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Contamination and Effects of Perfluorochemicals in Baikal Seal (Pusa sibirica). 2. Molecular Characterization, Expression Level, and Transcriptional Activation of Peroxisome Proliferator-Activated Receptor α

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journal contribution
posted on 01.04.2008, 00:00 by Hiroshi Ishibashi, Hisato Iwata, Eun-Young Kim, Lin Tao, Kurunthachalam Kannan, Shinsuke Tanabe, Valeriy B. Batoev, Evgeny A. Petrov
To investigate the biological effects of perfluorochemicals (PFCs) and to identify biomarkers of exposure to PFCs, this study focused on the effects mediated by peroxisome proliferator-activated receptor α (PPARα) in Baikal seals (Pusa sibirica). We cloned a full-length cDNA, encoding PPARα from the liver of Baikal seal, which has a deduced open reading frame of 468-amino acid residues with a predicted molecular mass of 52.2 kDa. Comparison of the amino-acid sequence of Baikal seal PPARα with that of other mammalian PPARα showed considerable similarities with PPARα of dog (97%), human (95%), rat (92%), and mouse (91%). The quantitative real-time RT-PCR analyses of tissues from Baikal seals revealed that PPARα mRNAs were primarily expressed in the liver, kidney, heart, and muscle. The hepatic expression levels of PPARα mRNA showed a positive correlation with the expression levels of immunochemically detected cytochrome P450 (CYP) 4A-like protein, indicating that the PPARα-CYP4A signaling pathway in Baikal seal is likely conserved. This study also developed an in vitro PPARα reporter gene assay using African green monkey kidney CV-1 cells transiently transfected with Baikal seal PPARα cDNA expression vector and a reporter vector containing a peroxisome proliferator-responsive element. The in vitro reporter gene assay displayed significant response to clofibrate, which is a known PPARα agonist in humans and rodents. Treatment with perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), perfluoroundecanoic acid (PFUnDA), or perfluorooctane sulfonate (PFOS) induced PPARα-mediated transcriptional activity in a dose-dependent manner, showing the lowest-observed-effect concentrations of 62.5, 125, 125, 62.5, and 125 µM, respectively. In the livers of wild Baikal seals, expression levels of PPARα mRNA showed a significant positive correlation with PFNA levels. Moreover, expression of hepatic CYP4A-like protein was significantly correlated with the hepatic concentrations of PFNA and PFDA. These results suggest modulation of the PPARα-CYP4A signaling pathway by PFCs in the wild Baikal seals. Our study demonstrates that the PPARα-mediated response may be a useful biomarker to evaluate potential biological effects of PFCs in wildlife.