Pathogenic mechanisms of F. alocis

University Of Louisville, Louisville KY

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Abstract

DESCRIPTION (provided by applicant): Periodontal diseases are one of the most common bacterial infections of humans and impose a significant burden on the health care system. Recent developments in molecular based bacterial detection technologies have identified a number of previously unrecognized or under-appreciated organisms as associated with periodontal lesions. One such emerging pathogen is the gram-positive anaerobe Filifactor alocis. This organism is a constituent of subgingival biofilms and is present in elevated numbers at site of periodontal disease as compared to health. F. alocis produces proteases and can invade gingival epithelial cells; however, little else is known regarding the potential pathogeniciy of the organism. In order to capitalize on the human microbiome studies it is necessary to functionally characterize the newly identified organisms. Hence, in this study we shall contribute to the process of assessing the pathogenic potential of F. alocis in vitro and in vivo. Our preliminary data indicate that F. alocis can induce apoptosis in gingival epithelial cells an event with relevance to disruption of periodontal tissue homeostasis. The first goal of the current proposal, therefore, is to begin to characterize the mechanistic basis of F. alocis-induced apoptosis. We will focus on the MEK-FOXO pathway as preliminary data show that F. alocis can inhibit MEK and potentially activate the pro-apoptotic transcriptional factor FOXO. The role of MEK will be assessed by gene knockin experiments with constitutively mutants, and the relevance of FOXO determined by reporter assays and siRNA knockdowns. Secondly, we will establish the in vivo characteristics of F. alocis in the mouse chamber model. The ability of F. alocis to survive and replicate in vivo will be established and, in addition, host cytokine responses, neutrophil recruitment and apoptosis will be assayed. These studies will generate both in vitro and in vivo data that will begin to establish the pathogenic credentials of F. alocis and provide a platform for future studies directed toward a more comprehensive understanding of molecular and cellular pathogenic mechanisms. Ultimately, the knowledge gained could be translated into novel diagnostic, therapeutic or preventive strategies for periodontal diseases. PUBLIC HEALTH RELEVANCE: Periodontal diseases afflict millions of Americans, and recent molecular techniques have identified a variety of bacteria associated with the disease that have yet to be studied in depth. In this project we will examine the interactions between F. alocis and the human cells that are colonized by the organism, and we will determine the ability of the organism to survive in mouse models of infection. The information to be gathered could result in a re-evaluation of the causes of periodontal disease, and ultimately be used to identify targets for novel therapeutic agents or diagnostic tests.