First evidence of induction of CYP1A in Caretta caretta skin biopsy slices treatedwithpolycyclic aromatic hydrocarbons, organochlorines and polybrominated diphenyl ethers

The aim of this study was to develop and validate the induction of CYP1A as a non-invasive biomarker of exposure to some classes oflipophilic contaminants in skin biopsy slices of Caretta caretta. A Western blot method to evaluate protein expression of CYP1A in this species was set up and applied in liver and skin sampled from two specimens of C. caretta that died in a turtle rescue center in Italy.We detected a strong band at about 59 kDa of molecularweight in both tissues, in accordance with the findings in liver of the freshwater C. picta picta. In order to semiquantify CYP1A, a series of CYP450 calibration curves from the liver (ranging from 0.5 to 5.0 pmol of CYP450) of Caretta caretta were analysed. In addition a skin standardwas analyzedwithin the curves and maintained as an internal standard in subsequent WBs. Semi quantitative analysis was performed for each WB with Quantity-One software (BioRad) using Adjusted Volume (Intensity*mm2) as a quantitative parameter. In the second part of the study, skin biopsy samples were taken from free-ranging specimens, and subaliquots (slices) were incubated in vitro for 24 h with increasing doses of PAHs, OCs and PBDEs in order to investigate the potential for CYP1A induction. Western blotting of CYP1A showed a dose-related induction in skin biopsy slices treated with the different compounds. A higher induction was found after treatment with PBDEs, indicating a strong potential of these chemicals to interfere with bioactivating enzyme systems in C. caretta. This study, which we believe is the first of its kind in the literature, demonstrated the presence and inducibility of CYP1A in skin biopsies of C. caretta, and validated this non destructive biomarker for subsequent application in field studies.