Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
9442808 | Experimental Parasitology | 2005 | 10 Pages |
Abstract
The humoral and cellular responses to DNA vaccination of BALB/c mice with a novel antigen from the Fasciola hepatica saposin-like protein family (FhSAP-2) have been investigated. Two constructs were produced containing the FhSAP-2 DNA sequence, one intended for extracellular secretion of FhSAP-2 protein, and one expressing FhSAP-2 in the cytoplasm of a transfected cell. The constructs were tested in HEK 293T cells, with the secretory construct producing less detectable FhSAP-2 relative to cytoplasmic construct when observed by fluorescence. The size of expressed protein was confirmed by Western blot of cell lysate, but FhSAP-2 was undetectable in cell supernatants. Both, secretory and cytoplasmic constructs as well as FhSAP-2 recombinant protein were tested in mice. The antibody response elicited in mice vaccinated with the rFhSAP-2 induced high levels of IgG1, IgG2, and IgE as well as high levels of IL-10 and IFNγ indicating a mixed Th1/Th2 response. Vaccination of mice intramuscularly with the cytoplasmic FhSAP-2 construct resulted in a dominant IgG2a isotype antibody as well as a dominant IFNγ cytokine, with significant IgE, IgG1, and IL-10 responses also present, suggesting a mixed Th1/Th2 profile. Isotype and cytokine profiles elicited by the FhSAP-2 secretory construct were similar to those obtained with the cytoplasmic construct but at levels that were significantly lower. The results demonstrate that FhSAP-2 can be delivered as a DNA vaccine construct and induces a stronger Th1 response than the recombinant protein alone. This could result in an improvement in the immunoprophylactic potential of this candidate vaccine against F. hepatica.
Keywords
IL-10TrematodePMSFE. coliIFNγGSTIFAtPATh1Th2Concanavalin ADBBSDSPBSCatLnatural killerDNAimmunofluorescence assayenzyme linked immunosorbent assayEDTAdeoxyribonucleic acidEscherichia coliInterleukin 10Con AELISAsodium dodecyl sulfateCMVcytomegalovirusIntramuscularHigh performancetissue plasminogen activatorLuria brothmajor histocompatibility complexMHCPhosphate-buffered salineDNA vaccinationpolymerase chain reactionPCRCathepsin Lglutathione S-transferase
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Authors
Ana M. Espino, Antonio Osuna, Ramón Gil, George V. Hillyer,