Skip to main content
Log in

Microcystin-LR induces toxic effects in differentiated and undifferentiated Caco-2 cells

  • Short Communication
  • Published:
Archives of Toxicology Aims and scope Submit manuscript

Abstract

Microcystins (MCs) are toxins of heptapeptidic structure produced by toxic cyanobacteria in surface eutrophic waters. MCs are known to be hepatotoxic in humans, but they are also able to induce gastrointestinal alterations, allergic reactions, irritation, and pneumonia-like symptoms. The impact of MC-LR, one of the most common cyanobacterial toxins, was studied on the Caco-2 cell line, a commonly used enterocytic model, established from a human colon carcinoma. Caco-2 cells were differentiated in order to compare the effect of MC-LR in differentiated and non-differentiated cells. They were seeded in a 96-well microtiter plate and treated with MC-LR pure standard (98% purity). The effects of different concentrations of this cyanotoxin (50, 100, 150, and 200 μM) were investigated at 24 and 48 h of exposure by morphological observation and biochemical changes (total protein content, neutral red uptake, and MTS metabolization). Differentiated Caco-2 cells were slightly more sensitive than undifferentiated cells. Moreover, toxic effects induced by MCs were higher at 48 h compared to those observed at 24 h. The most sensitive endpoint for the cell line was the reduction of total protein content. Morphological changes induced by MC-LR were reduction in the cell number and hydropic degeneration, being these alterations more evident 48 h after the exposure to MC-LR.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3

References

  • Baltrop JA, Owen TC, Cory AH, Cory JG (1991) 5-((carboxyphenyl)-3-(4, 5-dimethylthiazol)-3-(4-sulfophenyl) tetrazolium, inner salt (MTS) and related analogs of 2-(4, 5-dimethyl thiazolyl)-2, 5-diphenyltetrazolium bromide (MTT) reducing to purple water soluble formazans as cell viability indicators. Bioorg Med Chem Lett 1:611–614

    Article  Google Scholar 

  • Batista T, de Sousa G, Suput JS, Rahmani R, Suput D (2003) Microcystin-LR causes the collapse of actin filaments in primary human hepatocytes. Aquat Toxicol 65:85–91

    Article  CAS  PubMed  Google Scholar 

  • Boaru DA, Dragos N, Schirmer K (2006) Microcystin-LR induced cellular effects in mammalian and fish primary hepatocyte cultures and cell lines: a comparative study. Toxicology 218:134–148

    Article  CAS  PubMed  Google Scholar 

  • Borenfreund E, Puerner JA (1984) A simple quantitative procedure using monolayer cultures for cytotoxicity assays. J Tissue Cult Methods 9:7–9

    Article  Google Scholar 

  • Botha N, Gehringer MM, Downing T, van de Venter M, Shepard EG (2004) The role of microcystin-LR in the induction of apoptosis and oxidative stress in CaCo2 cells. Toxicon 43:85–92

    Article  CAS  PubMed  Google Scholar 

  • Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  • Carmichael WW (1992) Cyanobacteria secondary metabolites—the cyanotoxins. J Appl Bacteriol 72:445–459

    CAS  PubMed  Google Scholar 

  • Chong MWK, Gu D, Lam PKS, Yang M, Fong WF (2000) Study on the cytotoxicity of microcystin-LR on cultured cells. Chemosphere 41:143–147

    Article  CAS  PubMed  Google Scholar 

  • Dietrich D, Hoege S (2005) Guidance values for microcystins in water and cyanobacterial supplement products (blue-green algal supplements): a reasonable or misguided approach? Toxicol Appl Pharmacol 15:273–289

    Article  Google Scholar 

  • Eriksson JE, Toivola D, Meriluoto JAO, Karaki H, Han YG, Hartshorne D (1990) Hepatocyte deformation induced by cyanobacterial toxins reflects inhibition of protein phosphatases. Biochem Biophys Res Commun 173:1347–1353

    Article  CAS  PubMed  Google Scholar 

  • Fastner J, Heinze R, Humpage AR, Mischke U, Eaglesham GK, Chorus I (2003) Cylindrospermopsin occurrence in two German lakes and preliminary assessment of toxicity and toxin production of Cylindrospermopsis raciborskii (Cyanobacteria) isolates. Toxicon 42:313–321

    Article  CAS  PubMed  Google Scholar 

  • Fischer WJ, Altheimer S, Cattori V, Meier PJ, Dietrich DR, Hagenbuch B (2005) Organic anion transporting polypeptides expressed in liver and brain mediate uptake of microcystin. Toxicol Appl Pharmacol 203:257–263

    Article  CAS  PubMed  Google Scholar 

  • Fladmark KE, Serres MH, Larsen NL, Yasumoto T, Aune T, Doskeland SO (1998) Sensitive detection of apoptogenic toxins in suspension cultures of rat and salmon hepatocytes. Toxicon 36:1101–1114

    Article  CAS  PubMed  Google Scholar 

  • Hilgers AR, Conradi RA, Burton PS (1990) Caco-2 cell monolayers as a model for drug transport across the intestinal mucosa. Pharm Res 7:902–910

    Article  CAS  PubMed  Google Scholar 

  • Hoeger SJ, Hitzfeld BC, Dietrich DR (2005) Occurrence and elimination of cyanobacterial toxins in drinking water treatment plants. Toxicol Appl Pharmacol 203:231–242

    Article  CAS  PubMed  Google Scholar 

  • International Agency for Research on Cancer. http://www.iarc.fr

  • Li L, Xie P, Chen J (2005) In vivo studies on toxin accumulation in liver and ultrastructural changes of hepatocytes of the phytoplanktivorous bighead carp i.p.-injected with extracted microcystins. Toxicon 46:533–545

    Article  CAS  PubMed  Google Scholar 

  • Magalhaes VF, Marinho MM, Domingos P, Oliveira AC, Costa SM, Azevedo LO, Azevedo SMFO (2003) Microcystins (cyanobacteria hepatotoxins) bioaccumulation in fish and crustaceans from Sepetiba Bay (Brasil, RJ). Toxicon 42:289–295

    Article  CAS  PubMed  Google Scholar 

  • Pflugmacher S (2004) Promotion of oxidative stress in C. demersum due to exposure to cyanobacterial toxin. Aquat Toxicol 3:169–178

    Article  Google Scholar 

  • Pichardo S, Jos A, Zurita JL, Salguero M, Camean AM, Repetto G (2005) The use of the fish cell lines RTG-2 and PLHC-1 to compare the toxic effects produced by Microcystins LR and RR. Toxicol In Vitro 19:865–873

    Article  CAS  PubMed  Google Scholar 

  • Pichardo S, Jos A, Zurita JL, Salguero M, Camean AM, Repetto G (2007) Acute and subacute toxic effects produced by microcystin-YR on the fish cell lines RTG-2 and PLHC-1. Toxicol In Vitro 21:1460–1467

    Article  CAS  PubMed  Google Scholar 

  • Sai Y, Kaneko Y, Ito S, Mitsuoka K, Kato Y, Tamai I, Artursson P, Tsuji A (2006) Predominant contribution of organic anion transporting polypeptide OATP-B (OATP2B1) to apical uptake of estrone-3-sulfate by human intestinal Caco-2 cells. Drug Metab Dispos 34:1423–1431

    Article  CAS  PubMed  Google Scholar 

  • Sambuy Y, De Angelis I, Ranaldi G, Scarino ML, Stammati A, Zucco F (2005) The Caco-2 cell line as a model of the intestinal barrier: influence of cells and culture-related factors on Caco-2 cell functional characteristics. Cell Biol Toxicol 21:1–26

    Article  CAS  PubMed  Google Scholar 

  • Teneva I, Dzhambazov B, Koleva L, Mladenov R, Schirmer K (2005) Toxic potential of five freshwater Phormidium species (Cyanoprokaryota). Toxicon 45:711–725

    Article  CAS  PubMed  Google Scholar 

  • Zegura B, Volčič M, Lah TT, Filipič M (2008) Different sensitivities of human colon adenocarcinoma (CaCo-2), astrocytoma (IPDDC-A2) and lymphoblastoid (NCNC) cell lines to microcystin-LR induced reactive oxygen species and DNA damage. Toxicon 52:518–525

    Article  CAS  PubMed  Google Scholar 

  • Zhan L, Sakamoto H, Sakuraba M, Wu DS, Zhang LS, Suzuki T, Hayashi M, Honma M (2004) Genotoxicity of microcystin-LR in human lymphoblastoid TK6 cells. Mutat Res 557:1–6

    CAS  PubMed  Google Scholar 

  • Zhao M, Xie S, Zhu X, Yang Y, Gan N, Song L (2006) Effect of dietary cianobacteria on growth and accumulation of microcystins in Nile tilapia (Oreochromis niloticus). Aquaculture 261:960–966

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The authors wish to thank the Spanish CICYT (AGL2006-06523/ALI) for the financial support for this study and the Cell Culture Service of Centro de Innovación, Tecnología e Innovación de la Universidad de Sevilla (CITIUS) for providing technical assistance.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Silvia Pichardo.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Puerto, M., Pichardo, S., Jos, Á. et al. Microcystin-LR induces toxic effects in differentiated and undifferentiated Caco-2 cells. Arch Toxicol 84, 405–410 (2010). https://doi.org/10.1007/s00204-010-0513-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00204-010-0513-0

Keywords

Navigation