The female genital tract is the main route for heterosexual transmission of HIV-1. Studies with SIV in macaque model indicate that after crossing the mucosa of cervix and vagina, SIV infected cells migrate to draining lymph nodes where the virus expands before it gets disseminated into the gut and other organs of the body. We have developed a combined organ culture system composed of cervical tissue together with tonsil tissue (surrogate lymphoid tissue for lymph node) to study HIV-1 transmission across the mucosal barrier and to evaluate the fate of HIV-1 and HIV-1 infected cells after crossing cervical mucosa and migrating to regional lymph nodes. To study the transmission of virus, a defined amount of pretitered cell-free or cell-associated HIV-1BAL or HIV-1IIIB was added to the top of the cervix tissue and incubated at 37°C in a CO2 incubator. The top well was removed after 3-4 days and tonsil tissue or tonsil mononuclear cells in the bottom well were cultured for an additional 12 days. Replication of cell-free and cell-associated HIV-1BAL and HIV-1IIIB in tonsillar cells after crossing the cervix mucosa was confirmed by the presence of HIV-1 p24 in culture supernatant, the detection of HIV-1 DNA by real-time PCR, presence of p24 antigen by immunofluorescence assay and presence of HIV-1 RNA by simultaneous immunophenotyping and ultrasensitive fluorescence in situ hybridization. We have also characterized by immunofluorescence assay HIV-1 infected migratory cells exiting from the cervix tissue. CD4+ (T cells), CD11c+ (dendritic cells) and CD68+ (macrophages) cells were found to migrate from cervix tissue and were positive for HIV-1. The public health relevance of this model is that we may use the combined cervix and tonsil tissue/cell model to determine the mechanism of sexual transmission of HIV-1 in women at the cellular and molecular level and to evaluate anti-HIV microbicides.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-08012008-205429 |
| Date | 28 September 2008 |
| Creators | Soto-Rivera, Jackeline |
| Contributors | Velpandi Ayyavoo, PhD, Frank Jenkins, PhD, Phalguni Gupta, PhD |
| Publisher | University of Pittsburgh |
| Source Sets | University of Pittsburgh |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.pitt.edu/ETD/available/etd-08012008-205429/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Pittsburgh or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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