In this proposal, innovative carrageenan formulations combined either with zinc (Zn++) or with lignosulfonic acid (LSA), a byproduct of lignan purification, will be tested and compared to Carraguard for anti-HIV-1 activity in a human xenograft model for HIV-1 infection. The applicant is using this latest configuration of the human vaginal/xenograft model as a productive target for HIV infection. Virus replication is measured qualitatively and quantitatively using reverse transriptase/polymerase chain reaction (RT/PCR) to measure virus or provirus genomes and/or spliced HIV transcripts. Replication of virus can also be directly visualized and localized in the vaginal epithelial layer by in situ hybridization (ISH) for HIV RNA and concurrent immunocytochemistry for human lymphocyte or monocyte markers. Immunohistochemical analysis can also be used to detect the presence of HIV p24 antigen. This animal model system will be used to measure the microbicidal properties of formulated carrageenans in vivo.
The Specific Aims for this Project include: 1) Determining the sensitivity of vaginal epithelium to topically applied carrageenan formulations using freshly excised chips of human vagina as well as healed xenografts of human vaginal epithelium. Assessment will include histology and immunocytochemical techniques; 2) Premixing of carrageenan formulations with T -tropic, M-tropic and dual-tropic strains of HIV prior to inoculation of human vaginal xenografts in order to determine if selected compounds can directly inactivate HIV strains; and 3) Pre-applying non-toxic formulations of carrageenans into the lumen of tubular vaginal xenografts prior to HIV-1 inoculation in order to determine if pre-application of the microbicide prevents HIV-1 transmission.
