Semester

Fall

Date of Graduation

2012

Document Type

Dissertation

Degree Type

PhD

College

School of Pharmacy

Department

Pharmaceutical Sciences

Committee Chair

Bingyun Li

Committee Co-Chair

Kathleen Brundage

Committee Member

Patrick Callery

Committee Member

Peter Gannett

Committee Member

Yon Rojanasakul.

Abstract

Staphylococcus aureus (S. aureus) is the predominant cause of bone and joint infections worldwide. These infections may lead to severe bone and joint destruction and high mortality. Treatment of bone infections frequently requires long-term hospitalization for repeated debridement and prolonged antibiotic treatment. Bacteria including S. aureus, however, are often surprisingly successful in overcoming both host defense mechanisms and antibiotic treatments resulting in chronic and recurrent bone infections. Indeed, a high rate of recurrence or relapse of bone infections has been reported where ∼17% of infections related to traumatic extremity injuries recur or relapse. One thought is that bacteria like S. aureus used to elude host defenses and antibiotic treatments are able to "hide" themselves intracellularly within host cells.;S. aureus has conventionally been considered as an extracellular pathogen, but there is growing evidence showing that it can survive intracellularly within host cells. Once internalized into host cells, S. aureus may elicit a pro-inflammatory response often characterized by the secretion of interleukin 1beta (IL-1beta), IL-6, IL-12p70 (IL-12), and tumor necrosis factor-alpha (TNF-alpha) among other cytokines. Production of pro-inflammatory cytokines from infected osteoblasts may disrupt the balance of the immune system thereby leading to uncontrolled inflammation and bone destruction. On the other hand, IL-12 is released primarily by antigen-presenting cells and acts as a link between innate and acquired immunities by inducing the differentiation of the T helper 1 (Th1) phenotype and the release of interferon gamma (IFN-gamma) from activated T-cells as well as natural killer cells. T-cell-derived IFN-gamma is considered crucial to the activation of macrophages, the ultimate effector cells in host defense against intracellular infection. Studies have shown that both IL-12 and IFN-gamma are up-regulated in infection models of other bacteria, parasites, and viruses but little is known about how they may contribute to the outcome of infection.;The overall goal of this dissertation is to develop in vitro and in vivo infection models of intracellular S. aureus that can aid in understanding the pathogenesis of chronic and recurrent infections and to develop effective immunotherapy regimens to prevent or treat such infections. This dissertation starts with an overall literature review in Chapter 1 about the bacterium S. aureus, implant-associated infection, and finally IL-12 as a potential therapeutic agent against infections. Chapter 2 establishes and compares two S. aureus infection models utilizing a non-phagocytic cell (osteoblast) and a phagocyte (macrophage) and the corresponding cellular responses of each cell type upon infection. In Chapter 3, S. aureus within osteoblasts is used to infect an open femur fracture rat model. We have presented the first direct in vivo evidence that intracellular S. aureus could be sufficient to induce bone infection in animals and intracellular S. aureus could play an important role in the pathogenesis of S. aureus infections. Finally, a potential approach using IL-12 as a therapeutic agent to improve macrophage killing activities against intracellular S. aureus is investigated in Chapter 4..;Keywords: osteomyelitis, Staphylococcus aureus, infection, interleukin 12, cytokine, osteoblast, macrophage, rat, intracellular S. aureus..

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