Abstract
The lectins derived from Hippeastrum hybrid (Hippeastrum hybrid agglutinin, HHA) and from the stinging nettle Urtica dioica (UDA) have been investigated as anti-HIV microbicides. The present study was conducted to determine their diffusion through intact and de-epithelialised human vaginal epithelium. Both lectins were labelled with fluorescein isothiocyanate groups (FITC) and analysed by SDS-polyacrylamide gel electrophoresis. While UDA appeared to be fairly homogeneous with an average Mw of ∼ 8.5 kDa, HHA was a heterogeneous mixture of compounds with Mw's ranging between ∼ 13 and 52 kDa. Fresh human vaginal mucosa was snap-frozen in liquid nitrogen and stored at −85°C. Prior to an experiment, the tissue was defrosted to 20°C in PBS buffer, pH 7.4, and placed in the seven flow cells of a flow-through perfusion apparatus. Either FITC-labelled HHA or UDA was then pipetted into the donor chamber of the flow cell. Samples from each flow cell were collected every 2 hours (1.5 ml/h) over a 24-hour period and analysed by fluorospectrophotometry. Permeability experiments were repeated with vaginal mucosa specimens from which the epithelial layers had been removed by heat-stripping. Both lectins diffused through vaginal mucosa at rates proportionate to their average molecular weights, the flux rates of the smaller UDA being ∼ 5x higher than that of the larger HHA. Removal of the vaginal epithelium increased the flux rates of both HHA and UDA across the mucosa and this may have implications for a more rapid in vivo uptake of these lectins when used as anti-HIV microbicides.