Ellagic acid, a natural polyphenol protects rat peripheral blood lymphocytes against nicotine-induced cellular and DNA damage in vitro: with the comparison of N-acetylcysteine.
The present work is aimed at evaluating the protective effect of ellagic acid (EA), a natural polyphenolic compound that is widely distributed in fruits and nuts against nicotine-induced toxicity in rat peripheral blood lymphocytes. The effect of EA against nicotine toxicity was compared with N-acetylcysteine (NAC), a well-known antioxidant. Lymphocytes were exposed to nicotine at the doses of 0.125, 0.25, 0.5, 1, 2, 3 and 4 mM for 1h in culture media. Thiobarbituric acid reactive substances (TBARS), a lipid peroxidative marker and reduced glutathione (GSH), as indicative of endogenous antioxidant status were analyzed to fix the optimum dose. The lowest concentration eliciting significant damage was 1 mM nicotine and maximum damage was observed with 3 mM concentration, as evidenced by increased levels of TBARS and decreased levels of GSH. Hence, the test concentration was fixed at 3 mM nicotine. To establish most effective protective support we used five different concentrations of EA (10, 50, 100, 150 and 300 microM) against 3 mM nicotine. A dose-dependent inhibitory effect was observed with all doses of EA. Maximum protection was observed at the dose of 100 microM EA. So, 100 microM dose was used for further studies. We have tested five different concentrations of NAC-0.25, 0.5, 1, 2 and 4 mM to elucidate the optimum protective dose against nicotine toxicity. One millimolar NAC showed a significant protection against nicotine toxicity. Protective effect of EA against nicotine toxicity was elucidated by analyzing the lipid peroxidative index, viz., TBARS, hydroperoxides (HP) and endogenous antioxidant status, viz., superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), reduced glutathione (GSH), Vitamins A, E and C. DNA damage and repair were assessed by using alkaline single-cell microgel electrophoresis (Comet assay) and micronucleus assay. There was a significant increase in the levels of lipid peroxidative index, severity in DNA damage and micronuclei number in nicotine-treated group, which was positively modulated by EA treatment. Antioxidant status was significantly depleted in nicotine-treated group, which was effectively restored by EA treatment. The protection of EA against nicotine toxicity was equally effective to that of NAC. EA and NAC treatment alone did not produce any damage to the normal lymphocytes at their effective doses. These findings suggest the potential use and benefit of EA as a modifier of nicotine-induced genotoxicity.
Sudheer
Adluri Ram
AR
Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalainagar 608 002, Tamil Nadu, India. biosudheer99@yahoo.co.in
Muthukumaran
Shanmugavelu
S
Devipriya
Nagarajan
N
Menon
Venugopal Padmanabhan
VP
D000111:Acetylcysteine / Q000494:pharmacology*; D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D002469:Cell Separation; D020552:Comet Assay; D004249:DNA Damage / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D004610:Ellagic Acid / Q000494:pharmacology*; D005419:Flavonoids / Q000494:pharmacology*; D016166:Free Radical Scavengers / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism; D066298:In Vitro Techniques; D015227:Lipid Peroxidation / Q000187:drug effects; D008214:Lymphocytes / Q000187:drug effects*; D008297:Male; D015162:Micronucleus Tests; D009538:Nicotine / Q000037:antagonists & inhibitors* / Q000633:toxicity*; D018722:Nicotinic Agonists / Q000633:toxicity*; D010636:Phenols / Q000494:pharmacology*; D059808:Polyphenols; D051381:Rats; D017392:Thiobarbituric Acid Reactive Substances / Q000378:metabolism
D000975:Antioxidants; D005419:Flavonoids; D016166:Free Radical Scavengers; D018722:Nicotinic Agonists; D010636:Phenols; D059808:Polyphenols; D017392:Thiobarbituric Acid Reactive Substances; D004610:Ellagic Acid; D009538:Nicotine; D005978:Glutathione; D000111:Acetylcysteine