etters

S116 Abstracts / Toxicology L

(at least in our case) and a care is to be taken to avoid false positiveor negative results when measuring toxicity and proliferation withhelp of these test methods.

Acknowledgements: Supported by GACR (grant 301/06/P047).

doi:10.1016/j.toxlet.2008.06.459

V69ES-cell derived ready-to-use cardiomyocytes: An efficient andpredictive tool to detect cardiac specific cytotoxicity

Silke Schwengberg ∗, Ralf Kettenhofen, Heribert Bohlen

Axiogenesis AG, Cologne, Germany

Standardised in vitro cardiac cell and tissue models to determinecardiac-specific cytotoxicity are in short supply. Most scientistshave to use primary preparations of cardiomyocytes. Freshlyisolated primary cardiomyocytes, although showing good physio-logical properties, are time- and cost intensive to produce, difficultto standardise, and not suitable for long-term storage.

Here we present data that murine embryonic stem cell-derivedcardiomyocytes (Cor.At®) provide a standardized, homogenous andreproducible cell system for the in vitro classification of a com-pound’s cardio-cytotoxic potential. After incubation with knowncardiotoxic compounds, effects which directly affect the viabil-ity and integrity of cardiac cells can be determined using variousendpoints: morphological changes, cell viability, and release of Tro-ponin. Troponin levels in cell culture supernatant of Cor.At® cellstreated with 1 �M Emetine for 48 h were elevated 5 times com-pared to solvent controls. Comparison of viability of Cor.At® cellswith mouse embryonic fibroblasts after treatment with known car-diotoxic compounds prove the specificity and clinical relevanceof the assay. Cor.At® cells are available ready-to-use and can bestored frozen in different formats (vials, 96 well plates, coverslips),offering a time and cost effective tool for determination of cardiacspecific cytotoxicity even for medium to high throughput screen-ings at early stages of drug development.

The use of Cor.At® cells with a disease phenotype (i.e. hyper-trophic cardiomyocytes) in the above described systems willcomplete the picture of predicted adverse cardiotoxic effects of acompound even in patients with cardiac risk factors, thus increas-ing safety- and function profiles already in pre-clinical stages ofdevelopment.

doi:10.1016/j.toxlet.2008.06.460

V70Evaluation of cellular toxicity for cisplatin, arsenic andacetaminophen in the cancer and normal cell line

Mohammad Shokrzadeh ∗, Seyyed Soheil Saeedi Saravi

Mazandaran University of Medical Sciences, Sari, Mazandaran,Islamic Republic of Iran

Keywords: Cell toxicity; Clonogenic assay; Arsenic; Cisplatin;Acetaminophen

Purposes: Cell culture is a process in which, the cells were isolatedfrom original tissue are dispersed in the liquid media and then,placed in culture plate and the cells adhere together and propagate.Today this manner is used for assessment of cell toxicity and itsmechanisms and different compounds on intracellular organs.

180S (2008) S32–S246

Methods: Clonogenic assay was used for assessment of cell tox-icity and amount of cell death after a specific time that the cellwere exposed to different compounds. Thus, IC50 in caner celllines (HePG2, SKOV3 and A549) and normal cell (LLCPK1, CHO andHGF1) was assessed after exposure to cisplatin, acetaminophen andArsenic.

Results: Results showed that, Acetaminophen has maxi-mum resistance and minimum sensitivity in CHO line withIC50 = 16.7 ± 1.06 and regarding HePG2 with IC50 = 18.6 ± 1.29. Onthe other hand, cisplatin showed minimum resistance and maxi-mum Sensitivity in HePG2 with IC50 = 0.87 ± 0.07 and HGF1 withIC50 = 1.6 ± 0.21 and at last Arsenic showed minimum resistanceand maximum sensitivity in A549 with IC50 = 4.59 ± 0.29 andLLCPK1 with IC50 = 1 ± 0.37.

Conclusion: According to the evaluated IC50, it showed differ-ences between results of sensitivity of cell lines which exposed tothe three examinated drugs (P < 0.05). Entirely, resistance in cancercell lines is lower than normal cells, therefore, the results showedthe importance of cell defensive mechanisms encounting differentsubstances, such as glutathione.

doi:10.1016/j.toxlet.2008.06.461

V71Effect of P-Glycoprotein inducers on its expression and activityin Caco-2 cells

Renata Silva 1,∗, Anabela Cordeiro-da-Silva 2, Sofia A.C. Lima 3,Félix Carvalho 1, Maria Lourdes Bastos 1, Helena Carmo 1, FernandoRemião 1

1 REQUIMTE, Toxicology Department, Faculty of Pharmacy, Universityof Porto, Porto, Portugal, 2 Biochemistry Department, Faculty ofPharmacy, University of Porto, Porto, Portugal, 3 IBMC, Institute ofMolecular and Cellular Biology, University of Porto, Porto, Portugal

P-glycoprotein (P-gp) is a membrane-bound glycoprotein belong-ing to the ATP-binding cassette transporter superfamily expressedin several organs including, liver, brain, kidney and small intestine.P-gp acts as a defence mechanism by “effluxing” drugs and toxiccompounds from the cell membrane or cytoplasm to the extracellu-lar space, thus affecting the absorption, disposition, and eliminationof different xenobiotics. Caco-2 cells have been widely accepted as areliable in vitro model for the rapid screening of the intestinal drugabsorption and excretion mediated by P-gp. In fact, when grown

on permeable supports Caco-2 cells from epithelial monolayersexpress many features characteristic of the normal human intestinesuch as spontaneous morphological and biochemical enterocyticdifferentiation.

Thus, the main objective of the present work was to characterizethe P-gp expression levels in caco-2 cells during their differentia-tion processes. To evaluate potential changes in those expressionlevels, as well as changes in P-gp activity, when exposed to several P-gp inducers, such as dexamethasone, vinblastine and rifampin werethe other objectives. Alkaline phosphatase activity was measuredto assess the cell differentiation. P-gp expression levels were deter-mined by flow cytometry, using a CD243-PE conjugated antibody.The protein transport activity was evaluated using daunorubicinas a fluorescent transported P-gp subtract. Caco-2 cell P-gp ATPaseactivity was also evaluated by colorimetric assay.

The observed results are important to characterize these cells inorder to study the induction mechanism to protect cells from toxiccompounds.

doi:10.1016/j.toxlet.2008.06.462