Occurrence of cholesterol 7α- and 7β-hydroperoxides in rat skin as aging markers

Ozawa, Naok̆i; Yamazaki, Shinji; Chiba, Koji; Aoyama, H.; Tomisawa, Hiroki; Tateishi, Mitsuru; Watabe, Tadashi

doi:10.1016/0006-291x(91)91805-m

Cited by 36 publications

(20 citation statements)

References 13 publications

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“…We have demonstrated that 73-OOH-Chol exists in human atherosclerotic lesions; it has been identified previously (28) in vivo in rat skin and was observed to increase with age. It has been shown to be an intermediate in the nonenzymatic oxidation of cholesterol that leads to the formation of 7,-hydroxycholesterol, 7a-hydroxycholesterol, and 7-ketocholesterol (29,30), all of which have been observed in vivo (31).…”

mentioning

confidence: 88%

7 beta-hydroperoxycholest-5-en-3 beta-ol, a component of human atherosclerotic lesions, is the primary cytotoxin of oxidized human low density lipoprotein.

Chisolm

Irwin

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

193

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Modification of low density lipoprotein (LDL) by free radical oidat renders this molecular complex cytotoxic. Oxidized lipoproteins exist in vivo In atherosclerotic lesions and in the plasma of diabetic i , suggesting that Upoprotein-induced tissue damage may occur in cen diseases. We undertook purifiation and Identicati of the major cytotoxin In oxidized LDL. The lipid extract from oxidized LDL was subjected to multiple HPLC separations, and the fractions were assayed for cytotoxicity. Mass spectrometry and nuclear magnetic resonance Identified the purified toxin as 7p-hydroperoxycholest-5-en-3I3-ol Vascular cells are susceptible to the toxic effects of low density lipoprotein (LDL) or very low density lipoprotein (VLDL) after modification of the lipoproteins by free radical oxidation (1-4). Oxidation of LDL leads to cytotoxin formation whether oxidation is mediated by lipoxygenases (5), metal ions (6), or ultraviolet irradiation (7) in cell-free systems or by the action of free radicals from cultured endothelial cells (8), vascular smooth muscle cells (8), neutrophils (9), or stimulated human monocytes (10). The cytotoxic moiety of oxidized LDL (oxLDL) is extractable with organic solvents (4), and numerous candidate substances could be proposed to explain its toxic action (11).Oxidized lipoproteins occur in vivo in vascular lesions (12)(13)(14) and in plasma of certain diabetic subjects (15). OxLDL has been proposed to play a causal role in atherosclerosis (11,(16)(17)(18) is an important step toward testing evolving theories of atherogenesis that include lipoprotein oxidation (11,(16)(17)(18) and vascular injury (20,21) as putative early events.by dialysis against 2-6 /uM cupric sulfate for various times (23). OxLDL preparations were dialyzed against 0.15 M NaCl/0.5 mM EDTA, pH 8.5, to remove cupric ions. Relative oxidation, measured as thiobarbituric acid reactivity (6,24), was equivalent to 4-6 nmol of malondialdehyde (MDA) per mg of LDL cholesterol.Human foreskin fibroblasts were plated in a 1:1 (vol/vol) mixture of Dulbecco's minimal essential medium and Ham's F-12 medium (DME/F-12) supplemented with 5% (vol/vol) fetal bovine serum (6) Aliquots of5 mg ofnative LDL or oxLDL were lyophilized overnight, and the lipid was extracted with 5 ml of acetone. The mixture was sonicated for 10-20 sec, mixed for 1 min, and allowed to stand for 20 min. After centrifugation at 1000x g for 10-20 min, the residue was reextracted twice, as above. Preliminary experiments revealed that the recovery of the cytotoxic activity by this extraction procedure was equivalent or superior to other means, including chloroform/ methanol extraction ofaqueous lipoprotein solutions. Pooled extracts were dried under nitrogen and redissolved in isopropanol/acetonitrile, 1:1 (vol/vol). After centrifugation for 5 min at 1000 x g, the supernatant was analyzed by reversephase HPLC (Waters 1LBondapak C1s preparative column).The solvent gradient for elution consisted of water/ acetonitrile, 1:1 (vol/vol), which was increased over 5 ...

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mentioning

confidence: 88%

7 beta-hydroperoxycholest-5-en-3 beta-ol, a component of human atherosclerotic lesions, is the primary cytotoxin of oxidized human low density lipoprotein.

Chisolm

Irwin

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

193

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“…Our previous study using a high-performance liquid chromatography (HPLC)-chemiluminescence detector system demonstrated that cholesterol 7␣-and 7␤-hydroperoxides accumulate linearly with aging in rats from 1-45 weeks after birth, strongly suggesting the dermal steroid hydroperoxides to be good aging markers in these animals (6). However, rat livers contained no detectable levels of cholesterol 7-hydroperoxides even those from 45-week-old animals.…”

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confidence: 99%

Subunit Ya-specific Glutathione Peroxidase Activity toward Cholesterol 7-Hydroperoxides of Glutathione S-Transferases in Cytosols from Rat Liver and Skin

Hiratsuka

Yamane

Yamazaki³

et al. 1997

Journal of Biological Chemistry

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Dermal 7␣-and 7␤-hydroperoxycholest-5-en-3␤-ols (cholesterol 7␣-and 7␤-hydroperoxides), regarded as good aging markers in the rat (Ozawa, N., Yamazaki, S., Chiba, K., Aoyama, H., Tomisawa, H., Tateishi, M., and Watabe, T. (1991) Biochem. Biophys. Res. Commun. 178, 242-247), were reduced in the presence of glutathione (GSH) with concomitant formation of GSSG by cytosol from rat liver in which no detectable level of the hydroperoxides had been demonstrated to occur. The GSH peroxidase (GSH Px) activity toward the toxic steroid hydroperoxides was exerted to almost the same extent by both Alpha-class GSH S-transferases (GSTs), Ya-Ya and Ya-Yc, and by selenium-containing GSH Px (Se-GSH Px) in rat liver cytosol. None of three Mu-class GSTs, Yb1-Yb1, Yb1-Yb2, and Yb2-Yb2, and a Theta-class GST, Yrs-Yrs, from rat liver and a Pi-class GST, Yp-Yp, from rat kidney showed any appreciable GSH Px activity toward the hydroperoxides. The subunit Ya-bearing GSTs and Se-GSH Px purified from rat liver cytosol showed marked differences in apparent specific activity toward the cholesterol hydroperoxides (GSTs Ya-Ya > Ya-Yc > > Se-GSH Px). However, a kinetic study indicated that Se-GSH Px had a higher affinity for steroid hydroperoxides than did the GSTs, so that Se-GSH Px could catalyze the reduction of lower concentrations of cholesterol 7-hydroperoxides with approximately equal V max /K m values to those by the GSTs. Rat skin had no GST bearing the subunit Ya but contained only a very low concentration of Se-GSH Px, possibly resulting in the accumulation of cholesterol 7-hydroperoxides in the skin but not in the liver. From rat skin cytosol, GSTs Yc-Yc, Yb1-Yb1, Yb1-Yb2, Yb2-Yb2, and Yp-Yp were isolated, purified to homogeneity, and identified with the corresponding GSTs from liver and kidney. The GSTs accounted for 0.23% of total skin cytosolic protein, and the most abundant isoform of skin GSTs was Yb2-Yb2, followed by Yc-Yc, Yp-Yp, Yb1-Yb1, and Yb1-Yb2 in decreasing order.5␣-Hydroperoxycholest-6-en-3␤-ol (cholesterol 5␣-hydroperoxide) 1 and 7␣-hydroperoxycholest-5-en-3␤-ol (cholesterol 7␣-hydroperoxide) are intrinsic mutagens to Salmonella typhimurium TA 1537 (1). Cholesterol 7␣-hydroperoxide and its isomer, cholesterol 7␤-hydroperoxide, a cytotoxic steroid to human foreskin fibroblasts (2), are normal but very minor components of human serum lipoproteins (3) and increase upon oxidation of low density lipoprotein isolated from human sera (2, 4, 5).Our previous study using a high-performance liquid chromatography (HPLC)-chemiluminescence detector system demonstrated that cholesterol 7␣-and 7␤-hydroperoxides accumulate linearly with aging in rats from 1-45 weeks after birth, strongly suggesting the dermal steroid hydroperoxides to be good aging markers in these animals (6). However, rat livers contained no detectable levels of cholesterol 7-hydroperoxides even those from 45-week-old animals. In the present study on the mechanism of steroid hydroperoxide accumulation in the skin, we gave attention to glutathione (GSH)-dependent re...

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“…These cholesterol hydroperoxides are toxic and are aging markers in the rat skin (Ozawa et al 1991). Hiratsuka et al (1997) reported that in the rat skin Ya GSTs were below detection and Se-GSH Px were present at very low levels which in all likelihood leads to high levels of the toxic cholesterol 7-hydroperoxides.…”

Section: Xenobiotica-metabolizing Enzymes In the Rat Skinmentioning

confidence: 99%

Xenobiotica-metabolizing enzymes in the skin of rat, mouse, pig, guinea pig, man, and in human skin models

2018

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Studies on the metabolic fate of medical drugs, skin care products, cosmetics and other chemicals intentionally or accidently applied to the human skin have become increasingly important in order to ascertain pharmacological effectiveness and to avoid toxicities. The use of freshly excised human skin for experimental investigations meets with ethical and practical limitations. Hence information on xenobiotic-metabolizing enzymes (XME) in the experimental systems available for pertinent studies compared with native human skin has become crucial. This review collects available information of which—taken with great caution because of the still very limited data—the most salient points are: in the skin of all animal species and skin-derived in vitro systems considered in this review cytochrome P450 (CYP)-dependent monooxygenase activities (largely responsible for initiating xenobiotica metabolism in the organ which provides most of the xenobiotica metabolism of the mammalian organism, the liver) are very low to undetectable. Quite likely other oxidative enzymes [e.g. flavin monooxygenase, COX (cooxidation by prostaglandin synthase)] will turn out to be much more important for the oxidative xenobiotic metabolism in the skin. Moreover, conjugating enzyme activities such as glutathione transferases and glucuronosyltransferases are much higher than the oxidative CYP activities. Since these conjugating enzymes are predominantly detoxifying, the skin appears to be predominantly protected against CYP-generated reactive metabolites. The following recommendations for the use of experimental animal species or human skin in vitro models may tentatively be derived from the information available to date: for dermal absorption and for skin irritation esterase activity is of special importance which in pig skin, some human cell lines and reconstructed skin models appears reasonably close to native human skin. With respect to genotoxicity and sensitization reactive-metabolite-reducing XME in primary human keratinocytes and several reconstructed human skin models appear reasonably close to human skin. For a more detailed delineation and discussion of the severe limitations see the Conclusions section in the end of this review.

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Occurrence of cholesterol 7α- and 7β-hydroperoxides in rat skin as aging markers

Cited by 36 publications

References 13 publications

7 beta-hydroperoxycholest-5-en-3 beta-ol, a component of human atherosclerotic lesions, is the primary cytotoxin of oxidized human low density lipoprotein.

7 beta-hydroperoxycholest-5-en-3 beta-ol, a component of human atherosclerotic lesions, is the primary cytotoxin of oxidized human low density lipoprotein.

Subunit Ya-specific Glutathione Peroxidase Activity toward Cholesterol 7-Hydroperoxides of Glutathione S-Transferases in Cytosols from Rat Liver and Skin

Xenobiotica-metabolizing enzymes in the skin of rat, mouse, pig, guinea pig, man, and in human skin models

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