Oxidation of cholesterol in synaptosomes and mitochondria isolated from rat brains.
Cholesterol and alpha-tocopherol oxidations were studied in brain subcellular fractions isolated from cerebral hemispheres of 4-month-old, male Fischer 344 rats. The fractions were suspended in buffered media (pH 7.4, 37 degrees C0 and oxidized by adding (i) ferrous iron (Fe2+) with or without ascorbate or (ii) peroxynitrite (an endogenous oxidant produced by the reaction of superoxide and nitric oxide). Treatment of subcellular fractions with Fe2+ in the presence or absence of ascorbate produced primarily 7-keto- and 7-hydroxy-cholesterols and small amounts of 5 alpha, 6 alpha-epoxycholesterol. Since brain contains high levels of ascorbate, and release of iron could result in oxysterol formation. Peroxynitrite oxidized alpha-tocopherol but not cholesterol. Hence, the toxicity of peroxynitrite or nitric oxide could not be due to cytotoxic oxysterols. When synaptosomes were incubated for 5 min in the presence of 0.5 to 2 microM Fe2+ and ascorbate, alpha-tocopherol was oxidized while cholesterol remained unchanged. Thus, alpha-tocopherol is functioning as an antioxidant, protecting cholesterol. Diethylenetriaminepentaacetic acid blocked production of oxysterols, whereas citrate, ADP and EDTA did not. A significant percentage of mitochondrial cholesterol was oxidized by treatment with Fe2+ and ascorbate. Hence, mitochondrial membrane properties dependent on cholesterol could be particularly susceptible to oxidation. The oxysterols formed were retained within the membranes of synaptosomes and mitochondria. The 7-oxysterols produced are known to be inhibitors of membrane enzymes and also can modify membrane permeability. Hence, oxysterols may plan an important role in brain tissue damage during oxidative stress.
Vatassery
G T
GT
Research Service, VA Medical Center, Minneapolis, Minnesota 55417, USA. vatas001@maroon.tc.umn.edu
Quach
H T
HT
Smith
W E
WE
Ungar
F
F
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001205:Ascorbic Acid / Q000494:pharmacology; D001894:Borohydrides / Q000378:metabolism; D001921:Brain / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology; D002784:Cholesterol / Q000031:analogs & derivatives / Q000378:metabolism*; D002849:Chromatography, Gas; D005296:Ferrous Compounds / Q000494:pharmacology; D008297:Male; D008928:Mitochondria / Q000378:metabolism*; D009566:Nitrates / Q000494:pharmacology; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D051381:Rats; D011916:Rats, Inbred F344; D013574:Synaptosomes / Q000378:metabolism*; D014810:Vitamin E / Q000378:metabolism
D000975:Antioxidants; D001894:Borohydrides; D002614:Chelating Agents; D005296:Ferrous Compounds; D009566:Nitrates; D014810:Vitamin E; C039642:peroxynitric acid; C025364:sodium borohydride; D002784:Cholesterol; D001205:Ascorbic Acid