i) an inducer of TJPs at PDZ, which protects the developing embryo and (ii) a decision-maker protein between life and death in decidual cells.
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Affiliation(s)
- Swarnali Dey
- Division of Cell Biology and Physiology, CSIR-Indian Institute of Chemical Biology, Kolkata, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, UP, India
| | - Tamal Kanti Gope
- Division of Cell Biology and Physiology, CSIR-Indian Institute of Chemical Biology, Kolkata, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, UP, India
| | - Rupasri Ain
- Division of Cell Biology and Physiology, CSIR-Indian Institute of Chemical Biology, Kolkata, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, UP, India
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Trevisani M, Berselli A, Alberini G, Centonze E, Vercellino S, Cartocci V, Millo E, Ciobanu DZ, Braccia C, Armirotti A, Pisani F, Zara F, Castagnola V, Maragliano L, Benfenati F. A claudin5-binding peptide enhances the permeability of the blood-brain barrier in vitro. SCIENCE ADVANCES 2025; 11:eadq2616. [PMID: 39792664 PMCID: PMC11721574 DOI: 10.1126/sciadv.adq2616] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/05/2024] [Accepted: 12/09/2024] [Indexed: 01/12/2025]
Abstract
The blood-brain barrier (BBB) maintains brain homeostasis but also prevents most drugs from entering the brain. No paracellular diffusion of solutes is allowed because of tight junctions that are made impermeable by the expression of claudin5 (CLDN5) by brain endothelial cells. The possibility of regulating the BBB permeability in a transient and reversible fashion is in strong demand for the pharmacological treatment of brain diseases. Here, we designed and tested short BBB-active peptides, derived from the CLDN5 extracellular domains and the CLDN5-binding domain of Clostridium perfringens enterotoxin, using a robust workflow of structural modeling and in vitro validation techniques. Computational analysis at the atom level based on solubility and affinity to CLDN5 identified a CLDN5-derived peptide not reported previously called f1-C5C2, which was soluble in biological media, displayed efficient binding to CLDN5, and transiently increased BBB permeability. The peptidomimetic strategy described here may have potential applications in the pharmacological treatment of brain diseases.
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Affiliation(s)
- Martina Trevisani
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- Department of Experimental Medicine, Università degli Studi di Genova, Viale Benedetto XV, 3, 16132 Genova, Italy
| | - Alessandro Berselli
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132 Genova, Italy
| | - Giulio Alberini
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132 Genova, Italy
| | - Eleonora Centonze
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
| | - Silvia Vercellino
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132 Genova, Italy
| | - Veronica Cartocci
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132 Genova, Italy
| | - Enrico Millo
- Department of Experimental Medicine, Università degli Studi di Genova, Viale Benedetto XV, 3, 16132 Genova, Italy
| | - Dinu Zinovie Ciobanu
- Analytical Chemistry Facility, Istituto Italiano di Tecnologia, Via Morego 30, 16163 Genova, Italy
| | - Clarissa Braccia
- Analytical Chemistry Facility, Istituto Italiano di Tecnologia, Via Morego 30, 16163 Genova, Italy
| | - Andrea Armirotti
- Analytical Chemistry Facility, Istituto Italiano di Tecnologia, Via Morego 30, 16163 Genova, Italy
| | - Francesco Pisani
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- Department of Biosciences, Biotechnologies and Biopharmaceutics, University of Bari “Aldo Moro”, 70125 Bari, Italy
| | - Federico Zara
- Department of Neurosciences, Rehabilitation, Ophthalmology, Genetics, Maternal and Child Health (DINOGMI), University of Genova, 16132 Genova, Italy
- Medical Genetics Unit, IRCCS Giannina Gaslini Institute, 16147 Genova, Italy
| | - Valentina Castagnola
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132 Genova, Italy
| | - Luca Maragliano
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- Department of Life and Environmental Sciences, Polytechnic University of Marche, Via Brecce Bianche, 60131 Ancona, Italy
| | - Fabio Benfenati
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132 Genova, Italy
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Cazalla E, Cuadrado A, García-Yagüe ÁJ. Role of the transcription factor NRF2 in maintaining the integrity of the Blood-Brain Barrier. Fluids Barriers CNS 2024; 21:93. [PMID: 39574123 PMCID: PMC11580557 DOI: 10.1186/s12987-024-00599-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2024] [Accepted: 11/15/2024] [Indexed: 11/25/2024] Open
Abstract
BACKGROUND The Blood-Brain Barrier (BBB) is a complex and dynamic interface that regulates the exchange of molecules and cells between the blood and the central nervous system. It undergoes structural and functional throughout oxidative stress and inflammation, which may compromise its integrity and contribute to the pathogenesis of neurodegenerative diseases. MAIN BODY Maintaining BBB integrity is of utmost importance in preventing a wide range of neurological disorders. NRF2 is the main transcription factor that regulates cellular redox balance and inflammation-related gene expression. It has also demonstrated a potential role in regulating tight junction integrity and contributing to the inhibition of ECM remodeling, by reducing the expression of several metalloprotease family members involved in maintaining BBB function. Overall, we review current insights on the role of NRF2 in addressing protection against the effects of BBB dysfunction, discuss its involvement in BBB maintenance in different neuropathological diseases, as well as, some of its potential activators that have been used in vitro and in vivo animal models for preventing barrier dysfunction. CONCLUSIONS Thus, emerging evidence suggests that upregulation of NRF2 and its target genes could suppress oxidative stress, and neuroinflammation, restore BBB integrity, and increase its protection.
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Affiliation(s)
- Eduardo Cazalla
- Department of Biochemistry, School of Medicine, Autonomous University of Madrid (UAM), Madrid, Spain
- Instituto de Investigaciones Biomédicas "Sols-Morreale" (CSIC-UAM), C/ Arturo Duperier, 4, Madrid, 28029, Spain
- Instituto de Investigación Sanitaria La Paz (IdiPaz), Madrid, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain
| | - Antonio Cuadrado
- Department of Biochemistry, School of Medicine, Autonomous University of Madrid (UAM), Madrid, Spain
- Instituto de Investigaciones Biomédicas "Sols-Morreale" (CSIC-UAM), C/ Arturo Duperier, 4, Madrid, 28029, Spain
- Instituto de Investigación Sanitaria La Paz (IdiPaz), Madrid, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain
| | - Ángel Juan García-Yagüe
- Department of Biochemistry, School of Medicine, Autonomous University of Madrid (UAM), Madrid, Spain.
- Instituto de Investigaciones Biomédicas "Sols-Morreale" (CSIC-UAM), C/ Arturo Duperier, 4, Madrid, 28029, Spain.
- Instituto de Investigación Sanitaria La Paz (IdiPaz), Madrid, Spain.
- Centro de Investigación Biomédica en Red de Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.
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Kang J, Xie W, Wu L, Liu Y, Xu Y, Xu Y, Mai Y, Peng L, Huang B, Guo S, Luo S. The ethanolic extract of domesticated Amauroderma rugosum alleviated DSS-induced ulcerative colitis via repairing the intestinal barrier. Food Sci Biotechnol 2024; 33:3335-3345. [PMID: 39328223 PMCID: PMC11422322 DOI: 10.1007/s10068-024-01565-5] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2023] [Revised: 01/21/2024] [Accepted: 03/15/2024] [Indexed: 09/28/2024] Open
Abstract
Amauroderma rugosum (Blume and T. Nees) Torrend (Ganodermataceae) (A. rugosum) has been found to have anti-inflammatory ability in previous studies. The present study aimed to verify the therapeutic benefits of A. rugosum in the treatment of ulcerative colitis and to investigate its underlying mechanism of action. Acute experimental ulcerative colitis was induced by feeding the mice drinking water supplemented with dextran sodium sulfate (DSS). The findings indicated that the ethanolic extract of domesticated A. rugosum exhibited therapeutic efficacy comparable to Salazosulfapyridine (SASP) in mitigating clinical symptoms and the pathological score of the colon. Furthermore, A. rugosum exhibited the capacity to enhance the expression of tight junction (TJ) proteins, while concurrently decreasing the levels of TNF-ɑ and IL-6. A noteworthy finding is that it exhibited the capability to diminish the nuclear translocation of NF-κB p65. In conclusion, A. rugosum attenuates DSS-induced ulcerative colitis by enhancing intestinal barrier function and inhibiting mucosal inflammation. Supplementary Information The online version contains supplementary material available at 10.1007/s10068-024-01565-5.
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Affiliation(s)
- Jianyuan Kang
- Institute of Gastroenterology, Shenzhen Traditional Chinese Medicine Hospital, The Fourth Clinical Medical College of Guangzhou University of Chinese Medicine, No. 1 Fuhua Road, Futian District, Shenzhen, 518033 Guangdong China
| | - Weicang Xie
- Institute of Gastroenterology, Shenzhen Traditional Chinese Medicine Hospital, The Fourth Clinical Medical College of Guangzhou University of Chinese Medicine, No. 1 Fuhua Road, Futian District, Shenzhen, 518033 Guangdong China
| | - Lingping Wu
- School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, 510006 China
| | - Yuanyuan Liu
- Yantian District Maternity and Child Healthcare Hospital, Shenzhen, 518081 China
| | - Youcai Xu
- Institute of Gastroenterology, Shenzhen Traditional Chinese Medicine Hospital, The Fourth Clinical Medical College of Guangzhou University of Chinese Medicine, No. 1 Fuhua Road, Futian District, Shenzhen, 518033 Guangdong China
| | - Yifei Xu
- Institute of Gastroenterology, Shenzhen Traditional Chinese Medicine Hospital, The Fourth Clinical Medical College of Guangzhou University of Chinese Medicine, No. 1 Fuhua Road, Futian District, Shenzhen, 518033 Guangdong China
| | - Yanzhen Mai
- Huizhou Health Sciences Polytechnic, Huizhou, 516025 China
| | - Lisheng Peng
- Institute of Gastroenterology, Shenzhen Traditional Chinese Medicine Hospital, The Fourth Clinical Medical College of Guangzhou University of Chinese Medicine, No. 1 Fuhua Road, Futian District, Shenzhen, 518033 Guangdong China
| | - Bin Huang
- Institute of Gastroenterology, Shenzhen Traditional Chinese Medicine Hospital, The Fourth Clinical Medical College of Guangzhou University of Chinese Medicine, No. 1 Fuhua Road, Futian District, Shenzhen, 518033 Guangdong China
| | - Shaoju Guo
- Institute of Gastroenterology, Shenzhen Traditional Chinese Medicine Hospital, The Fourth Clinical Medical College of Guangzhou University of Chinese Medicine, No. 1 Fuhua Road, Futian District, Shenzhen, 518033 Guangdong China
| | - Shuang Luo
- Institute of Gastroenterology, Shenzhen Traditional Chinese Medicine Hospital, The Fourth Clinical Medical College of Guangzhou University of Chinese Medicine, No. 1 Fuhua Road, Futian District, Shenzhen, 518033 Guangdong China
- Guizhou Provincial Engineering Research Center of Ecological Food Innovation, School of Public Health, Guizhou Medical University, Guian New Area, Guiyang, 561113 Guizhou China
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Ma J, Zhao Y, Cui Y, Lin H. Hypoxia Postconditioning Attenuates Hypoxia-Induced Inflammation and Endothelial Barrier Dysfunction. J Surg Res 2024; 301:413-422. [PMID: 39042975 DOI: 10.1016/j.jss.2024.06.007] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2023] [Revised: 05/15/2024] [Accepted: 06/16/2024] [Indexed: 07/25/2024]
Abstract
INTRODUCTION In recent years, a number of studies have demonstrated that hypoxia reoxygenation (HR) induced by ischemia postconditioning (IPC) reduces endothelial barrier dysfunction and inflammation in various models. When HR occurs, the P38 mitogen-activated protein kinase (P38 MAPK) breaks down the endothelial barrier. But no study has clearly clarified the effect of hypoxia postconditioning (HPC) on P38 MAPK in human dermal microvascular endothelial cells. Therefore, we investigated the function of HPC on P38 MAPK during HR in vitro. METHODS Human dermal microvascular endothelial cells were cultured in a hypoxic incubator for 8 h. Then cells were reperfused for 12 h (reoxygenation) or postconditioned by 5 min of reoxygenation and 5 min of re-hypoxia 3 times followed by 11.5 h reoxygenation. SB203580 was used as an inhibitor of P38 MAPK. Cell counting kit-8 assay kits were employed to detect cell activity. The corresponding levels of IL-6, IL-8 and IL-1β were examined via Enzyme-Linked ImmunoSorbent Assay. The endothelial barrier was evaluated using fluorescein isothiocyanate-dextran leakage assay. Western blot was used to detect claudin-5, phosphorylation of P38 MAPK (P-P38 MAPK) and P38 MAPK expression. Claudin-5 localization was studied by immunofluorescence. RESULTS HR induced endothelial barrier hyperpermeability, elevated inflammation levels, and increased the P-P38 MAPK. But HPC reduced cell injury and maintained the integrity of the endothelial barrier while inhibiting P-P38 MAPK and increasing expression of claudin-5. HPC redistributed claudin-5 in a continuous and linear pattern on the cell membrane. CONCLUSIONS HPC protects against HR induced downregulation and redistribution of claudin-5 by inhibiting P-P38 MAPK.
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Affiliation(s)
- Jiaxing Ma
- Plastic and Reconstructive Surgery, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
| | - Yinhua Zhao
- Plastic and Reconstructive Surgery, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
| | - Yue Cui
- Plastic and Reconstructive Surgery, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
| | - Huang Lin
- Plastic and Reconstructive Surgery, Beijing Anzhen Hospital, Capital Medical University, Beijing, China.
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Marsch P, Rajagopal N, Nangia S. Biophysics of claudin proteins in tight junction architecture: Three decades of progress. Biophys J 2024; 123:2363-2378. [PMID: 38859584 PMCID: PMC11365114 DOI: 10.1016/j.bpj.2024.06.010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/17/2024] [Revised: 05/19/2024] [Accepted: 06/07/2024] [Indexed: 06/12/2024] Open
Abstract
Tight junctions are cell-cell adhesion complexes that act as gatekeepers of the paracellular space. Formed by several transmembrane proteins, the claudin family performs the primary gate-keeping function. The claudin proteins form charge and size-selective diffusion barriers to maintain homeostasis across endothelial and epithelial tissue. Of the 27 known claudins in mammals, some are known to seal the paracellular space, while others provide selective permeability. The differences in permeability arise due to the varying expression levels of claudins in each tissue. The tight junctions are observed as strands in freeze-fracture electron monographs; however, at the molecular level, tight junction strands form when multiple claudin proteins assemble laterally (cis assembly) within a cell and head-on (trans assembly) with claudins of the adjacent cell in a zipper-like architecture, closing the gap between the neighboring cells. The disruption of tight junctions caused by changing claudin expression levels or mutations can lead to diseases. Therefore, knowledge of the molecular architecture of the tight junctions and how that is tied to tissue-specific function is critical for fighting diseases. Here, we review the current understanding of the tight junctions accrued over the last three decades from experimental and computational biophysics perspectives.
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Affiliation(s)
- Patrick Marsch
- Department of Biomedical and Chemical Engineering, Syracuse University, Syracuse, New York
| | - Nandhini Rajagopal
- Department of Biomedical and Chemical Engineering, Syracuse University, Syracuse, New York
| | - Shikha Nangia
- Department of Biomedical and Chemical Engineering, Syracuse University, Syracuse, New York.
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Mao JX, Li JJ, Lu XY, Zhong HX, Zhao YY, Zhu LY, Fu H, Ding GS, Teng F, Chen M, Guo WY. Dichotomous roles of ADAR1 in liver hepatocellular carcinoma and kidney renal cell carcinoma: Unraveling the complex tumor microenvironment and prognostic significance. Int Immunopharmacol 2024; 136:112340. [PMID: 38820962 DOI: 10.1016/j.intimp.2024.112340] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2024] [Revised: 05/14/2024] [Accepted: 05/22/2024] [Indexed: 06/02/2024]
Abstract
BACKGROUND Adenosine deaminase acting on RNA 1 (ADAR1) is an RNA-editing enzyme that significantly impacts cancer progression and various biological processes. The expression of ADAR1 mRNA has been examined in multiple cancer types using The Cancer Genome Atlas (TCGA) dataset, revealing distinct patterns in kidney chromophobe (KICH), kidney renal clear cell carcinoma (KIRC), kidney renal papillary cell carcinoma (KIRP), and liver hepatocellular carcinoma (LIHC) compared to normal controls. However, the reasons for these differential expressions remain unclear. METHODS In this study, we performed RT-PCR and western blotting (WB) to validate ADAR1 expression patterns in clinical tissue samples. Survival analysis and immune microenvironment analysis (including immune score and stromal score) were conducted using TCGA data to determine the specific cell types associated with ADAR1, as well as the key genes in those cell types. The relationship between ADAR1 and specific cell types' key genes was verified by immunohistochemistry (IHC), using clinical liver and kidney cancer samples. RESULTS Our validation analysis revealed that ADAR1 expression was downregulated in KICH, KIRC, and KIRP, while upregulated in LIHC compared to normal tissues. Notably, a significant correlation was found between ADAR1 mRNA expression and patient prognosis, particularly in KIRC, KIRP, and LIHC. Interestingly, we observed a positive correlation between ADAR1 expression and stromal scores in KIRC, whereas a negative correlation was observed in LIHC. Cell type analysis highlighted distinct relationships between ADAR1 expression and the two stromal cell types, blood endothelial cells (BECs) and lymphatic endothelial cells (LECs), and further determined the signature gene claudin-5 (CLDN5), in KIRC and LIHC. Moreover, ADAR1 was inversely related with CLDN5 in KIRC (n = 26) and LIHC (n = 30) samples, verified via IHC. CONCLUSIONS ADAR1 plays contrasting roles in LIHC and KIRC, associated with the enrichment of BECs and LECs within tumors. This study sheds light on the significant roles of stromal cells within the complex tumor microenvironment (TME) and provides new insights for future research in tumor immunotherapy and precision medicine.
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MESH Headings
- Adenosine Deaminase/genetics
- Adenosine Deaminase/metabolism
- Humans
- Tumor Microenvironment
- Liver Neoplasms/genetics
- Liver Neoplasms/pathology
- Liver Neoplasms/metabolism
- Carcinoma, Hepatocellular/genetics
- Carcinoma, Hepatocellular/metabolism
- Carcinoma, Hepatocellular/pathology
- Carcinoma, Hepatocellular/mortality
- Kidney Neoplasms/genetics
- Kidney Neoplasms/pathology
- Kidney Neoplasms/metabolism
- Kidney Neoplasms/mortality
- Carcinoma, Renal Cell/genetics
- Carcinoma, Renal Cell/metabolism
- Carcinoma, Renal Cell/pathology
- RNA-Binding Proteins/metabolism
- RNA-Binding Proteins/genetics
- Prognosis
- Gene Expression Regulation, Neoplastic
- Female
- Male
- Biomarkers, Tumor/metabolism
- Biomarkers, Tumor/genetics
- Middle Aged
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Affiliation(s)
- Jia-Xi Mao
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China
| | - Jing-Jing Li
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China
| | - Xin-Yi Lu
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China
| | - Han-Xiang Zhong
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China
| | - Yuan-Yu Zhao
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China
| | - Li-Ye Zhu
- Department of Immunology and Medical Immunology State Key Laboratory, Naval Medical University, Shanghai 200433, China
| | - Hong Fu
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China
| | - Guo-Shan Ding
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China
| | - Fei Teng
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China.
| | - Ming Chen
- Department of Urology, Changzheng Hospital, Naval Medical University, Shanghai 200003, China.
| | - Wen-Yuan Guo
- Department of Liver Surgery and Organ Transplantation, Changzheng Hospital, Naval Medical University, Shanghai 200003, China.
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10
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Dithmer S, Blasig IE, Fraser PA, Qin Z, Haseloff RF. The Basic Requirement of Tight Junction Proteins in Blood-Brain Barrier Function and Their Role in Pathologies. Int J Mol Sci 2024; 25:5601. [PMID: 38891789 PMCID: PMC11172262 DOI: 10.3390/ijms25115601] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2024] [Revised: 03/10/2024] [Accepted: 03/28/2024] [Indexed: 06/21/2024] Open
Abstract
This review addresses the role of tight junction proteins at the blood-brain barrier (BBB). Their expression is described, and their role in physiological and pathological processes at the BBB is discussed. Based on this, new approaches are depicted for paracellular drug delivery and diagnostics in the treatment of cerebral diseases. Recent data provide convincing evidence that, in addition to its impairment in the course of diseases, the BBB could be involved in the aetiology of CNS disorders. Further progress will be expected based on new insights in tight junction protein structure and in their involvement in signalling pathways.
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Affiliation(s)
- Sophie Dithmer
- Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany (I.E.B.)
| | - Ingolf E. Blasig
- Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany (I.E.B.)
| | | | - Zhihai Qin
- Institute of Biophysics, Chinese Academy of Sciences, Beijing 100049, China
| | - Reiner F. Haseloff
- Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany (I.E.B.)
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11
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Han SH, Lee HD, Lee S, Lee AY. Taraxacum coreanum Nakai extract attenuates lipopolysaccharide-induced inflammatory responses and intestinal barrier dysfunction in Caco-2 cells. JOURNAL OF ETHNOPHARMACOLOGY 2024; 319:117105. [PMID: 37660957 DOI: 10.1016/j.jep.2023.117105] [Citation(s) in RCA: 2] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/05/2023] [Revised: 08/27/2023] [Accepted: 08/28/2023] [Indexed: 09/05/2023]
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE Taraxacum coreanum Nakai (TC) is a dandelion native to Korea that has long been used as a medicinal herb with antioxidant and anti-inflammatory properties. Intestinal inflammation is closely associated with intestinal epithelial barrier disruption, which leads to the progression of various intestinal diseases. AIM OF THE STUDY The aim of this study was to investigate the protective effects of TC extract on inflammatory responses and intestinal barrier dysfunction in lipopolysaccharide (LPS)-stimulated Caco-2 cells. MATERIALS AND METHODS The inhibitory effect of TC on nitric oxide (NO) and pro-inflammatory cytokines production were determined by Griess reagent and enzyme-linked immunosorbent assay, respectively. The epithelial permeability was evaluated by transepithelial electrical resistance (TEER) assay, and inflammation- and tight junction (TJ)-related protein expression were analyzed by Western blotting. In addition, the presence of ten active compounds was identified and quantified using UHPLC-ESI-MS and HPLC-DAD analyses. RESULTS Treatment with TC significantly reduced NO production and pro-inflammatory cytokines production [interleukin (IL)-6 and tumor necrosis factor (TNF)-α] compared to the group treated with LPS only, particularly at 100 μg/mL. TC significantly decreased monolayer permeability as detected by TEER. In addition, the transmission of fluorescein isothiocyanate-dextran 4 across the barrier was decreased after treatment with TC. Inflammation-related proteins (inducible NO synthase, cyclooxygenase-2, TNF-α, IL-6, and IL-1β) were down-regulated after treatment with TC. In contrast, TC significantly increased the protein levels of the TJ-related protein, claudin-5. Ten phytochemicals (protocatechuic acid, chlorogenic acid, caffeic acid, scopoletin, chicoric acid, hyperoside, nicotiflorin, luteoloside, sophoricoside, and luteolin) were identified by UHPLC-ESI-MS and HPLC-DAD analysis. CONCLUSION Our findings suggest that ethanolic extract of TC could attenuate the LPS-induced intestinal barrier dysfunction by increasing the TJ protein and suppressing inflammatory responses.
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Affiliation(s)
- Seok Hee Han
- Department of Food Science, Gyeongsang National University, Jinju, 52725, Republic of Korea.
| | - Hak-Dong Lee
- Department of Plant Science and Technology, Chung-Ang University, Anseong, 17546, Republic of Korea.
| | - Sanghyun Lee
- Department of Plant Science and Technology, Chung-Ang University, Anseong, 17546, Republic of Korea; Natural Product Institute of Science and Technology, Anseong, 17546, Republic of Korea.
| | - Ah Young Lee
- Department of Food Science, Gyeongsang National University, Jinju, 52725, Republic of Korea.
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12
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Stein L, Vollstaedt ML, Amasheh S. Cannabidiol Strengthening of Gastric Tight Junction Complexes Analyzed in an Improved Xenopus Oocyte Assay. MEMBRANES 2024; 14:18. [PMID: 38248708 PMCID: PMC10819461 DOI: 10.3390/membranes14010018] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/30/2023] [Revised: 12/25/2023] [Accepted: 01/03/2024] [Indexed: 01/23/2024]
Abstract
Cannabidiol (CBD), the non-psychoactive compound derived from the cannabis plant, has gained attention in recent years as a remedy against gastrointestinal disorders ranging from nausea and inflammation to abdominal pain. Recent advances demonstrated an effect on inflammatory pathways and barrier proteins. However, information on possible direct effects is scarce and needs to be addressed, as applications are currently increasing in popularity. To accomplish this, we have employed Xenopus laevis oocytes as a heterologous expression system for analysis of the direct effects on stomach-specific claudins and further developed tight junction (TJ) protein interaction assays. Human claudin-4, claudin-5, and claudin-18.2 were expressed in Xenopus oocytes, clustered in pairs to form contact areas, and analyzed in a two-cell model approach, including measurement of the contact area and contact strength. CLDN4/5/18 + CLDN4/5/18 oocyte pairs were incubated with 20 µM CBD or with 40 µM CBD and were compared to cells without CBD treatment (ctrl). For interaction analysis, the contact area was measured after 24 h and 48 h. Whereas CBD did not affect the size of the protein interaction area, Double Orbital Challenge experiments revealed an increased contact strength after 24 h incubation with CBD. In addition, the Xenopus oocyte experiments were accompanied by an analysis of claudin-4, -5, and -18 expression in gastric epithelium by immunoblotting and immunohistochemistry. Claudin-4, -5, and -18 were strongly expressed, indicating a major role for gastric epithelial barrier function. In summary, our study shows direct effects of 40 µM CBD on Xenopus oocytes heterologously expressing a stomach-specific claudin combination, indicating a supportive and beneficial effect of CBD on gastric TJ proteins.
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Affiliation(s)
| | | | - Salah Amasheh
- Institute of Veterinary Physiology, School of Veterinary Medicine, Freie Universität Berlin, 14163 Berlin, Germany; (L.S.); (M.-L.V.)
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13
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Boehm E, Droessler L, Amasheh S. Cannabidiol attenuates inflammatory impairment of intestinal cells expanding biomaterial-based therapeutic approaches. Mater Today Bio 2023; 23:100808. [PMID: 37779918 PMCID: PMC10539670 DOI: 10.1016/j.mtbio.2023.100808] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2023] [Revised: 09/06/2023] [Accepted: 09/19/2023] [Indexed: 10/03/2023] Open
Abstract
Cannabis-based biomaterials have the potential to deliver anti-inflammatory therapeutics specifically to desired cells, tissues, and organs, enhancing drug delivery and the effectiveness of anti-inflammatory treatment while minimizing toxicity. As a major component of Cannabis, Cannabidiol (CBD) has gained major attention in recent years because of its potential therapeutic properties, e.g., for restoring a disturbed barrier resulting from inflammatory conditions. The aim of this study was to test the hypothesis that CBD has beneficial effects under normal and inflammatory conditions in the established non-transformed intestinal epithelial cell model IPEC-J2. CBD induced a significant increase in transepithelial electrical resistance (TER) values and a decrease in the paracellular permeability of [³H]-D-Mannitol, indicating a strengthening effect on the barrier. Under inflammatory conditions induced by tumor necrosis factor alpha (TNFα), CBD stabilized the TER and mitigated the increase in paracellular permeability. Additionally, CBD prevented the barrier-disrupting effects of TNFα on the distribution and localization of sealing TJ proteins. CBD also affected the expression of TNF receptors. These findings demonstrate the potential of CBD as a component of Cannabis-based biomaterials used in the development of novel therapeutic approaches against inflammatory pathogenesis.
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Affiliation(s)
- Elisa Boehm
- Institute of Veterinary Physiology, School of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163, Berlin, Germany
| | - Linda Droessler
- Institute of Veterinary Physiology, School of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163, Berlin, Germany
| | - Salah Amasheh
- Institute of Veterinary Physiology, School of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163, Berlin, Germany
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14
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Raya-Sandino A, Lozada-Soto KM, Rajagopal N, Garcia-Hernandez V, Luissint AC, Brazil JC, Cui G, Koval M, Parkos CA, Nangia S, Nusrat A. Claudin-23 reshapes epithelial tight junction architecture to regulate barrier function. Nat Commun 2023; 14:6214. [PMID: 37798277 PMCID: PMC10556055 DOI: 10.1038/s41467-023-41999-9] [Citation(s) in RCA: 29] [Impact Index Per Article: 14.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2022] [Accepted: 09/26/2023] [Indexed: 10/07/2023] Open
Abstract
Claudin family tight junction proteins form charge- and size-selective paracellular channels that regulate epithelial barrier function. In the gastrointestinal tract, barrier heterogeneity is attributed to differential claudin expression. Here, we show that claudin-23 (CLDN23) is enriched in luminal intestinal epithelial cells where it strengthens the epithelial barrier. Complementary approaches reveal that CLDN23 regulates paracellular ion and macromolecule permeability by associating with CLDN3 and CLDN4 and regulating their distribution in tight junctions. Computational modeling suggests that CLDN23 forms heteromeric and heterotypic complexes with CLDN3 and CLDN4 that have unique pore architecture and overall net charge. These computational simulation analyses further suggest that pore properties are interaction-dependent, since differently organized complexes with the same claudin stoichiometry form pores with unique architecture. Our findings provide insight into tight junction organization and propose a model whereby different claudins combine to form multiple distinct complexes that modify epithelial barrier function by altering tight junction structure.
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Affiliation(s)
- Arturo Raya-Sandino
- Department of Pathology, University of Michigan Medical School, Ann Arbor, MI, USA
| | | | - Nandhini Rajagopal
- Department of Biomedical and Chemical Engineering, Syracuse University, Syracuse, NY, USA
| | | | - Anny-Claude Luissint
- Department of Pathology, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Jennifer C Brazil
- Department of Pathology, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Guiying Cui
- Department of Pediatrics, Emory + Children's Center for Cystic Fibrosis and Airways Disease Research, Emory University School of Medicine, Atlanta, GA, USA
| | - Michael Koval
- Departments of Medicine and Cell Biology, Emory University School of Medicine, Atlanta, GA, USA
| | - Charles A Parkos
- Department of Pathology, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Shikha Nangia
- Department of Biomedical and Chemical Engineering, Syracuse University, Syracuse, NY, USA.
| | - Asma Nusrat
- Department of Pathology, University of Michigan Medical School, Ann Arbor, MI, USA.
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15
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Zhang S, Chen A, Jiang L, Liu X, Chai L. Copper-mediated shifts in transcriptomic responses of intestines in Bufo gargarizans tadpoles to lead stress. ENVIRONMENTAL SCIENCE AND POLLUTION RESEARCH INTERNATIONAL 2023; 30:50144-50161. [PMID: 36790706 DOI: 10.1007/s11356-023-25801-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/19/2022] [Accepted: 02/04/2023] [Indexed: 04/16/2023]
Abstract
The differential transcriptomic responses of intestines in Bufo gargarizans tadpoles to Pb alone or in the presence of Cu were evaluated. Tadpoles were exposed to 30 μg/L Pb individually and in combination with Cu at 16 or 64 μg/L from Gosner stage (Gs) 26 to Gs 38. After de novo assembly, 105,107 unigenes were generated. Compared to the control group, 7387, 6937, and 11139 differentially expressed genes (DEGs) were identified in the treatment of Pb + Cu0, Pb + Cu16, and Pb + Cu64, respectively. In addition, functional annotation and enrichment analysis of DEGs revealed substantial transcriptional reprogramming of diverse molecular and biological pathways were induced in all heavy metal treatments. The relative expression levels of genes associated with intestinal epithelial barrier and bile acids (BAs) metabolism, such as mucin2, claudin5, ZO-1, Asbt, and Ost-β, were validated by qPCR. This study demonstrated that Pb exposure induced transcriptional responses in tadpoles, and the responses could be modulated by Cu.
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Affiliation(s)
- Siliang Zhang
- School of Water and Environment, Chang'an University, Xi'an, 710054, People's Republic of China
- Key Laboratory of Subsurface Hydrology and Ecological Effect in Arid Region of Ministry of Education, Chang'an University, Xi'an, 710054, People's Republic of China
| | - Aixia Chen
- School of Water and Environment, Chang'an University, Xi'an, 710054, People's Republic of China
- Key Laboratory of Subsurface Hydrology and Ecological Effect in Arid Region of Ministry of Education, Chang'an University, Xi'an, 710054, People's Republic of China
| | - Ling Jiang
- School of Water and Environment, Chang'an University, Xi'an, 710054, People's Republic of China
- Key Laboratory of Subsurface Hydrology and Ecological Effect in Arid Region of Ministry of Education, Chang'an University, Xi'an, 710054, People's Republic of China
| | - Xiaoli Liu
- School of Water and Environment, Chang'an University, Xi'an, 710054, People's Republic of China
- Key Laboratory of Subsurface Hydrology and Ecological Effect in Arid Region of Ministry of Education, Chang'an University, Xi'an, 710054, People's Republic of China
| | - Lihong Chai
- School of Water and Environment, Chang'an University, Xi'an, 710054, People's Republic of China.
- Key Laboratory of Subsurface Hydrology and Ecological Effect in Arid Region of Ministry of Education, Chang'an University, Xi'an, 710054, People's Republic of China.
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16
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Liman N. The abundance and localization of claudin-1 and -5 in the adult tomcats (Felis catus) testis, tubules rectus, rete testis, efferent ductules, and epididymis. Anat Rec (Hoboken) 2023. [PMID: 36688626 DOI: 10.1002/ar.25165] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2022] [Revised: 01/09/2023] [Accepted: 01/09/2023] [Indexed: 01/24/2023]
Abstract
Tight junctions (TJ) are the anatomical component of blood-testis (BTB) and blood-epididymis (BEB) barriers and contain many proteins, including claudins. The presence of claudins in domestic cat testis and epididymis has not been previously described. This study aimed to determine whether claudin-1 and claudin-5 participate in the structure of BTB and BEB and whether their amounts differ between the testis and epididymal segments of adult cats, using Western blotting (WB) and immunohistochemistry. WB results demonstrated that claudin-1 was significantly lower in the testis than in all epididymal segments and higher in the corpus epididymis than in the cauda, while claudin-5 in the testis was significantly lower than in the caput and corpus. Claudin-1 was absent at the Sertoli-Sertoli junctions, while claudin-5 was detected at the level of the BTB during stages I and VIII. Both claudins were observed in the pachytene spermatocytes and the developing acrosome of the round and elongating spermatids. Claudin-5 was also detected in the cytoplasm of some spermatogonia, Sertoli cells, and late spermatid acrosome. In the epididymal segments, both claudins were localized to the area of the tight junctions and along the entire length of the lateral plasma membranes of adjacent principal cells and between principal and basal cells. These results may indicate that in the domestic cat, claudin-1 and -5 participate as both tight junction proteins and adhesion molecules in the BEB's structure, claudin 5 is a component of the BTB, and both proteins may be involved in postmeiotic germ cell development, especially acrosome development.
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Affiliation(s)
- Narin Liman
- Department of Histology and Embryology, Faculty of Veterinary Medicine, Erciyes University, Kayseri, Turkey
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17
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Berselli A, Alberini G, Benfenati F, Maragliano L. The impact of pathogenic and artificial mutations on Claudin-5 selectivity from molecular dynamics simulations. Comput Struct Biotechnol J 2023; 21:2640-2653. [PMID: 37138900 PMCID: PMC10149405 DOI: 10.1016/j.csbj.2023.04.001] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2023] [Revised: 04/03/2023] [Accepted: 04/03/2023] [Indexed: 05/05/2023] Open
Abstract
Tight-junctions (TJs) are multi-protein complexes between adjacent endothelial or epithelial cells. In the blood-brain-barrier (BBB), they seal the paracellular space and the Claudin-5 (Cldn5) protein forms their backbone. Despite the fundamental role in brain homeostasis, little is known on Cldn5-based TJ assemblies. Different structural models were suggested, with Cldn5 protomers generating paracellular pores that restrict the passage of ions and small molecules. Recently, the first Cldn5 pathogenic mutation, G60R, was identified and shown to induce Cl--selective channels and Na+ barriers in BBB TJs, providing an excellent opportunity to validate the structural models. Here, we used molecular dynamics to study the permeation of ions and water through two distinct G60R-Cldn5 paracellular architectures. Only the so-called Pore I reproduces the functional modification observed in experiments, displaying a free energy (FE) minimum for Cl- and a barrier for Na+ consistent with anionic selectivity. We also studied the artificial Q57D and Q63D mutations in the constriction region, Q57 being conserved in Cldns except for cation permeable homologs. In both cases, we obtain FE profiles consistent with facilitated passage of cations. Our calculations provide the first in-silico description of a Cldn5 pathogenic mutation, further assessing the TJ Pore I model and yielding new insight on BBB's paracellular selectivity.
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Affiliation(s)
- Alessandro Berselli
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- Department of Experimental Medicine, Università degli Studi di Genova, Viale Benedetto XV, 3, 16132, Genova, Italy
| | - Giulio Alberini
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132, Genova, Italy
| | - Fabio Benfenati
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132, Genova, Italy
- Corresponding authors at: Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy.
| | - Luca Maragliano
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- Department of Life and Environmental Sciences, Polytechnic University of Marche, Via Brecce Bianche, 60131, Ancona, Italy
- Corresponding authors at: Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy.
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18
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Yadav R, Kumar Y, Dahiya D, Bhatia A. Claudins: The Newly Emerging Targets in Breast Cancer. Clin Breast Cancer 2022; 22:737-752. [PMID: 36175290 DOI: 10.1016/j.clbc.2022.09.001] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/13/2022] [Accepted: 09/04/2022] [Indexed: 01/25/2023]
Abstract
Claudin-low breast cancers are recently described entities showing low expression of certain claudins and cell adhesion molecules. Claudins constitute the backbone of tight junctions (TJs) formed between 2 cells. Their dysregulation plays a vital role in tumorigenesis. First part of the article focuses on the role of claudins in the TJ organization, their structural-functional characteristics, and post-transcriptional and translational modifications. The latter part of the review attempts to summarize existing knowledge regarding the status of claudins in breast cancer. The article also provides an overview of the effect of claudins on tumor progression, metastasis, stemness, chemotherapy resistance, and their crosstalk with relevant signaling pathways in breast cancer. Claudins can act as 2-edged swords in tumors. Some claudins have either tumor-suppressive/ promoting action, while others work as both in a context-dependent manner. Claudins regulate many important events in breast cancer. However, the intricacies involved in their activity are poorly understood. Post-translational modifications in claudins and their impact on TJ integrity, function, and tumor behavior are still unclear. Although their role in adverse events in breast cancer is recognized, their potential to serve as relevant targets for future therapeutics, especially for difficult-to-treat subtypes of the above malignancy, remains to be explored.
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Affiliation(s)
- Reena Yadav
- Department of Experimental Medicine and Biotechnology, Postgraduate Institute of Medical Education and Research, Chandigarh, India
| | - Yashwant Kumar
- Department of Immunopathology, Postgraduate Institute of Medical Education and Research, Chandigarh, India
| | - Divya Dahiya
- Department of General Surgery, Postgraduate Institute of Medical Education and Research, Chandigarh, India
| | - Alka Bhatia
- Department of Experimental Medicine and Biotechnology, Postgraduate Institute of Medical Education and Research, Chandigarh, India.
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19
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Berselli A, Benfenati F, Maragliano L, Alberini G. Multiscale modelling of claudin-based assemblies: a magnifying glass for novel structures of biological interfaces. Comput Struct Biotechnol J 2022; 20:5984-6010. [DOI: 10.1016/j.csbj.2022.10.038] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2022] [Revised: 10/24/2022] [Accepted: 10/24/2022] [Indexed: 11/03/2022] Open
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20
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Liu G, Guo B, Luo M, Sun S, Lin Q, Kan Q, He Z, Miao J, Du H, Xiao H, Cao Y. A comprehensive review on preparation, structure-activities relationship, and calcium bioavailability of casein phosphopeptides. Crit Rev Food Sci Nutr 2022; 64:996-1014. [PMID: 36052610 DOI: 10.1080/10408398.2022.2111546] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/03/2022]
Abstract
Calcium is one of the important elements for human health. Calcium deficiencies can lead to numerous diseases. Calcium chelating peptides have shown potential application in the management of calcium deficiencies. Casein phosphopeptides (CPP) are phosphoseryl-containing fragments of casein by enzymatic hydrolysis or fermentation during manufacture of milk products as well as during intestinal digestion. An increasing number of CPP with the ability to facilitate and enhance the bioavailability of calcium are being discovered and identified. In this review, 249 reported CPP derived from four types of bovine casein (αs1, αs2, β and κ) were collected, and the amino acid sequence and phosphoserine group information were sorted out. This review outlines the current enzyme hydrolysis, detection methods, purification, structure-activity relationship and mechanism of intestinal calcium absorption in vitro and in vivo as well as application of CPP.
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Affiliation(s)
- Guo Liu
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
- College of Horticulture, South China Agricultural University, Guangzhou, Guangdong, China
| | - Baoyan Guo
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
- College of Materials and Energy, South China Agricultural University, Guangzhou, China
| | - Minna Luo
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
- Department of Food Science, University of Massachusetts, Amherst, MA, USA
| | - Shengwei Sun
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
| | - Qianru Lin
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
| | - Qixin Kan
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
| | - Zeqi He
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
| | - Jianyin Miao
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
| | - Hengjun Du
- Department of Food Science, University of Massachusetts, Amherst, MA, USA
| | - Hang Xiao
- Department of Food Science, University of Massachusetts, Amherst, MA, USA
| | - Yong Cao
- Guangdong Provincial Key Laboratory of Nutraceuticals and Functional Foods, College of Food Science, South China Agricultural University, Guangzhou, China
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21
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Mucosal expression of Ca and P transporters and claudins in the small intestine of broilers is altered by dietary Ca:P in a limestone particle size dependent manner. PLoS One 2022; 17:e0273852. [PMID: 36048795 PMCID: PMC9436080 DOI: 10.1371/journal.pone.0273852] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2021] [Accepted: 08/16/2022] [Indexed: 12/01/2022] Open
Abstract
High calcium (Ca) intake and fine limestone reduces precaecal phosphorus (P) absorption independently of P solubility in broilers. This study aimed to determine whether dietary total Ca: total P ratio (Ca:P) and limestone particle size (LPS) affect gene expression of P transporters in the small intestine. A total of 384 one-day-old Ross 308 male broiler chickens received diets low (0.50), medium (1.00) or high (1.75) in Ca:P containing either fine (160 μm) or coarse (1062 μm) limestone, in a 3×2 factorial arrangement. Expression of Ca- and P-related genes were determined using real-time quantitative PCR (RT-qPCR) in duodenum and jejunum. Increasing dietary Ca:P decreased duodenal calcium-sensing receptor (CaSR), calbindin-D28k (CaBP-D28k), plasma membrane Ca-ATPase 1 (PMCA1) and sodium-coupled P cotransporter type IIb (NaPi-IIb), but not transient receptor potential canonical 1 (TRPC1) mRNA. This effect was greater with fine limestone when Ca:P increased from low to medium, but greater with coarse limestone when increased from medium to high. A similar inhibitory effect was observed for jejunal CaBP-D28k expression where increasing dietary Ca:P and fine limestone decreased CaSR mRNA, while dietary Ca:P decreased TRPC1 mRNA only for coarse limestone. It also decreased jejunal NaPi-IIb mRNA irrespective of LPS. Dietary treatments did not affect jejunal PMCA1 mRNA expression or that of inorganic phosphate transporter 1 and 2 and xenotropic and polytropic retrovirus receptor 1 in both intestinal segments. Dietary Ca increase reduced mucosal claudin-2 mRNA in both segments, and jejunal zonula occludens-1 (ZO-1) mRNA only for coarse limestone. In conclusion, increasing dietary Ca:P reduced expression of duodenal P transporters (NaPi-IIb) in a LPS dependent manner, hence Ca induced reduction in intestinal P absorption is mediated by decreasing P transporters expression. Dietary Ca reduces Ca digestibility by downregulating mRNA expression of both Ca permeable claudin-2 and Ca transporters (CaBP-D28k, PMCA1).
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22
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Stein L, Brunner N, Amasheh S. Functional Analysis of Gastric Tight Junction Proteins in Xenopus laevis Oocytes. MEMBRANES 2022; 12:membranes12080731. [PMID: 35893449 PMCID: PMC9330571 DOI: 10.3390/membranes12080731] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/29/2022] [Revised: 07/18/2022] [Accepted: 07/21/2022] [Indexed: 11/16/2022]
Abstract
The epithelial barrier is crucial for proper gastrointestinal function, preventing the unwanted passage of solutes and therefore representing a prerequisite for vectorial transport. Claudin-4 and claudin-18.2, two critical tight junction proteins of the gastric epithelium, seal neighboring cells in a physically and mechanically challenging environment. As the Xenopus laevis oocyte allows the functional and molecular analyses of claudin interaction, we have addressed the hypothesis that this interaction is not only dependent on mechanical force but also on pH. We expressed human claudin-4 and claudin-18 in Xenopus oocytes, and analyzed them in a two-cell model approach. Cells were clustered in pairs to form contact areas expressing CLDN18 + CLDN18, CLDN4/18 + CLDN4/18, and compared to controls, respectively. Contact areas in cells incubated in medium at pH 5.5 and 7.4 were quantified by employing transmitted light microscopy. After 24 h at pH 5.5, clustering of CLDN18 + CLDN18 and CLDN4/18 + CLDN4/18-expressing oocytes revealed a contact area reduced by 45% and 32%, compared with controls, respectively. A further approach, high-pressure impulse assay, revealed a stronger tight junction interaction at pH 5.5 in oocyte pairs expressing CLDN18 + CLDN18 or CLDN4/18 + CLDN4/18 indicating a protective role of claudin-18 for tight junction integrity during pH challenge. Thus, our current analysis of gastric tight junction proteins further establishes oocytes as an expression and two-cell screening model for tight junction integrity analysis of organ- and tissue-specific claudins by the characterization of homo- and heterophilic trans-interaction dependent on barrier effectors.
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Affiliation(s)
| | | | - Salah Amasheh
- Correspondence: ; Tel.: +49-30-838-62602; Fax: +49-30-838-462602
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23
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Li Y, Wang C, Zhang L, Chen B, Mo Y, Zhang J. Claudin-5a is essential for the functional formation of both zebrafish blood-brain barrier and blood-cerebrospinal fluid barrier. Fluids Barriers CNS 2022; 19:40. [PMID: 35658877 PMCID: PMC9164509 DOI: 10.1186/s12987-022-00337-9] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/21/2022] [Accepted: 05/04/2022] [Indexed: 11/25/2022] Open
Abstract
Background Mammalian Claudin-5 is the main endothelial tight junction component maintaining blood-brain barrier (BBB) permeability, while Claudin-1 and -3 seal the paracellular space of choroid plexus (CP) epithelial cells contributing to the blood-cerebrospinal fluid barrier (BCSFB). In zebrafish, two paralogs of claudin-5a and -5b are expressed while their roles in the formation of BBB and BCSFB are unclear. Methods The expression patterns of Claudin-5a and -5b in zebrafish brains were systematically analyzed by immunofluorescence (IF) assay. The developmental functions of Claudin-5a and -5b were characterized by generating of claudin-5a and -5b mutants respectively. Meanwhile, the cerebral inflammation and cell apoptosis in claudin-5a-/- were assessed by live imaging of transgenic zebrafish, RT-qPCR, IF, and TUNEL assay. The integrity of BBB and BCSFB was evaluated by in vivo angiographic and dye permeation assay. Finally, RT-qPCR, whole-mount RNA in situ hybridization (WISH), and transmission electron microscopy (TEM) analyses were performed to investigate the development of cerebral vessels and choroid plexus. Results We showed that Claudin-5a and -5b are both expressed in zebrafish cerebrovascular endothelial cells (ECs). In addition, Claudin-5a was strongly expressed in CP epithelial cells. Loss of Claudin-5b showed no effect on zebrafish vasculogenesis or BBB function. In contrast, the knockout of claudin-5a caused a lethal phenotype of severe whole-brain oedema, ventricular dilatation, and cerebral hernia in zebrafish larvae, although the cerebral vasculogenesis and the development of CP were not altered. In claudin-5a-/- , although ultrastructural analysis of CP and cerebral capillary showed intact integrity of epithelial and endothelial tight junctions, permeability assay indicated a disruption of both BBB and BCSFB functions. On the molecular level, it was found that ZO-1 was upregulated in the CP epithelium of claudin-5a-/-, while the notch and shh pathway responsible for CP development was not affected due to loss of Claudin-5a. Conclusions Our findings verified a non-functional role of zebrafish Claudin-5b in the BBB and identified Claudin-5a as the ortholog of mammalian Claudin-5, contributing to the development and the functional maintenance of both BBB and BCSFB. Supplementary Information The online version contains supplementary material available at 10.1186/s12987-022-00337-9.
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Affiliation(s)
- Yanyu Li
- Affiliated Hospital of Guangdong Medical University & Key Laboratory of Zebrafish Model for Development and Disease of Guangdong Medical University, Zhanjiang, 524001, China
| | - Chunchun Wang
- Affiliated Hospital of Guangdong Medical University & Key Laboratory of Zebrafish Model for Development and Disease of Guangdong Medical University, Zhanjiang, 524001, China
| | - Liang Zhang
- College of Life Sciences, Shandong Normal University, Jinan, 250014, China
| | - Bing Chen
- Affiliated Hospital of Guangdong Medical University & Key Laboratory of Zebrafish Model for Development and Disease of Guangdong Medical University, Zhanjiang, 524001, China
| | - Yuqian Mo
- Affiliated Hospital of Guangdong Medical University & Key Laboratory of Zebrafish Model for Development and Disease of Guangdong Medical University, Zhanjiang, 524001, China.,School of Public Health, Guangdong Medical University, Dongguan, 523808, China
| | - Jingjing Zhang
- Affiliated Hospital of Guangdong Medical University & Key Laboratory of Zebrafish Model for Development and Disease of Guangdong Medical University, Zhanjiang, 524001, China. .,The Marine Biomedical Research Institute of Guangdong Zhanjiang, Zhanjiang, 524023, China.
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24
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Egorova AV, Baranich TI, Brydun AV, Glinkina VV, Sukhorukov VS. Morphological and Histophysiological Features of the Brain Capillary Endothelium. J EVOL BIOCHEM PHYS+ 2022. [DOI: 10.1134/s0022093022030115] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
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25
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Barbara G, Barbaro MR, Fuschi D, Palombo M, Falangone F, Cremon C, Marasco G, Stanghellini V. Corrigendum: Inflammatory and Microbiota-Related Regulation of the Intestinal Epithelial Barrier. Front Nutr 2021; 8:790387. [PMID: 34790692 PMCID: PMC8591313 DOI: 10.3389/fnut.2021.790387] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/06/2021] [Accepted: 10/07/2021] [Indexed: 12/18/2022] Open
Affiliation(s)
- Giovanni Barbara
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Maria Raffaella Barbaro
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Daniele Fuschi
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Marta Palombo
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Francesca Falangone
- Medical-Surgical Department of Clinical Sciences and Translational Medicine, University Sapienza, Rome, Italy
| | - Cesare Cremon
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Giovanni Marasco
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Vincenzo Stanghellini
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
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26
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Barbara G, Barbaro MR, Fuschi D, Palombo M, Falangone F, Cremon C, Marasco G, Stanghellini V. Inflammatory and Microbiota-Related Regulation of the Intestinal Epithelial Barrier. Front Nutr 2021; 8:718356. [PMID: 34589512 PMCID: PMC8475765 DOI: 10.3389/fnut.2021.718356] [Citation(s) in RCA: 163] [Impact Index Per Article: 40.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2021] [Accepted: 08/12/2021] [Indexed: 12/19/2022] Open
Abstract
The intestinal epithelial barrier (IEB) is one of the largest interfaces between the environment and the internal milieu of the body. It is essential to limit the passage of harmful antigens and microorganisms and, on the other side, to assure the absorption of nutrients and water. The maintenance of this delicate equilibrium is tightly regulated as it is essential for human homeostasis. Luminal solutes and ions can pass across the IEB via two main routes: the transcellular pathway or the paracellular pathway. Tight junctions (TJs) are a multi-protein complex responsible for the regulation of paracellular permeability. TJs control the passage of antigens through the IEB and have a key role in maintaining barrier integrity. Several factors, including cytokines, gut microbiota, and dietary components are known to regulate intestinal TJs. Gut microbiota participates in several human functions including the modulation of epithelial cells and immune system through the release of several metabolites, such as short-chain fatty acids (SCFAs). Mediators released by immune cells can induce epithelial cell damage and TJs dysfunction. The subsequent disruption of the IEB allows the passage of antigens into the mucosa leading to further inflammation. Growing evidence indicates that dysbiosis, immune activation, and IEB dysfunction have a role in several diseases, including irritable bowel syndrome (IBS), inflammatory bowel disease (IBD), and gluten-related conditions. Here we summarize the interplay between the IEB and gut microbiota and mucosal immune system and their involvement in IBS, IBD, and gluten-related disorders.
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Affiliation(s)
- Giovanni Barbara
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Maria Raffaella Barbaro
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Daniele Fuschi
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Marta Palombo
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Francesca Falangone
- Medical-Surgical Department of Clinical Sciences and Translational Medicine, University Sapienza, Rome, Italy
| | - Cesare Cremon
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Giovanni Marasco
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
| | - Vincenzo Stanghellini
- IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, Italy.,Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
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27
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Endo S, Nishiyama T, Matuoka T, Miura T, Nishinaka T, Matsunaga T, Ikari A. Loxoprofen enhances intestinal barrier function via generation of its active metabolite by carbonyl reductase 1 in differentiated Caco-2 cells. Chem Biol Interact 2021; 348:109634. [PMID: 34506768 DOI: 10.1016/j.cbi.2021.109634] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2021] [Revised: 08/31/2021] [Accepted: 09/05/2021] [Indexed: 02/06/2023]
Abstract
Nonsteroidal anti-inflammatory drugs (NSAIDs) are used worldwide as antipyretic analgesics and agents for rheumatoid arthritis and osteoarthritis, but known to cause damage to the gastrointestinal mucosae as their serious adverse effects. Few studies showed the impairment of intestinal epithelial barrier function (EBF) by high concentrations (0.5-1 mM) of NSAIDs, but the underlying mechanism is not fully understood. This study is aimed at clarifying effects at a low concentration (50 μM) of three NSAIDs, loxoprofen (Lox), ibuprofen and indomethacin, on intestinal EBF using human intestinal epithelial-like Caco-2 cells. Among those NSAIDs, Lox increased the transepithelial electric resistance (TER) value, decreased the paracellular Lucifer yellow CH (LYCH) permeability, and upregulated claudin (CLDN)-1, -3 and -5, indicating that low doses of Lox enhanced EBF through increasing expression of CLDNs. Lox is known to be metabolized to a pharmacologically active metabolite, (2S,1'R,2'S)-loxoprofen alcohol (Lox-RS), by carbonyl reductase 1 (CBR1), which is highly expressed in human intestine. CBR1 was expressed in the Caco-2 cells, and the pretreatment with a CBR1 inhibitor suppressed both the Lox-evoked CLDN upregulation and EBF enhancement. In addition, the treatment of the cells with Lox-RS resulted in higher TER value and lower LYCH permeability than those with Lox. Thus, Lox-RS synthesized by CBR1 may greatly contribute to the improving efficacy of Lox on the barrier function. Since EBF is decreased in inflammatory bowel disease, we finally examined the effect of Lox on EBF using the Caco-2/THP-1 co-culture system, which is used as an in vitro inflammatory bowel disease model. Lox significantly recovered EBF which was impaired by inflammatory cytokines secreted from THP-1 macrophages. These in vitro observations suggest that Lox enhances intestinal EBF, for which the metabolism of Lox to Lox-RS by CBR1 has an important role.
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Affiliation(s)
- Satoshi Endo
- Laboratory of Biochemistry, Department of Biopharmaceutical Sciences, Gifu Pharmaceutical University, 501-1196, Gifu, Japan
| | - Tsubasa Nishiyama
- Laboratory of Biochemistry, Department of Biopharmaceutical Sciences, Gifu Pharmaceutical University, 501-1196, Gifu, Japan
| | - Tomoe Matuoka
- Laboratory of Biochemistry, Department of Biopharmaceutical Sciences, Gifu Pharmaceutical University, 501-1196, Gifu, Japan
| | - Takeshi Miura
- Pharmaceutical Education Support Center, School of Pharmacy and Pharmaceutical Sciences, Mukogawa Women's University, Nishinomiya, 663-8184, Japan
| | - Toru Nishinaka
- Laboratory of Biochemistry, Faculty of Pharmacy, Osaka Ohtani University, Tondabayashi, 584-8540, Japan
| | - Toshiyuki Matsunaga
- Education Center of Green Pharmaceutical Sciences, Gifu Pharmaceutical University, Gifu, 502-8585, Japan
| | - Akira Ikari
- Laboratory of Biochemistry, Department of Biopharmaceutical Sciences, Gifu Pharmaceutical University, 501-1196, Gifu, Japan.
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28
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Segers C, Mysara M, Claesen J, Baatout S, Leys N, Lebeer S, Verslegers M, Mastroleo F. Intestinal mucositis precedes dysbiosis in a mouse model for pelvic irradiation. ISME COMMUNICATIONS 2021; 1:24. [PMID: 36737646 PMCID: PMC9723693 DOI: 10.1038/s43705-021-00024-0] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 02/25/2021] [Revised: 05/12/2021] [Accepted: 05/25/2021] [Indexed: 02/07/2023]
Abstract
Pelvic radiotherapy is known to evoke intestinal mucositis and dysbiosis. Currently, there are no effective therapies available to mitigate these injuries, which is partly due to a lack of insight into the events causing mucositis and dysbiosis. Here, the complex interplay between the murine host and its microbiome following pelvic irradiation was mapped by characterizing intestinal mucositis along with extensive 16S microbial profiling. We demonstrated important morphological and inflammatory implications within one day after exposure, thereby impairing intestinal functionality and inducing translocation of intraluminal bacteria into mesenteric lymph nodes as innovatively quantified by flow cytometry. Concurrent 16S microbial profiling revealed a delayed impact of pelvic irradiation on beta diversity. Analysis of composition of microbiomes identified biomarkers for pelvic irradiation. Among them, members of the families Ruminococcaceae, Lachnospiraceae and Porphyromonadaceae were differentially affected. Altogether, our unprecedented findings showed how pelvic irradiation evoked structural and functional changes in the intestine, which secondarily resulted in a microbiome shift. Therefore, the presented in vivo irradiation-gut-microbiome platform allows further research into the pathobiology of pelvic irradiation-induced intestinal mucositis and resultant dysbiosis, as well as the exploration of mitigating treatments including drugs and food supplements.
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Affiliation(s)
- Charlotte Segers
- Interdisciplinary Biosciences group, Belgian Nuclear Research Centre SCK CEN, Mol, Belgium
- Department of Bioscience Engineering, University of Antwerp, Antwerp, Belgium
| | - Mohamed Mysara
- Interdisciplinary Biosciences group, Belgian Nuclear Research Centre SCK CEN, Mol, Belgium
| | - Jürgen Claesen
- Interdisciplinary Biosciences group, Belgian Nuclear Research Centre SCK CEN, Mol, Belgium
- Department of Epidemiology and Data Science, Amsterdam UMC, VU University Amsterdam, Amsterdam, The Netherlands
| | - Sarah Baatout
- Interdisciplinary Biosciences group, Belgian Nuclear Research Centre SCK CEN, Mol, Belgium
- Department of Biotechnology, University of Ghent, Ghent, Belgium
| | - Natalie Leys
- Interdisciplinary Biosciences group, Belgian Nuclear Research Centre SCK CEN, Mol, Belgium
| | - Sarah Lebeer
- Department of Bioscience Engineering, University of Antwerp, Antwerp, Belgium
| | - Mieke Verslegers
- Interdisciplinary Biosciences group, Belgian Nuclear Research Centre SCK CEN, Mol, Belgium
| | - Felice Mastroleo
- Interdisciplinary Biosciences group, Belgian Nuclear Research Centre SCK CEN, Mol, Belgium.
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29
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Petito V, Greco V, Laterza L, Graziani C, Fanali C, Lucchetti D, Barbaro MR, Bugli F, Pieroni L, Lopetuso LR, Sgambato A, Sanguinetti M, Scaldaferri F, Urbani A, Gasbarrini A. Impact of the Trophic Effects of the Secretome From a Multistrain Probiotic Preparation on the Intestinal Epithelia. Inflamm Bowel Dis 2021; 27:902-913. [PMID: 33300553 DOI: 10.1093/ibd/izaa298] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/03/2019] [Indexed: 12/13/2022]
Abstract
BACKGROUND Probiotics are defined as live, nonpathogenic bacteria that confer health benefits beyond their nutritional value. In particular, VSL#3 exhibits demonstrated efficacy in the management of diseases characterized by an increased intestinal permeability. Our study aimed to understand how VSL#3 promotes gut health by secreting bioactive factors and identify which human pathways are modulated by secretome derived from the VSL#3 formula. METHODS Two different lots of VSL#3 were used, and Caco-2 cell line was treated with conditioned media (CM) prepared using 1 g of the probiotic formula. We evaluated the effects of the probiotics on cellular proliferation and apoptosis by cytometry and the expression of tight junction proteins by western blotting. A proteomics analysis of both culture media and the whole proteome of Caco-2 cells treated with VSL#3-CM was performed by nano-ultra performance liquid chromatography - tandem mass (nUPLC MS/MS) spectrometry. RESULTS The probiotic formula increased cell proliferation, decreased cellular apoptosis cells, and increased re-epithelialization in the scratch assay. Several peptides specifically synthetized by all the species within the probiotic preparation were recognized in the proteomics analysis. Human proteins synthesized by CaCo-2 cells were also identified. CONCLUSIONS To our knowledge, this manuscript describes the first evaluation of the probiotic secretome, and the results showed that the improvement in intestinal barrier functions induced by probiotics seems to be accompanied by the modulation of some human cellular pathways.
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Affiliation(s)
- Valentina Petito
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy
| | - Viviana Greco
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy.,Università Cattolica del Sacro Cuore, Dipartimento Universitario di Scienze Biotecnologiche di Base, Cliniche Intensivologiche e Perioperatorie, Rome, Italy
| | - Lucrezia Laterza
- Fondazione Policlinico Universitario "A. Gemelli" IRCCS, Dipartimento di Scienze di Laboratorio e Infettivologiche, Rome, Italy
| | - Cristina Graziani
- Fondazione Policlinico Universitario "A. Gemelli" IRCCS, Dipartimento di Scienze di Laboratorio e Infettivologiche, Rome, Italy
| | - Caterina Fanali
- Fondazione Policlinico Universitario "A. Gemelli" IRCCS, Dipartimento di Scienze di Laboratorio e Infettivologiche, Rome, Italy
| | - Donatella Lucchetti
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy
| | - Maria Raffaella Barbaro
- Fondazione Policlinico Universitario "A. Gemelli" IRCCS, Dipartimento di Scienze Mediche e Chirurgiche, Rome, Italy
| | - Francesca Bugli
- Policlinico Sant'Orsola- Malpighi, Università di Bologna, Dipartimento di Scienze Mediche e Chirurgiche, Bologna, Italia
| | - Luisa Pieroni
- Fondazione Santa Lucia IRCCS, Unitá di Proteomica e Metabolomica, Rome, Italy
| | - Loris Riccardo Lopetuso
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy.,Fondazione Policlinico Universitario "A. Gemelli" IRCCS, Dipartimento di Scienze di Laboratorio e Infettivologiche, Rome, Italy
| | - Alessandro Sgambato
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy
| | - Maurizio Sanguinetti
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy.,Università Cattolica del Sacro Cuore, Dipartimento Universitario di Scienze Biotecnologiche di Base, Cliniche Intensivologiche e Perioperatorie, Rome, Italy
| | - Franco Scaldaferri
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy.,Fondazione Policlinico Universitario "A. Gemelli" IRCCS, Dipartimento di Scienze di Laboratorio e Infettivologiche, Rome, Italy
| | - Andrea Urbani
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy.,Università Cattolica del Sacro Cuore, Dipartimento Universitario di Scienze Biotecnologiche di Base, Cliniche Intensivologiche e Perioperatorie, Rome, Italy
| | - Antonio Gasbarrini
- Università Cattolica del Sacro Cuore, Dipartimento Universitario di Medicina e Chirurgia Traslazionale, Rome, Italy.,Fondazione Policlinico Universitario "A. Gemelli" IRCCS, Dipartimento di Scienze di Laboratorio e Infettivologiche, Rome, Italy
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30
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Luo T, Liu H, Chen B, Liu H, Abdel-Latif A, Kitakaze M, Wang X, Wu Y, Chou D, Kim JK. A novel role of claudin-5 in prevention of mitochondrial fission against ischemic/hypoxic stress in cardiomyocytes. Can J Cardiol 2021; 37:1593-1606. [PMID: 33838228 DOI: 10.1016/j.cjca.2021.03.021] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2019] [Revised: 03/20/2021] [Accepted: 03/29/2021] [Indexed: 11/27/2022] Open
Abstract
BACKGROUND Downregulation of claudin-5 in the heart is associated with the end-stage heart failure. However, the underlying mechanism of claudin-5 is unclear. Here we investigated the molecular actions of claudin-5 in perspective of mitochondria in cardiomyocytes to better understand the role of claudin-5 in cardioprotection during ischemia. METHODS AND RESULTS Claudin-5 was detected in the murine heart tissue and the neonatal rat cardiomyocytes (NRCM). Its protein level was severely decreased after myocardial ischemia/reperfusion (I/R; 30 min/24 h) or hypoxia/reoxygenation (H/R; 24 h/4 h). Claudin-5 was present in the mitochondria of NRCM as determined by confocal microscopy. H/R-induced downregulation of claudin-5 was accompanied by mitochondrial fragmentation. The protein level of mitofusin 2 (Mfn2) was dramatically decreased while the expression of dynamin-related protein (Drp) 1 was significantly increased after H/R. H/R-induced mitochondrial swelling and fission were observed by transmission electron microscope (TEM). Overexpression of claudin-5 by adenoviral infection reversed these structural disintegration of mitochondria. The mitochondria-centered intrinsic pathway of apoptosis triggered by H/R and indicated by the expression of cytochrome c and cleaved caspase 3 in the cytoplasm of NRCMs was also reduced by overexpressing claudin-5. Overexpression of claudin-5 in mouse heart also significantly decreased cleaved caspase 3 expression and the infarct size in ischemic heart with improved systolic function. CONCLUSION We demonstrated for the first time the presence of claudin-5 in the mitochondria in cardiomyocytes and provided the firm evidence for the cardioprotective role of claudin-5 in the preservation of mitochondrial dynamics and cell fate against hypoxia- or ischemia-induced stress.
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Affiliation(s)
- Tao Luo
- Department of Pathophysiology, Zhuhai Campus of Zunyi Medical University, Zhuhai, 519041, China; Division of Cardiology, Department of Medicine, School of Medicine, University of California Irvine, Irvine, CA 92697, USA.
| | - Haiqiong Liu
- Department of Cardiology, Zhujiang Hospital, Southern Medical University, Guangzhou, Guangdong 510282, China
| | - Baihe Chen
- Department of Pathophysiology, Zhuhai Campus of Zunyi Medical University, Zhuhai, 519041, China; Division of Cardiology, Department of Medicine, School of Medicine, University of California Irvine, Irvine, CA 92697, USA
| | - Han Liu
- Division of Cardiology, Department of Medicine, School of Medicine, University of California Irvine, Irvine, CA 92697, USA
| | - Ahmed Abdel-Latif
- Saha Cardiovascular Research Center, University of Kentucky, Lexington, 40536-0509, USA
| | - Masafumi Kitakaze
- Department of Clinical Research and Development, National Cerebral and Cardiovascular Center, 5-7-1 Fujishirodai, Suita, 5675-8565, Japan
| | - Xianbao Wang
- Department of Cardiology, Zhujiang Hospital, Southern Medical University, Guangzhou, Guangdong 510282, China
| | - Yuanzhou Wu
- Department of Cardiovascular Surgery, Zhujiang Hospital, Southern Medical University, Guangzhou, Guangdong, 510282, China
| | - Dylan Chou
- Department of Physiology, Zhuhai Campus of Zunyi Medical University, Zhuhai, 519041, China
| | - Jin Kyung Kim
- Division of Cardiology, Department of Medicine, School of Medicine, University of California Irvine, Irvine, CA 92697, USA.
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Tight Junction Modulating Bioprobes for Drug Delivery System to the Brain: A Review. Pharmaceutics 2020; 12:pharmaceutics12121236. [PMID: 33352631 PMCID: PMC7767277 DOI: 10.3390/pharmaceutics12121236] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2020] [Revised: 12/16/2020] [Accepted: 12/17/2020] [Indexed: 12/19/2022] Open
Abstract
The blood-brain barrier (BBB), which is composed of endothelial cells, pericytes, astrocytes, and neurons, separates the brain extracellular fluid from the circulating blood, and maintains the homeostasis of the central nervous system (CNS). The BBB endothelial cells have well-developed tight junctions (TJs) and express specific polarized transport systems to tightly control the paracellular movements of solutes, ions, and water. There are two types of TJs: bicellular TJs (bTJs), which is a structure at the contact of two cells, and tricellular TJs (tTJs), which is a structure at the contact of three cells. Claudin-5 and angulin-1 are important components of bTJs and tTJs in the brain, respectively. Here, we review TJ-modulating bioprobes that enable drug delivery to the brain across the BBB, focusing on claudin-5 and angulin-1.
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32
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Profaci CP, Munji RN, Pulido RS, Daneman R. The blood-brain barrier in health and disease: Important unanswered questions. J Exp Med 2020; 217:151582. [PMID: 32211826 PMCID: PMC7144528 DOI: 10.1084/jem.20190062] [Citation(s) in RCA: 426] [Impact Index Per Article: 85.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2019] [Revised: 09/21/2019] [Accepted: 11/21/2019] [Indexed: 12/11/2022] Open
Abstract
The blood vessels vascularizing the central nervous system exhibit a series of distinct properties that tightly control the movement of ions, molecules, and cells between the blood and the parenchyma. This "blood-brain barrier" is initiated during angiogenesis via signals from the surrounding neural environment, and its integrity remains vital for homeostasis and neural protection throughout life. Blood-brain barrier dysfunction contributes to pathology in a range of neurological conditions including multiple sclerosis, stroke, and epilepsy, and has also been implicated in neurodegenerative diseases such as Alzheimer's disease. This review will discuss current knowledge and key unanswered questions regarding the blood-brain barrier in health and disease.
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Affiliation(s)
- Caterina P Profaci
- Department of Neurosciences, University of California, San Diego, San Diego, CA.,Department of Pharmacology, University of California, San Diego, San Diego, CA
| | - Roeben N Munji
- Department of Neurosciences, University of California, San Diego, San Diego, CA.,Department of Pharmacology, University of California, San Diego, San Diego, CA
| | - Robert S Pulido
- Department of Neurosciences, University of California, San Diego, San Diego, CA.,Department of Pharmacology, University of California, San Diego, San Diego, CA
| | - Richard Daneman
- Department of Neurosciences, University of California, San Diego, San Diego, CA.,Department of Pharmacology, University of California, San Diego, San Diego, CA
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33
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Gericke B, Römermann K, Noack A, Noack S, Kronenberg J, Blasig IE, Löscher W. A face-to-face comparison of claudin-5 transduced human brain endothelial (hCMEC/D3) cells with porcine brain endothelial cells as blood-brain barrier models for drug transport studies. Fluids Barriers CNS 2020; 17:53. [PMID: 32843059 PMCID: PMC7449095 DOI: 10.1186/s12987-020-00212-5] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2020] [Accepted: 07/13/2020] [Indexed: 12/25/2022] Open
Abstract
BACKGROUND Predictive in vitro models of the human blood-brain barrier (BBB) are essential in early drug discovery and development. Among available immortalized human brain capillary endothelial cell lines (BCECs), the hCMEC/D3 cell line has become the most widely used in vitro BBB model. However, monolayers of hCMEC/D3 cells form only moderately restrictive barriers, most likely because the major tight junction protein, claudin-5, is markedly downregulated. Thus, hCMEC/D3 monolayers cannot be used for vectorial drug transport experiments, which is a major disadvantage of this model. METHODS Here we transduced hCMEC/D3 cells with a claudin-5 plasmid and compared the characteristics of these cells with those of hCMEC/D3 wildtype cells and primary cultured porcine BCECs. RESULTS The claudin-5 transduced hCMEC/D3 exhibited expression levels (and junctional localization) of claudin-5 similar to those of primary cultured porcine BCECs. The transduced cells exhibited increased TEER values (211 Ω cm2) and reduced paracellular mannitol permeability (8.06%/h), indicating improved BBB properties; however, the barrier properties of porcine BCECs (TEER 1650 Ω cm2; mannitol permeability 3.95%/h) were not reached. Hence, vectorial transport of a selective P-glycoprotein substrate (N-desmethyl-loperamide) was not observed in claudin-5 transduced hCMEC/D3 (or wildtype) cells, whereas such drug transport occurred in porcine BCECs. CONCLUSIONS The claudin-5 transduced hCMEC/D3 cells provide a tool to studying the contribution of claudin-5 to barrier tightness and how this can be further enhanced by additional transfections or other manipulations of this widely used in vitro model of the BBB.
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Affiliation(s)
- Birthe Gericke
- Department of Pharmacology, Toxicology, and Pharmacy, University of Veterinary Medicine Hannover, Hannover, Germany
| | - Kerstin Römermann
- Department of Pharmacology, Toxicology, and Pharmacy, University of Veterinary Medicine Hannover, Hannover, Germany
| | - Andreas Noack
- Department of Pharmacology, Toxicology, and Pharmacy, University of Veterinary Medicine Hannover, Hannover, Germany
| | - Sandra Noack
- Department of Trauma Surgery, Hannover Medical School, Hannover, Germany
| | - Jessica Kronenberg
- Department of Pharmacology, Toxicology, and Pharmacy, University of Veterinary Medicine Hannover, Hannover, Germany
| | | | - Wolfgang Löscher
- Department of Pharmacology, Toxicology, and Pharmacy, University of Veterinary Medicine Hannover, Hannover, Germany. .,Center for Systems Neuroscience, Hannover, Germany.
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34
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Liu M, Xie W, Wan X, Deng T. Clostridium butyricum protects intestinal barrier function via upregulation of tight junction proteins and activation of the Akt/mTOR signaling pathway in a mouse model of dextran sodium sulfate-induced colitis. Exp Ther Med 2020; 20:10. [PMID: 32934675 PMCID: PMC7471846 DOI: 10.3892/etm.2020.9138] [Citation(s) in RCA: 28] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2020] [Accepted: 07/01/2020] [Indexed: 12/14/2022] Open
Abstract
Clostridium butyricum (CB), a probiotic, is a gram-positive obligate anaerobic bacillus with acid and heat resistant properties. Previous studies have reported that CB has beneficial effects in intestinal diseases and regulates intestinal function. The aim of the present study was to investigate the protective effects and mechanisms of CB on the intestinal barrier function. Mice were randomly divided into three experimental groups (n=15 mice/group), including control, dextran sodium sulfate (DSS) and DSS + CB. In the DSS and DSS + CB groups colitis was induced with 3% DSS dissolved in drinking water for 7 days. DSS + CB group mice were co-treated daily with 200 µl (2x108 CFU) CB solution via gavage. The intestinal mucosal barrier function in mice was assessed by measuring FITC-labeled 4-kDa dextran (molecular weight, 4,000 Da) flux and by analyzing the expression of tight junction (TJ)-related proteins using western blot analysis. In addition, the secretion levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, IL-10 and IL-13, and the concentration of malondialdehyde, glutathione and superoxide dismutase were detected using ELISAs to determine inflammation and oxidative stress, respectively. The activation status of the Akt/mTOR signaling pathway was also investigated using western blot analysis. The results demonstrated that, in mice with DSS-induced colitis treatment, co-treatment with CB attenuated colitis symptoms and intestinal permeability, increased the expression levels of TJ-related proteins, decreased TNF-α, IL-1β and IL-13 secretion levels but increased those of IL-10, and reduced oxidative stress. Additionally, CB elevated the phosphorylation of Akt, mTOR and p70 ribosomal protein S6 kinase. Collectively, the present results indicated that CB protected intestinal barrier function and decreased intestinal mucosal permeability via upregulating the expression levels of TJ-related proteins in a mouse model of DSS-induced colitis. Moreover, the results suggested that the effects of CB could be mediated by the Akt/mTOR signaling pathway.
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Affiliation(s)
- Miao Liu
- Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, P.R. China.,Key Laboratory of Hubei Province for Digestive System Disease, Wuhan, Hubei 430060, P.R. China.,Central Laboratory of Renmin Hospital, Wuhan, Hubei 430060, P.R. China
| | - Wenjie Xie
- Central Laboratory of Renmin Hospital, Wuhan, Hubei 430060, P.R. China.,Department of Critical Care Medicine, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, P.R. China
| | - Xinyue Wan
- Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, P.R. China.,Key Laboratory of Hubei Province for Digestive System Disease, Wuhan, Hubei 430060, P.R. China.,Central Laboratory of Renmin Hospital, Wuhan, Hubei 430060, P.R. China
| | - Tao Deng
- Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, P.R. China.,Key Laboratory of Hubei Province for Digestive System Disease, Wuhan, Hubei 430060, P.R. China.,Central Laboratory of Renmin Hospital, Wuhan, Hubei 430060, P.R. China
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35
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Alanyl-Glutamine Restores Tight Junction Organization after Disruption by a Conventional Peritoneal Dialysis Fluid. Biomolecules 2020; 10:biom10081178. [PMID: 32823646 PMCID: PMC7464725 DOI: 10.3390/biom10081178] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2020] [Revised: 08/07/2020] [Accepted: 08/11/2020] [Indexed: 12/13/2022] Open
Abstract
Understanding and targeting the molecular basis of peritoneal solute and protein transport is essential to improve peritoneal dialysis (PD) efficacy and patient outcome. Supplementation of PD fluids (PDF) with alanyl-glutamine (AlaGln) increased small solute transport and reduced peritoneal protein loss in a recent clinical trial. Transepithelial resistance and 10 kDa and 70 kDa dextran transport were measured in primary human endothelial cells (HUVEC) exposed to conventional acidic, glucose degradation products (GDP) containing PDF (CPDF) and to low GDP containing PDF (LPDF) with and without AlaGln. Zonula occludens-1 (ZO-1) and claudin-5 were quantified by Western blot and immunofluorescence and in mice exposed to saline and CPDF for 7 weeks by digital imaging analyses. Spatial clustering of ZO-1 molecules was assessed by single molecule localization microscopy. AlaGln increased transepithelial resistance, and in CPDF exposed HUVEC decreased dextran transport rates and preserved claudin-5 and ZO-1 abundance. Endothelial clustering of membrane bound ZO-1 was higher in CPDF supplemented with AlaGln. In mice, arteriolar endothelial claudin-5 was reduced in CPDF, but restored with AlaGln, while mesothelial claudin-5 abundance was unchanged. AlaGln supplementation seals the peritoneal endothelial barrier, and when supplemented to conventional PD fluid increases claudin-5 and ZO-1 abundance and clustering of ZO-1 in the endothelial cell membrane.
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36
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Lochhead JJ, Yang J, Ronaldson PT, Davis TP. Structure, Function, and Regulation of the Blood-Brain Barrier Tight Junction in Central Nervous System Disorders. Front Physiol 2020; 11:914. [PMID: 32848858 PMCID: PMC7424030 DOI: 10.3389/fphys.2020.00914] [Citation(s) in RCA: 227] [Impact Index Per Article: 45.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2020] [Accepted: 07/08/2020] [Indexed: 12/16/2022] Open
Abstract
The blood-brain barrier (BBB) allows the brain to selectively import nutrients and energy critical to neuronal function while simultaneously excluding neurotoxic substances from the peripheral circulation. In contrast to the highly permeable vasculature present in most organs that reside outside of the central nervous system (CNS), the BBB exhibits a high transendothelial electrical resistance (TEER) along with a low rate of transcytosis and greatly restricted paracellular permeability. The property of low paracellular permeability is controlled by tight junction (TJ) protein complexes that seal the paracellular route between apposing brain microvascular endothelial cells. Although tight junction protein complexes are principal contributors to physical barrier properties, they are not static in nature. Rather, tight junction protein complexes are highly dynamic structures, where expression and/or localization of individual constituent proteins can be modified in response to pathophysiological stressors. These stressors induce modifications to tight junction protein complexes that involve de novo synthesis of new protein or discrete trafficking mechanisms. Such responsiveness of BBB tight junctions to diseases indicates that these protein complexes are critical for maintenance of CNS homeostasis. In fulfillment of this vital role, BBB tight junctions are also a major obstacle to therapeutic drug delivery to the brain. There is an opportunity to overcome this substantial obstacle and optimize neuropharmacology via acquisition of a detailed understanding of BBB tight junction structure, function, and regulation. In this review, we discuss physiological characteristics of tight junction protein complexes and how these properties regulate delivery of therapeutics to the CNS for treatment of neurological diseases. Specifically, we will discuss modulation of tight junction structure, function, and regulation both in the context of disease states and in the setting of pharmacotherapy. In particular, we will highlight how these properties can be potentially manipulated at the molecular level to increase CNS drug levels via paracellular transport to the brain.
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37
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Brunner N, Stein L, Cornelius V, Knittel R, Fallier-Becker P, Amasheh S. Blood-Brain Barrier Protein Claudin-5 Expressed in Paired Xenopus laevis Oocytes Mediates Cell-Cell Interaction. Front Physiol 2020; 11:857. [PMID: 32848831 PMCID: PMC7396581 DOI: 10.3389/fphys.2020.00857] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2020] [Accepted: 06/26/2020] [Indexed: 01/08/2023] Open
Abstract
Claudin-5 determines the sealing properties of blood-brain barrier tight junctions and its function is impaired in neurodegenerative and neuroinflammatory disorders. Focusing on the contribution of claudin-5 to the trans-interaction within the tight junction seal, we used Xenopus laevis oocytes as an expression system. Cells were clustered and challenged in a novel approach for the analysis of claudin interaction. We evaluated the strengthening effect of claudin-5 to cell-cell-connection in comparison to claudin-3. Application of a hydrostatic pressure impulse on clustered control oocyte pairs revealed a reduction of contact areas. In contrast, combinations with both oocytes expressing claudins maintained an enhanced connection between the cells (cldn5-cldn5, cldn3-cldn3). Strength of interaction was increased by both claudin-3 and claudin-5. This novel approach allowed an analysis of single claudins contributing to tight junction integrity, characterizing homophilic and hetrophilic trans-interaction of claudins. To test a new screening approach for barrier effectors, exemplarily, this 2-cell model of oocytes was used to analyze the effect of the absorption enhancer sodium caprate on the oocyte pairs.
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Affiliation(s)
- Nora Brunner
- Department of Veterinary Medicine, Institute of Veterinary Physiology, Freie Universität Berlin, Berlin, Germany
| | - Laura Stein
- Department of Veterinary Medicine, Institute of Veterinary Physiology, Freie Universität Berlin, Berlin, Germany
| | - Valeria Cornelius
- Department of Veterinary Medicine, Institute of Veterinary Physiology, Freie Universität Berlin, Berlin, Germany
| | - Ria Knittel
- Institute of Pathology and Neuropathology, University Hospital of Tuebingen, Eberhard Karls University of Tuebingen, Tuebingen, Germany
| | - Petra Fallier-Becker
- Institute of Pathology and Neuropathology, University Hospital of Tuebingen, Eberhard Karls University of Tuebingen, Tuebingen, Germany
| | - Salah Amasheh
- Department of Veterinary Medicine, Institute of Veterinary Physiology, Freie Universität Berlin, Berlin, Germany
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38
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Areco VA, Kohan R, Talamoni G, Tolosa de Talamoni NG, Peralta López ME. Intestinal Ca 2+ absorption revisited: A molecular and clinical approach. World J Gastroenterol 2020; 26:3344-3364. [PMID: 32655262 PMCID: PMC7327788 DOI: 10.3748/wjg.v26.i24.3344] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/26/2020] [Revised: 05/11/2020] [Accepted: 06/10/2020] [Indexed: 02/06/2023] Open
Abstract
Ca2+ has an important role in the maintenance of the skeleton and is involved in the main physiological processes. Its homeostasis is controlled by the intestine, kidney, bone and parathyroid glands. The intestinal Ca2+ absorption occurs mainly via the paracellular and the transcellular pathways. The proteins involved in both ways are regulated by calcitriol and other hormones as well as dietary factors. Fibroblast growth factor 23 (FGF-23) is a strong antagonist of vitamin D action. Part of the intestinal Ca2+ movement seems to be vitamin D independent. Intestinal Ca2+ absorption changes according to different physiological conditions. It is promoted under high Ca2+ demands such as growth, pregnancy, lactation, dietary Ca2+ deficiency and high physical activity. In contrast, the intestinal Ca2+ transport decreases with aging. Oxidative stress inhibits the intestinal Ca2+ absorption whereas the antioxidants counteract the effects of prooxidants leading to the normalization of this physiological process. Several pathologies such as celiac disease, inflammatory bowel diseases, Turner syndrome and others occur with inhibition of intestinal Ca2+ absorption, some hypercalciurias show Ca2+ hyperabsorption, most of these alterations are related to the vitamin D endocrine system. Further research work should be accomplished in order not only to know more molecular details but also to detect possible therapeutic targets to ameliorate or avoid the consequences of altered intestinal Ca2+ absorption.
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Affiliation(s)
- Vanessa A Areco
- Laboratorio “Dr. Fernando Cañas”, Cátedra de Bioquímica y Biología Molecular, Facultad de Ciencias Médicas, INICSA (CONICET-Universidad Nacional de Córdoba), Córdoba 5000, Argentina
| | - Romina Kohan
- Laboratorio “Dr. Fernando Cañas”, Cátedra de Bioquímica y Biología Molecular, Facultad de Ciencias Médicas, INICSA (CONICET-Universidad Nacional de Córdoba), Córdoba 5000, Argentina
| | - Germán Talamoni
- Laboratorio “Dr. Fernando Cañas”, Cátedra de Bioquímica y Biología Molecular, Facultad de Ciencias Médicas, INICSA (CONICET-Universidad Nacional de Córdoba), Córdoba 5000, Argentina
| | - Nori G Tolosa de Talamoni
- Laboratorio “Dr. Fernando Cañas”, Cátedra de Bioquímica y Biología Molecular, Facultad de Ciencias Médicas, INICSA (CONICET-Universidad Nacional de Córdoba), Córdoba 5000, Argentina
| | - María E Peralta López
- Laboratorio “Dr. Fernando Cañas”, Cátedra de Bioquímica y Biología Molecular, Facultad de Ciencias Médicas, INICSA (CONICET-Universidad Nacional de Córdoba), Córdoba 5000, Argentina
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39
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Irudayanathan FJ, Nangia S. Paracellular Gatekeeping: What Does It Take for an Ion to Pass Through a Tight Junction Pore? LANGMUIR : THE ACS JOURNAL OF SURFACES AND COLLOIDS 2020; 36:6757-6764. [PMID: 32450698 DOI: 10.1021/acs.langmuir.0c00877] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/11/2023]
Abstract
Tight junction pores are physiological gatekeepers of paracellular transport in epithelial tissues. Conventionally, tight junction permeability is determined via in vitro electrophysiology measurements; however, the macroscopic readout does not provide molecular-level understanding into the mechanism of ion permeation. Insight into the factors governing selectivity across the paracellular space is just emerging. In this study, we investigated tight junction pores comprising of claudin-2 and claudin-5 proteins that are structurally similar to subnanometer radii but have measurably different in vitro ion permeabilities. To evaluate the mechanistic differences in ion transport across the pores, we computed the free-energy profiles and relative rate constants for the transport of monovalent (Na+, K+, Cl-) and divalent (Mg2+ and Ca2+) ions through the pores using replica exchange metadynamics. In claudin-2, we demonstrate how a single residue dictates selective permeability of Na+ and K+ ions. In claudin-5, we found no clear preference for anion or cation selectivity; thus, pores formed by claudin-5 are indeed barriers to ion permeation. Mutations to claudin-5 that widen the pore's steric radius did not significantly impact pore selectivity, indicating that electrostatics dominate pore selectivity. The key takeaways from this work are as follows: (a) two pores that are similar in diameter and length can have dissimilar ion conductance, (b) existence of a physical pore does not guarantee ion permeability, and (c) the electrostatic environment created by the pore-lining residues dictates the ion conductivity. These mechanistic understandings of the tight junction pores are critical for the interpretation of tight junction physiology.
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Affiliation(s)
| | - Shikha Nangia
- Department of Biomedical and Chemical Engineering, Syracuse University, Syracuse, New York 13244, United States
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40
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Twarog C, Liu K, O'Brien PJ, Dawson KA, Fattal E, Illel B, Brayden DJ. A head-to-head Caco-2 assay comparison of the mechanisms of action of the intestinal permeation enhancers: SNAC and sodium caprate (C 10). Eur J Pharm Biopharm 2020; 152:95-107. [PMID: 32387703 DOI: 10.1016/j.ejpb.2020.04.023] [Citation(s) in RCA: 46] [Impact Index Per Article: 9.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2019] [Revised: 04/15/2020] [Accepted: 04/28/2020] [Indexed: 02/07/2023]
Abstract
Salcaprozate sodium (SNAC) and sodium caprate (C10) are the two leading intestinal permeation enhancers (PEs) in oral peptide formulations in clinical trials. There is debate over their mechanism of action on intestinal epithelia. The aims were: (i) to compare their effects on the barrier function by measuring transepithelial electrical resistance (TEER), permeability of FITC-4000 (FD4) across Caco-2 monolayers, and on immunohistochemistry of tight junction (TJ)-associated proteins; and (ii) to compare cellular parameters using conventional end-point cytotoxicity assays and quantitative high content analysis (HCA) of multiple sub-lethal parameters in Caco-2 cells. C10 (8.5 mM) reversibly reduced TEER and increased FD4 permeability across monolayers, whereas SNAC had no effects on either parameter except at cytotoxic concentrations. C10 exposure induced reorganization of three TJ proteins, whereas SNAC only affected claudin-5 localization. High concentrations of C10 and SNAC were required to cause end-point toxicology changes in vitro. SNAC was less potent than C10 at inducing lysosomal and nuclear changes and plasma membrane perturbation. In parallel, HCA revealed that both agents displayed detergent-like features that reflect initial membrane fluidization followed by changes in intracellular parameters. In conclusion, FD4 permeability increases in monolayers in response to C10 were in the range of concentrations that altered end-point cytotoxicity and HCA parameters. For SNAC, while HCA parameters were also altered in a similar overall pattern as C10, they did not lead to increased paracellular flux. These assays show that both agents are primarily surfactants, but C10 has additional TJ-opening effects. While these in vitro assays illucidate their epithelial mechanism of action, clinical experience suggests that they over-estimate their toxicology in the dynamic intestinal environment.
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Affiliation(s)
- Caroline Twarog
- UCD School of Veterinary Medicine, University College Dublin, Belfield, Dublin 4, Ireland; UCD Conway Institute, University College Dublin, Belfield, Dublin 4, Ireland
| | - Kai Liu
- UCD Conway Institute, University College Dublin, Belfield, Dublin 4, Ireland; UCD School of Chemistry, University College Dublin, Belfield, Dublin 4, Ireland
| | - Peter J O'Brien
- UCD School of Veterinary Medicine, University College Dublin, Belfield, Dublin 4, Ireland
| | - Kenneth A Dawson
- UCD Conway Institute, University College Dublin, Belfield, Dublin 4, Ireland; UCD School of Chemistry, University College Dublin, Belfield, Dublin 4, Ireland
| | - Elias Fattal
- School of Pharmacy, Institut Galien, CNRS, Univ. Paris-Sud, Univ. Paris-Saclay, 92290 Châtenay-Malabry, France
| | - Brigitte Illel
- Drug Product Development, Small Molecules Oral Platform, Sanofi Research and Development, Montpellier, France
| | - David J Brayden
- UCD School of Veterinary Medicine, University College Dublin, Belfield, Dublin 4, Ireland; UCD Conway Institute, University College Dublin, Belfield, Dublin 4, Ireland.
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41
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Litak J, Grochowski C, Litak J, Osuchowska I, Gosik K, Radzikowska E, Kamieniak P, Rolinski J. TLR-4 Signaling vs. Immune Checkpoints, miRNAs Molecules, Cancer Stem Cells, and Wingless-Signaling Interplay in Glioblastoma Multiforme-Future Perspectives. Int J Mol Sci 2020; 21:ijms21093114. [PMID: 32354122 PMCID: PMC7247696 DOI: 10.3390/ijms21093114] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/20/2020] [Revised: 04/23/2020] [Accepted: 04/24/2020] [Indexed: 02/06/2023] Open
Abstract
Toll-like-receptor (TLR) family members were detected in the central nervous system (CNS). TLR occurrence was noticed and widely described in glioblastomamultiforme (GBM) cells. After ligand attachment, TLR-4 reorients domains and dimerizes, activates an intracellular cascade, and promotes further cytoplasmatic signaling. There is evidence pointing at a strong relation between TLR-4 signaling and micro ribonucleic acid (miRNA) expression. The TLR-4/miRNA interplay changes typical signaling and encourages them to be a target for modern immunotherapy. TLR-4 agonists initiate signaling and promote programmed death ligand-1 (PD-1L) expression. Most of those molecules are intensively expressed in the GBM microenvironment, resulting in the autocrine induction of regional immunosuppression. Another potential target for immunotreatment is connected with limited TLR-4 signaling that promotes Wnt/DKK-3/claudine-5 signaling, resulting in a limitation of GBM invasiveness. Interestingly, TLR-4 expression results in bordering proliferative trends in cancer stem cells (CSC) and GBM. All of these potential targets could bring new hope for patients suffering from this incurable disease. Clinical trials concerning TLR-4 signaling inhibition/promotion in many cancers are recruiting patients. There is still a lot to do in the field of GBM immunotherapy.
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Affiliation(s)
- Jakub Litak
- Department of Neurosurgery and Pediatric Neurosurgery, Medical University of Lublin, 20-954 Lublin, Poland
- Department of Immunology, Medical University of Lublin, 20-093 Lublin, Poland
| | - Cezary Grochowski
- Department of Anatomy, Medical University of Lublin, 20-090 Lublin, Poland
- Laboratory of Virtual Man, Department of Anatomy, Medical University of Lublin, 20-090 Lublin, Poland
- Correspondence:
| | - Joanna Litak
- St. John‘s Cancer Center in Lublin, 20-090 Lublin, Poland
| | - Ida Osuchowska
- Department of Anatomy, Medical University of Lublin, 20-090 Lublin, Poland
| | - Krzysztof Gosik
- Department of Immunology, Medical University of Lublin, 20-093 Lublin, Poland
| | | | - Piotr Kamieniak
- Department of Immunology, Medical University of Lublin, 20-093 Lublin, Poland
| | - Jacek Rolinski
- Department of Immunology, Medical University of Lublin, 20-093 Lublin, Poland
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42
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Milatz S. A Novel Claudinopathy Based on Claudin-10 Mutations. Int J Mol Sci 2019; 20:ijms20215396. [PMID: 31671507 PMCID: PMC6862131 DOI: 10.3390/ijms20215396] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2019] [Revised: 10/26/2019] [Accepted: 10/27/2019] [Indexed: 02/06/2023] Open
Abstract
Claudins are key components of the tight junction, sealing the paracellular cleft or composing size-, charge- and water-selective paracellular channels. Claudin-10 occurs in two major isoforms, claudin-10a and claudin-10b, which constitute paracellular anion or cation channels, respectively. For several years after the discovery of claudin-10, its functional relevance in men has remained elusive. Within the past two years, several studies appeared, describing patients with different pathogenic variants of the CLDN10 gene. Patients presented with dysfunction of kidney, exocrine glands and skin. This review summarizes and compares the recently published studies reporting on a novel autosomal-recessive disorder based on claudin-10 mutations.
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Affiliation(s)
- Susanne Milatz
- Institute of Physiology, Kiel University, Christian-Albrechts-Platz 4, 24118 Kiel, Germany.
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43
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Castro Dias M, Mapunda JA, Vladymyrov M, Engelhardt B. Structure and Junctional Complexes of Endothelial, Epithelial and Glial Brain Barriers. Int J Mol Sci 2019; 20:E5372. [PMID: 31671721 PMCID: PMC6862204 DOI: 10.3390/ijms20215372] [Citation(s) in RCA: 97] [Impact Index Per Article: 16.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2019] [Revised: 10/25/2019] [Accepted: 10/26/2019] [Indexed: 01/04/2023] Open
Abstract
The homeostasis of the central nervous system (CNS) is ensured by the endothelial, epithelial, mesothelial and glial brain barriers, which strictly control the passage of molecules, solutes and immune cells. While the endothelial blood-brain barrier (BBB) and the epithelial blood-cerebrospinal fluid barrier (BCSFB) have been extensively investigated, less is known about the epithelial and mesothelial arachnoid barrier and the glia limitans. Here, we summarize current knowledge of the cellular composition of the brain barriers with a specific focus on describing the molecular constituents of their junctional complexes. We propose that the brain barriers maintain CNS immune privilege by dividing the CNS into compartments that differ with regard to their role in immune surveillance of the CNS. We close by providing a brief overview on experimental tools allowing for reliable in vivo visualization of the brain barriers and their junctional complexes and thus the respective CNS compartments.
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Affiliation(s)
| | | | | | - Britta Engelhardt
- Theodor Kocher Institute, University of Bern, 3012 Bern, Switzerland.
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44
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Nunes C, Freitas V, Almeida L, Laranjinha J. Red wine extract preserves tight junctions in intestinal epithelial cells under inflammatory conditions: implications for intestinal inflammation. Food Funct 2019; 10:1364-1374. [PMID: 30735221 DOI: 10.1039/c8fo02469c] [Citation(s) in RCA: 78] [Impact Index Per Article: 13.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
The altered expression and subcellular distribution of tight junction (TJ) proteins, leading to a dysfunctional intestinal barrier, is a key mechanistic feature of inflammatory bowel disease (IBD). Therefore, increasing the integrity of the intestinal barrier by manipulating the TJ may constitute an innovative and effective therapeutic strategy in IBD. In this context, recent studies showed that dietary polyphenols are able to protect the intestinal TJ barrier integrity. Here, using a cellular model of intestinal inflammation, consisting of cytokine-stimulated HT-29 colon epithelial cells, we show that a polyphenolic extract obtained from Portuguese red wine (RWE) decreased the paracellular permeability across the cell monolayer compared with the control cells, even in the presence of pro-inflammatory cytokines. The beneficial effect of RWE was exerted at three complementary levels: (1) by promoting a significant increase of the mRNA of key barrier-forming TJ proteins, including occludin, claudin-5 and zonnula occludens (ZO)-1 above the levels observed in the control cells; (2) by preventing the decrease in the expression of these proteins under inflammatory conditions and (3) by averting the increase in claudin-2 mRNA, a channel-forming TJ protein induced by pro-inflammatory cytokines. Taken together, these results strongly suggest that polyphenols presented and consumed in red wine as a mixture can reinforce and protect the intestinal barrier against inflammatory stimulus by affecting the TJ protein expression and, thus, without the need for purifying individual compounds, might represent a readily available therapeutic intervention against IBD and intestinal inflammation.
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Affiliation(s)
- Carla Nunes
- Center for Neurosciences and Cell Biology and Faculty of Pharmacy, University of Coimbra, Health Sciences Campus, Azinhaga de Santa Comba, 3000-548 Coimbra, Portugal.
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45
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Caprate Modulates Intestinal Barrier Function in Porcine Peyer's Patch Follicle-Associated Epithelium. Int J Mol Sci 2019; 20:ijms20061418. [PMID: 30897851 PMCID: PMC6471651 DOI: 10.3390/ijms20061418] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2019] [Revised: 03/09/2019] [Accepted: 03/19/2019] [Indexed: 12/29/2022] Open
Abstract
Background: Many food components influence intestinal epithelial barrier properties and might therefore also affect susceptibility to the development of food allergies. Such allergies are triggered by increased antibody production initiated in Peyer’s patches (PP). Usually, the presentation of antigens in the lumen of the gut to the immune cells of the PP is strongly regulated by the follicle-associated epithelium (FAE) that covers the PP. As the food component caprate has been shown to impede barrier properties in villous epithelium, we hypothesized that caprate also affects the barrier function of the PP FAE, thereby possibly contributing a risk factor for the development of food allergies. Methods: In this study, we have focused on the effects of caprate on the barrier function of PP, employing in vitro and ex vivo experimental setups to investigate functional and molecular barrier properties. Incubation with caprate induced an increase of transepithelial resistance, and a marked increase of permeability for the paracellular marker fluorescein in porcine PP to 180% of control values. These effects are in accordance with changes in the expression levels of the barrier-forming tight junction proteins tricellulin and claudin-5. Conclusions: This barrier-affecting mechanism could be involved in the initial steps of a food allergy, since it might trigger unregulated contact of the gut lumen with antigens.
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Greene C, Hanley N, Campbell M. Claudin-5: gatekeeper of neurological function. Fluids Barriers CNS 2019; 16:3. [PMID: 30691500 PMCID: PMC6350359 DOI: 10.1186/s12987-019-0123-z] [Citation(s) in RCA: 330] [Impact Index Per Article: 55.0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2018] [Accepted: 01/11/2019] [Indexed: 02/07/2023] Open
Abstract
Tight junction proteins of the blood–brain barrier are vital for maintaining integrity of endothelial cells lining brain blood vessels. The presence of these protein complexes in the space between endothelial cells creates a dynamic, highly regulated and restrictive microenvironment that is vital for neural homeostasis. By limiting paracellular diffusion of material between blood and brain, tight junction proteins provide a protective barrier preventing the passage of unwanted and potentially damaging material. Simultaneously, this protective barrier hinders the therapeutic effectiveness of central nervous system acting drugs with over 95% of small molecule therapeutics unable to bypass the blood–brain barrier. At the blood–brain barrier, claudin-5 is the most enriched tight junction protein and its dysfunction has been implicated in neurodegenerative disorders such as Alzheimer’s disease, neuroinflammatory disorders such as multiple sclerosis as well as psychiatric disorders including depression and schizophrenia. By regulating levels of claudin-5, it is possible to abrogate disease symptoms in many of these disorders. This review will give an overview of the blood–brain barrier and the role of tight junction complexes in maintaining blood–brain barrier integrity before focusing on the role of claudin-5 and its regulation in homeostatic and pathological conditions. We will also summarise therapeutic strategies to restore integrity of cerebral vessels by targeting tight junction protein complexes.
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Affiliation(s)
- Chris Greene
- Trinity College Dublin, Smurfit Institute of Genetics, Dublin 2, Ireland
| | - Nicole Hanley
- Trinity College Dublin, Smurfit Institute of Genetics, Dublin 2, Ireland
| | - Matthew Campbell
- Trinity College Dublin, Smurfit Institute of Genetics, Dublin 2, Ireland.
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Alberini G, Benfenati F, Maragliano L. Molecular Dynamics Simulations of Ion Selectivity in a Claudin-15 Paracellular Channel. J Phys Chem B 2018; 122:10783-10792. [DOI: 10.1021/acs.jpcb.8b06484] [Citation(s) in RCA: 34] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Affiliation(s)
- Giulio Alberini
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- Department of Experimental Medicine, Università degli Studi di Genova, Viale Benedetto XV, 3, 16132 Genova, Italy
| | - Fabio Benfenati
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132 Genova, Italy
| | - Luca Maragliano
- Center for Synaptic Neuroscience and Technology (NSYN@UniGe), Istituto Italiano di Tecnologia, Largo Rosanna Benzi, 10, 16132 Genova, Italy
- IRCCS Ospedale Policlinico San Martino, Largo Rosanna Benzi, 10, 16132 Genova, Italy
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48
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Alves NG, Yuan SY, Breslin JW. Sphingosine-1-phosphate protects against brain microvascular endothelial junctional protein disorganization and barrier dysfunction caused by alcohol. Microcirculation 2018; 26:e12506. [PMID: 30281888 DOI: 10.1111/micc.12506] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/31/2018] [Revised: 09/26/2018] [Accepted: 09/27/2018] [Indexed: 12/11/2022]
Abstract
OBJECTIVE S1P has known endothelial barrier-protective properties, but whether this extends to the BBB is unclear. We hypothesized that alcohol-induced disruption of brain microvascular endothelial barrier function and junctional protein organization can be ameliorated by S1P treatment. METHODS Cultured primary HBMEC monolayers were used to characterize endothelial-specific mechanisms of BBB regulation. TER and apparent permeability coefficients for albumin, dextran-4 kDa, and sodium fluorescein were used as indices of barrier function. Junctional localization of Claudin-5, VE-cadherin, and β-catenin was determined by immunofluorescence confocal microscopy. S1P was applied following treatment with alcohol. RESULTS Alcohol significantly impaired HBMEC TER. Application of S1P after alcohol treatment resulted in a hastened recovery to the baseline HBMEC TER. Alcohol-treated HBMEC had a significantly higher mean permeability than control that was reversed by S1P. Alcohol caused the formation of gaps between cells. Treatment with S1P (after alcohol) increased junctional localization of VE-Cadherin, Claudin-5, and β-catenin. CONCLUSIONS Alcohol impairs the barrier function and junctional organization of HBMEC monolayers. S1P enhanced barrier function and restored junctions in the presence of alcohol, and thus may be useful for restoring BBB function during alcohol intoxication.
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Affiliation(s)
- Natascha G Alves
- Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, Florida
| | - Sarah Y Yuan
- Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, Florida
| | - Jerome W Breslin
- Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, Florida
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49
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Li X, Wang X, Xie J, Liang B, Wu J. Suppression of Angiotensin-(1-7) on the Disruption of Blood-Brain Barrier in Rat of Brain Glioma. Pathol Oncol Res 2018; 25:429-435. [PMID: 30229380 DOI: 10.1007/s12253-018-0471-z] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/20/2018] [Accepted: 09/13/2018] [Indexed: 01/03/2023]
Abstract
Glioblastoma multiforme (GBM) is the most primary brain tumor, specially characterized with the damage of blood-brain barrier (BBB). The Ang-(1-7) was proven to have an inhibitory effect on glioblastoma growth. However, its role on blood-brain barrier (BBB) and the underlying molecular mechanism remains unclear. In this study, Ang-(1-7) significantly relieved the damage of blood-brain barrier in rats with intracranial U87 gliomas as evaluated by magnetic resonance imaging (MRI). Furthermore, its treatment attenuated BBB permeability, tumor growth and edema formation. Similarly, Ang-(1-7) also decreased U87 glioma cells barrier permeability in vitro. Further analysis showed that Ang-(1-7) could effectively restore tight junction protein (claudin-5 and ZO-1) expression levels both in rats and U87 glioma cells by affecting the activation of JNK pathway. SP600125, an inhibitor of JNK, significantly enhanced the expression of Claudin-5 and ZO-1, and decreased the disruption of BBB and enhanced the efficiency of Ang-(1-7) in glioma rats. Taken together, this study demonstrated a protective role of Ang-(1-7) in glioma-induced blood-brain barrier damage by regulating tight junction protein expression. Accordingly, Ang-(1-7) may become a promising therapeutic agent against glioma.
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Affiliation(s)
- Xiaohui Li
- Department of Neurosurgery, The Fifth Affiliated Hospital of Zhengzhou University, No.3 Kangfuqian Street, Erqi District, Zhengzhou, 450052, China
| | - Xinjun Wang
- Department of Neurosurgery, The Fifth Affiliated Hospital of Zhengzhou University, No.3 Kangfuqian Street, Erqi District, Zhengzhou, 450052, China.
| | - Jingwei Xie
- Department of Neurosurgery, The Fifth Affiliated Hospital of Zhengzhou University, No.3 Kangfuqian Street, Erqi District, Zhengzhou, 450052, China
| | - Bo Liang
- Department of Neurosurgery, The Fifth Affiliated Hospital of Zhengzhou University, No.3 Kangfuqian Street, Erqi District, Zhengzhou, 450052, China
| | - Jianheng Wu
- Department of Neurosurgery, The Fifth Affiliated Hospital of Zhengzhou University, No.3 Kangfuqian Street, Erqi District, Zhengzhou, 450052, China
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50
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Nakayama H, Kitagawa N, Otani T, Iida H, Anan H, Inai T. Ochratoxin A, citrinin and deoxynivalenol decrease claudin-2 expression in mouse rectum CMT93-II cells. Microscopy (Oxf) 2018; 67:99-111. [PMID: 29474583 DOI: 10.1093/jmicro/dfy005] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2017] [Accepted: 01/16/2018] [Indexed: 01/01/2023] Open
Abstract
Intestinal epithelial cells are the first targets of ingested mycotoxins, such as ochratoxin A, citrinin and deoxynivalenol. It has been reported that paracellular permeability regulated by tight junctions is modulated by several mycotoxins by reducing the expression of specific claudins and integral membrane proteins in cell-cell contacts, accompanied by increase in phosphorylation of mitogen-activated protein kinases, including extracellular signal-related kinase (ERK) 1/2, p38 and c-Jun NH2-terminal protein kinase. Claudin-2 is expressed in the deep crypt cells, but not in the villus/surface cells in vivo. While Caco-2, T84 and IPEC-J2 cells, which are widely used intestinal epithelial cell lines to assess the influence of mycotoxins, do not express claudin-2, CMT93-II cells express claudin-2. We previously reported that inhibition of the ERK pathway reduced claudin-2 levels in cell-cell contacts in CMT93-II cells. In this study, we examined whether ochratoxin A, citrinin and deoxynivalenol affect claudin-2 expression and ERK1/2 phosphorylation in CMT93-II cells. We found that all mycotoxins reduced claudin-2 expression in cell-cell contacts, with reduction (by citrinin and deoxynivalenol) or no change (by ochratoxin A) in phosphorylated ERK1/2. All mycotoxins increased transepithelial electrical resistance, but did not affect flux of fluorescein. While ochratoxin A and citrinin are known to be nephrotoxic, only deoxynivalenol reduced claudin-2 expression in MDCK II cells derived from the renal tubule. These results suggest that claudin-2 expression is regulated not only by the ERK pathway, but also by other pathways in an organ-specific manner.
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Affiliation(s)
- Hideaki Nakayama
- Department of Odontology, Fukuoka Dental College, 2-15-1 Tamura, Sawara-ku, Fukuoka 814-0193, Japan
| | - Norio Kitagawa
- Department of Morphological Biology, Fukuoka Dental College, 2-15-1 Tamura, Sawara-ku, Fukuoka 814-0193, Japan
| | - Takahito Otani
- Department of Morphological Biology, Fukuoka Dental College, 2-15-1 Tamura, Sawara-ku, Fukuoka 814-0193, Japan
| | - Hiroshi Iida
- Laboratory of Zoology, Graduate School of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan
| | - Hisashi Anan
- Department of Odontology, Fukuoka Dental College, 2-15-1 Tamura, Sawara-ku, Fukuoka 814-0193, Japan
| | - Tetsuichiro Inai
- Department of Morphological Biology, Fukuoka Dental College, 2-15-1 Tamura, Sawara-ku, Fukuoka 814-0193, Japan
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