Nitric oxide inhibits lipopolysaccharide-induced apoptosis in pulmonary artery endothelial cells

• Published in: American journal of physiology. Lung cellular and molecular physiology Vol. 275; no. 4; pp. 717 - L728
• Main Authors: Ceneviva, Gary D, Tzeng, Edith, Hoyt, Dale G, Yee, Emily, Gallagher, Alicia, Engelhardt, John F, Kim, Young-Myeong, Billiar, Timothy R, Watkins, Simon A, Pitt, Bruce R
• Format: Journal Article
• Language: English
• Published: United States 01.10.1998
• Subjects: Adenoviridae; Animals; Apoptosis - drug effects; Apoptosis - physiology; Caspase 3; Caspases - metabolism; Cell Nucleus - drug effects; Cell Nucleus - physiology; Cell Nucleus - ultrastructure; Cells, Cultured; Endothelium, Vascular - cytology; Endothelium, Vascular - drug effects; Endothelium, Vascular - physiology; Genetic Vectors; Humans; Lipopolysaccharides - antagonists & inhibitors; Lipopolysaccharides - pharmacology; Liver - enzymology; Mitochondria - drug effects; Mitochondria - physiology; Mitochondria - ultrastructure; Moloney murine leukemia virus; Nitric Oxide - physiology; Nitric Oxide Synthase - genetics; Nitric Oxide Synthase - metabolism; Nitric Oxide Synthase Type II; Nitrites - metabolism; omega-N-Methylarginine - pharmacology; Pulmonary Artery; Recombinant Proteins - biosynthesis; Recombinant Proteins - metabolism; Sheep; Superoxide Dismutase - metabolism; Transfection; Vacuoles - ultrastructure
• DOI: 10.1152/ajplung.1998.275.4.L717
• ISSN: 1040-0605; 0002-9513

3  Department of Pharmacology; 1  Division of Pediatric Critical Care Medicine, Department of Anesthesiology; 2  Department of Surgery; 4  Division of Pulmonary, Allergy, and Critical Care Medicine; and 6  Department of Cell Biology and Physiology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261; and 5  Department of Anatomy and Cell Biology, University of Iowa Medical Center, Iowa City, Iowa 52242 Our group recently reported that cultured sheep pulmonary artery endothelial cells (SPAECs) became resistant to lipopolysaccharide (LPS)-induced apoptosis several days after constitutive synthesis of nitric oxide (NO) after adenoviral (Ad) transfer of inducible NO synthase (iNOS) or exposure to the NO donor S -nitroso- N -acetylpenicillamine (SNAP) (E. Tzeng, Y.-M. Kim, B. R. Pitt, A. Lizonova, I. Kovesdi, and T. R. Billiar. Surgery 122: 255-263, 1997). In the present study, we confirmed this observation by establishing stable transfectants after retroviral gene transfer [replication-deficient retrovirus (DFG)] of human iNOS (DFG-iNOS) SPAECs and then used all three approaches (Ad, DFG, and SNAP) to determine underlying mechanisms of this phenomenon. Continuous endogenous production of NO in itself did not cause apoptosis as assessed by phase-contrast microscopy, nuclear morphology, and internucleosomal DNA fragmentation. Prolonged (72-96 h) synthesis of NO, however, after DFG- or replication-deficient adenovirus (Ad.CMV)-iNOS or SNAP (100 µM, 96 h) inhibited LPS-induced apoptosis. The kinetics of such protection suggested that NO may be inducing other gene products. Ad-mediated transfer of manganese superoxide dismutase (MnSOD) decreased the sensitivity of wild-type SPAECs to LPS-induced apoptosis. MnSOD, however, was not induced in an N G -monomethyl- L -arginine ( L -NMMA)-sensitive time-dependent fashion after Ad.CMV-iNOS. Other inducible genes that may be affected by NO and that may protect against potential oxidant-mediated LPS-induced apoptosis including 70-kDa heat shock protein, heme oxygenase-1, metallothionein, and Bcl-2 also were not elevated in an L -NMMA-sensitive, time-dependent fashion. Although the candidate gene product underlying NO-induced protection remains unclear, we did note that prolonged synthesis of NO inhibited LPS-induced activation of an interleukin-1 -converting enzyme-like cysteine protease (cysteine protease protein-32-like) in a dithiothreitol-sensitive fashion, suggesting that S-nitrosylation of an important downstream target of convergence of apoptotic signals may contribute to the sensitivity of SPAECs to LPS. cysteine protease protein-32-like protease; manganese superoxide dismutase; mitochondria; heme oxygenase; Bcl-2