Intracellular pathogens like <i>Salmonella</i> have developed various mechanisms to evade host defenses, and they can establish infections. Treatment and eradication are difficult due to our inability in achieving the optimum concentrations of cell-impermeable aminoglycosides like gentamicin within these cells. In this dissertation, we hypothesize that developing a novel core-shell methodology for incorporating high amounts of gentamicin into the cores with either hydrophilic or amphiphilic shell will be more effective than the free gentamicin in clearing intracellular <i>Salmonella</i> infection.
Hydrophilic core-shell nanostructures (N1) were made with block co-polymers of poly (ethylene oxide-<i>b</i>-sodium acrylate) blended with sodium polyacrylate (PAA<sup>-+</sup>Na) and complexed with the polycationic antibiotic gentamicin. N1 showed 20-25 fold higher gentamicin loading than the currently existing materials and reduced numbers of viable <i>Salmonella</i> in the liver and spleen compared to free gentamicin. To further improve the rate and route of uptake, the shell of the nanostructures were made amphiphilic by incorporating pluronics F68 (PPO)₆₈ in the block copolymer. We showed that core-shell nanostructures encapsulating gentamicin having (PPO)₆₈ in the shell (N2) enhances the rate and modulates the route of uptake into macrophages, thus promoting significant reduction in the intracellular <i>Salmonella in-vitro</i> and <i>in-vivo</i>. The main drawback of N2 was its poor stability at physiological pH of 7.4, 0.1 M NaCl. Therefore, core-shell nanostructures encapsulating gentamicin containing pluronic P85 (PPO)₈₅ in the shell (N3) with improved colloidal and ionic stability were designed. N3 achieved significant intracellular reduction of vacuolar <i>Salmonella</i> (0.53 log₁₀) and cytoplasm resident <i>Listeria</i> (3.11 log₁₀) compared to free gentamicin in-vitro. However, greater reduction of <i>Listeria</i> suggested that sub-cellular localization of bacterium influences targeting by N3. Even though oral administration of N3 was not effective compared to free gentamicin, parenteral (I.P.) administration significantly reduced the intracellular <i>Salmonella</i> from liver and spleen compared to free gentamicin and appeared to have no abnormal <i>in-vivo</i> toxicity.
In summary, core-shell nanostructures encapsulating gentamicin (N) with improved encapsulation efficiency and different shell chemistry (N1, N2 and N3) were developed with enhanced efficacy against intracellular Salmonella. The novel gentamicin delivery approach developed in this study may be applicable for therapy of many intracellular infections. / Ph. D.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/29082 |
| Date | 21 October 2009 |
| Creators | Ranjan, Ashish |
| Contributors | Veterinary Medical Sciences, Kasimanickam, Ramanathan, Sriranganathan, Nammalwar, Riffle, Judy, Pelzer, Kevin D., Pickrell, Gary R., Swecker, William S. Jr. |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Detected Language | English |
| Type | Dissertation |
| Format | application/pdf, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
| Relation | Ranjan_Ashish_D_2009.pdf, CERTIF.pdf |
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