Effect Of Ts-cystatin Recombinant Attenuated Salmonella On Immune Response In Mice

Trichinosis is caused by ingesting undercooked meat containing the larvae of thenematode Trichinella. The infection rate in swine remains high in prevalent areas andposes a great threat to human health. Salmonella-carried targeted genes are extremelyeffective at inducing both cellular and humoral immunity. Administering aSalmonella-carried targeted gene orally can mimic the natural process of Trichinellainfection, which targets the intestinal mucosa, where professional antigen-presentingcells (APCs) are abundant in the intestinal Peyer’s patches (PPs). In S. typhimurium,one such regulon is modulated by the PhoP (transcriptional activator) and PhoQ(sensor kinase) proteins, which are essential to S. typhimurium pathogenesis andsurvival within macrophages. These regulators control the transcription of multipleunlinked phoP-activated and phoP-repressed genes. PhoP/PhoQ null mutants aremarkedly attenuated in BALB/c mice and are effective vaccines in these animals. SophoP/phoQ deletion mutants are attenuated strain of Salmonella and provide securityguarantees for host. Currently, phoP/phoQ mutants were widely used in a variety ofpharmacology experimental studies in mice and immune models.Trichinella has a complex life cycle that involves both enteral and parenteralphases, which stimulate both mucosal and systemic immune responses. Althoughmuch remains unknown about the mechanism of Trichinella invasion, recent studieshave successfully focused on some of the immune regulate antigens of this parasite. Inour previous works, the cDNA encoding a cystatin-like protein (Ts-cystatin) wasidentified by immunoscreening intestinal muscle larvae cDNA libraries with serumfrom pigs experimentally infected with20000T. spiralis muscle larvae. Further,immunoscreening cDNA libraries of all Trichinella life stages that it is a highlyimmunogenic/antigenic gene. The main goal of this work was to determine the abilityof this gene to induce an immune response, when expressed by live attenuatedSalmonella. Although Salmonella is reported to be competent at carrying both prokaryotic and eukaryotic plasmids and then provoking an immune response, manyresearch groups have targeted gene expression in E. coli and have collected the fusionprotein. Although this method can provide a large amount of protein, the expressedprotein may differ from the native active protein in terms of structure and function.We chose a eukaryotic expression system based on several comparisons of theimmunogenicities of two Salmonella administration schemes, which indicated that theeukaryotic expression system is superior. Humoral IgG and mucosal IgA weremeasured to examine the antibody response, and a splenocyte proliferation assay wasperformed to evaluate cellular responses. To explore whether the Th1or Th2responsewas induced, Th1-and Th2-specific cellular transcription factors were examined inaddition to the cytokine profile. Furthermore, changes in T lymphocyte andmacrophage populations were detected by flow cytometry. Lastly, parasitologicalexamination was examined. The results showed that Ts-cystatin induced aTh1/Th2-mixed type of immune response and decreased STAT6transcription. Theintestinal adult recovery increased by15.8%in the Ts-cystatin group, the Ts-cystatingroup fecundity rate was decreased by89%. Furthermore, the number of musclelarvae did not change compared with the control group. In conclusion, our resultssuggest that Ts-cystatin plays an important role in Trichinella resistance to rapidexpulsion by the host and is worth further study.