Date of Award

Spring 1992

Project Type

Dissertation

Program or Major

Microbiology

Degree Name

Doctor of Philosophy

First Advisor

Thomas G Pistole

Abstract

The macrophage has been shown to bind potentially pathogenic bacteria in the absence of serum components, but the mechanism is not well understood. The macrophage is a key component of our non-inducible defenses and this form of innate immunity is of particular importance for individuals in which the immune system may be sub-optimally functional, e.g., neonatal and geriatric individuals, and for sites in the body in which serum enhancement through opsonins is thought to be insufficient, e.g., lungs. To understand the mechanism by which mammalian innate host defenses respond to potential microbial aggression by opportunistic microorganisms, I developed two assays to quantify the in vitro, serum-free adherence of group B Streptococcus to murine peritoneal macrophages. The first is a modified direct microscopic assay, while the second is based upon an enzyme-linked assay using bacterial specific antibody.

Sugar inhibition assays as well as studies with isogenic strains and neuraminidase-treated streptococci could not confirm a mechanism proposed by other investigators that macrophages recognize these streptococci by a lectin-like receptor for galactose. I observed that binding characteristics of the macrophage, such as temperature- and divalent cation-dependence, protease sensitivity, and enhancement by fibronectin and phorbol esters, were similar to those of the $\beta\sb2$ integrins, a heterodimeric family of leukocyte receptors. Monoclonal antibodies M1/70, which recognizes the $\alpha$-subunit of one member of the family, complement receptor 3 (CR3), and M18/2, which recognizes the $\beta$-subunit, common to all three members, were each shown to reduce bacterial adherence to down-modulation, flow cytometric and soluble inhibition studies. Using similar approaches I have additionally shown that the purified streptococcal membrane phosphosugar, lipoteichoic acid, can inhibit the binding of these bacteria to macrophages. These results support roles for both lipoteichoic acid and the $\beta\sb2$ integrins in the innate recognition of group B streptococci by macrophages.

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