Tissue distribution of a plasmid DNA encoding Hsp65 gene is dependent on the dose administered through intramuscular delivery

In order to assess a new strategy of DNA vaccine for a more complete understanding of its action in immune response, it is important to determine the in vivo biodistribution fate and antigen expression. In previous studies, our group focused on the prophylactic and therapeutic use of a plasmid DNA encoding the Mycobacterium leprae 65-kDa heat shock protein (Hsp65) and achieved an efficient immune response induction as well as protection against virulent M. tuberculosis challenge. In the present study, we examined in vivo tissue distribution of naked DNA-Hsp65 vaccine, the Hsp65 message, genome integration and methylation status of plasmid DNA. The DNA-Hsp65 was detectable in several tissue types, indicating that DNA-Hsp65 disseminates widely throughout the body. The biodistribution was dose-dependent. In contrast, RT-PCR detected the Hsp65 message for at least 15 days in muscle or liver tissue from immunized mice. We also analyzed the methylation status and integration of the injected plasmid DNA into the host cellular genome. The bacterial methylation pattern persisted for at least 6 months, indicating that the plasmid DNA-Hsp65 does not replicate in mammalian tissue, and Southern blot analysis showed that plasmid DNA was not integrated. These results have important implications for the use of DNA-Hsp65 vaccine in a clinical setting and open new perspectives for DNA vaccines and new considerations about the inoculation site and delivery system.     

Analysis of binding properties and specificity through identification of the interface forming residues (IFR) for serine proteases <Emphasis Type="Italic">in silico</Emphasis> docked to different inhibitors

Background  

Enzymes belonging to the same super family of proteins in general operate on variety of substrates and are inhibited by wide selection of inhibitors. In this work our main objective was to expand the scope of studies that consider only the catalytic and binding pocket amino acids while analyzing enzyme specificity and instead, include a wider category which we have named the Interface Forming Residues (IFR). We were motivated to identify those amino acids with decreased accessibility to solvent after docking of different types of inhibitors to sub classes of serine proteases and then create a table (matrix) of all amino acid positions at the interface as well as their respective occupancies. Our goal is to establish a platform for analysis of the relationship between IFR characteristics and binding properties/specificity for bi-molecular complexes.     

Permeability of the urban matrix to arboreal gliding mammals: Sugar gliders in Melbourne,Australia

Habitat corridors that facilitate functional connectivity are a fundamental component of wildlife conservation in fragmented landscapes. However, the landscape matrix separating suitable habitat is not uniformly impermeable to movement and management to increase matrix permeability could be an alternative means to maintain connectivity. Gliding mammals are particularly sensitive to fragmentation because their movements are constrained by glide distance thresholds. Populations of gliders in cities are at risk of being isolated by increasing habitat loss and urban development, yet little is known about how the urban matrix affects glider movement. Here we investigate how the level of urbanization and tree cover in the matrix influence matrix permeability to sugar gliders (Petaurus breviceps) within suburban forest reserves. Twenty‐two sugar gliders were radio‐tracked over winter and summer at four reserves. Boundary crossing behaviour was measured as the number of times each glider crossed into the matrix, and matrix permeability was determined as the maximum distance travelled by gliders into the matrix. The majority of gliders (81%) were located in the matrix at least once, and rates of boundary crossing were consistent across urbanization and tree cover levels. Matrix permeability was negatively affected by matrix urbanization, but not by matrix tree cover, and no interaction effects were found. Although distances travelled by gliders into the matrix did not exceed 180 m, they were comparable with typical movement distances by gliders in reserves. Our results demonstrate that the urban matrix can provide suitable habitat for gliding mammals to move and forage, but that increased urbanization may inhibit glider use of the matrix irrespective of tree cover. This finding has implications for conservation planning and suggests that structurally connected areas may not be used if movement behaviour is inhibited. Conversely, management of matrix permeability could be used to maintain connectivity without needing to construct physical corridors.