Listeriolysin O Causes ENaC Dysfunction in Human Airway Epithelial Cells.

Pulmonary permeability edema is characterized by reduced alveolar Na⁺ uptake capacity and capillary barrier dysfunction and is a potentially lethal complication of listeriosis. Apical Na⁺ uptake is mainly mediated by the epithelial sodium channel (ENaC) and initiates alveolar liquid clearance. Here we examine how listeriolysin O (LLO), the pore-forming toxin of Listeria monocytogenes, impairs the expression and activity of ENaC. To that purpose, we studied how sub-lytic concentrations of LLO affect negative and positive regulators of ENaC expression in the H441 airway epithelial cell line. LLO reduced expression of the crucial ENaC-α subunit in H441 cells within 2 h and this was preceded by activation of PKC-α, a negative regulator of the channel's expression. At later time points, LLO caused a significant reduction in the phosphorylation of Sgk-1 at residue T256 and of Akt-1 at residue S473, both of which are required for full activation of ENaC. The TNF-derived TIP peptide prevented LLO-mediated PKC-α activation and restored phospho-Sgk-1-T256. The TIP peptide also counteracted the observed LLO-induced decrease in amiloride-sensitive Na⁺ current and ENaC-α expression in H441 cells. Intratracheally instilled LLO caused profound pulmonary edema formation in mice, an effect that was prevented by the TIP peptide; thus indicating the therapeutic potential of the peptide for the treatment of pore-forming toxin-associated permeability edema.

Malignant but not naïve hepatocytes of human and rodent origin are killed by TNF after metabolic depletion of ATP by fructose.

BACKGROUND & AIMS: TNF was the first cytokine employed for cancer therapy, but its use was limited due to its insufficient selectivity towards malignant cells. Fructose induces transient hepatic ATP depletion in humans and rodents due to the liver-specific fructose metabolism via fructokinase, while other cells e.g. Muscle cells metabolize fructose via hexokinase. Under ATP depleted conditions hepatocytes are protected against TNF-induced apoptosis. Our aim was to identify metabolic differences between normal and malignant liver cells that can be exploited for selective immunotherapy. METHODS: We analyzed the expression and activities of enzymes involved in fructose metabolism in primary hepatocytes and hepatoma cell lines. Furthermore, we studied the influence of hexokinase II (HKII) on fructose-mediated ATP depletion and cytoprotection in murine hepatocytes. RESULTS: Primary mouse, rat and human hepatocytes depleted of ATP by fructose were fully protected against TNF-induced cytotoxicity. By contrast, hepatic tumor cell lines showed increased HKII expression, lack of fructose-mediated ATP depletion and, therefore, remained susceptible to TNF/ActD-induced apoptosis. Inhibition of hexokinases restored fructose-induced ATP depletion in hepg2 cells. Finally, hypoxia-inducible factor1 (HIF1)-mediated up-regulation of HKII prevented fructose-induced ATP depletion and overexpression of HKII inhibited fructose-mediated cytoprotection against TNF-induced apoptosis in primary murine hepatocytes. CONCLUSION: Increased expression of HKII in malignant cells of hepatic origin shifts the fructose metabolism from liver- to muscle-type, thereby preventing ATP depletion and subsequent cytoprotection of the target cells. Therefore, healthy liver cells are transiently protected from TNF-mediated cell death by fructose-induced ATP depletion, while malignant cells can be selectively eliminated through TNF-induced apoptosis.