1141180267Mechanism of HIV transmission across female primary genital epithelial cells

Our lab is utilizing human primary genital epithelial cultures to investigate the mechanism of HIV transmission across the female genital mucosa. Primary endometrial epithelial cells (ECs) grown on matrigel‐ coated cell inserts were used for HIV infection studies. EC's were infected with cell‐ free or cell‐ associated R5 and X4 virus strains. Infections were performed in the presence or absence of the appropriate macrophage (U937) or T‐cell (Jurkat) target cell‐ line in basolateral compartments of cultures. Virus was quantified from supernatants using p24 ELISA. U373‐CXCR4 and CCR5 magi cell lines were utilized to measure infectious virus and tropism. Sybrgreen real time PCR was used to detect viral RNA in cell lysates. Infectious HIV particles were found only in basolateral supernatants of EC's infected with X4 strains of HIV in the presence of appropriate target cells. This indicated that EC's may not be productively infected, but were only able to transmit HIV in the presence of target cells. These results were supported by the detection of higher levels of gag gene (CT‐15) in X4 target cells compared to EC's (CT‐25) using real‐time PCR. No infectious virus was present in primary EC's infected with cell‐free R5 strain, although gag gene product was detected by real time PCR in both R5 target cells as well as EC's. P24 levels in supernatants were not indicative of infectious virus. Our data indicates that R5 and X4 virus strains have differential abilities to cross the female genital mucosa to infect target cells. The presence of target cells appears to be critical for the production of infectious HIV particles under these culture conditions. These studies are providing important information regarding the ability of HIV‐1 to cross the female genital mucosa.