Ethinylestradiol administration selectively alters liver sinusoidal membrane lipid fluidity and protein composition

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Abstract
Administration of high-dose ethinylestradiol to rats decreases bile flow, Na,K-ATPase specific activity, and liver plasma membrane fluidity. By use of highly purified sinusoidal and bile canalicular membrane fractions, the effect of ethinylestradiol administration on the protein and lipid composition and fluidity of plasma membrane fractions was examined. In sinusoidal fractions, ethinylestradiol (EE) administration decreased Na,K-ATPase activity (32%) and increased activities of alkaline phosphatase (254%), Mg2+-ATPase (155%), and a 160-kDa polypeptide (10-fold). Steady-state and dynamic fluorescence polarization was used to study membrane lipid structure. Steady-state polarization of diphenylhexatriene (DPH) was significantly higher in canalicular compared to sinusoidal membrane fractions. Ethinylestradiol (5 mg/kg per day for 5 days) selectively increased sinusoidal polarization values. Similar changes were demonstrated with the probes 2- and 12-anthroyloxystearate. Time-resolved fluorescence polarization measurements indicated that EE administration for 5 days did not change DPH lifetime but increased the order component (r.infin.) and decreased the rotation rate (R). However, 1 and 3 days after EE administration and with low doses (10-100 .mu.g/kg per day for 5 days) the Na,K-ATPase, bile flow, and order component were altered, but the rotation rate was unchanged. Vesicles prepared from total sinusoidal membrane lipids of EE-treated rats, as well as phospholipid vesicles, demonstrated increased DPH polarization, as did intact plasma membrane fractions. Liver plasma membrane fractions showed no change in free cholesterol or cholesterol/phospholipid molar ratios, while esterified cholesterol content was increased with high-dose but not low-dose ethinylestradiol. High-dose ethinylestradiol treatment produced minor changes in phospholipid polar head groups and fatty acids; however, neither the sphingomyelin/phosphatidylcholine ratio nor the percent of saturated fatty acids was altered. In summary, the sinusoidal layer plasma membrane order component was selectively increased with low- and high-dose ethinylestradiol treatment. This structural change is due to changes in polar rather than neutral lipid composition. These results indicate estrogens may selectively alter plasma membrane lipid and protein domains, and these changes may be integral to the pathogenesis of intrahepatic cholestasis.