The glycoprotein fibronectin is a component of the molecular mesh known as the extracellular matrix, which not only provides physical support for our cells but also directs cellular activities during embryonic development, tissue repair, and other processes. High levels of soluble fibronectin (300 μg/ml) are also found in our bloodstream, where it quietly circulates until it is recruited to stabilize clots and promote wound repair.
Fibronectin has a modular organization consisting of binding sites for various matrix and cell surface molecules. Examples include attachment sites for integrins (labeled "Cell" in the figure below) and glycosaminoglycans (labeled "Heparin"), which are exposed on the surface of the endothelial cells that line our blood vessels.
B. burgdorferi is injected into the skin by an infected tick and spreads outward within the dermis, causing the familiar "bulls-eye" rash in some Lyme disease patients. The spirochete may eventually enter the bloodstream so that it can spread to other tissues. While in the bloodstream, the spirochete is surrounded by fibronectin. In fact, B. burgdorferi attaches to fibronectin in vitro, suggesting a role for fibronectin in Lyme disease.
It turns out that B. burgdorferi exploits the adhesive properties of plasma fibronectin to bind to the vessel wall before escaping into the surrounding tissue. Like their earlier work, which I described in my last post, this follow-up study by Norman and colleagues was conducted with fluorescent B. burgdorferi injected into the veins of live mice. The interactions of the spirochete with the capillary wall were observed by fluorescent intravital microscopy. The earlier study revealed that most of the interactions were transient, lasting for less than a second. B. burgdorferi was also seen crawling (dragging) along the vessel wall, which was followed by escape into the tissue or by stationary adhesion, a more intimate association with the vessel wall. Stationary adhesion could also be followed by extravasation and escape of the spirochete into the tissue. Both stationary adhesion and escape usually occurred between the endothelial cells lining the vessel wall. Each type of interaction (transient, dragging, and stationary adhesion) was quantitated by counting.
In their follow-up study, the authors demonstrated that coinjection of anti-fibronectin antibody and B. burgdorferi into the bloodstream of the mice diminished all catagories of interactions (transient, dragging, and stationary adhesion) by at least 90%. Control antibody (goat IgG) had no effect. These results indicate that fibronectin has a key role in mediating the attachment of B. burgdorferi to the microvasculature. Since fibronectin could potentially bind to GAGs and integrins on endothelial cells, the investigators also coinjected B. burgdorferi with peptides or antibodies known to block attachment of fibronectin to these targets. They found that the GAG-specific peptide reduced the interaction of B. burgdorferi with the vessel wall, whereas the integrin-specific peptide and antibodies had little effect. Thus, fibronectin may serve as a molecular bridge linking B. burgdorferi to GAGs displayed on the endothelial cells lining the blood vessel.
Which B. burgdorferi factor is involved in adherence to the vessel wall in vivo? Past studies had shown that the borrelial protein BBK32, a known fibronectin binding protein, mediated attachment of B. burgdorferi to fibronectin in vitro. Therefore, BBK32 was a logical candidate. To determine whether BBK32 was involved in vascular interactions in the mouse model, the research team employed a noninfectious B. burgdorferi strain that had lost bbk32 and other genes during long-term culture. The noninfectious strain failed to interact with the vasculature in the mouse. However, expression of BBK32 restored the ability of the noninfectious strain to transiently interact and drag along the vessel wall but only partly restored stationary adhesion. This results suggest that although BBK32 plays a role in vascular adherence, other bacterial factors are also involved.
Many pathogens have been shown to interact with fibronectin and GAGs in vitro. However, this study is highly significant as it is the first to demonstrate a role for these host molecules in bacterial adherence to the microvasculature in a living animal. Other spirochetes such as Treponema pallidum and Leptospira also express fibronectin binding proteins. Hence, the mechanism employed by B. burgdorferi to escape from the bloodstream may be similar for all disease-causing spirochetes. Moreover, other microbial pathogens have been shown to stick to fibronectin and GAGs in vitro. Thus, a large number of invasive pathogens may employ similar mechanisms to spread to different tissues via the bloodstream.
M. Ursula Norman, Tara J. Moriarty, Ashley R. Dresser, Brandie Millen, Paul Kubes, George Chaconas (2008). Molecular Mechanisms Involved in Vascular Interactions of the Lyme Disease Pathogen in a Living Host PLoS Pathogens, 4 (10) DOI: 10.1371/journal.ppat.1000169