RESUMO
Schistosomiasis is considered the most important human helminthiasis in terms of morbidity and mortality. In this study, comparative soluble proteomic analysis of normal cercariae and ultraviolet-irradiated attenuated cercariae (UVAC) from Schistosoma japonicum were carried out in view of the high efficiency of irradiation-attenuated cercariae vaccine. The results revealed that some proteins showed significant differential expression in the parasite after treatment with ultraviolet light. Total 20 protein spots were identified by mass spectrometry, corresponded to five groups according to their functions in the main that were structural and motor proteins (actin, et al.), energy metabolism associated enzymes (glyceraldehydes-3-phosphage dehydrogenase, et al.), signaling transduction pathway-associated molecules (14-3-3 protein, et al.), heat shock protein families (HSP 70 family, et al.), and other functional proteins (20S proteasome). Furthermore, our results indicated that the differential expression of the proteins by ultraviolet irradiation may be, at least partially, acquired by regulating the mRNA levels of corresponding proteins. These results may provide new clues for further exploring the mechanism of protective immunity induced by UVAC and may shed some light on the development of vaccines against schistosomiasis.
Assuntos
Proteínas de Helminto/análise , Proteoma/análise , Schistosoma japonicum/química , Schistosoma japonicum/efeitos da radiação , Raios Ultravioleta , Animais , Eletroforese em Gel Bidimensional , Perfilação da Expressão Gênica , Espectrometria de MassasRESUMO
Researches on genes expressed in a cercarial stage-specific manner may help us understand the molecular events and functions during schistosome invasion of skin. A genomic clone encoding 8-kDa calcium-binding protein (SjCa8) specifically expressed in cercariae and skin-stage schistosomulum (transformed within 3 h) was obtained from cercariae. Recombinant protein was expressed in vector pET32a (+) and purified using a Ni-NTA purification system. The target protein SjCa8 was determined by matrix-assisted laser desorption/ionization time-of-flight (TOF)/TOF mass spectrometer after thrombin digestion and dialysis. Reverse transcriptase polymerase chain reaction and Western blot revealed SjCa8 can be detected in cercaria and skin-stage schistosomulum but not lung-stage schistosomulum, adult, or egg and was localized to head gland, penetration gland tubes, and penetration glands where Ca(2+) was abundant, and the cercarial tegument (but not tegument of tail) and body-tail junction. Furthermore, SjCa8 was interestingly detected in cercarial secretions. The characterization of SjCa8 indicated that it may undergo structural and physiological modifications, including repair of the surface membrane, changes in permeability that account for the loss of water tolerance, activities of calcium-depending enzymes, and immune signaling, etc. Furthermore, vaccination with rSjCa8 plus adjuvant induced protective effect with 50.39% worm reduction rate and significantly high hepatic and intestine egg reduction rates (54.16%, 50.63%, respectively), which is possibly mediated through an apparent induction of Th1-type immune response for strikingly high level of IgG2a and IgG2b developed in immunized C57BL/6 mice.
Assuntos
Antígenos de Helmintos , Proteínas de Ligação ao Cálcio , Schistosoma japonicum/imunologia , Esquistossomose Japônica , Vacinas Sintéticas , Animais , Anticorpos Anti-Helmínticos/sangue , Antígenos de Helmintos/genética , Antígenos de Helmintos/imunologia , Antígenos de Helmintos/metabolismo , Western Blotting , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/imunologia , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Helminto/genética , Proteínas de Helminto/imunologia , Proteínas de Helminto/metabolismo , Imunização , Imuno-Histoquímica , Estágios do Ciclo de Vida , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Schistosoma japonicum/genética , Schistosoma japonicum/crescimento & desenvolvimento , Schistosoma japonicum/metabolismo , Esquistossomose Japônica/imunologia , Esquistossomose Japônica/parasitologia , Esquistossomose Japônica/prevenção & controle , Células Th1/imunologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologiaRESUMO
OBJECTIVE: To study the immunomodulation effect of the recombination Sj16 from Schistosoma japonicum (reSj16) on inflammation response of host. METHODS: reSj16 expressed in pGEX-4T-1 was purified by GST purification kit and detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectra (MS). The immunomodulation effect of reSj16 was observed by means of dimethylbenzene inducing mouse ear edema model, carrageenan inducing rat voix pedis swell model and acetic acid inducing mouse experiment peritonitis model. RESULTS: The soluble protein of reSjl6 was obtained and identified by SDS-PAGE and MS. reSjl6 1.0, 5.0, 10.0 microg/kg evidently suppressed the mouse ear edema induced by dimethyl-benzene, significantly mitigated the rat voix pedis swelling induced by carrageenan, and remarkably suppressed the increase of the capillary permeability of abdominal cavity in experiment peritonitis mouse model. CONCLUSION: The results further prove that Sj16 may be a potential immunosuppressive molecule and may have a notable effect on immunomodulation.
