Modification of the Woodruff-Stamper assay demonstrates binding of Toxoplasma gondii tachyzoites to retinal vascular endothelium.

In the human host, infection with the protozoan parasite, Toxoplasma gondii, most commonly involves the eye and/or the brain. Previous work indicates a relative susceptibility of the human retinal vascular endothelium to infection with the T. gondii tachyzoite, which may contribute to this tissue localization. To facilitate the investigation of potential adhesive interactions between parasite and endothelium in the retina, we have modified the Woodruff-Stamper assay, originally described to study lymphocytic-endothelial binding. Vascular endothelium was identified in sections of human retina by Alexa Fluor 594-tagged anti-von Willebrand factor antibody. Binding of yellow fluorescent protein-expressing tachyzoites to endothelium under conditions of flow, simulated by rotation on an orbital shaker, was quantified in a masked fashion using imaging software. We observed multiple yellow spots in contact with Alexa Fluor 594-positive retina, indicating binding of T. gondii tachyzoites to retinal vascular endothelium. This modification of the Woodruff-Stamper assay provides an opportunity to evaluate potential host receptors for T. gondii on the retinal vascular endothelium. In addition, the assay suggests a methodology that could be used to examine adhesion of other microbes to microvasculature in different tissues.

Susceptibility to endotoxin induced uveitis is not reduced in mice deficient in BLT1, the high affinity leukotriene B4 receptor.

AIM: To investigate the role of arachidonic acid derived chemotactic factor, LTB(4), in the development of endotoxin induced uveitis (EIU), using mice deficient in the BLT1 gene which encodes the high affinity LTB(4) receptor. METHODS: BLT1 gene deficient and wild type BALB/c mice were injected intravitreally with Escherichia coli 055:B5 lipopolysaccharide (250 ng/2 microl). Number of leukocytes invading the anterior chamber 24 hours later were counted on tissue cross sections. RESULTS: In all mice, EIU was characterised by a polymorphonuclear and mononuclear cell infiltrate. Numbers of infiltrating cells did not differ significantly between control and BLT1 gene knockout mice. CONCLUSION: Chemotactic factors other than LTB(4) are primarily responsible for leukocyte migration into the eye during murine EIU.