The biochemical and immunological characterization of two serpins from Clonorchis sinensis.

Serpins (serine proteinase inhibitors) are evidenced to regulate numerous biological processes such as immunoregulation in parasitic helminths. The functions of serpins from Clonorchis sinensis remain unclear to date. In this study, two serpin genes, respectively denominated as CsproSERPIN and CsSERPIN2, had been selected from metacercaria cDNA library of C. sinensis. The biochemical activities of both recombinant proteins (rCsproSERPIN and rCsSERPIN2) were analyzed by assays of inhibition on some serine or cysteine proteases, the results showed that rCsproSERPIN significantly inhibited trypsin, chymotrypsin and thrombin, while rCsSERPIN2 inhibited only chymotrypsin. Moreover, cytokine and antibody measurements indicated that rats subcutaneously immunized with rCsproSERPIN and rCsSERPIN2 respectively developed a strong IFN-γ production and IgG2a levers of sera were higher than IgG1. Besides, immunoblot assays revealed that the rCsproSERPIN and rCsSERPIN2 could be recognized by the sera of rats infected with C. sinensis and the sera of rabbits immunized by excretory/secretory products. Furthermore, immunofluorescence assays illuminated the two were similarly localized in the reproductive organs such as vitelline glands, testis and eggs in adult stage. In short, all the results collectively indicated that CsproSERPIN and CsSERPIN2 might play important role in the parasite development by preventing the parasite from digestion by exogenous serine proteases, as well as CsproSERPIN and CsSERPIN2 probably involved in immunoregulation of host by inducing Th1-biased type cytokines in rats.