HSV enhances HIV-1 transmigration through the genital epithelial barrier.

<p>A. Model for the HIV-1 transwell assay. B. PGEC were seeded onto polycarbonate membrane transwells and cultured until formation of tight junctions was achieved. HIV-1 (wild-type JR-CSF, NL4.3 or gp160-deficient (ΔE) NL4.3) was then added to the apical surface of PGEC and amounts of transcytosed viruses were collected at various time points (2, 4, 8 and 16 h) and quantified by p24 ELISA of the lower chamber corresponding to the basal surface. Data are expressed in percentage of the original viral inoculum (top panel). Infectivity of transcytosed viruses: medium from the basal chamber was collected at different intervals of time, filtered, standardized for p24 content (20 pg), and added to TZM indicator cells. Infection was measured 48 h post-infection by determining levels of beta-galactosidase activity in cell lysates (bottom panel). Results are representative of the results of four independent experiments using PGEC derived from each of the 4 donors. Error bars represent standard deviations. C. PGEC isolated from 2 donors were incubated with HSV-1 or HSV-2 (MOI of 0.005). Two days post-infection, infection of PGEC was quantified by FACS by anti-HSV gD IgG immunostaining. D. PGEC seeded onto the transwells were exposed to HSV-1 or HSV-2 (MOI of 0.005) in the presence or absence of ACV (50 µmol/L). Two days post-infection, the integrity of the PGEC barrier was analyzed by measuring the paracellular passage of the extracellular marker inulin (top panel) as well as the TER of each monolayer (bottom panel) as we described previously <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0018917#pone.0018917-Bobardt2" target="_blank">[19]</a>.