Calcium Channels and Oxidative Stress Mediate a Synergistic Disruption of Tight Junctions by Ethanol and Acetaldehyde in Caco-2 Cell Monolayers

• Published in: Scientific reports 2016-12, Vol.6 (1), p.38899-38899, Article 38899
• Main Authors: Samak, Geetha, Gangwar, Ruchika, Meena, Avtar S., Rao, Roshan G., Shukla, Pradeep K., Manda, Bhargavi, Narayanan, Damodaran, Jaggar, Jonathan H., Rao, RadhaKrishna
• Format: Article
• Language: English
• Subjects: 14; 14/1; 14/19; 631/80/79/1987; 692/4020/2199; 82; 82/29; 96; 96/106; 96/109; Acetaldehyde; Acetaldehyde - metabolism; Acetaldehyde - pharmacology; Acetylcysteine; Alcohol dehydrogenase; Alcohol Dehydrogenase - metabolism; Aldehyde dehydrogenase; Aldehyde Dehydrogenase, Mitochondrial - metabolism; Caco-2 Cells; Calcium (intracellular); Calcium channels; Calcium Channels - metabolism; Caveolin-1; Cells, Cultured; Cyclosporins; Cysteine; Dehydrogenase; Dehydrogenases; Diltiazem; Ethanol; Ethanol - metabolism; Ethanol - pharmacology; Humanities and Social Sciences; Humans; multidisciplinary; Myosin-light-chain kinase; Oxidative stress; Oxidative Stress - drug effects; Reactive oxygen species; Science; Small intestine; Src protein; Synergistic effect; Tight junctions; Tight Junctions - drug effects; Tight Junctions - metabolism; Zonula Occludens-1 Protein - metabolism
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/srep38899

Ethanol is metabolized into acetaldehyde in most tissues. In this study, we investigated the synergistic effect of ethanol and acetaldehyde on the tight junction integrity in Caco-2 cell monolayers. Expression of alcohol dehydrogenase sensitized Caco-2 cells to ethanol-induced tight junction disruption and barrier dysfunction, whereas aldehyde dehydrogenase attenuated acetaldehyde-induced tight junction disruption. Ethanol up to 150 mM did not affect tight junction integrity or barrier function, but it dose-dependently increased acetaldehyde-mediated tight junction disruption and barrier dysfunction. Src kinase and MLCK inhibitors blocked this synergistic effect of ethanol and acetaldehyde on tight junction. Ethanol and acetaldehyde caused a rapid and synergistic elevation of intracellular calcium. Calcium depletion by BAPTA or Ca 2+ -free medium blocked ethanol and acetaldehyde-induced barrier dysfunction and tight junction disruption. Diltiazem and selective knockdown of TRPV6 or Ca V 1.3 channels, by shRNA blocked ethanol and acetaldehyde-induced tight junction disruption and barrier dysfunction. Ethanol and acetaldehyde induced a rapid and synergistic increase in reactive oxygen species by a calcium-dependent mechanism. N-acetyl-L-cysteine and cyclosporine A, blocked ethanol and acetaldehyde-induced barrier dysfunction and tight junction disruption. These results demonstrate that ethanol and acetaldehyde synergistically disrupt tight junctions by a mechanism involving calcium, oxidative stress, Src kinase and MLCK.