Evaluation of Protective Immune Response Induced by a DNA Vaccine Encoding GRA8 against Acute Toxoplasmosis in a Murine Model

Toxoplasma gondii is an apicomplexan zoonotic protozoan parasite that infects most species of warm-blooded animals, including humans. The heavy incidence and severe or lethal damage caused by T. gondii infection clearly indicate a need for the development of an effective vaccine. T. gondii GRA8 is a member of the dense granules protein family and is used as a marker of acute infection. In the present study, we evaluated the protective immunity induced by DNA vaccination based on a recombinant eukaryotic plasmid, pDsRed2-GRA8, against acute toxoplasmosis in mice. BALB/c mice were intramuscularly immunized with the pDsRed2-GRA8 plasmid and then challenged by infection with the highly virulent GFP-RH strain of T. gondii. The specific immune responses and protective efficacy against T. gondii of this vaccine were analyzed by measuring cytokine and serum antibody titers, splenocyte proliferation assays, and the survival times of mice after challenge. Our results showed that mice immunized with pDsRed2-GRA8 demonstrated specific humoral and cellular responses, induced higher IgG antibody titers with predominant IgG2a production; increased levels of IL-10, IL-12 (p70), IFN-γ, TNF-α, and splenocyte proliferation; and prolonged survival times compared to those of control mice. The present study showed that DNA immunization with pDsRed2-GRA8 induced humoral and cellular immune responses, and all immunized mice showed greater Th1-type immune responses and longer survival times than those of control mice. These results indicated that T. gondii GRA8 DNA immunization induces a partial protective effect against acute toxoplasmosis.

Effects of miR-192 on the expression of ZEB2 and the abilities of migration and invasion of colorectal cancer cells / 肿瘤

Objective: To investigate the effects of miR-192 on the expression of zinc-finger E-box binding homeobox 2 (ZEB2) and the abilities of migration and invasion of colorectal cancer (CRC) SW480 and SW620 cells. Methods: After transfection of miR-192 mimics into SW480 and SW620 cells, the expression levels of miR-192 and ZEB2 mRNA and ZEB2 protein were detected by real-time fluorescence quantitative RT-PCR and Western blotting, respectively. The abilities of migration and invasion of SW480 and SW620 cells after transfection with miR-192 mimics were determined by wound healing and Transwell assays, respectively. The recombinant vector pmiR-ZEB2-wt and miR-192 mimics were co-transfected into SW480 cells. The binding site of 3′-untranslated region (3′-UTR) of ZEB2 gene with miR-192 was verified by Dual Luciferase™ Reporter Gene Assay. Results: As compared with the negative control (NC) group (transfection with miR-192 mimics-NC), the expression levels of miR-192 in SW480 and SW620 cells after transfection with miR-192 mimics were increased (P < 0.05), and the expression levels of ZEB2 mRNA and protein were decreased (P < 0.05), as well as the abilities of migration and invasion of SW480 and SW620 cells were inhibited (P < 0.05). The Dual Luciferase™ Reporter Gene Assay revealed that the luciferase activity in SW480 cells after co-transfection with recombinant vector pmiR-ZEB2-wt and miR-192 mimics was inhibited (P < 0.05), which indicated that the 3′-UTR of ZEB2 harbored a binding site for miR-192. Conclusion: ZEB2 may be one of the target genes for miR-192. Overexpression of miR-192 may down-regulate the ZEB2 expression and inhibit the migration and invasion of SW480 and SW620 cells. Copyright© 2014 by TUMOR.