Effects of Nrf2-Keap1 signaling pathway on antioxidant defense system and oxidative damage in the clams Ruditapes philippinarum exposure to PAHs.

NF-E2-related factor 2 (Nrf2) is a master regulator of antioxidant defense system which can maintain the oxidation balance in the cell. In our previous study, we first cloned the Nrf2 gene in clams and preliminarily explored the role of the Nrf2 at the transcription level. In this study, RNA interference (RNAi) technology was used to interfere with the expression of Nrf2 after being exposed to benzo(a)pyrene (BaP) for 5 days to verify the role of Nrf2 in the antioxidant defense system. Besides, we examined the mRNA expression and enzyme activities of antioxidases and the oxidative damage. The positive correlations between the Nrf2 with the mRNA expression and the enzyme activities of antioxidases indicated that Nrf2 was required for the induction of these antioxidant genes. Additionally, the mRNA expression and the enzyme activities of the glutathione peroxidase (GPx) in the Nrf2-dsRNA group were significantly higher than those in the control groups on the fifth day, indicating that the GPx is more sensitive to oxidative stress. Moreover, the oxidative damage in the RpNrf2-dsRNA group was markedly increased than control groups, indicating that Nrf2 transcriptional regulation may play an essential role in defending against oxidative damage. This study provides a foundation for further research on the mechanism of detoxification and antioxidation of polycyclic aromatic hydrocarbons (PAHs) in the clams at the transcription level and the protein level.

The molecular mechanism of AhR-ARNT-XREs signaling pathway in the detoxification response induced by polycyclic aromatic hydrocarbons (PAHs) in clam Ruditapes philippinarum.

The aryl hydrocarbon receptor (AhR) has been known primarily for its role in the regulation of several drug and xenobiotic metabolizing enzymes to mitigate environmental stresses. In this study, we interfere the expression of AhR gene to investigate the mechanism of AhR signaling pathway in the detoxification and antioxidation defense system that induced by Polycyclic Aromatic Hydrocarbons (PAHs) exposure by RNA interference (RNAi). The gene expressions of aryl hydrocarbon receptor nuclear translocator (ARNT), heat shock protein 90 (Hsp90) were evaluated after being exposed to benzo(a)pyrene (BaP) (4 µg/L) for 5 days and the positive correlations between AhR, ARNT, HSP90 indirectly indicating that AhR may have the ability to bind to ligands such as PAHs in Ruditapes philippinarum (R. philippinarum). Besides, the activities of detoxification enzymes were determined to investigate the role of AhR signaling pathway played in the metabolic detoxification. What's more, the gene expressions of protein kinase C (PKC) signaling pathway, mitogen-activated protein kinase (MAPKs) signaling pathway, NF-E2-related factor 2 (Nrf2) signaling pathway and antioxidant defense system indicated that AhR may regulate the Nrf2-Keap1 signaling pathway through Kelch-like ECH-associated protein-1 (Keap1) and MAPKs, PKC signaling pathways. In conclusion, adoption of RNA interference technology to explore the role of RpAhR gene played in the detoxification and antioxidation defense system under the PAHs stress at different time points can informe molecular endpoints for application towards ecotoxicology monitoring of bivalves.

Using multi-integrated biomarker indexes approach to assess marine quality and health status of marine organism: a case study of Ruditapes philippinarum in Laizhou Bay, China.

With the progress of technology and the deepening of understanding of biological monitoring, much more attention has been paid to the multiple evaluation of marine pollution monitoring. In view of this, our study aimed at establishing a multi-integrated biomarker indexes approach to evaluate marine condition systematically and comprehensively. In the current study, sampling was conducted in Laizhou Bay, China (S1, S2, and S3) in May, August, and October of 2015. And then, multi-integrated biomarker indexes approach was applied to assess marine PAHs pollution, select appropriate biomarkers, and evaluate marine environmental quality and health status of the clams of Ruditapes philippinarum. As the results showed, S2 was the most PAHs-polluted site while S1 was the least polluted site, and the levels of tPAHs in seawater and sediments ranged from 69.78 to 315.30 ng/L and 163.19 to 565.17 ng/g d.w., respectively. And all three sampling sites had different sources of PAHs. IBR represented DNA damage (F value), the expression of SOD, EROD activity, GST activity, and LPO could be served as biomarkers to monitor the PAHs pollution in Laizhou Bay. And MPI suggested the quality of all three sites: S1 was generally favorable, S2 was moderately polluted, and S3 was lightly polluted. BRI values showed that the order of health status of R. philippinarum was S1 > S3 > S2.

Multi-biomarker approach in the scallop Chlamys farreri to assess PAHs pollution in Qingdao coastal areas of China.

A multi-biomarker approach was conducted in the scallop Chlamys farreri from three sites, denoted here as S1, S2, and S3, in Qingdao coastal areas of China in March, June, September and December 2014 to assess pollution from polycyclic aromatic hydrocarbons (PAHs) and to select appropriate biomarkers. A suite of biological responses of the gills and digestive glands of the scallops was assayed, including: (i) phase I detoxification enzymes of 7-ethoxyresorufin-O-deethylase (EROD), epoxide hydrolase (EH), and dihydrodiol dehydrogenase (DD) and phase II detoxification enzymes of glutathione-S-transferase (GST) and sulfotransferase (SULT); (ii) antioxidant enzymes: catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx); (iii) oxidative damage parameters: lipid peroxidation (LPO) expressed by malondialdehyde (MDA) contents, protein carbonylation (PC) and DNA damage (F value); and (iv) the metabolism-related genes of EH, DD, GST, SULT and SOD. Simultaneously, the concentrations of total PAHs along with 16 types of PAHs previously identified by the US Environmental Protection Agency (USEPA) and environmental parameters, including temperature and salinity together with pH, were measured. Using Principle Component Analysis (PCA), it was revealed that S2 was the most PAH-contaminated site, while S1 was identified as the least PAH-polluted site, which was consistent with the results utilizing the Biomarker Response Index (BRI); in other words, the biological health status of S2 was worse than S1 and S3. Moreover, the most suitable biomarkers to assess PAH pollution in Qingdao coastal areas proved to be DD mRNA expression and the F value in both the gills and digestive glands for the total PAHs, DD activity and PC contents or PC and MDA contents in the gills or digestive glands for 5 + 6 rings PAHs and DD mRNA expression in both the gills and digestive glands for 2 + 3 rings and 4 rings PAHs. Moreover, this study highlighted the possible use of the scallop Chlamys farreri for studying contamination due to PAHs and provided valuable information on environmental assessment.

A multi-biomarker approach in scallop Chlamys farreri to assess the impact of contaminants in Qingdao coastal area of China.

A multi-biomarker approach was carried out to classify the environmental quality and the adverse effects of contaminants on scallop Chlamys farreri. The scallops were collected from three sampling stations in Qingdao coastal area of China in March, May, August and October of 2015. A suite of environmental factors and biomarkers, including temperature, salinity, pH, the concentrations of polycyclic aromatic hydrocarbons (PAHs), tetrabromobisphenol A (TBBPA) and metals (Cr, Mn, Cu, Zn, Cd, Pb, As) in seawater and soft tissue, mRNA expression of aryl hydrocarbon receptor (AhR) and P-glycoprotein (P-gp), 7-ethoxyresorufin O-deethylase (EROD), glutathione-S-transferase (GST), uridine-diphosphate-glucuronyl-transferase (UGT), sulfotransferase (SULT), metallothionein (MT), Superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), lipid peroxidation (LPO) and protein carbonyl (PC) contents and DNA strand breaks, were measured in the gill and digestive gland. The results showed that S2 was the most polluted while S1 was identified the least polluted. Despite the differentiation of pollution levels and environmental parameters the selected biomarkers responded efficiently to contaminants. Principal component analysis (PCA) revealed that EROD for PAHs, AhR for TBBPA, MT for Cr, Pb and Mn, LPO and PC for Zn were the effective biomarkers respectively. This study demonstrated that the application of multi-biomarker approach in conjunction with the traditional analysis of environmental parameters and contaminants provided valuable information in environmental risk assessment.