Study On The Function Of Serine Protease Inhibitors Serpin B6 And Serpin 4848 In Cysticercus Cellulosae

Cysticercosis is a zoonotic foodborne parasitic disease caused by Cysticercus cellulose,a larvae of Taenia solium,which seriously endangers the health of humans and animals and can even lead to death.An important reason why porcine cysticercosis is still prevalent worldwide is the immune evasion phenomenon during the invasion of insects,serine protease inhibitors(serpin)play an important role in the invasion process of insects,and the serpin expressed by porcine cysticercosis makes the worm body evade the immune attack of the host by regulating or inhibiting the endogenous and exogenous protease action.In this study,serpin B6 and serpin 4848 proteins were studied,and the possibility of serpin participating in or mediating immune evasion was explored through molecular biology,bioinformatics,immunology and other techniques,with the goal of elucidating the mechanism of immune evasion in insects.Analysis of serpin B6 and serpin 4848 genes by bioinformatics analysis;Using molecular biology methods and techniques,the recombinant soluble proteins of serpin B6 and serpin 4848 of Cysticercosis were expressed and purified.Enzyme activity inhibition test was verified by coloring substrate method.Through coagulation tests,the possibility of serpin B6 and serpin 4848 proteins participating in the coagulation cascade was explored.Histological localization of serpin B6 and serpin 4848 proteins by immunohistochemical methods.Bioinformatics analysis showed that serpin B6 and serpin 4848 did not contain signal peptide sequences and transmembrane structures,and evolutionary tree analysis showed that serpin affinity with Polyadenos echinococcus serpin may have a similar biological function recently.Prokaryotic expression and purification of serpin B6 and serpin 4848 proteins.The western blotting results showed that it can have a specific reaction with the positive serum of porcine cysticercosis,has good reactivity,and canprovide new ideas for the screening and diagnosis of porcine cysticercosis.Enzyme activity inhibition tests have shown that both serpin B6 and serpin 4848 can react with and inhibit the activity of trypsin,a mammalian digestive enzyme,and are concentration-dependent.At a concentration of 14 μg/m L,the inhibition rate of serpin B6 can reach 79.36 %,and the inhibition rate of serpin 4848 can reach 87.56 %.It is speculated that these two proteins can play an immune evasion role in the stage when the parasite enters the intestine.The results of APTT and PT data from the coagulation test showed that neither serpin B6 nor serpin 4848 participated in the coagulation cascade.Rabbit polyclonal antibody serum was successfully prepared,and the antibody titer was determined by ELISA method,and the antibody titer reached 1:128000.Immunohistochemical tests showed that both serpin B6 and serpin 4848 were expressed in porcine cysticercosis,but the expression was not high.It was expressed in the suction cup and fiber layer region,respectively,and the expression of serpin 4848 at the suction cup was higher than that of serpin B6.It is speculated that these two proteins may be involved in the intake of nutrients in the insect body and promote the metabolism of the insect body.