When stimulated by the environmental stressors, the dynamic balance between the generation and removal of reactive oxygen species (ROS) in organisms will be disturbed. Excessive ROS can be removed by the enzymatic reaction of antioxidant enzymes. Moreover, peroxidation mostly occurs on the polyunsaturated fatty acids in the plasma membrane phospholipids, and the change of ROS can be judged by detecting MDA, the product of lipid peroxidation (Diguiseppi et al., 1984). The purpose of this study is to clarify the antioxidant response in the gonad of marine mussel M. coruscus under ocean acidification and hypoxia and its connection with gonad development. By exposing mussels to acidified and hypoxic seawater, a series of antioxidant changes were observed.
The relative contribution of each antioxidant enzyme to protect against oxidative stress during exposure period is not well known, and the relationships among reactive oxygen species (ROS) levels and activities of antioxidant enzymes are complex. In this study, the decrease of CAT and GST and the increase of GPX, GSH and MDA under acidification and hypoxia were observed, but no significant changes in SOD.
As a vital scavenger of H2O2, CAT is considered as a second line of antioxidant defense (Chelikani et al., 2004). Although CAT activity often increases as a result of the increased H2O2 under oxidative stress (Marcelo Hermes-Lima., 2004), multiple stressors may change this trend (Matozzo et al., 2013). As we found, CAT activity was significantly reduced during short-term acidification and hypoxia exposure. Woo et al. (2013) also found that CAT activity decreased when mussels Mytilus galloprovincialis were exposed to hypoxia. The decline may be GPX-related.
GPX not only participates in the conversion of H2O2 to water and molecular oxygen, but also plays a vital role in the use of GSH as a reducing agent to convert other lipids to non-toxic products (Sies et al., 1997). In our study, GPX activity was significantly increased by acidification and hypoxia. In some other researches, the effect of pH or hypoxia was quite different, mainly due to different matching stresses and test subjects. Lima et al. (2019) found that the GPX activity in oyster Crassostrea gasar exposed to combined effects of acute pH changes and phenanthrene for 96 h showed no significant change. Johannsson et al. (2018) found GPX rose in the brain and gills of Characid fish Cyphocharax abramoides during hypoxia. Although both GPX and CAT could catalyze the decomposition of H2O2, their different changes suggested that GPX is more capable of scavenging free radicals than CAT, which is consistent with previous research (Dorval et al., 2003).
GSH is a major thiol compound that acts as a protective agent for a variety of toxic substances through thiol groups. Its functions include the direct removal of oxy compounds and catalysis of organic hydrogen peroxide or H2O2, maintaining membrane protein thiols and acting as a substrate for GPX and glutathione reductase (Habig et al., 1974; Moreno et al., 2005). Some studies have shown that OA and hypoxia or the combination with other stressors either decrease or increase GPX activity, depending on the species and specific tissue (Huang et al., 2018; Khan & Ringwood., 2016). Hu et al. (2015) found GSH in gill and digestive gland of mussels exposed to acidification increased significantly. Similar in our experiment, the GSH activity was increased by pH under DO 6 mg L− 1.
In the process of oxidative stress, GST greatly increases on the basis of sulfhydryl reaction metabolites and the reduction of oxygen species, playing a fundamental role in the process of chemical detoxification (Aniya et al., 1993; Sheehan et al., 2001). Lopes et al. (2018) found GST activities in the soft coral Veretillum cynomorium did not change significantly under warming and acidification. The GST activity in our experiment significantly decreased under pH 7.3, indicating that extreme low pH disrupted the balance of GST. The similar phenomenon was also found in the study of Lima et al. (2019).
MDA content is an important parameter reflecting the body's potential antioxidant capacity, which can reflect the body's lipid peroxidation rate and strength, and indirectly reflect the degree of tissue peroxidation damage. In our results, MDA level was significantly increased, meaning that oxidative damage occurred to mussels.
At the end of the recovery period, all enzyme activities returned to normal except GPX and GST, which coincidentally were also the two enzymes that were significantly affected by the combination of acidification and hypoxia during the exposure period. When acting on GPX and GST, acidification and hypoxia coordinated, preventing the mussels from recovering in the short term. Whether long-term recovery will be curable remains to be determined.
As enzyme activities are inhibited and induced under pollution conditions, and the responses of various enzymes to biotoxicity exposure are not synchronous, different enzymes have different sensitivity to pollution. Therefore, the analysis of several enzymes and even other biomarkers can be combined to evaluate the pollution situation more effectively. The IBR index is such a powerful tool. It has been widely used in many studies to assess stress responses and ecological risks (Cao et al., 2019; Damiens et al., 2007; Xie et al., 2016). In this study, among the exposure groups, the group DO 2 mg L− 1 and pH 7.7 had the highest IBR/n value, which reflected this group was the most highly impacted one. When put sights on the GSA, it seemed more reasonable. GSA decreased the most at extreme pH level. There is a hypothesis that mussels exhibit physiological trade-offs, and that, under increased pressure from climate change scenarios, they may allocate energy from reproduction to costly physiological defenses (Petes et al., 2008; Béguel et al., 2013). The combined stress of extreme pH and hypoxia put severely negative effects on mussel and threatened their survival, so they had to spend a large part of the energy originally used for gonadal development to cope with the stress. Taking all these results into account, we can confirm that acidification and hypoxia exposure have adverse effects in mussel gonadal antioxidant system, and consequently impact mussel gonad development.