Anti-viral and anti-tumor vaccines aim to induce cytotoxic CD8+ T cells (CTL) and antibodies. Conserved protein antigens, such as p24 from human immunodeficiency virus, represent promising component for elicitation CTLs, nevertheless with suboptimal immunogenicity, if formulated as recombinant protein. To enhance immunogenicity and CTL response, recombinant proteins may be targeted to dendritic cells (DC) for cross presentation on MHCI, where mannose receptor and/or other lectin receptors could play an important role. Here, we constructed liposomal carrier-based vaccine composed of recombinant p24 antigen bound by metallochelating linkage onto surface of nanoliposomes with surface mannans coupled by aminooxy ligation. Generated mannosylated proteonanoliposomes were analyzed by dynamic light scattering, isothermal titration, and electron microscopy. Using murine DC line MutuDC and murine bone marrow derived DC (BMDC) we evaluated their immunogenicity and immunomodulatory activity. We show that p24 mannosylated proteonanoliposomes activate DC for enhanced MHCI, MHCII and CD40, CD80, and CD86 surface expression both on MutuDC and BMDC. p24 mannosylated liposomes were internalized by MutuDC with p24 intracellular localization within 1 to 3 h. The combination of metallochelating and aminooxy ligation could be used simultaneously to generate nanoliposomal adjuvanted recombinant protein-based vaccines versatile for combination of recombinant antigens relevant for antibody and CTL elicitation.
- MeSH
- antigeny MeSH
- dendritické buňky MeSH
- HIV-1 * MeSH
- lidé MeSH
- liposomy metabolismus MeSH
- mannany metabolismus MeSH
- myši MeSH
- rekombinantní proteiny metabolismus MeSH
- vakcíny proti AIDS * imunologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Detection of serum galactomannan (GM) and (1,3)-β-d-glucan (BG) is considered useful for non-culture diagnosis of invasive pulmonary aspergillosis (IPA) in neutropenic patients. Only few studies evaluated these seromarkers in non-neutropenic patients suspected of having IPA. The aim of this study was to evaluate both tests together with the Aspergillus fumigatus-specific serum IgG and IgA (IgAG) test for serological IPA diagnosis in non-neutropenic patients. Sera from 87 patients suspected of having IPA were retrospectively analysed. Patients were categorised into groups of proven IPA (n = 10), putative IPA (n = 31) and non-IPA colonisation (n = 46). When the GM, BG and IgAG assays were used for patients included in the study, the sensitivity/specificity/positive predictive value (PPV)/negative predictive value (NPV) were 48.8%/91.3%/83.3%/66.7%, 82.9%/73.9%/73.9%/82.9% and 75.6%/95.7%/93.9%/81.5%, respectively. Thus, the highest specificity and PPV were confirmed for the IgAG assay. Improvements in the sensitivity and NPV were achieved by "at least one positive" analysis with the GM and BG assays, with the sensitivity/specificity/PPV/NPV values being 85.0%/69.6%/71.4%/84.2%. Nevertheless, the highest sensitivity and NPV were achieved by the "at least one positive" analysis combining the GM, BG and IgAG tests (97.6% and 96.8%, respectively). The involvement of the IgAG assay could improve IPA diagnosis in non-neutropenic patients by increasing the sensitivity and NPV when combined with the GM or BG assays. Furthermore, improvement was achieved by combining the GM, BG and IgAG assays using the "at least one positive test" strategy, especially if doubt exists.
- MeSH
- Aspergillus fumigatus chemie imunologie MeSH
- beta-glukany krev MeSH
- dospělí MeSH
- imunoglobulin A krev MeSH
- imunoglobulin G krev MeSH
- invazivní plicní aspergilóza diagnóza MeSH
- lidé středního věku MeSH
- lidé MeSH
- mannany krev MeSH
- mladiství MeSH
- mladý dospělý MeSH
- prediktivní hodnota testů MeSH
- protilátky fungální krev MeSH
- retrospektivní studie MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- senzitivita a specificita MeSH
- sérum chemie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- multicentrická studie MeSH
Heat shock proteins hsp70 and gp96 have been confirmed as adjuvants enabling induction of cell- and antibody-mediated immunity specific to associated protein or peptide antigens due to the activation of naive dendritic cells and supporting cross-presentation of associated antigen. An efficacious vaccine preventing HIV-1 infection should induce (1) antibodies neutralizing HIV-1 Env protein, preventing virus spreading and (2) CD4(+) Th1 and CD8(+) T cells specific to viral proteins, especially gag p24, important for elimination of HIV-1 infected cells. As p24 is relatively poorly recognized by dendritic cells, its targeting to DC is important for enhancement of vaccine efficacy. In this study, a p24 protein fused to the C- or N-terminus of murine hsp70 was produced as a recombinant protein and administered without any adjuvant to experimental BALB/c mice. Consequently, p24-specific cellular and humoral immune responses were measured. To minimize the effect of bacterial endotoxin, each protein was subjected to a repeated endotoxin phase extraction until each preparation contained less than 2.5 endotoxin unit (EU) per mg of antigen. In addition, endocytosis of p24 fused to hsp70 by dendritic cells and their activation were characterized. The fusion to hsp70 protein enhanced endocytosis of p24 as well as activation of dendritic cells in vitro. After immunization of mice, hsp70-p24 fusion protein induced the strongest p24-specific CD4(+) and CD8(+) T cells (IFN-γ production) and humoral (IgG2b) responses corresponding to Th1 type dominance, whereas p24-hsp70 or p24 itself induced weaker responses.
- MeSH
- antigeny virové imunologie MeSH
- CD8-pozitivní T-lymfocyty imunologie MeSH
- dendritické buňky imunologie MeSH
- endocytóza imunologie MeSH
- endotoxiny imunologie MeSH
- HIV korový protein p24 imunologie metabolismus MeSH
- imunizace MeSH
- myši inbrední BALB C MeSH
- myši MeSH
- nepřímá aktivace imunologie MeSH
- proteiny tepelného šoku HSP70 imunologie metabolismus MeSH
- rekombinantní fúzní proteiny imunologie MeSH
- Th1 buňky imunologie MeSH
- vakcíny proti AIDS imunologie MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- endopeptidasy biosyntéza MeSH
- reakce na tepelný šok MeSH
- Trichophyton MeSH
- Publikační typ
- techniky in vitro MeSH
- MeSH
- adjuvancia imunologická imunologie terapeutické užití MeSH
- alergeny imunologie izolace a purifikace terapeutické užití MeSH
- alergie imunologie terapie MeSH
- Bacteria imunologie MeSH
- desenzibilizace imunologická metody MeSH
- myši MeSH
- pyl imunologie MeSH
- rostlinné extrakty imunologie izolace a purifikace terapeutické užití MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- srovnávací studie MeSH
