Studies On Regulation Of CpG ODNs On The Immune Response Of Chinese Mitten Crab Eriocheir Sinensis

CpG oligodeoxynucleotides (CpG ODN), which can mimic the immune stimulatoryactivity of bacterial DNA, is a kind of synthetic oligodeoxynucleotides (ODN) containingunmethylated CpG dinucleotides motifs. Fingding out that CpG ODN was able to activatethe immune system of human, mamarian and fish etc., many studies had proved that it wasa kind of good non-specificity immunostimulants and-specific immunology adjuvant. Yetthere was no relevant report about the effects of immune response or action mechnisomson Chinese mitten crab regulated by CpG ODN.In order to determine the immunomodulatory effects of CpG ODNs in the Chinesemitten crab (Eriocheir sinensis), the experiment one was processed. Crab were coelominjected with 1, 5, 25μg ODN-1670(containing 1 CpG motif) and ODN-R (containing 1inverted CpG motif, GpC) kg^-1crab weight. A series ,of immunology parameters werecollected at 0, 1, 3, 5, and 8 days after administration of CpG ODNs. At the same time, onegroup injected with 0.1mL saline and another blank group were set as controls. Crab werechallenged at 1.2×10⁷ colony forming units (cfu)/kg crab weight with a virulent strain ofAeromonas hydrophila (CL99920) of E. sinensis at the sencond sampling time (3d), andmortalities were recorded over a 7-day period.The results demonstrated that groupsinjected with ODN-1670 showed a remarkble higher number of THC , hyaline cells (HC)and granular cells (GC) than controls; while the haemocyte phagocytic activity and therespiratory burst (release of superoxide anion) increased significantly; also the serumparameters, including phenoloxidase (PO), peroxidase (POD), superoxide dismutase(SOD), acid phosphatase (ACP), alkaline phosphatase (ALP), lysozyme (LSZ), homolysin,antimicrobial activity(Ua) were assayed to be significantly increased than the controls. Atthe same time, the ODN-1670 treated crab's ability of anti-infected by Aeromonas hydrophila was remarkably enhanced. Besides, excepted the PO activity in serum, no moresignificant difference of the parameters was detected after the administration of ODN-R.Therefore, the present study results indicated that ODN-1670 could enhance thenon-specific immune response and resistance to potential infection of E. sinensis .So it is anew potential type of immunity-stimulant for E. sinensis. Furthermore, the inducement ofimmunity responses of ODN- 1670 is dose-dependent.In an attempt to determine which signal transduction pathway is involved in the CpGODNs activating proPO system in Chinese mitten crab, the experiment two was carried out.The exsomalized crab haemocytes, which were cultured on M199 medium, were separatelytreated with certain concentrations of CpG ODN and activators or inhibitors for specificsignalling components. The CpG ODN induced proPO system activation signaltransduction pathway was appraised by surveying the variety of PO activity in the crabhaemocytes. The results showed that both ODN-1670 and ODN-R could activate theproPO system. The signaling pathway of the proPO system activation stimulated by thetwo ODNs appeared to be some similar to each other, both the signaling passways mayinclude G-protein mediated PKC way, and ODN-1670 induced proPO activation wasnegatively regulated via the tyrosine kinase pathway. In addition, the cAMP-dependentprotein kinase (cAPK) pathway maybe was not involved in the ODNs-induced proPOsystem activation.