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Objectives: Previous studies have shown that HIV can be transcytosed across epithelial cell line barriers; however, there is no information concerning primary epithelial cells. Our objectives were to determine if primary epithelial cells have the ability to harbor and transmit HIV and to determine if primary epithelial cells can transcytose HIV.
Methods: For HIV transmission, primary prostate epithelial cells (PrEC) and two epithelial cervical carcinoma cell lines, ME-180 and CaSKI, were inoculated with HI and washed. Various concentrations of resting or activated CD8-depleted PBMCs were added before or after the epithelial cells were trypsinized.
Supernatants were monitored every 2 days for HIV expression using a p24 ELISA. DNA PCR was performed on the pot-trypsinized epithelial cells to evaluate proviral integration. For HIV transcytosis, the epithelial cells were cultured on 0.4 mM transwell filters until confluent (day 7). Cell-free HIV (LAI; MOI .001) or cell-associated (18 hours, TNF a-induced OM10.1 cell line) HIV was added to the apical side. The basolateral medium was sampled for HIV p24.
Results: Without trypsinization, HIV was recovered by day 3 from ME-180 and CaSKI cell lines and by day 7 from the PrEC cells y activated PBMCs but not by resting PBMCs. For all epithelial cells, at least 5 [times] 105 activated PBMCs (2 PBMCs to 1 epithelial cell) were required for HIV recovery. Trypsinization of the epithelial cells resulted in a loss of recoverable HIV from PEC, but not ME-180 and CaSKI cells, even though all transiently had provirus. We next explored HIV transcytosis. PrEC developed a tight-junction monolayer as seen by high transepithelial resistance (433 W [times] cm2). CaSKI cells developed a moderate tight-junction monolayer (50 W [times] cm2), while the ME-180 cells failed to make a tight-junction monolayer. Consequently, cell-free HIV was readily transcytosed through ME-180 cells by 1 hour and through CaSKI cells by 2 hours. Cell-associated virus began to transcytose through ME-180 and CaSKI cells by 24 hours. Importantly, PrEC did not transcytose cell-free or cell-associated HIV.
Conclusions: Both primary and immortalized epithelial cells have the capacity to transiently sequester HIV, but primary (PrEC) cells are incapable of transmissin. Further, formation of a tight-junction monolayer by PrEC did not allow transcytosis of cell-free or cell-associated HIV. Collectively, these data suggest that the in vivo mucosal epithelial barrier protects against HIV transmission, and that factors, such as STDs, affecting the integrity of transepithelial tight-junctions may allow viral entry and thus have implications for sexual transmission.
Choice 1: Track B Clinical Science and Care, Organ Systems, Genitourinary
Choice 2: [A24]
Presentation Date, Time, Room: 30-Jun-98, 3:00P, Hall IV
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