A single immunization with HA DNA vaccine by electroporation induces early protection against H5N1 avian influenza virus challenge in mice

<p>Abstract</p> <p>Background</p> <p>Developing vaccines for the prevention of human infection by H5N1 influenza viruses is an urgent task. DNA vaccines are a novel alternative to conventional vaccines and should contribute to the prophylaxis of emerging H5N1 virus. In this study, we assessed whether a single immunization with plasmid DNA expressing H5N1 hemagglutinin (HA) could provide early protection against lethal challenge in a mouse model.</p> <p>Methods</p> <p>Mice were immunized once with HA DNA at 3, 5, 7 days before a lethal challenge. The survival rate, virus titer in the lungs and change of body weight were assayed to evaluate the protective abilities of the vaccine. To test the humoral immune response induced by HA DNA, serum samples were collected through the eye canthus of mice on various days after immunization and examined for specific antibodies by ELISA and an HI assay. Splenocytes were isolated after the immunization to determine the antigen-specific T-cell response by the ELISPOT assay.</p> <p>Results</p> <p>Challenge experiments revealed that a single immunization of H5N1 virus HA DNA is effective in early protection against lethal homologous virus. Immunological analysis showed that an antigen-specific antibody and T-cell response could be elicited in mice shortly after the immunization. The protective abilities were correlated with the amount of injected DNA and the length of time after vaccination.</p> <p>Conclusion</p> <p>A single immunization of 100 μg H5 HA DNA vaccine combined with electroporation was able to provide early protection in mice against homologous virus infection.</p

Amiloride Enhances Antigen Specific CTL by Faciliting HBV DNA Vaccine Entry into Cells

The induction of relatively weak immunity by DNA vaccines in humans can be largely attributed to the low efficiency of transduction of somatic cells. Although formulation with liposomes has been shown to enhance DNA transduction of cultured cells, little, if any, effect is observed on the transduction of somatic tissues and cells. To improve the rate of transduction, DNA vaccine delivery by gene gun and the recently developed electroporation techniques have been employed. We report here that to circumvent requirement for such equipment, amiloride, a drug that is prescribed for hypertension treatment, can accelerate plasmid entry into antigen presenting cells (APCs) both in vitro and in vivo. The combination induced APCs more dramatically in both maturation and cytokine secretion. Amiloride enhanced development of full CD8 cytolytic function including induction of high levels of antigen specific CTL and expression of IFN-γ+perforin+granzymeB+ in CD8+ T cells. Thus, amiloride is a facilitator for DNA transduction into host cells which in turn enhances the efficiency of the immune responses

Dendritic Cells Transfected with scFv from Mab 7.B12 Mimicking Original Antigen gp43 Induces Protection against Experimental Paracoccidioidomycosis

Paracoccidioidomycosis (PCM), endemic in Latin America, is a progressive systemic mycosis caused by Paracoccidioides brasiliensis (P. brasiliensis), which primarily attacks lung tissue. Dendritic cells (DCs) are able to initiate a response in naïve T cells, and they also participate in Th-cell education. Furthermore, these cells have been used for therapy in several disease models. Here we transfected DCs with a plasmid (pMAC/PS-scFv) encoding a single chain variable fragment (scFv) of an anti-Id antibody that is capable of mimicking gp43, the main antigenic component of P. brasiliensis. First, Balb/c mice were immunized subcutaneously with pMAC/PS-scFv and, after seven days, scFv protein was presented to the regional lymph nodes cells. Moreover, we showed that the DCs transfected with scFv were capable of efficiently activating proliferation of total lymph node cells and inducing a decrease in lung infection. Therefore, our results suggested that the use of scFv-transfected DCs may be a promising therapy in the paracoccidioidomycosis (PCM) model

Toll-like receptor-9 induced by physical trauma mediates release of cytokines following exposure to CpG motif in mouse skin

The skin plays a crucial role in defence against microbial infection via the innate immune system, but the exact cellular mechanisms of this defence are not well understood. Toll-like receptors (TLRs), a newly recognized 10-member family of vertebrate pattern recognition receptors (PRRs), have been identified as crucial mediators of innate immune recognition. Although both TLR2 and TLR4 have been detected in normal human skin, little is known about the expression and function of TLR9, a CpG motif receptor, in skin. In this study, reverse transcription–polymerase chain reaction and in situ hybridization analysis were used to identify TLR9 mRNA expression in mouse skin. Results showed that TLR9 mRNA was not detected in normal mouse skin, but its presence in skin could be induced by intradermal injection of either normal saline, or the bacteria-based CpG motif in a time- and volume-dependent manner. Furthermore, intradermal injection of CpG motif induced increased expression of mRNAs for proinflammatory cytokines such as interleukin (IL)-1, IL-6, IL-12 and tumour necrosis factor α. This suggests that TLR9, while not present basally in skin, can be induced by physical trauma and then mediate responses to CpG motif. In conclusion, TLR9 is involved in the innate immune response in skin and that it may have a role in secondary inflammation following physical trauma such as epidermal damage or microbial infection. This role of TLR9 may help explain the previously identified enhancement of DNA immunization by CpG ODN

CpG motif acts as a ‘danger signal’ and provides a T helper type 1-biased microenvironment for DNA vaccination

The method of delivering a DNA vaccine can influence the type of immune response induced by the vaccine. Application of a DNA vaccine by gene gun typically induces a T helper type 2 (Th2)-type reaction, whereas needle inoculation triggers a Th1 response. In the present study, we found that physical trauma, gold-particle bombardment and the CpG motif can act as ‘danger signals’ that recruit inflammatory cells to damaged tissues. Analysis of the cytokine profiles of draining lymph nodes or lymph-node-derived mononuclear cells from different groups by real-time reverse transcription–polymerase chain reaction revealed that, while gene-gun-bombardment induced a Th2-type cytokine microenvironment with increased interleukin-4 (IL-4) and IL-10 mRNA expression and almost no increase in IL-12 and interferon-γ mRNA expression in draining lymph nodes, intradermal injection as well as subcutaneous injection of muscle induced the opposite. We further studied whether the addition of the CpG motif can switch the Th2-type cytokine microenvironment produced by gene-gun bombardment in draining lymph nodes. Results showed that the addition of the CpG motif can increase IL-12 mRNA expression in draining lymph nodes whether induced by intradermal injection, intramuscular injection, or gene-gun bombardment. These data suggest that delivery of the CpG motif induced a Th1-biased microenvironment in draining lymph nodes. Taken together, the CpG motif can act as a ‘danger signal’ and Th1 immune response enhancer in DNA vaccination. These results may help to explain the mechanism of different types of immune response induced by DNA vaccines delivered by different routes and facilitate the application of DNA vaccines

DNA vaccines and allergic diseases

10.1111/j.1440-1681.2006.04405.xClinical and Experimental Pharmacology and Physiology335-6546-55