Membrane lipid fluidity affects the nitroxide radical decay of 5-doxyl stearic acids in isolated rat hepatocytes.
We investigated the effect of membrane fluidity on the nitroxide radical decay rate of 5-doxyl stearic acid in spin-labeled rat hepatocytes. The half-time (t1/2) for the EPR signal decay of 5-doxyl stearic acids incorporated into the membranes of isolated rat hepatocytes was 12 min (mean value). When spin-labeled hepatocytes were separated into membrane and cytosol fractions, the t1/2 of the membrane fraction was prolonged by more than 2 hrs. However, when the cytosolic fraction was added to the membrane fraction, the radical decay reaction recovered (t1/2 was 27 min). Incubation of hepatocytes with a stream of 95% O2 at 37 degrees C for 2 hrs prolonged t1/2 by 106% and was associated with a 18% decrease in water-soluble antioxidant content. When the measurement temperature was changed from 24 degrees C to 37 degrees C, t1/2 was shortened with a decrease in the order parameter (S). The t1/2 and S in hepatocytes treated with phosphatidylcholine (PC) were reduced by 14% and 0.008, respectively. Conversely, after treatment with phosphatidylethanolamine (PE), PC+cholesterol and PE+cholesterol, t1/2 and S increased by 14% and 0.014, 20% and 0.018 and 29% and 0.040, respectively. These findings suggest that the nitroxide radical decay of 5-doxyl stearic acids incorporated into hepatocyte membranes is mediated by the antioxidants in the cytosol fraction, and that the nitroxide radical decay rate is affected not only by water-soluble antioxidant content but also by the membrane lipid fluidity of the hepatocytes.
Shima
T
T
Third Department of Internal Medicine, Kyoto Prefectural University of Medicine, Japan.
Nakashima
T
T
Kashima
K
K
Nishikawa
H
H
D000818:Animals; D002462:Cell Membrane / Q000737:chemistry; D003497:Cyclic N-Oxides / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D005609:Free Radicals; D066298:In Vitro Techniques; D008099:Liver / Q000737:chemistry*; D008297:Male; D008560:Membrane Fluidity; D051381:Rats; D017208:Rats, Wistar; D013113:Spin Labels; D013696:Temperature
D003497:Cyclic N-Oxides; D005609:Free Radicals; D013113:Spin Labels; C005072:5-doxylstearic acid