Acetaminophen exposure alters the DNA methylation pattern of Mugilogobius chulae, along with the changes in the Nrf2-Keap1 signaling pathway.

DNA methylation can dynamically regulate multiple physiological processes in organisms in response to changes of the external environment. The effects of acetaminophen (APAP) on DNA methylation in aquatic organisms and its toxic mechanisms is an interesting issue. In the present study, Mugilogobius chulae (Approximately 225 individual), a small benthic native fish, were employed to assess the toxic effects of APAP-exposure on non-target organisms. First, under APAP exposure (0.5 µg/L and 500 µg/L) for 168 h, 17,488 and 14,458 differentially methylated regions (DMRs) were identified in liver of M. chulae, respectively, which were involved in energy metabolism, signaling transduction, and cellular processes etc. The modification of lipid metabolism by DNA methylation was particularly prominent and the increased fat vacuoles in the sections were observed. Some key nodes associated with oxidative stress and detoxification such as Kelch-1ike ECH-associated protein l (Keap1) and fumarate hydratase (FH) were modified by DNA methylation. Meanwhile, changes in DNA methyltransferase and Nrf2-Keap1 signaling pathways at different concentrations of APAP (0.5 µg/L, 5 µg/L, 50 µg/L and 500 µg/L) for different time (24 h and 168 h) were addressed at the transcriptional level. Results showed that ten eleven translocation enzymes 2 (TET2) transcript expression was upregulated 5.7-folds after being exposed to 500 µg/L APAP for 168 h, indicating the urgent need for active demethylation in the exposed organism. The elevated DNA methylation levels of Keap1 led to repression of its transcriptional expression so as to promote recovery or reactivation of Nrf2, which displayed negatively relationship with Keap1 gene. Meanwhile, P62 was significantly positively correlated with Nrf2. Downstream genes in the Nrf2 signaling pathway changed synergistically except for Trx2, in which GST and UGT were highly significantly upregulated. This work illustrated that APAP exposure altered the DNA methylation processes, together with the Nrf2-Keap1 signaling pathway, and affected the stress responses of M. chulae to pharmaceuticals exposure.

Hypoxia aggravates the burden of yellowstripe goby (Mugilogobius chulae) under atorvastatin exposure.

In the present study, an estuarine benthic fish, Mugilogobius chulae (M. chulae), was exposed to hypoxia, atorvastatin (ATV), a highly used and widely detected lipid-lowering drug in aquatic environment, and the combination of hypoxia and ATV for 7 days, respectively, so as to address and compare the effects of the combination of hypoxia and ATV exposure on M. chulae. The results showed that lipid metabolism in M. chulae was greatly affected: lipid synthesis was blocked and catabolism was enhanced, exhibiting that lipids content were heavily depleted. The combined exposure of hypoxia and ATV caused oxidative stress and induced massive inflammatory response in the liver of M. chulae. Signaling pathways involving in energy metabolism and redox responses regulated by key factors such as HIF, PPAR, p53 and sirt1 play important regulatory roles in hypoxia-ATV stress. Critically, we found that the response of M. chulae to ATV was more sensitive under hypoxia than normoxia. ATV exposure to aquatic non-target organisms under hypoxic conditions may make a great impact on the detoxification and energy metabolism, especially lipid metabolism, and aggravate the oxidative pressure of the exposed organisms.

Assessment of ecotoxicity effects of aspirin on non-target organism (Daphnia magna) via analysis of the responses of oxidative stress, DNA methylation-related genes expressions and life traits changes.

Aspirin (acetylsalicylic acid, ASA), a widely used non-steroidal anti-inflammatory drug, was frequently detected in aquatic environments around the world. However, information on the potential toxic effects of aspirin on non-target aquatic invertebrates is limited. In the present study, we investigated the effects of ASA on the transcriptional expressions of antioxidant genes (Nrf2, Keap1, HO-1, GCLC, GPx, TRX, TrxR and Prx1) and DNA methylation genes (DNMT1, DNMT3 and TET2) in Daphnia magna (D. magna)for 24, 48 and 96 h and the changes of antioxidant enzymatic activity and GSH, MDA content for 48 h. The effects of ASA on the life traits of D. magna were also addressed via a 21-days chronic toxicity test. Results showed that the expressions of Nrf2 and its target genes (HO-1, GPx and TrxR, GCLC, TRX and Prx1) were induced to different degrees at 48 h and/or 96 h. The activity of antioxidant enzymes (SOD, CAT, GST and GPx) and MDA content increased but GSH content decreased, indicating that ASA caused oxidative stress in D. magna. ASA also changed the expression of DNA methylation genes, such as DNMT and TET2, in D. magna. We speculated that ASA may affect the antioxidant system responses through regulation of Nrf2/Keap1 signaling pathway, and/or through indirectly influencing DNA methylation levels by DNMT and TET gene expression, but the detailed mechanism needs further investigations. Chronic exposure to ASA for 21 days caused inhibitions on the growth, reproduction and behavior of D. magna (e.g., delaying days to the first brood and shortening the body length). In summary, ASA significantly affected the antioxidant responses of D. magna, and negatively disturbed its life traits in growth, development and reproduction.

Simvastatin affects the PPARα signaling pathway and causes oxidative stress and embryonic development interference in Mugilogobius abei.

Simvastatin (SV) is a common hypolipidemic drug in clinical medicine that can reduce endogenous cholesterol biosynthesis by inhibiting hydroxyl-methyl-glutaryl coenzyme A reductase. SV took a large market share in the lipid-lowering drugs and it is frequently detected in various water bodies due to its increasing consumption in past years. In the present investigation, we selected a native fish species in the Pearl River Basin in China, Mugilogobius abei (M. abei), to study the effects of SV on non-target aquatic organisms. Results showed that a significant decrease in the volume of adipocytes under SV exposure were observed on oil red O section, and the expression of HMG-CoAR decreased significantly. The mRNA and protein expression of PPARα were significantly up-regulated, the expressions of other genes related to lipid metabolism were up-regulated to varying degrees as well. There was a positive correlation between the concentrations of SV and the protein expressions of plasma phospholipid transfer protein (PLTP) and cholesterolester transfer protein (CETP). In addition, the frozen sections showed that SV led to ROS accumulation in liver in a time and concentration dependent manner. The mRNA and protein expressions of Nrf2 were significantly up-regulated after 24 hours of SV exposure. Some biomarkers associated with antioxidant such as Trx2, TrxR and MDA content were positively correlated with the exposure concentration and time, while the content of GSH decreased sharply. It is noteworthy that the environmentally relevant concentration (0.5 µg/L) of SV exposure caused delayed embryonic development and deformations, decreased hatching rates. We conclude that SV promotes fat metabolism, gives rise to oxidative stress and has significant toxicity on embryo development in M. abei.

Atorvastatin causes oxidative stress and alteration of lipid metabolism in estuarine goby Mugilogobius abei.

The potential effects of the environmental residues of Atorvastatin (ATV) as a widely used antilipemic agent on aquatic organisms deserve more investigations because of its high detection frequency in environment. The responses of Nrf2/Keap1 signaling pathway (including the transcriptional expression of Nrf2, Keap1, GCLC, GPx, GST, SOD, CAT, Trx2, TrxR, HMG-CoAR and PGC-1α) in Mugilogobius abei were investigated under acute and sub-chronic exposure of ATV in the simulated laboratory conditions. The changes of related enzymatic activity (GST, GPx, SOD, CAT and TrxR) and the content of GSH and MDA combining with the observation of histology sections of liver in M. abei were also addressed. The results show Nrf2 and its downstream antioxidant genes were induced to different degrees under ATV exposure. The activities of antioxidant enzymes were inhibited at 24 h and 72 h but induced/recovered at 168 h. Correspondingly, negatively correlated to GSH, MDA increased first but reduced then. Notably, with the increase of exposure concentration/time, the volume of lipid cells in liver decreased, suggesting more lipid decomposition. Therefore, lipid metabolism was suppressed (down-regulation of PGC-1α) and cholesterol biosynthesis was induced (up-regulation of HMG-COAR) at 168 h. In short, ATV brings oxidative stress to M. abei in the initial phase. However, with the increase of exposure time, ATV activates Nrf2/Keap1 signaling pathway and improves the antioxidant capacity of M. abei to reverse this adverse effect. ATV also affects lipid metabolism of M. abei by reducing cholesterol content and accelerating lipid decomposition.