Rheumatoid arthritis (RA) is an autoimmune disease affecting nearly 1% of the population. The joints of RA patients become inflamed and a pannus of synovial tissue invades cartilage and destroys bone. Current treatments, though moderately successful, are often administered systemically and are associated with significant side effects. Local treatment of arthritis by intra-articular injection of gene transfer vectors has the potential to overcome some of the obstacles of other treatment regimens. In particular, adeno-associated virus (AAV) is attractive as a gene transfer vector because it is non-pathogenic in humans, has low immunogenicity and broad tissue tropism, and provides for long-term transgene expression. The primary target of gene transfer in the joint is the fibroblast-like synoviocytes (FLS), which make up part of the pannus of tissue responsible for cartilage and bone destruction. However, FLS are only minimally transduced by AAV and provide only low levels of transgene expression. For local treatment with AAV vectors to be successful, improvement in the transduction of FLS is necessary. Several agents have been shown to increase AAV transduction in other cell types. The specific effect of inflammation and proteasome inhibition on AAV-mediated transgene expression in FLS has not been examined. Here we show that both inflammatory cytokines and the proteasome inhibitor zLLL can increase transgene expression in FLS dramatically. Remarkably, both agents are also able to regulate transgene expression. The cytokines appear to act in part by promoting uncoating of the virus, while proteasome inhibition works by both increasing the nuclear trafficking of the virus and by increasing transcription of the viral DNA. In addtion, we screened 3 serotypes of AAV (2, 2.5, and 5) for their ability to transduce FLS from 4 different species: human, mouse, horse, and rabbit. AAV2 provided the highest transgene expression in human and horse FLS, while AAV2.5 was better suited for mouse and rabbit FLS. These results have implications in the future use of AAV vectors in the treatment of RA.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-08292006-134427 |
| Date | 30 August 2006 |
| Creators | Traister, Russell Scott |
| Contributors | Paul Robbins, Penelope Morel, Xiao Xiao, Jay Kolls, Raphael Hirsch |
| 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-08292006-134427/ |
| 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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