Faculty Collaboration Database

Lipid peroxidation in photodynamically stressed mammalian cells: use of cholesterol hydroperoxides as mechanistic reporters. Free Radic Biol Med 1997;23(1):57-68

Date

01/01/1997

Pubmed ID

9165297

DOI

10.1016/s0891-5849(96)00587-4

Scopus ID

2-s2.0-17144467109 (requires institutional sign-in at Scopus site)   51 Citations

Abstract

Photodynamic action of merocyanine 540, an antileukemic sensitizing dye, on murine L1210 cells results in the formation of lipid hydroperoxides and loss of cell viability. High-performance liquid chromatography with mercury cathode electrochemical detection was used for determining lipid oxidation products, including the following cholesterol-derived hydroperoxides: 5 alpha-OOH, 6 alpha-OOH, 6 beta-OOH, and unresolved 7 alpha, 7 beta-OOH. Among these species, 5 alpha-, 6 alpha-, and 6 beta-OOH (singlet oxygen adducts) were predominant in the early stages of photooxidation, whereas 7 alpha- and 7 beta-OOH (products of free radical reactions) became so after prolonged irradiation or during dark incubation after exposure to a light dose. These mechanistic changes were studied in a unique way by monitoring shifts in the peroxide ratio, i.e., 7-OOH/5 alpha-OOH, or 7-OOH/6-OOH. When cells (10(7)/ml) were exposed to a visible light fluence of 0.6 J/cm2 in the presence of 10 microM merocyanine 540, 7-OOH/5 alpha-OOH increased by approximately 100% after 2 h of dark incubation at 37 degrees C. The increase was much larger (approximately 250%) when cells were photooxidized after treatment with 1 microM ferric-8-hydroxyquinoline, a lipophilic iron donor, whereas no increase was observed when cells were pretreated with 100 microM desferrioxamine, an avid iron chelator/redox inhibitor. Correspondingly, postirradiation formation of thiobarbituric acid-reactive material was markedly enhanced by ferric-8-hydroxyquinoline and suppressed by desferrioxamine, as was the extent of cell killing. When added to cells after a light dose, chain-breaking antioxidants such as butylated hydroxytoluene and alpha-tocopherol strongly protected against cell killing and slowed the increase in 7-OOH/5 alpha-OOH ratio. It is apparent from these results that (1) the 7-OOH/5 alpha-OOH or 7-OOH/6-OOH ratio can be used as a highly sensitive index of singlet oxygen vs. free radical dominance in photodynamically stressed cells; and (2) that postirradiation chain peroxidation plays an important role in photodynamically initiated cell killing.

Author List

Geiger PG, Korytowski W, Lin F, Girotti AW

Author

Albert W. Girotti PhD Adjunct Professor in the Biochemistry department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Animals
Butylated Hydroxytoluene
Cell Survival
Cholesterol
Chromatography, High Pressure Liquid
Deferoxamine
Free Radicals
Iron Compounds
Leukemia L1210
Light
Lipid Peroxidation
Lipid Peroxides
Mice
Photosensitizing Agents
Pyrimidinones
Reactive Oxygen Species
Thiobarbituric Acid Reactive Substances
Tumor Cells, Cultured