RT Journal Article
SR Electronic
T1 Dependence of Reactive Oxygen Species and FLICE Inhibitory Protein on Lipofectamine-Induced Apoptosis in Human Lung Epithelial Cells
JF Journal of Pharmacology and Experimental Therapeutics
JO J Pharmacol Exp Ther
FD American Society for Pharmacology and Experimental Therapeutics
SP 969
OP 977
DO 10.1124/jpet.107.136077
VO 325
IS 3
A1 Lalana Kongkaneramit
A1 Narong Sarisuta
A1 Neelam Azad
A1 Yongju Lu
A1 Anand Krishnan V. Iyer
A1 Liying Wang
A1 Yon Rojanasakul
YR 2008
UL http://jpet.aspetjournals.org/content/325/3/969.abstract
AB Cationic liposomes such as Lipofectamine (LF) are widely used as nonviral gene delivery vectors; however, their clinical application is limited by their cytotoxicity. These agents have been shown to induce apoptosis as the primary mode of cell death, but their mechanism of action is not well understood. The present study investigated the mechanism of LF-induced apoptosis and examined the role of reactive oxygen species (ROS) in this process. We found that LF induced apoptosis of human epithelial H460 cells through a mechanism that involves caspase activation and ROS generation. Inhibition of caspase activity by pan-caspase inhibitor (z-VAD-fmk) or by specific caspase-8 inhibitor (z-IETD-fmk) or caspase-9 inhibitor (z-LEHD-fmk) inhibited the apoptotic effect of LF. Overexpression of FLICE-inhibitory protein (FLIP) or B-cell lymphoma-2, which are known inhibitors of the extrinsic and intrinsic death pathways, respectively, similarly inhibited apoptosis by LF. Induction of apoptosis by LF was shown to require ROS generation because its inhibition by ROS scavengers or by ectopic expression of antioxidant enzyme superoxide dismutase and glutathione peroxidase strongly inhibited the apoptotic effect of LF. Electron spin resonance studies showed that LF induced multiple ROS; however, superoxide was found to be the primary ROS responsible for LF-induced apoptosis. The mechanism by which ROS mediate the apoptotic effect of LF involves down-regulation of FLIP through the ubiquitination pathway. In demonstrating the role of FLIP and ROS in LF death signaling, we document a novel mechanism of apoptosis regulation that may be exploited to decrease cytotoxicity and increase gene transfection efficiency of cationic liposomes. The American Society for Pharmacology and Experimental Therapeutics