Differential tissue targeting and pathogenesis of verotoxins 1 and 2 in the mouse animal model

Background. Both verotoxin (VT) 1 and VT2 share the same receptor, globotriaosyl ceramide (Gb(3)). Although VT1 is slightly more cytotoxic in vitro and binds Gb(3) with higher affinity, VT2 is more toxic in mice and may be associated with greater pathology in human infections. In this study we have compared the biodistribution of iodine 125 (I-125)-VT1 and I-125-VT2 versus pathology in the mouse. Methods. I-125-VT1 whole-body autoradiography defined the tissues targeted. VT1 and VT2 tissue distribution, clearance, and tissue binding sites were compared. The effect of a soluble receptor analogue, adamantylGb(3), on VT2/Gb(3) binding and in vivo pathology was assessed. Results. I-125-VT1 autoradiography identified the lungs and nasal turbinates as major, previously unrecognized, targets, while kidney cortex and the bone marrow of the spine, long bones, and ribs were also significant targets. VT2 did not target the lung, but accumulated in the kidney to a greater extent than VT1. The serum half-life of VT1 was 2.7 minutes with 90% clearance at 5 minutes, while that of VT2 was 3.9 minutes with only 40% clearance at 5 minutes. The extensive binding of VT1, but not VT2, within the lung correlated with induced lung disease. Extensive hemorrhage into alveoli, edema, alveolitis and neutrophil margination was seen only after VT1 treatment. VT1 targeted lung capillary endothelial cells. Identical tissue binding sites (subsets of proximal/distal tubules and collecting ducts) for VT1 and VT2 were detected by toxin overlay of serial frozen kidney sections. Glucosuria was found to be a new marker of VT1- and VT2-induced renal pathology and positive predictor of outcome in the mouse, consistent with VT-staining of proximal tubules. Lung Gb(3) migrated on thin-layer chromatography (TLC) faster than kidney Gb(3), suggesting a different lipid composition. AdamantylGb(3), a soluble Gb(3) analogue, competed effectively for Gb(3) binding by VT1 and VT2 in vitro. However, the effect in the mouse model (only measured against VT2, due to the lower LD50, a concentration required for 50% lethality) was to increase, rather than reduce, pathology and further reduce the VT2 serum clearance rate. Additional renal pathology was seen in VT2 + adamantylGb(3)-treated mice. Conclusions. The lung is a preferential (Gb(3)) "sink "for VT1, which explains the relatively slower clearance of VT2 and subsequent increased VT2 renal targeting and VT2 mortality in this animal model.