http://www.loc.gov/mods/v3 http://www.loc.gov/standards/mods/v3/mods-3-2.xsd
role of TNF-α in the resistance of guinea pigs to Mycobacterium tuberculosis infection
ABSTRACT: TNF-α plays an important role in the host immune response to infection with the intracellular pathogen Mycobacterium tuberculosis. It is essential for the formation of protective tuberculous granulomas and regulates the expression of other cytokines which contribute to a protective immune response. In this study, using recombinant guinea pig (rgp) TNF-α or neutralizing polyclonal anti-rgpTNF-α antibody, we have investigated the role of guinea pig TNF-α in guinea pig macrophage cultures as well as cocultures of immune lymphocytes and macrophages infected with M. tuberculosis. Resident alveolar and peritoneal macrophages were isolated from guinea pig and stimulated with rgpTNF-α or infected with either the attentuated H37Ra or virulent H37Rv strains of M. tuberculosis in the presence of rgpTNF-α. IL-12 p40 mRNA was up-regulated in a dose-dependent manner by rgp TNF-α alone. In infected macrophages, a lower dose of rgpTNF-α intensified the mRNA levels of TNF-α and IL-12 p40. However, higher doses of rgpTNF-α suppressed TNF-α and IL-12 p40 mRNA, which suggests that the dose of TNF-α is crucial to the stimulation of optimal expression of protective cytokines. Infected resident alveolar and peritoneal macrophages treated with anti-gpTNF-α antibody to block endogenous TNF-α activity exhibited increased intracellular mycobacterial growth. TNF-α is known to contribute to the activation of innate immunity and the transition to antigen-specific adaptive immunity in tuberculosis. Therefore, we have examined the effect of TNF-α on lymphocyte activation in unvaccinated and BCG-vaccinated guinea pigs. Splenocytes were stimulated with PPD or ConA in the presence or absence of rgpTNF-α for 96 hours. rgpTNF-α alone was able to stimulate a significant degree of proliferation in splenocytes. The addition of rgpTNF-α enhanced the proliferation of PPD-stimulated splenocytes from BCG-vaccinated guinea pigs, in conjunction with upregulation of the levels of Type 1 cytokine mRNA (IL-12p40 and IFN-γ). The effect of neutralization of endogenous TNF-α on anti-mycobacterial immune function was examined using cocultures of purified immune T cells and macrophages obtained from BCG-vaccinated guinea pigs. Resident peritoneal macrophages (PM) were infected with virulent M. tuberculosis H37Rv and nylon-wool purified autologous splenic T lymphocytes were added in the presence or absence of polyclonal rabbit anti rgp TNF-α and cultured for 4-7 days. Anti-rgp TNF-α antibody significantly suppressed T cell proliferation in cocultures with higher number of PM. Neutralization of TNF-α downregulated the expression of IL-12p40 and IFN-γ mRNA significantly. Most importantly, neutralization of endogenous TNF-α significantly enhanced the intracellular survival of virulent mycobacteria. Taken together, these results suggest that TNF-α plays a crucial role in the activation of a Type 1 cytokine profile and the control of intracellular mycobacteria in the guinea pig.
"Submitted to the Graduate School of Biomedical Sciences of The Texas A&M University System Health Science Center in partial fulfillment for the requirements for the degree of Doctor of Philosophy December 2005."
Approved as to style and content by: David N. McMurrary, Roger Smith III, James E. Samuel, Vernon L Tesh, John M. Quarles.
Ph. D. Texas A&M University System Health Science Center 2005.
Includes bibliographical references (leaves 87-103).