Study On Oxidative Stress And Mechanism In The Freshwater Mussel Anodonta Woodiana Exposed To Subchronic Cadmium

Cadmium(Cd)is a toxic and non-essential heavy metal element which can cause adverse effects on animal tissues and organs.Cd pollution in aquatic environments around the world has become an increasingly serious environmental problem.Anodonta woodiana has been proposed as a suitable indicator of "freshwater mussel watch" to monitor pollution in freshwater environments.Traditionally,the gills and digestive gland are the main target organs for studying the adverse effects of Cd on bivalves,and the kidney also has a strong ability to accumulate Cd.This thesis studied the Cd bioaccumulation and elimination in eight main tissues and organs such as kidney,gills and digestive gland of the freshwater mussel A.woodiana.The antioxidant responses and histopathological changes were analyzed in the kidney,gills and digestive gland of A.woodiana exposed to subchronic Cd.The m RNA expressions of antioxidants related genes were explored in the kidney and gills of A.woodiana exposed to subchronic Cd.The aim of this thesis was to select the suitable tissue and corresponding biomarkers of A.woodiana to monitor and assess Cd pollution in a freshwater environment.The results of this study were as followed:Firstly,experiments were carried out to investigate the bioaccumulation and elimination of Cd in tissues(kidney,gills,digestive gland,mantle,visceral mass,foot,adductor muscle and hemolymph)from the freshwater mussel,A.woodiana.The mussels were exposed to Cd at concentrations of 0.168 and 0.675 mg·L-1 for 28 d of bioaccumulation and 28 d of elimination.During the bioaccumulation phase,Cd bioaccumulations increased in all tissues.The highest bioaccumulation of Cd was found in the kidney.The second-highest and third-highest bioaccumulations of Cd were found in the digestive gland and gills,respectively.The Cd bioaccumulations in the tissues of A.woodiana increased with exposure time and concentration,except for hemolymph,which reached the highest value on d 14.The bioconcentration factors(BCFs)increased with exposure time,but an inverse relationship was observed between BCFs and exposure concentration.During the elimination phase,the visceral mass showed the highest Cd elimination rate.In the kidney,digestive gland and gills,the elimination rates almost reached 40%,but their Cd concentrations were still higher than those in other tissues.Secondly,we detected and analyzed seven types of biochemical biomarkers(superoxide dismutase,SOD;catalase,CAT;glutathione peroxidase,GPx;glutathione S-transferase,GST;glutathione,GSH;total antioxidant capacity,TAC;malondialdehyde,MDA)and histopathological alterations in three important target organs gills,digestive gland and kidney of freshwater mussel A.woodiana exposed to 0.168 and 0.675 mg·L-1 Cd for 28 d.These three tissues showed tissue-specific antioxidant responses.The significant inductions various of biomarkers were observed in the gills(GST and MDA),digestive gland(GPx,GSH,TAC and MDA)and kidney(TAC and MDA).And the significant inhibitions were occurred in the gills(SOD and CAT)and kidney(SOD,CAT,GPx,and GST).Many histopathological alterations were also observed in the three tissues and were consistent with the MDA results.The tissue damages in the gills and digestive gland were increased as the Cd exposure time extending.Notably,the kidney showed significant damages on d 7 more than on d 28 exposed to Cd.And thirdly,we analyzed the expressions of mt,nrf2,keap1,cu/znsod,cat,gst and trx m RNA in the gills and kidney of A.woodiana to 0.168 and 0.675 mg·L-1 Cd for 28 d.The results showed that the expression of these seven genes in the gills were too low.The trusted Ct value could not be detected by q PCR,and the Sq-PCR showed that the bands of the gene fragments were mostly dark.After Cd exposure,the kidney gst m RNA decreased significantly.After the 0.168 mg·L-1 Cd exposure,kidney mt m RNA was up-regulated at d 7 and down-regulated at d 28;kidney nrf2 m RNA was up-regulated,accompanied by keap1 m RNA down-regulated,and cu/znsod,cat and trx m RNA up-regulated.After 0.675 mg·L-1 Cd exposure,kidney mt m RNA was up-regulated,kidney nrf2 m RNA was unchanged.The expressions of keap1,cu/znsod,cat and trx m RNA were all down-regulated.Therefore,we conclude that:1.The kidney,gills and digestive gland of A.woodiana are target tissues for subchronic Cd toxicity.Cd accumulation in the gills can reflect short-term Cd exposure,while those in the kidney and digestive gland indicate longer Cd exposure.2.The visceral mass,kidney,gills and mantle of A.woodiana are important tissues for to Cd elimination.3.The IBR/n index integrated SOD,CAT,GPx,GST and GSH in the gills of A.woodiana could be used to monitor short-term and high-dose of Cd exposure,while the kidney could be used to monitor long-term and low-dose of Cd exposure.Histopathology and GPx activity in the digestive gland,GST activity and MDA content in the gills and SOD activity in the kidney are suitable biomarkers for Cd contamination in aquatic ecosystems.4.The expression of gst m RNA in the kidney are suitable molecular biomarkers for Cd pollution in aquatic environment.