CROI 2016 Abstract eBook

Abstract Listing

Poster Abstracts

240

In Vivo Evidence for a Role of Pyroptosis in HIV Pathogenesis Gilad Doitsh ; Stefanie Sowinski; Caroline Miller;Warner C. Greene Gladstone Insts, San Francisco, CA, USA

Background: 30% of HIV-infected subjects fail to increase their CD4 T-cell lymphocyte counts during combination antiretroviral therapy (cART) and remain both inflamed and immunodeficient. These individuals are termed “immunological non-responders.” Systemic inflammation persists in these HIV-infected patients despite the absence of detectable viremia. The reason for this persistent inflammation is unknown. We have found that that CD4 T cells in lymphoid tissues are naturally primed to mount inflammatory responses–– these cells constitutively express high levels of cytoplasmic pro-IL-1b, ASC (caspase-1 adaptor protein), and NLRP3 inflammasome. The release of intracellular ATP by pyroptotic CD4 T cells may provide a second inflammatory stimulus to induce activation of caspase-1 by the NLRP3 inflammasome in surrounding uninfected CD4 T cells that are already primed. Thus, pyroptosis initiated by HIV may result in an avalanche of new rounds of pyroptosis in primed CD4 T cells by the repeated release of intracellular ATP in a virus-independent manner . Such an “auto-inflammation” scenario could generate persistent rounds of pyroptosis, chronic inflammation, and loss of CD4 T cells even when viral replication is reduced by antiretroviral therapy Methods: Longitudinal analyses of plasma and peripheral lymphoid tissues from subject with three distinct clinical backgrounds: uninfected, infected untreated, infected treated virological suppressed. Results: We now show extensive caspase-1 activity and secretion of IL-18 in large areas of paracortical zones in lymph nodes freshly recovered from infected untreated patients. Interestingly, while caspase-1 activity is decreased in some subjects following treatment with antiretroviral therapy (ART), other patients exhibit persistent caspase-1 activity even when viral replication was undetectable. Moreover, in tissues where active caspase-1 was detected, we observed a significant loss of myeloid cells including resident macrophages and dendritic cells. This loss was consistent with high levels of extracellular bioactive IL-18, suggesting cell death by caspase-1-mediated pyroptosis. These findings highlight a role for pyroptosis beyond CD4 T cell death, and underscore the involvement of myeloid cells in promoting chronic inflammation and secretion of IL-18. Conclusions: These preliminary findings further underscore the role of pyroptosis during R5-tropic viral infection in vivo , raising the possibility for a new class of “anti-AIDS” therapeutics.

Poster Abstracts

241 Caspase Inhibition Prevents HIV Replication and Cell Death in Human Lymphoid Tissue Elisabet Gómez-Mora 1 ; Elisabet Garcia 1 ; PeterWienberg Ludwig 2 ; Maria Dolores Guerrero Gilabert 3 ; Bonaventura Clotet 4 ; Julià Blanco 5 ; Cecilia Cabrera 6 1 IrsiCaixa Inst for AIDS Rsr, Badalona, Spain; 2 Hosp Sant Joan de Déu, Barcelona, Spain; 3 Hosp Universitari Germans Trias i Pujol, Badalona, Spain; 4 Lluita Contra la SIDA Fndn, Germans Trias i Pujol Univ Hosp, Barcelona, Spain; 5 Univ de Vic, Barcelona, Spain; 6 IrsiCaixa Inst for AIDS Rsr, Barcelona, Spain Background: The pathway causing CD4 T-cell death in HIV-infected hosts remains poorly understood although apoptosis has been shown as a key mechanism. Using an ex vivo human lymphoid aggregate culture, it has been proposed that most of the quiescent CD4 T cells die by caspase-1-mediated pyroptosis, although the mechanisms of cell death in ex vivo lymphoid tissue has not been explored. Methods: Human tonsil histocultures were prepared from 4 healthy individuals and infected ex vivo with a X4 virus in the presence or absence of the entry inhibitor JM-2987 (1μg/ mL), the integrase inhibitor raltegravir (1μg/mL) and the pan-caspase inhibitor Q-VD-OPh (30μM). Drugs were added to the cultures 2 hours before infection. Culture mediumwas collected and replace every 3 days during 12 days and the p24 production was measured in the supernatant by ELISA. Tissue cells were isolated and CD4 T cell depletion (CD3+CD8-), p24 cellular content, and caspase-1 and 3/7 activity (FLICA assay) was analyzed by flow cytometry.

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CROI 2016

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