294 Antibody to LFA-1 Blocks HIV-1 Infection in Primary Cells through the Activation of Caspase 8 L. M. Cimakasky*, J. E. K. Hildreth Johns Hopkins Univ Sch of Med, Baltimore, MD
Background: We have shown that an anti-LFA-1 monoclonal antibody (MAb) blocks HIV-1 infection of primary cells through a post-entry mechanism. Investigation of the mechanism by which anti-LFA-1 MAb blocks infection suggested that apoptosis may be involved. We hypothesized that caspase-dependent apoptosis is induced by inhibition of LFA-1 on newly infected cells.
Methods: We used PHA-stimulated peripheral blood mononuclear cells (PBMC) from normal donors and the laboratory strain HIV-IIIB. We examined the effect of anti-LFA-1 MAb on HIV-IIIB infection, attachment, entry, and output. Virus was measured by p24 ELISA. Caspase activity was measured using a conjugated caspase substrate that fluoresces upon cleavage. The pan-caspase inhibitor z-VAD-fmk and the caspase 8 inhibitor z-IETD-fmk, both cell-permeable, were used to block caspase activity.
Results: Anti-LFA-1 MAb blocked HIV-IIIB infection in PHA-stimulated PBMC. However, anti-LFA-1 MAb did not block virus attachment, entry, or output. Instead, caspase 8 activity was associated with anti-LFA-1 MAb treatment of infected cultures. The pan-caspase inhibitor z-VAD-fmk and the caspase 8 inhibitor z-IETD-fmk reversed anti-LFA-1 MAb inhibition of infection.
Conclusions: Our data suggest that anti-LFA-1 MAb blocks HIV-1 infection by the induction of caspase-dependent apoptosis.
