1460VOLUME 115 | NUMBER 10 | October 2007•Environmental Health PerspectivesResearchFormaldehyde (FA) is an industrial chemicalused for manufacturing building materialsand household products. It is also found inautomobile emissions and tobacco smoke.Inhalation of FA induces sensory irritation atrelatively low concentrations in experimentalanimals as well as in humans (Alexanderssonand Hedenstierna 1988; Kane and Alarie1977). Although animal studies investigatingthe genotoxic potential of FA did not generatedefinitive results (Dallas et al. 1992; Miglioreet al. 1989), FA is carcinogenic at the site ofcontact as a consequence of epithelial cellregenerative proliferation resulting from cyto-toxicity and mutation (reviewed by Naya andNakanishi 2005). In 2004 the InternationalAgency for Research on Cancer (IARC) con-cluded that FA is a known human carcinogenbased on sufficient evidence in humans andexperimental animals (IARC 2004).Sick building syndrome (SBS) is the suiteof adverse health effects caused by seriousindoor air quality problems in homes, offices,and other work places. The syndrome is gen-erally characterized by mucosal irritation andnonspecific hypersensitivity. Although thespecific cause of the syndrome has not beenidentified, the accumulation of chemical andbiological contaminants from indoor sourcesbecause of inadequate ventilation is regardedas the main factor inducing SBS. Along withmany other factors, exposure to FA indoorscould contribute to the syndrome. It has beensuggested that the recent increase in allergicdiseases such as asthma and allergic rhinitis isrelated to exposure to environmental pollu-tants such as FA (Takafuji and Nakagawa2000).FA is metabolized into formic acid byaldehyde dehydrogenase, which is furtheroxidized to carbon dioxide or may serve as aone-carbon donor in tetrahydrofolate-depen-dent synthesis of purine, pyrimidine, andamino acids. Because of its rapid metabolismand exhalation, however, exposure to FAdoes not always result in an increased bloodor urine concentration of parent compoundin animals or humans (Gottschling et al.1984; Heck et al. 1985). Therefore, thedevelopment of biomarkers for FA exposureand toxicity is necessary for risk assessment.Thiazolidine-4-carboxylate (TZCA) isformed by a nonenzymatic condensationreaction of L-cysteine with a variety of amine-derived aldehydes or carbonyl compounds,including FA (Ratner and Clarke 1937). Werecently found that the urinary concentrationof TZCA reflects the degree of FA exposurevery well (Shin et al., in press). Therefore, wemeasured the urinary concentration ofTZCA as a marker for FA exposure.The identification of differentiallyexpressed genes or patterns of gene expressionusing a microarray hybridization assay pro-vides a logical approach to developing poten-tial biomarkers of toxicity. Hundreds ofpublications have suggested the use of bio-markers for the diagnosis and prognosis ofdisease, as well as for the assessment of expo-sure to xenobiotics in cells and animal tissues.However, only a very limited number of can-didate gene markers can ultimately be used inhuman samples. The reasons for this includethe inaccuracy of microarray data, discrepan-cies among in vitro, in vivo, and human stud-ies, and the lack of validation of the in vitroor in vivo data in human samples. To thisend, we investigated the effects of FA on geneexpression in human tracheal fibroblast andvalidated the results in human samples withvarying degrees of FA exposure. Our overallgoal was to identify global changes in geneexpression following FA exposure to producemechanistic insight into FA-induced toxicityand to provide potential biomarkers that canbe used in risk assessment for FA exposure.Materials and MethodsCells and chemicals.The Hs 680.Tr humantracheal fibroblast cell line was maintained inRPMI-1640 medium supplemented with10% fetal bovine serum (FBS) in a 5% CO2atmosphere at 37°C. The culture mediumdoes not contain cysteine. FA, TZCA, and2-methyl-TZCA were purchased fromSigma Chemical Co. (St. Louis, MO, USA).Cell viability measurement.We incu-bated the cells with various concentrations(0, 20, 50, 100, and 200 µM) of FA for 4and 24 hr. Aqueous solution of FA (37%)was added directly to the incubation media.Cell viability was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetra-zolium bromide (MTT) cell proliferationassay.RNA isolation.Total RNA was extractedusing an Easy-Blue total RNA extraction kit(Intron Biotech, Sungnam, Korea), purifiedusing Qiagen RNeasy Mini Kits (Qiagen,Basel, Switzerland), and examined forintegrity using an Agilent 2100 Bioanalyzer(Ambion, Austin, TX, USA).Address correspondence to B-H. Lee, College ofPharmacy, Seoul National University, San 56-1,Sillim-dong, Gwanak-gu, Seoul 151-742, Republicof Korea. Telephone: +82-2-880-7843. Fax: +82-2-874-7843. E-mail: [email protected] Material is available online (http://www.ehponline.org/docs/2007/10180/suppl.pdf).This work was supported by the Ministry ofEnvironment as “The Eco-Technopia 21 Project.”The authors declare they have no competingfinancial interests.Received 20 February 2007; accepted 12 July 2007.Identification of Gene Markers for Formaldehyde Exposure in HumansGuang-Yong Li,1Hye-Young Lee,1Ho-Sang Shin,2Hyeon-Young Kim,3Cheol-Hong Lim,3and Byung-Hoon Lee11College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea;2Department of Environmental Education and Abuse Drug Research Center, Kongju National University, Kongju, Republic of Korea;3Occupational Safety and Health Research Institute, Korea Occupational Safety and Health Agency, Daejeon, Republic of KoreaBACKGROUND: Formaldehyde (FA) is classified as a human carcinogen and has been linked toincreased leukemia rates in some epidemiologic studies. Inhalation of FA induces sensory irritationat relatively low concentrations. However, little is known concerning the cellular alterationsobserved after FA exposure in humans.OBJECTIVES: Our aim was to profile global gene expression in Hs 680.Tr human tracheal fibroblastsexposed to FA and to develop biomarkers for the evaluation of FA exposure in humans.METHODS AND RESULTS: We used gene expression analysis, and identified 54 genes designated asFA responsive. On the basis of these data, we conducted an exploratory analysis of the expression ofthese genes in human subjects exposed to high or low levels of FA. We monitored FA exposure bymeasuring
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