FLOW CYTOMETRY BASED MICRONUCLEUS ASSAY FOR EVALUATION OF GENOTOXIC POTENTIAL OF 2-ACBs IN HEPATIC CELLS HepG2

Angélica Bueno Barbezan

doi:10.15392/bjrs.v7i2A.684

Authors

Angélica Bueno Barbezan

DOI:

https://doi.org/10.15392/bjrs.v7i2A.684

Keywords:

flow cytometry, 2-ACBs, food irradiation

Abstract

Food irradiation is approved for use in more than 60 countries for applications and purposes in a wide variety of foods, being an effective and safe method for preservation and long-term storage. 2-Alkylcyclobutanones (2-ACBs) are the only known radiolytic products generated from foods that contain fatty acids (Triglycerides) when irradiated. The acids analyzed in this study are palmitic and stearic, which when irradiated form 2-Dodecylcyclobutanones (2-dDCB) and 2-Tetradecylcyclobutanone (2-tDCB). Part of the 2-ACBs ingested is excreted through feces and part is deposited in adipose tissues. In vitro studies so far have been only in colon cells. The work used a human hepatoma cell line (HepG2) since the accumulation of fat in this organ is quite common. Micronucleus test was selected to evaluate possible genotoxic effects of 2-dDCB and 2-tDCB compounds when exposed to high concentrations (447, 1422 and 2235 μM) for 4 and 24 hours. Tests were performed in quadriplicates using flow cytometric analysis. None detectable genotoxic damage was observed after 4 hours of exposure to the compounds, and cytotoxic effects were only significant at the highest concentration (2235 μM) of 2-dDCB. After 24 hours of exposure, slight genotoxic damage was observed at all concentrations evaluated, and cytotoxic effects were only present when exposed to compound 2-tDCB. Although there is a genotoxic and cytotoxic effect in some of the situations tested, the two compounds predominantly induced proliferation reduction effects of this hepatic tumor cell line.

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References

FARKAS, J., EHLERMANN, D. A. E., & FARKAS, C. M. “Food Technologies: Food Irradiation. Reference Module in Food Science” – Encyclopedia Food Safety, v. 3, pp.178-186 (2014).

IAEA International Atomic Energy Agency (2017). “Gamma Irradiators for Radiation Processing”, http://www-naweb.iaea.org/napc/iachem/Brochgammairradd.pdf

ROBERTS, P. B. “Food irradiation is safe: Half a century of studies”, Radiation Physics and Chemistry. v. 105, pp. 78-82 (2014).

GADGIL, P., HACHMEISTER, K., SMITH, J. S., & KROPF, D. H. “2-Alkylcyclobutanones as Irradiation Dose Indicators in Irradiated Ground Beef Patties”. Journal Agricultural and Food Chemistry, v. 50, pp. 5746-5750 (2002).

SONG, B. S., CHOI, S. J., JIN, Y. B., PARK, J. H., KIM, J. K., BYUN, E. B., KIM, J. H., …, MARCHIONI, E. “A critical review on toxicological safety of 2-alkylcyclobutanones”. Radiation Physics and Chemistry. v. 103, pp. 188-193 (2014).

DELINCEÉ, H., & POOL-ZOBEL, B.L. “Genotoxic properties of 2-dodecylcylobutanone, a compound formed on irradiation of food containing fat.” Radiation Physical and Chemistry, v. 52, pp. 39-42 (1998).

CHINTHALAPALLY, V. R. “Do Irradiated Foods Cause or Promote Colon Cancer?” Nutrition and Cancer, v. 46, pp. 107-109 (2003).

TICE, R. R., E. AGURELL, D. ANDERSON, B. BURLINSON, A. HARTMANN, H. KOBAYASHI, Y. MIYAMAE, …, Y. F. SASAKI. “Single cell gel/comet assay: guidelines for in vitro and in vivo genetic toxicology testing”. Environmental and Molecular Mutagenesis. v. 35, pp. 206–221 (2000).

FLORES, M.; YAMAGUCHI, M.U. “Teste de micronúcleo: uma triagem para avaliação genotóxica”, Revista Saúde e Pesquisa, v.1, n.3, pp. 337-340, (2008).

HAWLEY, T. S.; HAWLEY, R. G. Flow cytometry protocols. 2. ed. pp. 434. Humana Press Inc., Totowa, New Jersey (2004).

SETÚBAL, A.M.G.; REIS, S. R. A.; ROBINSON, W. M.; BORGES-OSÓRIO, M. R. “Micronúcleo: um importante marcador biológico intermediário na prevenção do câncer bucal.” Revista Odonto Ciência- Faculdade de Odontologia/PUCRS, v.20, n.28, (2005).

I. Z. OCAMPO, P.Q.S. PASSOS, L.R. DE CARVALHO, C.A.L. DA CRUZ, N.M. ESTEVES-PEDRO, F.M. DA SILVA, O.Z. HIGA, L.A. DIAS, K. OKAZAKI, D.P. VIEIRA, “In vitro cytotoxic and genotoxic evaluation of peptides used in nuclear medicine (DOTATATE and Ubiquicidin (29-41)) in

CHO-K1 cells,” Cytotechnology. 68, pp. 2301-2310 (2016).

ESTEVES-PEDRO, N. M. “Avaliação in vitro da toxicidade de óleos essenciais da flora Latino-americana candidatos ao uso em cosméticos. 2013”. Dissertação (Mestrado) - Faculdade de Ciências Farmacêuticas da Universidade de São Paulo – pp. 42-47, http://www.teses.usp.br/teses/disponiveis/9/9139/tde-24092015-120519/pt-br.php (2016).

BRYCE, S., SHI, J., NICOLETTE, J., DIEHL, M., SONDERS, P., AVLASEVICH, S., RAJA, S., BEMIS, J. C., DERTINGER, S. D. High content flow cytometric micronucleus scoring methods is applicable to attachment cell lines. Environ Mol Mutagen. v. 51, pp 260-266. (2010).

YAMAKAGE, K.; SUI, H.; OHTA, R.; TYOIZUMI, T.; KAWAKAMI, K.; MATSUMOTO, H.; TAKAHASHI, T.; SASAKI, K.; IKEZUMI, M.; NEGISHI, S.; IZUMI, K.; TODORIKI, S.; TAKASHI, K.; FURUTA, M. “Genotoxic potencial and in vitro tumour-promoting potential of 2-dodecylcyclobutanone and 2-tetradecylcyclobutanone, two radiolytic products of fatty acids”. Mutation Research, v. 770, pp. 94-104, (2014).

BREIDBACH, A.; ULBERTH, F. “Comparative evaluation of methods for the detection of 2-alkylcyclobutanones as indicators for irradiation treatment of cashew nuts and nutmeg”. Food Chemistry, v. 201, pp. 52-58 (2016).