Cadmium (Cd) pollution is a serious global problem, which causes irreversible toxic effects on animals. Freshwater crab, Sinopotamon henanense, is a useful environmental indicator since it is widely distributed in benthic habitats whereby it tends to accumulate Cd and other toxicants. However, its molecular responses to Cd toxicity remain unclear. In this study, we performed transcriptome sequencing and gene expression analyses of its hepatopancreas with and without Cd treatments. A total of 7.78 G clean reads were obtained from the pooled samples and 68,648 unigenes with an average size of 622 bp were assembled, in which 5,436 were metabolism-associated and 2,728 were stimulus response-associated that include 380 immunity-related unigenes. Expression profile analysis demonstrated that most genes involved in macromolecular metabolism, oxidative phosphorylation, detoxification and anti-oxidant defense were up-regulated by Cd exposure, whereas immunity-related genes were down-regulated, except the genes involved in phagocytosis were up-regulated. The current data indicate that Cd exposure alters gene expressions in a concentration-dependent manner. Therefore, our results provide the first comprehensive S.henanense transcriptome dataset, which is useful for biological and ecotoxicological studies on this crab and its related species at molecular level and some key Cd-responsive genes may provide candidate biomarkers for monitoring aquatic pollution by heavy metals.

We studied here the short-term toxicity effects of Cd on the oxidative state and cell death in the gill of freshwater crab Sinopotamon henanense. Crabs were exposed to Cd that resulted in Cd accumulation and a significant increase in the metallothionein (MT) level in the gill, but MT level increased disproportionally compared to the Cd accumulation with an extension of exposure time. Significant changes in the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) were observed. An increase in the levels of reactive oxygen species (ROS) and lipid peroxidation (LPO) was detected that will cause oxidative stress. Histological abnormalities of the gills were discovered, including the expansion of gill cavity, a decrease in the numbers of connection of the upper and the lower of the gill lamellae and epithelial cells, and an increase in the number of hemocytes. The results of a TUNEL test and transmission electron microscope (TEM) showed that more gill cells had apoptotic characteristics after 48 h of Cd treatment compared to the control, but epithelial cell necrosis and inflammatory response appeared only after 72 h. It was concluded that (1) Cd induced the ROS production and accumulation through inhibiting antioxidant enzyme activities and exceeding the saturation values of MT binging; (2) Cd led to lipid peroxidation and histopathological alternations; and (3) Cd induced apoptotic response at short time exposure, followed by necrotic features and inflammatory reaction after longer time exposure.

Calcium ion (Ca²⁺) is one of the key intracellular signals, which is implicated in the regulation of cell functions such as impregnation, cell proliferation, differentiation and death. Cadmium (Cd) is a toxic environmental pollutant that can disturb cell functions and even lead to cell death. Recently, we have found that Cd induced apoptosis in gill cells of the freshwater crab Sinopotamon henanense via caspase activation. In the present study, we further investigated the role of calcium signaling in the Cd-induced apoptosis in the animals. Our data showed that Cd triggered gill cell apoptosis which is evidenced by apoptotic DNA fragmentation, activations of caspases-3, -8 and -9 and the presence of apoptotic morphological features. Moreover, Cd elevated the intracellular concentration of Ca²⁺, the protein concentration of calmodulin (CaM) and the activity of Ca²⁺-ATPase in the gill cells of the crabs. Pretreatment of the animals with ethylene glycol-bis-(b-aminoethyl ether)-N,N,N’,N’-tetraacetic acid (EGTA), Ca²⁺ chelator, inhibited Cd-induced activation of caspases-3, -8 and -9 as well as blocked the Cd-triggered apoptotic DNA fragmentation. The apoptotic morphological features were no longer observed in gill cells pretreated with the Ca²⁺ signaling inhibitors before Cd treatment. Our results indicate that Cd evokes gill cell apoptosis through activating Ca²⁺-CaM signaling transduction pathway.