Im Hosub, Oh Eunha, Mun Joohee, Khim Jin-Young, Lee Eunil, Kang Hyung-Sik, Kim Eunmi, Kim Hyunsuk, Won Nam-Hee, Kim Young-Hwan, Jung Woon-Won, Sul Donggeun
School of Public Health, Korea University, Anamdong 5, Sungbukku, Seoul, 136-705, Korea.
J Proteome Res. 2006 Jun;5(6):1354-66. doi: 10.1021/pr050437b.
Formaldehyde (FA) is known as a low molecule weight organic compound and one of major components that causes sick building syndrome (SBS), and it has been reported that FA has cytotoxic, hemotoxic, immunotoxic, and genotoxic properties. The International Agency for Research on Cancer (IARC) has characterized FA as a carcinogen. In this study, we investigated the effects of FA on rat plasma proteins by using proteomic approach. Rats were exposed to three different concentrations of FA (0, 5, 10 ppm) for 2 weeks at 6 hours/day and 5 days/week in an inhalation chamber. Malondialdehyde (MDA) assay and carbonyl spectrometric assay were conducted to determine lipid peroxidation and protein oxidation levels and Comet assays were used for genotoxicity evaluation. Level of MDA, carbonyl insertion and DNA damage in plasma, livers, and in the lymphocytes of rats exposed to FA were found to be dose dependently increased. Proteomic analysis using three different pI ranges (3.5-5.6, 5.3-6.9, 6-9) and large size two-dimensional gel electrophoresis (2-DE) showed the presence of 3491 protein spots. A total of 32 (19 up- and 13 down-regulated) proteins were identified as biomarkers of FA, all showed dose dependent expressions in the plasma of rats exposed to FA and of these, 27 protein spots were identified by MALDI-TOF/MS. Several differentiated protein groups were found. Proteins involved in apoptosis, transportation, signaling, energy metabolism, and cell structure and motility were found to be up- or down-regulated. Among these, the identities of SNAP 23, apolipoprotein A-1 and E, clusterin, kinesin, and fibrinogen gamma were confirmed by Western blot assay, and apo E was further analyzed by using 2-DE immunoblot assays to determine isoform patterns. Two cytokine including IL4 and INF-gamma were measured in plasma with respect to fibrinogen gamma changes. In summary, cytotoxicity, and genotoxicity assays, namely MDA lipid peroxidation assay, the carbonyl protein oxidation assay, and Comet genotoxic assay showed that these effects increased on increasing FA levels. Proteomic analysis with three different pI ranges and long size 2-DE gel electrophoresis showed that 32 protein spots were up-or down-regulated. Of these 32 proteins, 7 proteins were confirmed by western blot assay. They could be potential biomarkers for human diseases associated with FA exposure.
甲醛（FA）是一种低分子量有机化合物，是导致病态建筑综合征（SBS）的主要成分之一，据报道，FA具有细胞毒性、血液毒性、免疫毒性和遗传毒性。国际癌症研究机构（IARC）已将FA列为致癌物。在本研究中，我们采用蛋白质组学方法研究了FA对大鼠血浆蛋白的影响。将大鼠置于吸入舱中，每天6小时、每周5天，暴露于三种不同浓度的FA（0、5、10 ppm）下2周。进行丙二醛（MDA）测定和羰基光谱测定以确定脂质过氧化和蛋白质氧化水平，并使用彗星试验进行遗传毒性评估。发现暴露于FA的大鼠血浆、肝脏和淋巴细胞中的MDA水平、羰基插入和DNA损伤呈剂量依赖性增加。使用三种不同的pH范围（3.5 - 5.6、5.3 - 6.9、6 - 9）和大尺寸二维凝胶电泳（2-DE）进行蛋白质组学分析，共显示出3491个蛋白点。总共32种（19种上调和13种下调）蛋白质被鉴定为FA的生物标志物，它们在暴露于FA的大鼠血浆中均呈剂量依赖性表达，其中27个蛋白点通过基质辅助激光解吸电离飞行时间质谱（MALDI-TOF/MS）鉴定。发现了几个不同的蛋白组。参与细胞凋亡、转运、信号传导、能量代谢以及细胞结构和运动的蛋白质被发现上调或下调。其中，通过蛋白质免疫印迹法（Western blot）确认了突触融合蛋白23（SNAP 23）、载脂蛋白A-1和E、簇集蛋白、驱动蛋白和纤维蛋白原γ的身份，并通过2-DE免疫印迹测定法进一步分析载脂蛋白E以确定其亚型模式。针对纤维蛋白原γ的变化，检测了血浆中的两种细胞因子，即白细胞介素4（IL4）和γ干扰素（INF-γ）。总之，细胞毒性和遗传毒性试验，即MDA脂质过氧化试验、羰基蛋白氧化试验和彗星遗传毒性试验表明，这些效应随着FA水平的增加而增强。使用三种不同pH范围和长尺寸2-DE凝胶电泳进行的蛋白质组学分析表明，32个蛋白点上调或下调。在这32种蛋白质中，7种蛋白质通过蛋白质免疫印迹法得到确认。它们可能是与FA暴露相关的人类疾病的潜在生物标志物。
