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Published on: January 30, 2018
Comparative study of Domoic Acid and Okadaic Acid induced-chromosomal abnormalities in the Caco-2 cell line
Pinto-Silva Carvalho1, R Catian, Serge Moukha
1Department of Environmental Engineering, Toxicological Laboratory of the Federal University of Santa Catarina, Brazil Campus Universitário, Trindade 88040-900, Brazil.
Abstract:
Okadaic Acid (OA) the major diarrheic shellfish poisoning (DSP) toxin is known as a tumor promoter and seems likely implicated in the genesis of digestive cancer. Little is known regarding genotoxicity and carcinogenicity of Domoic Acid (DA), the major Amnesic Shellfish Poisoning (ASP) toxin. Both OA and DA occur in seafood and are of human health concerns. Micronuclei (MN) arise from abnormalities in nuclear division during mitosis due to a failure of the mitotic spindle or by complex chromosomal configurations that pose problems during anaphase. In order to evaluate the ability of okadaic acid (OA) and domoic acid (DA) to induce DNA damage we performed the micronucleus assay using the Caco-2 cell line. To discriminate between a clastogenic or aneugenic effect of OA and DA, the micronucleus assay was conducted by cytokinesis-block micronucleus assay using cytochalasin B with Giemsa staining and/or acridine orange staining, in parallel to fluorescence in situ hybridization (FISH) using a concentrated human pan-centromeric chromosome paint probe. Our results showed that OA and DA significantly increased the frequency of MN in Caco-2 cells. The MN caused by OA are found in mononucleated cells and binucleated cells, whereas those caused by DA are mainly in binucleated cells. The results of FISH analysis showed that OA induced centromere-positive micronuclei and DA increased the percentage of MN without a centromeric signal. In conclusion, both OA and DA bear mutagenic potential as revealed in Caco-2 cells by induction of MN formation. Moreover, OA induced whole chromosome loss suggesting a specific aneugenic potential, whereas DA seems simply clastogenic. At present, one cannot rule out possible DNA damage of intestinal cells if concentrations studied are reached in vivo, since this may happen with concentrations of toxins just below regulatory limits in case of frequent consumption of contaminated shell fishes.
Insights
Okadaic acid (OA) and domoic acid (DA), toxins found in seafood, can cause DNA damage and form micronuclei in intestinal cells. OA shows aneugenic potential, while DA appears clastogenic, raising concerns about frequent consumption of contaminated shellfish.
Area of Science:
- Toxicology
- Genetics
- Cell Biology
Background:
- Okadaic acid (OA), a diarrheic shellfish poisoning (DSP) toxin, is a known tumor promoter linked to digestive cancers.
- Domoic acid (DA), an amnesic shellfish poisoning (ASP) toxin, has largely uncharacterized genotoxicity and carcinogenicity.
- Both OA and DA are prevalent in seafood, posing significant human health risks.
Purpose of the Study:
- To investigate the DNA damaging potential of OA and DA using the micronucleus assay in Caco-2 cells.
- To differentiate between clastogenic (chromosome breakage) and aneugenic (chromosome loss) effects induced by OA and DA.
Main Methods:
- Micronucleus (MN) assay in Caco-2 cells, utilizing cytokinesis-block method with cytochalasin B.
- Staining techniques including Giemsa and acridine orange.
- Fluorescence in situ hybridization (FISH) with a pan-centromeric probe to analyze MN characteristics.
Main Results:
- Both OA and DA significantly elevated MN frequency in Caco-2 cells.
- OA-induced MN were observed in both mononucleated and binucleated cells, with FISH indicating centromere-positive signals (aneugenic).
- DA-induced MN were predominantly in binucleated cells, with FISH showing a higher percentage of centromere-negative signals (clastogenic).
Conclusions:
- OA and DA exhibit mutagenic potential in intestinal Caco-2 cells, evidenced by MN formation.
- OA demonstrates aneugenic effects (whole chromosome loss), while DA shows clastogenic effects (chromosome breakage).
- Potential DNA damage to intestinal cells from OA and DA at concentrations near regulatory limits warrants further investigation, especially with frequent seafood consumption.
