Figure 3.

Spermidine prevents ethanol and LPS-induced NFκB nuclear translocation, IκB activation by phosphorylation and pro-inflammatory cytokine expression. Nuclear (A) and cytosolic (B) fractions were isolated from livers of mice treated in the absence (control) or presence of ethanol (EtOH), LPS and/or spermidine (SPN) as indicated and immunoblotted for NFκB. Representative immunoblots from both fractions are shown. Densitometric comparison analysis of the immunoreactive species was performed and normalized to Lamin B1 levels (for the nuclear fractions) or β-actin (for the cytosolic fractions). The relative distributions were determined and plotted as fold-increase over control. (C) Total protein isolated from control or treated mice was immunoblotted for total and phosphorylated IκB (pIκB). The ratio of phospho-IκB vs. total IκB was calculated from densitometric analysis of immunoreactive species. Representative immunoblots are shown. Values are plotted as fold-change. Total mRNA isolated from livers of control or treated mice was processed for qRT-PCR to determine expression levels of TNFα (D), IL1β (E) or IL6 (F). For each experimental condition, four mice were analyzed (n = 4). The values are means of triplicate experiments ± SE.