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4 Abstracts / Toxicology

1hemical exposure-related inflammation and cancer

01-1lcohol and hepatocarcinogenesis: Interaction with other liveriseases

elmut Seitz

Salem Medical Centre, Germany

Chronic alcohol consumption may lead to alcoholic liver diseaseALD) including hepatocellular cancer (HCC). The mechanisms byhich alcohol stimulates hepatocarcinogenesis are complex and

nclude the presence of hepatic cirrhosis, oxidative stress mainlyhrough the induction of cytochrome P-4502E1 (CYP2E1), the acti-ation of acetaldehyde, epigenetic changes, the lack of retinoiccid and severe effects on intercellular signal transduction. Chroniclcohol consumption results in the induction of CYP2E1 whicheads to the generation of reactive oxygen species (ROS). As aesult lipid peroxidation occurs and lipid peroxidation productsuch as 4-hydroxynonenal may bind to DNA forming highly car-inogenic exocyclic etheno DNA-adducts. Alcohol also stimulatesepatocarcinogenesis in other inflammatory liver diseases. In non-lcoholic steatohepatitis (NASH) even small amounts of alcoholurther increase hepatic CYP2E1, deteriorate the liver disease andesults in an increased risk for HCC. Other inflammatory hepaticisease such as hepatitis B and C are also negatively influenced byhronic alcohol consumption even at smaller doses. It has been con-incingly shown that HCC occurs almost 10 years earlier in hepatitiswhen alcohol is consumed. In hepatitis C not only the progressionf fibrosis and cirrhosis is accelerated under alcohol but also the risko develop HCC is significantly enhanced. In conclusion, chroniclcohol consumption by itself is a risk factor for HCC. If alcohol isonsumed in the presence of other inflammatory liver diseases suchs hepatitis B, hepatitis C and NASH, inflammation and fibrosis aretimulated and the risk for HCC is significantly enhanced.

oi:10.1016/j.toxlet.2012.03.024

01-3he role of transforming growth factor beta signallingathways in tumour biology

arene Landström

Umeå University, Sweden

Transforming growth factor � (TGF�) is a pluripotent cytokinehich promotes epithelial cell plasticity during embryogenesis

nd tumour progression. TGF� binding to type II and type I ser-ne/threonine kinase receptors (T�RII and T�RI) leads to activationf different intracellular signalling pathways, i.e. Smad and non-mad signalling pathways, where we have shown that Smad7lays an important role for activation of the TAK1-p38 MAPK path-ay in prostate cancer cells (Landström 2010). Activation of theon-Smad signalling pathway is explained by the fact that the�RI is associated with the ubiquitin ligase tumour necrosis factoreceptor (TNFR)-associated factor 6 (TRAF6), as we recently haveeported (Sorrentino et al., 2008). Furthermore, we have recently

emonstrated that TGF�, utilize TRAF6, to promote Lys63-linkedolyubiquitination of T�RI. This event in turn is associated withNF-alpha converting enzyme (TACE)-dependent cleavage of T�RI,n a PKCz-dependent manner. We show also that the generated

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intracellular domain (ICD) of T�RI associates with the transcrip-tional co-regulator p300 to cause activation of genes involved intumour cell invasiveness, such as Snail and MMP2. Investigationof a number of different cancer cell lines demonstrated that theobserved TGF�-induced invasion of cancer cells where the T�RIICD is localized in the nucleus of cancer cells is likely to be part ofgeneral mechanism and is dependent on TACE and PKCz. Thus, ourdata reveal a novel route in which TGF� use its own receptor in thenucleus to promote transcription of pro-invasive genes and tumourinvasion.

doi:10.1016/j.toxlet.2012.03.025

S01-4Oxidative stress and inflammation: The link between COPDand lung cancer

Ian Adcock

Imperial College, London, United Kingdom

Chronic obstructive pulmonary disease (COPD) and lung can-cer are major causes of death and their close association suggestseither common susceptibilities to cigarette smoking or that abnor-malities in COPD favour the development of lung cancer beyondthe effect of smoking alone. A recent large observational studydemonstrated a high (>10-fold) increased prevalence of lung can-cer in COPD, particularly emphysematous, patients compared withnormal smokers.

Oxidative stress-induced chronic inflammation may favourtumourigenesis through increased expression of inflammatorymediators and growth factors including TGF-�, EGFR, IL-1, IL-8 andG-CSF and this inflammation may persist for decades after smokingcessation. A reduction in Nrf2 activity in COPD may enhance oxida-tive stress possibly due to reduced HDAC2 activity, which is alsolinked to glucocorticoid resistance and amplified inflammation.

TGF� and MMPs produced by epithelial cells in COPD maypromote epithelial–mesenchymal transition, leading to malignanttransformation of the respiratory epithelium. In addition, thereis impairment of double-stranded DNA repair machinery withreduced nuclear expression of Ku86 in bronchiolar epithelial cellsof patients with COPD, suggesting that oxidant-induced DNA dam-age would be more likely to result in mutations and carcinogenesis.Recent evidence also links aberrant DNA repair processes withreduced HDAC expression. Understanding the common signallingpathways involved in COPD and lung cancer may allow the devel-opment of novel treatments for COPD and also reduce the high riskof developing lung cancer in these patients.

doi:10.1016/j.toxlet.2012.03.026

S01-5Molecular mechanisms linking chronic inflammation tocancer in the intestinal mucosa

Pablo Steinberg

University of Veterinary Medicine, Hannover, Germany

For a number of years now it has been postulated that chronicinflammation, which occurs in patients with inflammatory bowel

disease, predisposes to colorectal cancer. In this presentation therole of inflammation in colon cancer induced by various compoundsincluding food contaminants/constituents will be highlighted. Inthe case of heterocyclic aromatic amines, which are formed in

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Abstracts / Toxicology

eat when cooked at high temperatures for a long time or overn open flame, and iron it has been shown that the incidencef colorectal neoplasia can be increased and its progression tonvasive cancer can be accelerated by chemical induction of inflam-

ation in rodents. When considering the role of inflammation inhemically-induced colon cancer, various possibilities exist. First,nflammation might enhance the permeability of the colon mucosa,o that a higher concentration of a colon carcinogen may reachhe stem cell compartment in the colonic crypts. Second, reac-ive oxygen and nitrogen species released by macrophages andeutrophils in the immediate vicinity of colonic epithelial cellsould per se damage DNA damage and increase cellular turnover,hereby leading to a fixation of the genetic alterations induced byhe above-mentioned compounds. Third, the genetic alterationsnduced by chronic inflammation and those evoked by the above-

entioned chemicals differ, not only regarding the genes affectedut also regarding when the alterations occur. The genetic alter-tions induced by inflammation and chemicals could “sum up”,esulting in an acceleration of colon carcinogenesis. Data support-ng the different roles of inflammation in chemically-induced colonarcinogenesis will be presented.

oi:10.1016/j.toxlet.2012.03.027

2nnovative testing strategies to identify chemical respiratoryensitizers: Present and future

02-1lassification and labeling of chemical respiratory sensitizers:he way forward

anine Ezendam

RIVM, Netherlands

Chemical-induced respiratory allergy is a common occupationalealth problem associated with considerable morbidity. The dis-ase can be caused by low-molecular weight chemicals with theapacity to induce sensitization of the respiratory tract, for exampleiisocyanates and acid anhydrides. After sensitization, contin-ed exposure can lead to either allergic rhinitis, asthma or both.ccording to the UN globally harmonized system of classificationnd labeling of chemicals (GHS), respiratory sensitizers should beabeled with the risk phrase R42: may cause sensitization by inhala-ion. This classification and labeling is based on human evidencehat the substance can lead to specific respiratory hypersensitivitynd/or on a positive result in an animal test. The lack of validatednd accepted animal tests hinders identification of new substancesith the capacity to induce respiratory sensitization. Classifica-

ion is solely based on human evidence. Progress in this area islow, not at least due to uncertainties about the immunologicalechanisms and the relevant route of exposure for the induc-

ion of sensitization. This symposium will focus on recent progressn this area, including research in experimental animals focusingn improving our understanding of the immunological mecha-isms involved in the induction of sensitization. Future prospects

or the applicability of alternative test methods, such as in silicond in vitro methods, for hazard identification will be discussed asell.

oi:10.1016/j.toxlet.2012.03.029

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S02-2Cells and tissues encountered in the airways after inhalation ofchemical respiratory sensitizers

C. Frieke Kuper

TNO Research Group Quality and Safety, Netherlands

A wide range of cell types can potentially come into contactupon inhalation of respiratory sensitizers. Especially the airwaysare lined by a complex epithelium, which consists of at least fiveepithelial cell types and houses antigen-presenting cells and lym-phocytes. In addition, the epithelium contains an intricate networkof unmyelinated nerves running parallel to the basal membranewith branches of free nerve endings going up to tight junctions ofthe epithelial cells. The alveoli are lined by a somewhat less com-plex cell system. Together with the epithelium, fibroblasts, bloodvessels, nerves and extracellular matrix, which all lie underneaththe epithelium, form the respiratory mucosa. Mucus and surfac-tant are on top of the mucosa and form the first barrier of inhaledsensitizers. Key cells and their actions should be identified in theinduction and provocation of respiratory allergy, in order to designthe proper in vitro tests and establish the adverse outcome pathway(AOP). In addition, the contribution of cells and events that modifythe induction and provocation processes may help to understandthe variety of respiratory allergies and may facilitate the distinctionbetween allergy and irritation.

doi:10.1016/j.toxlet.2012.03.030

S02-3Genomic profiling in bronchial epithelial cells to identifyrespiratory sensitizers

Sylvie Remy 1, Inge Nelissen 2, Jef Hooyberghs 2, Rosette Van DenHeuvel 2, Greet Schoeters 2

1 VITO, Belgium, 2 Belgium

Purpose: Chemical respiratory allergy is an important occupa-tional health problem with considerable morbidity and relatedeconomic and social costs. No well-validated or widely acceptedtest methods are currently available to assess the respiratory sen-sitizing potential of chemical compounds. The aim of the studywas to identify gene markers for the identification of respiratorysensitizers.

Methods: Human bronchial epithelial BEAS-2B cells wereexposed during 10 h to seven respiratory sensitizing (RS) and ninerespiratory non-sensitizing (nRS) chemicals. To identify specificgene transcripts, the concentration yielding 20% inhibition of cellviability (IC20, after 24 h) was chosen for exposure of the cells. Inhi-bition in cell viability could not be demonstrated for 1RS and 1nRS,which were applied at highest soluble concentration. Discriminat-ing gene expression was evaluated using Agilent Whole HumanGenome 4 × 44 K oligonucleotide arrays.

Results and conclusion: The results indicate that a pre-testing cri-terion is needed to enable identification of respiratory sensitizersusing BEAS-2B cells. To discriminate chemicals that can be testedfrom chemicals that cannot, 10 gene transcripts could be identi-fied among which six are associated to the NRF-2 oxidative stresspathway. Respiratory sensitization could be assessed by two gene

markers, i.e. DNAJB4 and HSPA5, for all chemicals that meet thepre-testing criterion (4RS and 6nRS).

Based on these results can be concluded that induction of NRF-2related genes is an aspecific prerequisite for identification of respi-