Antioxidant Therapy: Rheumatoid Diseases, Diseases of the Eye 187

20.1 OXYGEN RADICALS AND RHEUMATOID DISEASE Barry Halliwell Department of Biochemistry, King's College, Strand, London WC2R 2LS, UK

My group is especially interested in the devel- opment of assays that can be used to study the rate of free radical reactions in vivo, especially in humans. Assays involving measure- ment of products of free radical attack upon uric acid, salicylate and the L- and D-isomers of phenylalanine will be described. "Finger- printing" of oxidative damage to DNA by using gas chromatography-mass spectrometry with selected ion monitoring will also be discussed. Rheumatoid arthritis is an especially-useful disease in which to evaluate putative markers of free radical damage, since it is possibleto sample directly from the site of increased oxidant generation and also to vary the rate of oxidant generation in vivo over short time periods. Some representative results will be presented.

RELATIONSHIPS BETWEEN EYE LENS PROTEIN OXIDATION, AGING, NUTRITION, CATARACT AND PROTEOLYSIS Allen Taylor, Ruth D. Lipman, Koko Murakami, and Jessica Jahngen Hodge USDA Human Nutrition Research Center on Aging at Tufts University, 711 Washington St., Boston, MA 02111, U.S.A.

The solid mass of the eye lens is proteins called crystallins. Crystallins must remain intact to transmit and focus light on the retina. Upon aging and/or (photo)oxidative stress, lens proteins oxidize, aggregate, and precipitate in opacities--or cataracts. In many cell types proteases remove damaged proteins, in part, via the highly selective, efficient ubiquitin/ATP-dependent proteolytic pathway. An important component of this system is the lens high molecular weight protease--or proteasome. Proteasome was isolated, and oxidative damage to crystallin was performed as described (Free Rad. [1990] 86217-222). Rates of proteolysis of

Co-irradiated alpha crystallin were 20>10>5>0 krad. Hydrolysis of these substrates by purified proteasome did not require energy. However, ATP- dependence and formation of ubiquitin- crystallin conjugates were detected in a bovine lens epithelial supernatant. Support: USDA, contract #53-3K06-5-I0 and Hoffmann La-Roche Inc.

20.2

20.3 OXIDANTS AND ANTIOXIDANTS IN RHEUMATIC DISEASES Ingrid Emerit Institut biomedical des Cordeliers, CNRS and University Paris VI, France

Rheumatic disorders include rheumatoid arthritis (RA), systemic lupus erythematosus, progressive systemic sclerosis, dermatomyositis and periarteritis nodosa. RA, the most frequent of these diseases, is also the most studied with respect to oxygen free radical generation, antioxidant tissue levels and possibilities of antioxidant treatment. The literature on these topics is reviewed and data from the author's laboratory are presented. Chromosome damaging material, so called clastogenic factors (CF) are released by cells in response to oxidative stress. Lipid peroxidation products and unconventional inosine nucleotides are components of these CF, which may be isolated from plasma, synovial fluid and cell culture supernatants. CF are not only at the origin of chromosome damage and DNA antibodies in these diseases, but also stimulate superoxide production by inflammatory cells. This leads to an autosustained process, which can be interrupt- ed by superoxide dismutase and possibly by other antioxidants.

INFLUENCES OF DISEASE MODIFYING ANTI RREUMATIC DRUGS ON SUPEROXIDE GENERA- TION F R O M THE HYPOXANTHINE-XANTHINE OXIDASE SYSTEM OR POLYHORPHONUCLEAR LEUKOCYTES T.Kaneko, T.Yoshikawa, H.Ichikawa, Y.Naito, H . T a k a n o , S . T a k a h a s h i a n d M.Kondo First Department of Medicine, Kyoto Prefectural University of Medicine, Kyoto 602, Japan

The effects of disease modifying anti rheumatic drugs. (DMARDs) On superoxide re±ease from human polymorphonuclear leukocytes (PMNs) were investigated by C y p r i d a n a luciferin analog (CLA)- dependent chemiluminescence assay. Auranofin, D-penicillamine, buci!iamine and sulphasalazopyridine had the in- hibitory effects against superoxide release from PMNs stimulated by various stimulants. Especially auranofin had the most strongest inhibition of all. Gold sodium thiomalate and lobenzarit had littIe effects on superoxide re- lease from PMNs. Furthermore, the influences of DMARDs on superoxide generation from the hypoxanthine- xanthine oxidase system were studied by an electron spin resonance assay using 5,5-dimethyl-l-pyrroline-N-oxide (DMPO) as a spln trapper. The intensity of DMPO~OOH signal generated from the hypoxanthine-xanthlne oxidase system was decreased in the presence of D- penicillamine and bucillamine. These results suggest that some DMARDs may have anti-inflammatory and anti-oxida- tive action due to scavenging super- oxide radicals or due to inhibiting superoxide production.

20.4