Ethanol induced NF-κB activation protects against cell injury in cultured rat gastric mucosal epithelium

Author:

Mustonen Harri,Hietaranta Antti,Puolakkainen Pauli,Kemppainen Esko,Paimela Hannu,Kiviluoto Tuula,Kivilaakso Eero

Abstract

Ethanol is a well-established irritant inducing inflammation in gastric mucosa, but the effects at the cellular level remain unclear. This study investigates NF-κB activation in gastric mucosal cells by ethanol and assesses the effects of heat shock pretreatment in this ulcerogenic situation. Rat gastric mucosal epithelia were exposed to ethanol for different time periods. Heat shock was induced by incubating the cells at 42°C for 1 h prior to the experiments. For evaluation of NF-κB activation, the nuclear fraction of the cell lysates was analyzed with an EMSA or an ELISA-based assay. Caspase-3 (a promoter of apoptosis) activity was measured with a time-resolved fluorescence based assay, cell viability with a tetrazolium assay, and cell membrane integrity with a LDH assay. Ethanol (1–5%) induced NF-κB activation, reaching a maximum after 3 h, and also led to moderately increased COX-2 expression. Heat shock pretreatment and the intracellular calcium chelator BAPTA were able to inhibit ethanol-induced NF-κB activation. Heat shock pretreatment decreased ethanol-induced caspase-3 activation, decreased cell membrane damage, and retained cellular viability. Inhibition of NF-κB activation by NEMO-binding peptide, by decreasing RelA expression, or by inhibiting COX-2 activity by CAY-14040 promoted the effects of ethanol, such as increased caspase-3 activity and decreased cell viability. In conclusion, ethanol induces NF-κB activation via a calcium-dependent pathway and induces COX-2 expression. Inhibition of the NF-κB activation or COX-2 activity potentiates apoptosis and cell damage induced by ethanol, suggesting a protective role for NF-κB activation and COX-2 expression.

Publisher

American Physiological Society

Subject

Physiology (medical),Gastroenterology,Hepatology,Physiology

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