Opposing Effect by Cytokines on Fas-Mediated Apoptosis in A549 Lung Epithelial Cells

Abstract

Tissue repair is determined by many signals provided in the local environment. Central to this process is the commitment of the parenchymal cell to undergo apoptosis, survive, or proliferate following inflammation. We hypothesize that lung epithelial cell apoptosis is influenced by exposure to cytokines released into the alveolar microenvironment during the inflammatory process. In this investigation we demonstrate that interferon (IFN)-gamma and interleukin (IL)-1beta have opposing effects on Fas-mediated apoptosis in A549 cells, a human lung epithelial cell line. Exposure to IFN-gamma before Fas activation significantly increased caspase activity, caspase processing of CK-18, a key cytoskeletal protein in epithelial cells, and increased the appearance of apoptotic nuclei. Induction of Fas-mediated death by IFN-gamma was 3-fold higher than with Fas activation alone. In contrast, pretreatment with IL-1beta before Fas activation completely inhibited apoptosis. Furthermore, our results demonstrate that IFN-gamma and IL-1beta induce opposite effects at multiple checkpoints during Fas-mediated apoptosis. Most striking, IL-1beta prevented the activation of caspases involved in Fas-mediated death by inducing an anti-apoptotic effect proximal to or at the point of caspase-8 activation. Finally, our investigation demonstrates that the differential impact of IL-1beta and IFN-gamma on Fas-mediated apoptosis are in part dependent on modulation of the PI 3-K/Akt survival pathway.

Activated T killer cells induce apoptosis in lung epithelial cells and the release of pro‐inflammatory cytokine TNF‐α

“…Activation of CD95 led to apoptosis of IFN-+ -pretreated A549 cells (Fig. 4A, D) in accordance with previous reports [23,24]. When protein A was utilized to cross-link anti-APO-1 Ab bound to CD95, apoptosis was induced not only in IFN-+ -pretreated but also in A549 cells that had not been pretreated with IFN-+ (Fig.…”

“…1C). As IFN-+ has been reported to sensitize A549 cells to agonistic anti-CD95 Ab-or TRAIL-induced apoptosis [23][24][25], analysis of cell surface markers was performed on IFN-+ -treated A549 cells as well as on non-stimulated cells. Both nonstimulated and IFN-+ -treated A549 cells expressed CD95, TRAIL-R1, and TRAIL-R2.…”

Lack of matrix metalloproteinase-9 worsens ventilator-induced lung injury

“…IL-1␤ can promote pulmonary injury (29), and IL-4 is a pleiotropic cytokine that has been related to cell recruitment in models of airway disease in Mmp9 Ϫ/Ϫ (33) and Mmp8 Ϫ/Ϫ (19) mice. Interestingly, both IL-1␤ and IL-4 can delay neutrophil apoptosis (10,17), thus impairing the resolution of inflammation. However, we cannot discard other molecules as responsible for our results.…”

Zinc modulates cytokine-induced lung epithelial cell barrier permeability

“…The presence of active caspases was determined by 7-amido-4-trifluoromethylcoumarin (AFC) assay using specific fluorosubstrates. For all AFC preparations, cells (3 ϫ 10 6 ) were collected by centrifugation and washed with potassium phosphate buffer containing magnesium sulfate and lysed by four cycles of freeze-thawing as previously described (10). Lysates were incubated with cyto-buffer (10% glycerol, 50 mM PIPES, pH 7.0, 1 mM EDTA) containing 1 mM DTT and 20 M DEVD-AFC (Enzyme Systems Products).…”

“…Cells were detached with a nonenzymatic cell dissociation solution (Sigma, St. Louis, MO) and pooled with cells already suspended during culture followed by cytopsin preparation onto silane-treated slides. Cells were then washed and stained with the M30 CytoDEATH-FITC antibody (Boehringer Mannheim, Indianapolis, IN), a monoclonal antibody that specifically detects caspase-cleaved human cytokeratin-18 (CK-18), as we have previously reported (10). Cells were also stained with 0.5 mg/ml 4Ј,6-diamidino-2-phenylindole dihydrochloride (DAPI; Roche Molecular Biochemicals, Indianapolis, IN).…”