Helmus, Ruth Anne
(2008)
Generation and Characterization of the Cellular Immune Response to a Clostridium perfringens anti-SIV Mucosal Vaccine.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
Most new human immunodeficiency virus (HIV) infections are acquired through vaginal or rectal mucosa, and gut mucosal tissue is a primary target of HIV infection. To generate mucosal immunity against HIV or its simian counterpart simian immunodeficiency virus (SIV), the Gram positive bacterium Clostridium perfringens was used to develop a vaccine that delivers SIV p27 to the gut and induces local T cell immunity.Under in vitro conditions, Clostridium perfringens expressing SIV p27 (Cp-p27) was found to induce dendrite cell (DC) maturation and stimulate p27-specific T cell responses. To improve intracellular delivery of p27 to DCs and thereby enhance immune priming, Cp-p27 variants expressing p27 conjugated with protein transduction domains (PTDs) at the 5' end were constructed. While internalization of p27 by DCs and gut epithelial cells was improved following exposure to the PTD-Cp-p27 variants, cellular p27-specific immune stimulation was not significantly improved compared with wild-type Cp-p27.The Cp-p27 vaccine was then tested in vivo in mice for its ability to prime gut mucosal T cell responses. First, an adjuvant optimization study with three mucosal adjuvants, cholera toxin (CT), mutant E. coli heat-labile enterotoxin (LT(R192G)), and unmethylated cytosine-phosphate-guanine oligodinucleotides (CpG ODNs) was performed to determine the best T cell immune response in the gut. While the combination of CpG ODNs and (LT(R192G)) induced the highest T cell immune response, (LT(R192G)) alone provided the best multifunctional CD8+ T cell response in the gut.Oral Cp-p27 vaccination was then tested for induction of T cell immunity in vivo in a prime-boost model by combining Cp-p27 with systemic immunization with an adenovirus expressing p27 (Ad-p27). Cp-p27 vaccination primed a strong multifunctional T cell immune response in gut lamina propria, although it could not stimulate a systemic immune response. In contrast, Ad-p27 vaccination stimulated strong systemic immunity but limited gut mucosal immunity. By sequentially delivering Cp-p27 and Ad-p27, immunity in both the gut and systemic tissues was achieved.Altogether, this study demonstrates that Cp-p27 can deliver p27 to gut T cells through dendritic cells to prime a strong, multifunctional immune response in the gut effector tissue.
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| Item Type: |
University of Pittsburgh ETD
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| Status: |
Unpublished |
| Creators/Authors: |
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| ETD Committee: |
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| Date: |
16 April 2008 |
| Date Type: |
Completion |
| Defense Date: |
4 April 2008 |
| Approval Date: |
16 April 2008 |
| Submission Date: |
10 April 2008 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
School of Medicine > Molecular Virology and Microbiology |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
mouse; cellular immunity; HIV; mucosal immunology; polyfunctional; vaccine; flow cytometry; SIV |
| Other ID: |
http://etd.library.pitt.edu/ETD/available/etd-04102008-114211/, etd-04102008-114211 |
| Date Deposited: |
10 Nov 2011 19:35 |
| Last Modified: |
15 Nov 2016 13:39 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/6963 |
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