فایل ورد کامل اشباح باکتریایی یک سیستم تحویل موثر برای واکسن های DNA می باشد

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تعداد صفحات این فایل: ۲۵ صفحه

بخشی از مقاله انگلیسیعنوان انگلیسی:Bacterial Ghosts Are an Efficient Delivery System for DNA Vaccines~~en~~

Mass implementation of DNA vaccines is hindered by the requirement of high plasmid dosages and poor immunogenicity. We evaluated the capacity of Mannheimia haemolytica ghosts as delivery system for DNA vaccines. In vitro studies showed that bacterial ghosts loaded with a plasmid carrying the green fluorescent protein-encoding gene (pEGFP-N1) are efficiently taken up by APC, thereby leading to high transfection rates (52–۶۰%). Vaccination studies demonstrated that ghost-mediated delivery by intradermal or i.m. route of a eukaryotic expression plasmid containing the gene coding for
-galactosidase under the control of the CMV immediate early gene promoter (pCMV
) stimulates more efficient Ag-specific humoral and cellular (CD4 and CD8) immune responses than naked DNA in BALB/c mice. The use of ghosts also allows modulating the major Th response from a mixed Th1/Th2 to a more dominant Th2 pattern. Intravenous immunization with dendritic cells loaded ex vivo with pCMV
- containing ghosts also resulted in the elicitation of
-galactosidase-specific responses. This suggests that dendritic cells play an important role in the stimulation of immune responses when bacterial ghosts are used as a DNA delivery system. Bacterial ghosts not only target the DNA vaccine construct to APC, but also provide a strong danger signal, acting as natural adjuvants, thereby promoting efficient maturation and activation of dendritic cells. Thus, bacterial ghosts constitute a promising technology platform for the development of more efficient DNA vaccines.

Nucleic1 acid vaccination has emerged as a powerful technology, which can be applied for the development of either prophylactic or therapeutic vaccines (1). The genes encoding the vaccine Ags are cloned into a eukaryotic expression plasmid, which is generally administered by i.m. injection or via biolistic skin bombardment with a gene gun. Then the biosynthetic machinery of the vaccinee’s cell is responsible for the in vivo expression of the corresponding gene. The presence of immunostimulatory motifs in the DNA further contributes to the elicitation of an immune response. However, routine implementation of this approach in humans still does not seem to be feasible. This is mainly due to poor immunogenicity and the requirement of extremely high plasmid dosages. The low efficiency of traditional naked DNA vaccination can be due, at least in part, to the fact that APC are not specifically targeted and the encoded Ag is not delivered in the context of an adequate danger signal. Bacterial ghosts are a novel nonliving vaccination technology platform, which is based on the conditional expression of the lethal lysis gene E from bacteriophage PhiX174 in Gram negatives (2– ۶). This leads to the formation of a transmembrane tunnel through the bacterial cellular envelope (Fig. 1D) (2). Due to the high internal osmotic pressure, the cytoplasm content is expelled through the tunnel (see Fig. 1D), resulting in an empty bacterial cell envelope (7). Bacterial ghosts retain all morphological, structural, and antigenic features of the cell wall and can be used as vaccine candidate per se. Alternatively, they can be exploited as a delivery system for proteins, which are either expressed and anchored to the envelopes before lysis or subsequently loaded (8). Bacterial ghosts can target APC and microvascular endothelial cells (9–۱۱). The envelope components might provide a danger signal through the activation of pattern recognition receptors, thereby acting as natural adjuvants (12). However, the endotoxic effects of free LPS are not observed, because the LPS is associated to the ghost envelopes (13). In this study, we evaluated the capacity of Mannheimia haemolytica ghosts as delivery system for DNA vaccines. In vitro studies demonstrate, for the first time, that ghosts are efficiently taken up by APC, thereby leading to high transfection efficiencies. Ghostmediated DNA delivery resulted in the elicitation of more efficient immune responses than using naked DNA, allowing also modulation of the obtained immune response from a mixed Th1/Th2 to a more dominant Th2 response pattern. Intravenous immunization with dendritic cells (DC)4 loaded ex vivo with plasmid-containing ghosts also resulted in the elicitation of specific humoral and cellular immune responses. Further in vitro studies demonstrated that bacterial ghosts promote efficient maturation and activation of DC. Thus, bacterial ghosts act as natural adjuvants, constituting a promising technology for the development of DNA vaccines.

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