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- Amasheh, Maren, et al.
(författare)
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Regulation of mucosal structure and barrier function in rat colon exposed to tumor necrosis factor alpha and interferon gamma in vitro : A novel model for studying the pathomechanisms of inflammatory bowel disease cytokines
- 2009
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Ingår i: SCANDINAVIAN JOURNAL OF GASTROENTEROLOGY. - : Informa UK Limited. - 0036-5521 .- 1502-7708. ; 44:10, s. 1226-1235
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Tidskriftsartikel (refereegranskat)abstract
- Objective. In Inflammatory bowel disease (IBD), elevated cytokines are responsible for disturbed intestinal transport and barrier function. The mechanisms of cytokine action have usually been studied in cell culture models only; therefore the aim of this study was to establish an in vitro model based on native intestine to analyze distinct cytokine effects on barrier function, mucosal structure, and inherent regulatory mechanisms. Material and methods. Rat colon was exposed to tumor necrosis factor alpha (TNF alpha) and interferon gamma (IFN gamma) in Ussing chambers. Transepithelial resistance (R-t) and H-3-mannitol fluxes were measured for characterization of the paracellular pathway. Transcellular transport was analyzed by horseradish peroxidase (HRP) flux measurements. Expression and distribution of tight junction proteins were characterized in immunoblots and by means of confocal laser-scanning microscopy (LSM). Results. Colonic viability could be preserved for 20 h in a specialized in vitro set-up. This was sufficient to alter mucosal architecture with crypt surface reduction. R-t was decreased (101 +/- 10 versus 189 +/- 10 Omega . cm(2)) with a parallel increase in mannitol permeability after cytokine exposure. Tight junction proteins claudin-1, -5, -7, and occludin decreased (45 +/- 10%, 16 +/- 7%, 42 +/- 8%, and 42 +/- 13% of controls, respectively), while claudin- 2 increased to 208 +/- 32%. Occludin and claudin- 1 translocated from the plasma membrane to the cytoplasm. HRP flux increased from 0.73 +/- 0.09 to 8.55 +/- 2.92 pmol . h(-1) . cm(-2). Conclusions. A new experimental IBD model with native colon in vitro is presented. One-day exposure to TNFa and IFNg alters mucosal morphology and impairs epithelial barrier function by up-regulation of the paracellular pore-former claudin-2 and down-regulation of the barrier-builders claudin-1, -5, and -7. These alterations resemble changes seen in IBD and thus underline their prominent role in IBD pathogenicity.
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| 2. |
- Klar, Joakim, 1974-, et al.
(författare)
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Altered paracellular cation permeability due to a rare CLDN10B variant causes anhidrosis and kidney damage
- 2017
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Ingår i: PLOS Genetics. - : Public Library of Science (PLoS). - 1553-7390 .- 1553-7404. ; 13:7
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Tidskriftsartikel (refereegranskat)abstract
- Claudins constitute the major component of tight junctions and regulate paracellular permeability of epithelia. Claudin-10 occurs in two major isoforms that form paracellular channels with ion selectivity. We report on two families segregating an autosomal recessive disorder characterized by generalized anhidrosis, severe heat intolerance and mild kidney failure. All affected individuals carry a rare homozygous missense mutation c.144C>G, p.(N48K) specific for the claudin-10b isoform. Immunostaining of sweat glands from patients suggested that the disease is associated with reduced levels of claudin-10b in the plasma membranes and in canaliculi of the secretory portion. Expression of claudin-10b N48K in a 3D cell model of sweat secretion indicated perturbed paracellular Na+ transport. Analysis of paracellular permeability revealed that claudin-10b N48K maintained cation over anion selectivity but with a reduced general ion conductance. Furthermore, freeze fracture electron microscopy showed that claudin-10b N48K was associated with impaired tight junction strand formation and altered cis-oligomer formation. These data suggest that claudin-10b N48K causes anhidrosis and our findings are consistent with a combined effect from perturbed TJ function and increased degradation of claudin-10b N48K in the sweat glands. Furthermore, affected individuals present with Mg2+ retention, secondary hyperparathyroidism and mild kidney failure that suggest a disturbed reabsorption of cations in the kidneys. These renal-derived features recapitulate several phenotypic aspects detected in mice with kidney specific loss of both claudin-10 isoforms. Our study adds to the spectrum of phenotypes caused by tight junction proteins and demonstrates a pivotal role for claudin-10b in maintaining paracellular Na+ permeability for sweat production and kidney function.
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