Oxidative Stress Facilitates IFN-γ-Induced Mimic Extracellular Trap Cell Death in A549 Lung Epithelial Cancer Cells.

We previously demonstrated that IFN-γ induces an autophagy-regulated mimic extracellular trap cell death (ETosis) in A549 human lung cancer cells. Regarding reactive oxygen species (ROS) are involved in ETosis, this study investigated the role of oxidative stress. After IFN-γ stimulation, a necrosis-like cell death mimic ETosis occurred accompanied by the inhibition of cell growth, aberrant nuclear staining, and nucleosome release. ROS were generated in a time-dependent manner with an increase in NADPH oxidase component protein expression. STAT1-mediated IFN regulatory factor-1 activation was essential for upregulating ROS production. By genetically silencing p47phox, IFN-γ-induced ROS and mimic ETosis were significantly attenuated. This mechanistic study indicated that ROS may mediate DNA damage followed by histone H3 citrullination. Furthermore, ROS promoted IFN-γ-induced mimic ETosis in cooperation with autophagy. These findings further demonstrate that ROS regulates IFN-γ-induced mimic ETosis in lung epithelial malignancy.

IFN-γ Induces Mimic Extracellular Trap Cell Death in Lung Epithelial Cells Through Autophagy-Regulated DNA Damage.

Treatment of interferon-γ (IFN-γ) causes cell growth inhibition and cytotoxicity in lung epithelial malignancies. Regarding the induction of autophagy related to IFN-γ signaling, this study investigated the link between autophagy and IFN-γ cytotoxicity. In A549 human lung cancer cells, IFN-γ treatment induced concurrent apoptotic and nonapoptotic events. Unexpectedly, the nonapoptotic cells present mimic extracellular trap cell death (ETosis), which was regulated by caspase-3 and by autophagy induction through immunity-related GTPase family M protein 1 and activating transcription factor 6. Furthermore, IFN-γ signaling controlled mimic ETosis through a mechanism involving an autophagy- and Fas-associated protein with death domain-controlled caspase-8/-3 activation. Following caspase-mediated lamin degradation, IFN-γ caused DNA damage-associated ataxia telangiectasia and Rad3-related protein (ATR)/ataxia telangiectasia mutated (ATM)-regulated mimic ETosis. Upon ATR/ATM signaling, peptidyl arginine deiminase 4 (PAD4)-mediated histone 3 citrullination promoted mimic ETosis. Such IFN-γ-induced effects were defective in PC14PE6/AS2 human lung cancer cells, which were unsusceptible to IFN-γ-induced autophagy. Due to autophagy-based caspase cascade activation, IFN-γ triggers unconventional caspase-mediated DNA damage, followed by ATR/ATM-regulated PAD4-mediated histone citrullination during mimic ETosis in lung epithelial malignancy.

Autophagy facilitates cytokine-induced ICAM-1 expression.

ICAM-1 can be induced by inflammatory cytokines such as IFN-γ and TNF-α. This study investigated whether autophagy regulates ICAM-1 given that autophagy facilitates signaling of these two cytokines. Exogenous IFN-γ induced ICAM-1 in human lung epithelial A549 cells carrying wild type p53, a transcription factor reported for ICAM-1, but not in PC14PE6/AS2 (AS2) cells carrying mutated p53. However, IFN-γ also induced ICAM-1 in A549 cells with short hairpin RNA-silenced p53. No changes in IFN-γ receptor expression were observed in AS2 cells, but IFN-γ-activated Jak2/STAT1/IFN regulatory factor 1 was markedly decreased. In AS2 cells, increased levels of reactive oxygen species induced the activation of Src homology domain-containing phosphatase 2 (SHP2), while SHP2 was essential for IFN-γ resistance. AS2 cells showed autophagy resistance, and the manipulation of the autophagy pathway altered IFN-γ resistance. Aberrant Bcl-2 expression and mammalian target of rapamycin activation contributed to both autophagy resistance and IFN-γ resistance. Autophagy, but not p53, also modulated TNF-α-induced NF-κB activation and ICAM-1 expression. Inhibiting autophagy decreased the adhesion of human monocytic U937 cells to IFN-γ-treated A549 cells. These results demonstrated that IFN-γ and TNF-α induced ICAM-1 expression through a common pathway that was regulated by autophagy, but not p53.