cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived Organoids
Claudins regulate paracellular permeability, contribute to epithelial polarization and are dysregulated during inflammation and carcinogenesis. Variants of the claudin-binding domain of <i>Clostridium perfringens</i> enterotoxin (cCPE) are highly sensitive protein ligands for generic det...
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MDPI AG
2023-07-01
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| Online Access: | https://www.mdpi.com/1999-4923/15/7/1980 |
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| author | Ayk Waldow Laura-Sophie Beier Janine Arndt Simon Schallenberg Claudia Vollbrecht Philip Bischoff Martí Farrera-Sal Florian N. Loch Christian Bojarski Michael Schumann Lars Winkler Carsten Kamphues Lukas Ehlen Jörg Piontek |
| author_facet | Ayk Waldow Laura-Sophie Beier Janine Arndt Simon Schallenberg Claudia Vollbrecht Philip Bischoff Martí Farrera-Sal Florian N. Loch Christian Bojarski Michael Schumann Lars Winkler Carsten Kamphues Lukas Ehlen Jörg Piontek |
| author_sort | Ayk Waldow |
| collection | DOAJ |
| description | Claudins regulate paracellular permeability, contribute to epithelial polarization and are dysregulated during inflammation and carcinogenesis. Variants of the claudin-binding domain of <i>Clostridium perfringens</i> enterotoxin (cCPE) are highly sensitive protein ligands for generic detection of a broad spectrum of claudins. Here, we investigated the preferential binding of YFP- or GST-cCPE fusion proteins to non-junctional claudin molecules. Plate reader assays, flow cytometry and microscopy were used to assess the binding of YFP- or GST-cCPE to non-junctional claudins in multiple in vitro and ex vivo models of human and rat gastrointestinal epithelia and to monitor formation of a tight junction barrier. Furthermore, YFP-cCPE was used to probe expression, polar localization and dysregulation of claudins in patient-derived organoids generated from gastric dysplasia and gastric cancer. Live-cell imaging and immunocytochemistry revealed cell polarity and presence of tight junctions in glandular organoids (originating from intestinal-type gastric cancer and gastric dysplasia) and, in contrast, a disrupted diffusion barrier for granular organoids (originating from discohesive tumor areas). In sum, we report the use of cCPE fusion proteins as molecular probes to specifically and efficiently detect claudin expression, localization and tight junction dysregulation in cell lines, tissue explants and patient-derived organoids of the gastrointestinal tract. |
| first_indexed | 2024-03-11T00:44:20Z |
| format | Article |
| id | doaj.art-dd7af9b4ff8a4d9d825bc64bed3e2388 |
| institution | Directory Open Access Journal |
| issn | 1999-4923 |
| language | English |
| last_indexed | 2024-03-11T00:44:20Z |
| publishDate | 2023-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Pharmaceutics |
| spelling | doaj.art-dd7af9b4ff8a4d9d825bc64bed3e23882023-11-18T20:56:32ZengMDPI AGPharmaceutics1999-49232023-07-01157198010.3390/pharmaceutics15071980cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived OrganoidsAyk Waldow0Laura-Sophie Beier1Janine Arndt2Simon Schallenberg3Claudia Vollbrecht4Philip Bischoff5Martí Farrera-Sal6Florian N. Loch7Christian Bojarski8Michael Schumann9Lars Winkler10Carsten Kamphues11Lukas Ehlen12Jörg Piontek13Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité—Universitätsmedizin Berlin, 12203 Berlin, GermanyClinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité—Universitätsmedizin Berlin, 12203 Berlin, GermanyBerlin Institute of Health (BIH), Charité—Universitätsmedizin Berlin, BIH Center for Regenerative Therapies (BCRT), 13353 Berlin, GermanyCharité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Berlin Institute of Health, Institute of Pathology, 10117 Berlin, GermanyCharité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Berlin Institute of Health, Institute of Pathology, 10117 Berlin, GermanyCharité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Berlin Institute of Health, Institute of Pathology, 10117 Berlin, GermanyBerlin Institute of Health (BIH), Charité—Universitätsmedizin Berlin, BIH Center for Regenerative Therapies (BCRT), 13353 Berlin, GermanyDepartment of General and Visceral Surgery, Campus Benjamin Franklin, Charité—Universitätsmedizin Berlin, 12203 Berlin, GermanyMedical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité—Universitätsmedizin Berlin, 12203 Berlin, GermanyMedical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité—Universitätsmedizin Berlin, 12203 Berlin, GermanyExperimental Pharmacology & Oncology Berlin-Buch GmbH, 13125 Berlin, GermanyPark-Klinik Weißensee, Department of General-Visceral and Minimally-Invasive Surgery, 13086 Berlin, GermanyBerlin Institute of Health (BIH), Charité—Universitätsmedizin Berlin, BIH Center for Regenerative Therapies (BCRT), 13353 Berlin, GermanyClinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité—Universitätsmedizin Berlin, 12203 Berlin, GermanyClaudins regulate paracellular permeability, contribute to epithelial polarization and are dysregulated during inflammation and carcinogenesis. Variants of the claudin-binding domain of <i>Clostridium perfringens</i> enterotoxin (cCPE) are highly sensitive protein ligands for generic detection of a broad spectrum of claudins. Here, we investigated the preferential binding of YFP- or GST-cCPE fusion proteins to non-junctional claudin molecules. Plate reader assays, flow cytometry and microscopy were used to assess the binding of YFP- or GST-cCPE to non-junctional claudins in multiple in vitro and ex vivo models of human and rat gastrointestinal epithelia and to monitor formation of a tight junction barrier. Furthermore, YFP-cCPE was used to probe expression, polar localization and dysregulation of claudins in patient-derived organoids generated from gastric dysplasia and gastric cancer. Live-cell imaging and immunocytochemistry revealed cell polarity and presence of tight junctions in glandular organoids (originating from intestinal-type gastric cancer and gastric dysplasia) and, in contrast, a disrupted diffusion barrier for granular organoids (originating from discohesive tumor areas). In sum, we report the use of cCPE fusion proteins as molecular probes to specifically and efficiently detect claudin expression, localization and tight junction dysregulation in cell lines, tissue explants and patient-derived organoids of the gastrointestinal tract.https://www.mdpi.com/1999-4923/15/7/1980claudintight junctionclostridium perfringens enterotoxincolonadenomatumor |
| spellingShingle | Ayk Waldow Laura-Sophie Beier Janine Arndt Simon Schallenberg Claudia Vollbrecht Philip Bischoff Martí Farrera-Sal Florian N. Loch Christian Bojarski Michael Schumann Lars Winkler Carsten Kamphues Lukas Ehlen Jörg Piontek cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived Organoids Pharmaceutics claudin tight junction clostridium perfringens enterotoxin colon adenoma tumor |
| title | cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived Organoids |
| title_full | cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived Organoids |
| title_fullStr | cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived Organoids |
| title_full_unstemmed | cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived Organoids |
| title_short | cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived Organoids |
| title_sort | ccpe fusion proteins as molecular probes to detect claudins and tight junction dysregulation in gastrointestinal cell lines tissue explants and patient derived organoids |
| topic | claudin tight junction clostridium perfringens enterotoxin colon adenoma tumor |
| url | https://www.mdpi.com/1999-4923/15/7/1980 |
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