Abstract
Ischemia causes severe damage in the gastrointestinal tract. Therefore, it is interesting to study how the barrier and transport functions of intestinal epithelium change under hypoxia and subsequent reoxygenation. For this purpose we simulated hypoxia and reoxygenation on mucosa-submucosa preparations from rat distal colon in Ussing chambers and on isolated crypts. Hypoxia (N2 gassing for 15 min) induced a triphasic change in short-circuit current (Isc): a transient decrease, an increase and finally a long-lasting fall below the initial baseline. During the subsequent reoxygenation phase, Isc slightly rose to values above the initial baseline. Tissue conductance (Gt) showed a biphasic increase during both the hypoxia and the reoxygenation phases. Omission of Cl− or preincubation of the tissue with transport inhibitors revealed that the observed changes in Isc represented changes in Cl− secretion. The radical scavenger trolox C reduced the Isc response during hypoxia, but failed to prevent the rise of Isc during reoxygenation. All changes in Isc were Ca2+-dependent. Fura-2 experiments at loaded isolated colonic crypts revealed a slow increase of the cytosolic Ca2+ concentration during hypoxia and the reoxygenation phase, mainly caused by an influx of extracellular Ca2+. Surprisingly, no changes could be detected in the fluorescence of the superoxide anion-sensitive dye mitosox or the thiol-sensitive dye thiol tracker, suggesting a relative high capacity of the colonic epithelium (with its low O2 partial pressure even under physiological conditions) to deal with enhanced radical production during hypoxia/reoxygenation.
Highlights
Intestinal hypoxia, e.g., caused by arterial or venous thromboembolism or mechanical compression, is a severe gastroenterologic disease associated with a high lethality (Haglund and Bergqvist, 1999)
Two pore-forming isoforms (Kir 6.1 and Kir 6.2) and two regulatory subunits (SUR1, SUR2B) have been identified on the mRNA as well as the protein level in colonic epithelium (Pouokam et al, 2013). As these channels play a central role in the protection of excitable tissues against hypoxia (e.g., Hund and Mohler, 2011), we investigated the impact of hypoxia/reoxygenation on the barrier and ion transport functions of the intestinal epithelium and the contribution of these channels to the changes induced
Baseline in short-circuit current (Isc), which is a measure of net ion movement across the epithelium, at the end of the normoxic period amounted to 1.55 ± 0.98 μEq·h−1·cm−2 (n = 8)
Summary
Intestinal hypoxia, e.g., caused by arterial or venous thromboembolism or mechanical compression, is a severe gastroenterologic disease associated with a high lethality (Haglund and Bergqvist, 1999). A reduction in mucosal blood flow impairs energy supply to the high energy demanding intestinal epithelium leading to severe mucosal damage and a loss of the barrier function of the epithelium. This damage may be even exaggerated after reperfusion/reoxygenation due to the production of reactive oxygen species. The generation of these oxidants begins already during the hypoxic phase due to the accumulation of hypoxanthine and the proteolytic conversion of xanthine dehydrogenase into xanthine oxidase. Hypoxanthine is oxidized to xanthine, which is further oxidized supply is troesutroircedaciddu;rbinogthrerepaecrtfiuosniosnl/eraedoxtoygtehneatpiroond,urcatdioicnalopf rsoudpuecrotixoindeis(Oex−2ag)g.eWratheednboyxyOg−2en
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