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ß-Carotene and Lutein Protect HepG2 Human Liver Cells against Oxidant-Induced Damage^1,2,3,

Department of Food, Nutrition and Food Service Management, The University of North Carolina at Greensboro, Greensboro, North Carolina 27412-5001 ^* Carotenoids Research Unit, Beltsville Human Nutrition Research Center, USDA, Beltsville, MD 20705

Numerous epidemiological studies support a strong inverse relationship between consumption of carotenoid-rich fruits and vegetables and the incidence of some degenerative diseases. One proposed mechanism of protection by carotenoids centers on their putative antioxidant activity, although direct evidence in support of this contention is limited at the cellular level. The antioxidant potential of ß-carotene (BC) and lutein (LUT), carotenoids with or without provitamin A activity, respectively, was evaluated using the human liver cell line HepG2. Pilot studies showed that a 90-min exposure of confluent cultures to 500 µmol/L tert-butylhydroperoxide (TBHP) at 37°C significantly (P < 0.05) increased lipid peroxidation and cellular leakage of lactate dehydrogenase (LDH), and decreased the uptake of ³H--aminoisobutyric acid and ³H-2-deoxyglucose. Protein synthesis, mitochondrial activity and glucose oxidation were not affected by TBHP treatment, suggesting that the plasma membrane was the primary site of TBHP-induced damage. Overnight incubation of cultures with 1 µmol/L dl--tocopherol protected cells against oxidant-induced changes. In parallel studies, overnight incubation of HepG2 in medium containing micelles with either BC or LUT (final concentrations of 1.1 and 10.9 µmol/L, respectively), the cell content of the carotenoids increased from <0.04 to 0.32 and 3.39 nmol/mg protein, respectively. Carotenoid-loaded cells were partially or completely protected against oxidant-induced changes in lipid peroxidation, LDH release and amino acid and deoxyglucose transport. These data demonstrate that BC and LUT or their metabolites protect HepG2 cells against oxidant-induced damage and that the protective effect is independent of provitamin A activity.

KEY WORDS: • carotenoids • antioxidant • amino acid transport • glucose transport • HepG2 human cell line

¹ Preliminary reports of this research were presented at Experimental Biology 95, April 9–13, 1995, Atlanta, GA [Martin, K. R., Failla, M. L. & Smith, J. C., Jr. (1995) ß-carotene and lutein protect HepG2 human liver cells against tert-butylhydroperoxide-induced damage. FASEB J. 9: A1000 (abs.) and Experimental Biology 96, April 14–17, 1996, Washington, DC [Martin, K. R., Failla, M. L. & Smith, J. C., Jr. (1996) ß-Carotene and lutein attenuate oxidant-induced damage to plasma membrane in HepG2 human liver cells. FASEB J. 10: A732 (abs.)] and in the AIN Graduate Student Research Award Abstract Competition.

² Supported in part by the North Carolina Agricultural Experiment Station and USDA NRI 94-37207-1035.

³ The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 USC section 1734 solely to indicate this fact.

⁴ Current address: Nutritional Immunology Laboratory, USDA Human Nutrition Research Center on Aging at Tufts University, Boston, MA 02111.

⁵ To whom correspondence should be addressed.

Manuscript received 18 March 1996. Revision accepted 4 June 1996.

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