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Abstracts / Toxicology L

erbicide in male albino rats. Thirty male rats were divided intohree groups, 10 animals per group. The first and second groupsere administered 125 mg/kg bw or 250 mg/kg bw diuron daily by

avage for 28 days.The third group left as control. Rats were euth-nized by diethyl ether anesthesia, blood samples were obtainedsing vacutainer tubes containing EDTA for hematological evalua-ion. Liver, kidneys, lungs, brain, spleen and testis were collected,eighted and divided into two parts, one for estimation of lipid per-

xidation and antioxidant status while the other portion was fixedn neutral formalin buffer for pathological examination. Diuronxposure produced macrocytic anemia, enlarged liver in treatedroups, increased spleen and testis weight at 250 mg/kg group.ipid peroxides increased in serum, liver, testis and kidneys whilehe total antioxidants decreased in serum, spleen and kidneys.athological examination of organs revealed that at 125 mg/kg theiver showed hydropic degeneration, edema and thrombus forma-ion. The kidneys showed degeneration and swelling of the renalubular epithelium. The testes showed congestion, edema, degen-ration of spermatocytes and presence of spermatid giant cells.he spleen showed congestion, edema and extensive hemosidrosis.onclusion: Diuron produced toxic effects in blood and tissues ofale rats, in addition to increased lipid peroxidation and reduction

f total antioxidants.

oi:10.1016/j.toxlet.2012.03.790

27-22CP pesticide metabolite: A groove binder molecule

oheila Kashanian 1, Zohreh Shariati 1, Hamideh Roshanfekr 2,aliheh Paknejad 3

Razi University, Iran, 2 Islamic Azad University, Ilam Branch, Iran,Tehran University of Medical Sciences, Iran

Pesticides are known genotoxicants, as they have capacity tonteract with and damage the structure of the DNA molecule.,5,6-Trichloro-2-pyridinol (TCP) is a toxic metabolite of triclopyrerbicide and chlorpyrifos insecticide which are widely used in theorld. TCP potential health hazard is identified due to its high affin-

ty to DNA molecule. The interaction of native calf thymus DNA withCP has been investigated to find the binding mode of TCP-DNAomplex. Thus we designed the competitive binding experimentetween Hoechst and TCP for DNA to understand whether TCP is aroove binder.

Fluorescence intensities were measured using a Cary lumines-ence spectrometer, all solutions prepared using Tris–HCl bufferpH = 7.4). Concentrations of DNA and Hoechst were kept constant5 × 10−5 and 5 × 10−6 M−1, respectively) while varying the TCPoncentration from 0 to 5 × 10−4 M−1.

Hoechst fluorescence emission spectra are significantlynhanced by increasing the DNA concentration, but decreasedfter addition of TCP. Since Hoechst is a strong groove binderhich binds strongly to the DNA minor groove, this indicated that

CP is able to replace Hoechst from DNA grooves. Thus TCP is aotential groove binder molecule which can interact with DNA anday cause structural changes in DNA which leads to genotoxicity.lso the concentration which was used in these experiments isuch less than that of currently is used in agriculture and industry.

herefore, there should be more consideration in using pesticideso repel pests.

oi:10.1016/j.toxlet.2012.03.791

211S (2012) S43–S216 S175

P27-23Immortalized rat brain endothelial cells are highly resistant toparaquat toxic effect

Vânia Vilas-Boas 1, Renata Silva 2, Maria de Lourdes Bastos 2,Fernando Remião 2

1 REQUIMTE – Laboratório de Toxicologia, Portugal, 2 REQUIMTE,Portugal

This work aimed at evaluating paraquat (PQ) cytotoxicity in theimmortalized rat brain endothelial cell line, RBE4. For that purposetwo methodologies were performed: the MTT reduction and theneutral red (NR) uptake assays. The intensity of the cytotoxic effectwas determined 24 and 48 h after exposure to PQ concentrationsbetween 0.5 and 50 mM.

The NR assay was more sensitive to assess the toxic effect ofthe intermediate PQ concentrations, than the MTT assay. There-fore, for the 48 h time-point, the NR was the only assay performed.After 24 h, the NR uptake assay detects significant differences inthe percentage of viable cells at the concentration of 5 mM andabove, whilst for the MTT reduction assay those differences are onlysignificant for concentrations higher than 10 mM. The percentageof viable cells reached about 40% at the concentration of 50 mM.After 48 h, the toxic effect of PQ is highlighted in a concentration-dependent manner, reaching statistic significant from 2.5 mM PQon. Cell viability does not exceed the 7% at the maximum concen-tration tested.

These results indicate that RBE4 cell line is highly resistant to PQtoxic effect, as compared to the already published results for Caco-2 cell line [1]. This result shows the presence of specific protectivemechanisms in rat brain endothelial cell line.

Acknowledgements: FCT grant (Project PTDC/SAU-OSM/101437/2008).

[1] R. Silva, et al. In vitro study of P-glycoprotein induction asan antidotal pathway to prevent cytotoxicity in Caco-2 cells, Arch.Toxicol. 85 (April (4)) (2011) 315–326.

doi:10.1016/j.toxlet.2012.03.792

P27-25ETU fungicide metabolite: A DNA cleaver molecule

Soheila Kashanian, Zohreh Shariati, Soyebeh Askari

Razi University, Iran

Pesticides have been extensively applied in recent decadesthroughout the world. They are powerful toxicants that areabsorbed and accumulate onto the soil, plants, food, and surfacewaters, which have caused severe health hazards. Ethyle-lenethiourea (ETU) is metabolite of Ethylenebisdithiocarbamate(EBDC). ETU is hydrosoluble and has raised concern because of itsthyroid toxicity and tumorigenicity. So it seems necessary to knowabout the mechanism of its toxicity. Many genotoxicants may causeDNA strand breakage by the formation of free radicals or abasic siteswhich can result in the breakage of phosphodiester linkages withinthe DNA molecule. So, DNA cleavage activity of ETU was studied tounderstand whether it can cleave DNA.

DNA cleavage activity of ETU was studied by gel electrophoresisusing supercoiled (SC) plasmid pUC18 DNA. Thus, pUC18 plas-

mid DNA was treated with ETU in TE Buffer (pH = 8), and waseletrophoresed on agarose gel.

When circular plasmid DNA is subjected to electrophoresis, rel-atively fast migration will be observed for the intact supercoiled