ANTHRAX TOXIN: IMMUNITY AND RECEPTOR ACTIVITY
Bacillus anthracis is a high priority biothreat agent because of two virulence factors, an anti-phagocytic capsule and an exotoxin, consisting of protective antigen (PA), lethal factor (LF), and edema factor (EF). These virulence factors inhibit the host immune responses, allowing the bacilli to rapidly multiply and overwhelm the host. While the capsule is nonimmunogenic, the PA component of anthrax toxin has been shown to induce a protective antibody response and is the main component of human anthrax vaccines, including the United States' anthrax vaccine adsorbed (AVA). This dissertation aimed to further characterize anthrax toxin activity, examine human antibody responses to AVA vaccination, and investigate the influence of toxin receptor type on susceptibility to anthrax toxin. Initial studies characterized the activity of LF and EF using the mouse J774A.1 cell line and examined if human antibody-mediated neutralization of toxin could be more efficiently assessed using human cells. A variety of human and animal cells lines, and human primary cells were examined for susceptibility to anthrax toxin. J774A.1 cells were the only cells found to be susceptible to LF-induced lysis, but all cells tested were susceptible to EF-induced increases in cAMP. The activity of LF and EF antibodies was also assessed utilizing the LF- and EF-susceptible J774A.1 cells. The results revealed that LF and EF antibodies do not significantly contribute to anthrax toxin neutralization in humans, and that antibodies to PA are sufficient to neutralize toxin activity. Lastly, the influence of receptor type on susceptibility to anthrax toxin components using Chinese hamster ovary cells (CHO), CHO cells lacking functional toxin receptors (CHO-R1.1), and CHO-R1.1 cells expressing the human forms of two PA receptors, TEM8 (CHO-TEM8) or CMG2 (CHO-CMG2) was examined. Unexpectedly, PA alone, previously believed to only mediate entry of LF or EF, was found to be toxic to CHO-TEM8 cells. CHO-TEM8 cells treated with PA alone displayed reduced cell growth and decreased metabolic activity. PA induced immediate cell swelling, and the cells became permeable to membrane-excluded dye, suggesting that PA-TEM8 binding leads to pore-formation on the cell surface. These unanticipated results reveal that PA alone might mediate toxic activity in disease.
School:University of Cincinnati
School Location:USA - Ohio
Source Type:Master's Thesis
Keywords:bacillus anthracis anthrax toxin ava
Date of Publication:01/01/2007