Death of HIV-infected lymphoid CD4 T cells and release of bioactive IL-1β are controlled by caspase-1. a, b. Caspase-1 inhibitors are sufficient to prevent CD4 T-cell death in HIV-infected HLACs. Viable CD4 T cells were counted by flow cytometry, and supernatants were analyzed for levels of cytoplasmic LDH enzyme release. c, Infection with CCR5-dependent HIV-1 induces pyroptosis of lymphoid CD4 T cells. Death of CCR5-expressing CD4 T cells is prevented by caspase-1 inhibitors and TAK779, but not buy the CXCR4 antagonist, AMD3100. Due to the small number of target CCR5-expressing cells, this experiment was performed by overlaying tonsil cells on a monolayer of 293T cells that had been transfected with an R5-tropic proviral HIV-1 clone, as previously described. The co-culture conditions for the R5 virus experiment induced no activation of the overlaid cells. d. Efficient repression of target genes by shRNA-coding lentiviral vectors. e. shRNA LV designed to silence either caspase-1 or ASC, key components of the pyroptotic pathway, protect lymphoid CD4 T cells from death by nigericin or HIV-1 infection. To specifically assess non-productively infected cells, cultures were treated with AZT before infections with HIV-1. f. Caspase-1 cleavage in HIV-infected CD4 T cells is blocked by specific caspase-1 inhibitors. g. Inhibitors of caspase-1, but not NLRP3, prevent release of bioactive IL-1β from HIV-infected lymphoid CD4 T cells. Error bars represent SD/√n of at least three independent experiments utilizing tonsil cells from at least three different donors. Protein analyses represent results from three independent experiments utilizing tonsillar CD4 T cells from three different donors.