Humic acid induced genotoxicity in human peripheral blood lymphocytes using comet and sister chromatid exchange assay.
Humic acid (HA) in well water used by the inhabitants for drinking is one of the possible etiological factors for blackfoot disease (BFD). Moreover, within BFD endemic areas cancers occur at significantly higher rates than in areas free of BFD. In this study, the genotoxic potential of HA is assessed using human peripheral blood lymphocytes. The cells were exposed to HA (0-200 microg/mL for 2 h), and the induction of DNA primary damage in cellular DNA was evaluated by single-cell gel electrophoresis (comet assay). HA-induced DNA damage was decreased by superoxide (O(2)(-)), hydrogen peroxide (H(2)O(2)), and reactive oxygen species (ROS) scavengers (superoxide dismutase, catalase, and Trolox), and nitric oxide (NO) synthase inhibitors (N(G)-nitro-l-arginine methyl ester and N(G)-methyl-l-arginine). Moreover, formamidopyrimidine-DNA glycosylase (Fpg) and endonuclease III (Endo III), known to catalyze the excision of oxidized bases, increase the amount of DNA migration in HA-treated cells. Pretreatment of the cells with both the Ca(2+)-chelator BAPTA and EGTA completely inhibited HA-induced DNA damage, indicating that HA-induced changes in Ca(2+)-homeostasis are the predominant pathways for the HA induction of genotoxicity. Furthermore, sister chromatid exchange was found in the HA-treated lymphocytes. Our findings suggest that HA can induce oxidative DNA damage and genotoxicity in human lymphocytes.
Hseu
You-Cheng
YC
Department of Cosmeceutics, China Medical University, Taichung, Taiwan.
Chen
Ssu-Ching
SC
Chen
Ya-Lai
YL
Chen
Jing-Yi
JY
Lee
Mei-Ling
ML
Lu
Fung-Jou
FJ
Wu
Fang-Yang
FY
Lai
Jim-Shoung
JS
Yang
Hsin-Ling
HL
D002374:Catalase / Q000494:pharmacology; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D002839:Chromans / Q000494:pharmacology; D020552:Comet Assay; D004249:DNA Damage / Q000187:drug effects; D043382:DNA-Formamidopyrimidine Glycosylase / Q000494:pharmacology; D043224:Deoxyribonuclease (Pyrimidine Dimer) / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D029968:Escherichia coli Proteins / Q000494:pharmacology; D006801:Humans; D006812:Humic Substances / Q000633:toxicity*; D008214:Lymphocytes / Q000187:drug effects; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D010618:Phenanthrolines / Q000494:pharmacology; D012854:Sister Chromatid Exchange; D013482:Superoxide Dismutase / Q000494:pharmacology; D014873:Water Pollutants / Q000633:toxicity*; D019323:omega-N-Methylarginine / Q000494:pharmacology
D002614:Chelating Agents; D002839:Chromans; D029968:Escherichia coli Proteins; D006812:Humic Substances; D010618:Phenanthrolines; D014873:Water Pollutants; D019323:omega-N-Methylarginine; D004533:Egtazic Acid; D002374:Catalase; D013482:Superoxide Dismutase; D043224:Deoxyribonuclease (Pyrimidine Dimer); C106403:NTH protein, E coli; D043382:DNA-Formamidopyrimidine Glycosylase; C010643:6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid; D019331:NG-Nitroarginine Methyl Ester; C025205:1,10-phenanthroline