66 research outputs found
Altering an Artificial Gagpolnef Polyprotein and Mode of ENV Co-Administration Affects the Immunogenicity of a Clade C HIV DNA Vaccine
HIV-1 candidate vaccines expressing an artificial polyprotein comprising Gag, Pol and Nef (GPN) and a secreted envelope protein (Env) were shown in recent Phase I/II clinical trials to induce high levels of polyfunctional T cell responses; however, Env-specific responses clearly exceeded those against Gag. Here, we assess the impact of the GPN immunogen design and variations in the formulation and vaccination regimen of a combined GPN/Env DNA vaccine on the T cell responses against the various HIV proteins. Subtle modifications were introduced into the GPN gene to increase Gag expression, modify the expression ratio of Gag to PolNef and support budding of virus-like particles. I.m. administration of the various DNA constructs into BALB/c mice resulted in an up to 10-fold increase in Gag- and Pol-specific IFNγ+ CD8+ T cells compared to GPN. Co-administering Env with Gag or GPN derivatives largely abrogated Gag-specific responses. Alterations in the molar ratio of the DNA vaccines and spatially or temporally separated administration induced more balanced T cell responses. Whereas forced co-expression of Gag and Env from one plasmid induced predominantly Env-specific T cells responses, deletion of the only H-2d T cell epitope in Env allowed increased levels of Gag-specific T cells, suggesting competition at an epitope level. Our data demonstrate that the biochemical properties of an artificial polyprotein clearly influence the levels of antigen-specific T cells, and variations in formulation and schedule can overcome competition for the induction of these responses. These results are guiding the design of ongoing pre-clinical and clinical trials
Der IT-gestützte Behandlungspfad als Routinearbeitsinstrument in der Hüftendoprothetik Neue Wege in der Qualitätssicherung
Studies in Heterobasidiomycetes, Part 38 Sphacelotheca polygoni-persicariae G. Demi & Oberw. spec. nov.
The DNA damaging drug cyproterone acetate causes gene mutations and induces glutathione-S-transferase P in the liver of female Big Blue<sup>TM</sup> transgenic F344 rats.
The gestagenic and antiandrogenic drug cyproterone acetate (CPA) is mitogenic, tumorigenic and induces DNA-adducts and DNA-repair synthesis in rat liver. Thus CPA is expected to be mutagenic. However in vitro mutagenicity test systems were negative. To examine whether CPA induces mutations in rat liver, the in vivo mutation assay based on Big Blue transgenic F344 rats was employed. Single oral doses of 25, 50, 75, 100 and 200 mg CPA/kg b.w. respectively were administered to female Big Blue rats. Six weeks after treatment, liver DNA was assayed for mutations. At the highest dose, 200 mg CPA/kg b.w., the frequency of (17 +/- 4) x 10(-6) spontaneous mutations was increased to a maximum of (80 +/- 8) x 10(-6) mutations. One-hundred and 75 mg CPA/kg b.w. resulted in mutation frequencies of (35 +/- 5) and (27 +/- 5) x 10(-6), respectively. The mutation frequency at doses of 50 and 25 mg CPA/kg b.w. was similar to that of vehicle treated controls. Statistical analysis of the dose-effect relationship revealed that it was not possible to decide whether a threshold dose exists or not. DNA adducts were analyzed by the 32P-postlabelling technique. The total level of the major and the two minor adducts observed in the autoradiograms increased between doses of 25 to 75 mg CPA/kg b.w. to a maximum of approximately 12,000 +/- 3000 adducts per 10(9) nucleotides. The level did not further increase significantly with 100 and 200 mg CPA/kg b.w. After CPA treatment no preneoplastic liver foci were observed. However, single glutathione-S-transferase placental form (GST-P) positive hepatocytes were observed and the frequency was dependent on the dose. These cells are not supposed to represent initiated cells, since they occurred only transiently after 6 weeks and disappeared thereafter completely. In conclusion, our results demonstrate that CPA is mutagenic in vivo. The mutation frequency increased at high CPA doses, when the increase of the DNA adduct formation had already ceased. This suggests that the mitogenic activity of CPA is required to express the mutations
Diethylnitrosamine induces long-lasting re-expression of insulin-like growth factor II during early stages of liver carcinogenesis in mice.
The expression of the insulin-like growth factor II (IGF-II) gene (lgf2) in rodents is completely abrogated in almost all adult tissues. A prominent exception are neoplasms in which IGF-II frequently serves as an autocrine growth factor. We have investigated the potential role of lgf2 expression during liver carcinogenesis. After application of diethylnitrosamine (DEN) preneoplastic foci and adenomas emerged in liver tissue of wildtype and phosphoenolpyruvate carboxykinase (PEPCK)-IGF-II transgenic mice. Surprisingly, number and size of preneoplastic foci were not significantly increased in PEPCK-IGF-II mice as compared with wild-type animals. In situ preparation showed that early adenomas expressed lgf2 transcripts. Reverse transcriptase polymerase chain reaction (RT-PCR) and restriction enzyme analysis confirmed that DEN treatment had indeed reactivated the hepatic expression of murine lgf2 in control mice in a dose-dependent manner. This re-expression of lgf2 persisted for at least 18 months. Species-specific RT-PCR analyses also revealed the presence of murine lgf2 mRNAs in some PEPCK-IGF-II mice. A similar reactivation of lgf2 was detected in bovine growth hormone transgenic mice which develop hepatocellular neoplasms with high frequency. Our results suggest that reactivation of lgf2 is an early event during hepatocarcinogenesis in mice. Its appearance in two independent animal models suggests that lgf2 may be important at pivotal checkpoints of hepatocarcinogenesis
Studies in Heterobasidiomycetes. Part 24. On Ustilago violacea (Pers.) Rouss. from Saponaria officinalis L.
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