[36 Antisense Oligodeoxyribonucleotides A Better Way to Inhibit Monocyte Superoxide Anion Production

By Erik A. Bey and Martha K. Cathcart

Introduction

Reduced nicotinamide-adenine dinucleotide phosphate (NADPH) oxidases are a group of plasma membrane-associated enzymes, among which the phagocytic leukocyte NADPH oxidase has been most often studied.1 ~3 During phagocytosis or exposure to other activating agonists, the NADPH oxidase of neutrophils and other professional phagocytic cells (including monocytes) catalyzes the production of superoxide anion (O2-) by the one-electron reduction of molecular oxygen. The enzyme complex uses NADPH, provided by the pentose phosphate pathway, as the electron donor.1'2'4,5 C>2~, generated by the NADPH oxidase of monocytes and neutrophils, is readily converted to other potent reactive oxidants, such as hydrogen peroxide (H2O2), hydroxyl radical (OH ), hypochlorite (OCl~), and singlet oxygen ('Oz).1'6'7 These more potent oxidants are directly responsible for killing bacterial and fungal pathogens. The production of 02~ and the more potent oxidants by these cells leads to an abrupt rise in oxygen consumption. For this reason the NADPH oxidase is often referred to as the respiratory burst oxidase.

The leukocyte NADPH oxidase complex consists of a membrane-associated Mype cytochrome ¿>553 that copurifies with a GTP-binding protein, RAP1A8;

2 R. A. Clark, J. Infect. Dis. 179 (Suppl. 2), S309 (1999).

3 T. Leto, in "Inflammation: Basic Principles and Clinical Correlates" (J. I. Gallin and R. Snyderman, eds.), p. 769. Lippincott Williams & Wilkins, Philadelphia, PA, 1999.

4 S. J. Chanock, J. el Benna, R. M. Smith, and B. M. Babior, J. Biol. Chem. 269, 24519 (1994).

6 J. M. Robinson and J. A. Badwey, Histochem. Cell Biol. 103, 163 (1995).

8 M. T. Quinn, C. A. Parkos, L. Walker, S. H. Orkin, M. C. Dinauer, and A. J. Jesaitis, Nature (London) 342, 198 (1989).

Fig. 1. (A) Components of the phagocytic NADPH oxidase lie unassembled in unactivated monocytes. The NADPH oxidase components are both membrane associated and cytosolic in unactivated monocytes. (B) NADPH oxidase components assemble on monocyte activation. After activation by appropriate stimuli, cytosolic components of the phagocytic NADPH oxidase translocate and assemble with membrane-associated NADPH oxidase components and form an active complex resulting in O2- production.

Fig. 1. (A) Components of the phagocytic NADPH oxidase lie unassembled in unactivated monocytes. The NADPH oxidase components are both membrane associated and cytosolic in unactivated monocytes. (B) NADPH oxidase components assemble on monocyte activation. After activation by appropriate stimuli, cytosolic components of the phagocytic NADPH oxidase translocate and assemble with membrane-associated NADPH oxidase components and form an active complex resulting in O2- production.

a cytosolic complex consisting of three components, pAlphox, p67ph"\ and p4Q''h"A: and a small cytosolic GTP-binding protein, Rac 1/2.4 The components of the leukocyte NADPH oxidase are unassembled in unactivated cells (Fig. 1A). In response to appropriate stimuli, the cytosolic components translocate and assemble at the membrane. The proper assembly of the NADPH oxidase components activates the oxidase and C>2_ is produced (Fig. IB).

Our laboratory previously reported that 02~ production was required for human monocyte-mediated low-density lipoprotein (LDL) oxidation9 because monocyte-mediated LDL oxidation was inhibited by superoxide dismutase (SOD), a scavenger of 02~ . However, to definitively show that 02~ was indeed required for monocyte-mediated LDL oxidation, we thought that it was necessary to perform additional experiments that did not rely on the specificity of action of SOD. Pharmacologic agents have been reported to inhibit NADPH oxidase activity, such as diphenylene iodonium (DPI) and phenylbutazone.10-12 Studies in our laboratory have shown that both of these agents dose dependently inhibit NADPH oxidase activity while also causing dose-dependent toxicity to the cells.13 As an alternative to

9 M. K. Cathcart, A. K. McNally, D. W. Morel, and G. M. Chisolm III,./. Immunol. 142,1963 (1989).

10 A. R. Cross and O. T. Jones, Biochem. J. 237, 111 (1986).

11 J. T. Hancock and O. T. Jones, Biochem. J. 242, 103 (1987).

12 D. G. Hafeman and Z. J. Lucas, J. Immunol. 123, 55 (1979).

13 V. F. Nivar, Ph.D. Thesis. Department of Regulatory Biology, Cleveland State University, Cleveland, OH, 1993.

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