title: The Application of a Modified d-ROMs Test for Measurement of Oxidative Stress and Oxidized High-Density Lipoprotein.
abstract: Reactive oxygen species (ROS) are involved in the initiation and progression of atherosclerosis. ROS-derived hydroperoxides, as an indicator of ROS production, have been measured by using the diacron reactive oxygen metabolites (d-ROMs) test, which requires iron-containing transferrin in the reaction mixture. In this study we developed a modified d-ROMs test, termed the Fe-ROMs test, where iron ions were exogenously added to the reaction mixture. This modification is expected to exclude the assay variation that comes from different blood iron levels in individuals. In addition, this Fe-ROMs test was helpful for determining the class of plasma lipoproteins that are hydroperoxidized. Low-density lipoprotein/very low-density lipoprotein (LDL/VLDL) and high-density lipoprotein (HDL) were purified by use of an LDL/VLDL purification kit and the dextran sulfate-Mg2+precipitation method, respectively; their hydroperoxide contents were assessed by performing the Fe-ROMs test. The majority of the hydroperoxides were detected only in the HDL fraction, not in the LDL/VLDL. Further detailed analysis of HDLs by size-exclusion high-performance liquid chromatography revealed that the hydroperoxide-containing molecules were small-sized HDLs. Because HDL was shown to be the principal vehicle for the plasma hydroperoxides, this Fe-ROMs test is a beneficial method for the assessment of oxidized-HDL levels. Indeed, Fe-ROMs levels were strongly associated with the levels of oxidized HDL, which were determined by performing the malondialdehyde-modified HDL enzyme immunoassay. In conclusion, the Fe-ROMs test using plasma itself or the HDL fraction after dextran sulfate-Mg2+precipitation is useful to assess the functionality of HDL, because the oxidation of HDL impairs its antiatherogenic capacity.
authors.affiliation: Institute of Health Sciences, Sunstar Inc., Osaka 569-1195, Japan. fumiaki.ito@jp.sunstar.com.
authors.forename: Fumiaki
authors.initials: F
authors.lastname: Ito
authors.affiliation: Department of Rehabilitation Medicine, Kyoto Prefectural University of Medicine, Kyoto 602-8566, Japan. rinito@par.odn.ne.jp.
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Ito
authors.affiliation: Department of Pediatric Cardiology and Nephrology, Kyoto Prefectural University of Medicine, Kyoto 602-8566, Japan. suzuki@koto.kpu-m.ac.jp.
authors.forename: Chinatsu
authors.initials: C
authors.lastname: Suzuki
authors.affiliation: Department of Pediatric Cardiology and Nephrology, Kyoto Prefectural University of Medicine, Kyoto 602-8566, Japan. tomoyo@koto.kpu-m.ac.jp.
authors.forename: Tomoyo
authors.initials: T
authors.lastname: Yahata
authors.affiliation: Department of Pediatric Cardiology and Nephrology, Kyoto Prefectural University of Medicine, Kyoto 602-8566, Japan. ikeda@koto.kpu-m.ac.jp.
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Ikeda
authors.affiliation: Department of Pediatric Cardiology and Nephrology, Kyoto Prefectural University of Medicine, Kyoto 602-8566, Japan. khamaoka@koto.kpu-m.ac.jp.
authors.forename: Kenji
authors.initials: K
authors.lastname: Hamaoka
mesh_terms: D050197:Atherosclerosis / Q000097:blood / Q000378:metabolism; D015415:Biomarkers; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D006861:Hydrogen Peroxide / Q000097:blood; D008075:Lipoproteins, HDL / Q000097:blood / Q000737:chemistry / Q000378:metabolism*; D008077:Lipoproteins, LDL; D008079:Lipoproteins, VLDL; D046911:Macromolecular Substances / Q000097:blood; D055432:Metabolomics / Q000379:methods*; D008970:Molecular Weight; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000097:blood / Q000737:chemistry / Q000378:metabolism*
chemical_list: D015415:Biomarkers; D008075:Lipoproteins, HDL; D008077:Lipoproteins, LDL; D008079:Lipoproteins, VLDL; D046911:Macromolecular Substances; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide
keywords: atherosclerosis; diacron-reactive oxygen metabolites (d-ROMs) test; high-density lipoprotein (HDL); oxidative stress; oxidized high-density lipoprotein; reactive oxygen species (ROS)