Gram-positive bacterial infections have risen over recent years and current antibiotic treatments are not always sufficient to control these infections. Specifically, antibiotics target bacteria themselves, but not the bacterially secreted proteins that contribute to bacterial pathogenesis and host tissue damage (i.e. virulence factors). These virulence factors may linger after bacteria are eradicated, making their interaction with the host important to understand for the development of novel therapeutics to supplement antibiotics. One class of virulence factors studied in our laboratory is a large pore-forming toxin family known as the cholesterol-dependent cytolysins (CDC). These exotoxins are secreted by over twenty species of gram-positive bacteria and have been shown to contribute to the virulence of the bacteria that secrete them. We are interested in exploring the pathways initiated by CDC in host innate immune cells such as macrophages and dendritic cells. These cells would be expected to first encounter CDC after bacterial infection and therefore, pathways initiated in these cells by CDC could be targeted for the benefit of the host.
We have characterized the mechanism of mature IL-1beta secretion induced by CDC tetanolysin O (TLO) from LPS-primed murine bone marrow-derived macrophages (BMDM). This process is dependent on TLO dose and relies on the caspase-1-containing NLRP3 inflammasome as well as associated signaling pathways, which include ion fluxes and iPLA2 and cathepsin B activities. Furthermore, TLO induces different cell death programs in BMDM that are dependent on TLO dose. High TLO doses induce conventional necrotic cell death while low TLO doses cause NLRP3 inflammasome-dependent and cathepsin B-dependent necrotic cell death that is characterized by lactate dehydrogenase (LDH) and high mobility group box 1 (HMGB1) release. Both IL-1beta and HMGB1 are pro-inflammatory cytokines that contribute to inflammation and may be useful therapeutic targets, in addition to the inflammasome. Finally, susceptibility to CDC-induced cell killing varies based on cell type. In order to determine pathways that might explain these differences, we created a variant murine dendritic cell line resistant to pore formation. Though this cell line has been characterized to some degree, future studies will be needed to pinpoint the pathways responsible for the phenotype observed.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-02072010-153303 |
| Date | 04 March 2010 |
| Creators | Chu, Jessica |
| Contributors | Adriana T. Larregina, M.D., Ph.D., Penelope A. Morel, M.D., Bruce A. McClane, Ph.D., Lawrence P. Kane, Ph.D., Russell D. Salter, Ph.D. |
| 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-02072010-153303/ |
| 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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