Dynamics of Simian Immunodeficiency Virus Two-Long-Terminal-Repeat Circles in the Presence and Absence of CD8+ Cells.

CD8+ cells play a key role in human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) infection, but their specific mechanism(s) of action in controlling the virus is unclear. Two-long-terminal-repeat (2-LTR) circles are extrachromosomal products generated upon failed integration of HIV/SIV. To understand the specific effects of CD8+ cells on infected cells, we analyzed the dynamics of 2-LTR circles in SIVmac251-infected rhesus macaques (RMs) treated with an integrase inhibitor (INT). Twenty RMs underwent CD8+ cell depletion and received raltegravir (RAL) monotherapy or a combination of both. Blood, lymph nodes (LNs), and gut biopsy specimens were routinely sampled. Plasma viral loads (pVLs) and 2-LTR circles from peripheral blood mononuclear cells (PBMCs) and LN lymphocytes were measured with quantitative reverse transcription-PCR (qRT-PCR). In the CD8 depletion group, an ∼1-log increase in pVLs and a slow increase in PBMC 2-LTRs occurred following depletion. In the INT group, a strong decline in pVLs upon treatment initiation and no change in 2-LTR levels were observed. In the INT and CD8+ cell depletion group, an increase in pVLs following CD8 depletion similar to that in the CD8 depletion group was observed, with a modest decline following INT initiation, and 2-LTR circles significantly increased in PBMCs and LNs. Analyzing the 2-LTR data across all treatment groups with a mathematical model indicates that the data best support an effect of CD8+ cells in killing cells prior to viral integration. Sensitivity analyses of these results confirm that effect but also allow for additional effects, which the data do not discriminate well. Overall, we show that INT does not significantly increase the levels of 2-LTR circles. However, CD8+ cell depletion increases the 2-LTR levels, which are enhanced in the presence of an INT.IMPORTANCE CD8+ T cells play an essential role in controlling HIV and SIV infection, but the specific mechanisms involved remain poorly understood. Due to failed viral infection, HIV and SIV can form 2-LTR extrachromosomal circles that can be quantified. We present novel data on the dynamics of these 2-LTR forms in a SIV-infected macaque model under three different treatment conditions: depletion of CD8+ cells, administration of the integrase inhibitor in a monotherapy, which favors the formation of 2-LTR circles, and a combination of the two treatments. We used a new mathematical model to help interpret the data, and the results suggest that CD8+ cells exert a killing effect on infected cells prior to virus integration. These results provide new insights into the mechanisms of action of CD8+ cells in SIV infection. Confirmation of our results would be an important step in understanding immune control of HIV.

Inflammatory monocytes expressing tissue factor drive SIV and HIV coagulopathy.

In HIV infection, persistent inflammation despite effective antiretroviral therapy is linked to increased risk of noninfectious chronic complications such as cardiovascular and thromboembolic disease. A better understanding of inflammatory and coagulation pathways in HIV infection is needed to optimize clinical care. Markers of monocyte activation and coagulation independently predict morbidity and mortality associated with non-AIDS events. We identified a specific subset of monocytes that express tissue factor (TF), persist after virological suppression, and trigger the coagulation cascade by activating factor X. This subset of monocytes expressing TF had a distinct gene signature with up-regulated innate immune markers and evidence of robust production of multiple proinflammatory cytokines, including interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), and IL-6, ex vivo and in vitro upon lipopolysaccharide stimulation. We validated our findings in a nonhuman primate model, showing that TF-expressing inflammatory monocytes were associated with simian immunodeficiency virus (SIV)-related coagulopathy in the progressive [pigtail macaques (PTMs)] but not in the nonpathogenic (African green monkeys) SIV infection model. Last, Ixolaris, an anticoagulant that inhibits the TF pathway, was tested and potently blocked functional TF activity in vitro in HIV and SIV infection without affecting monocyte responses to Toll-like receptor stimulation. Strikingly, in vivo treatment of SIV-infected PTMs with Ixolaris was associated with significant decreases in D-dimer and immune activation. These data suggest that TF-expressing monocytes are at the epicenter of inflammation and coagulation in chronic HIV and SIV infection and may represent a potential therapeutic target.

Critical Role for the Adenosine Pathway in Controlling Simian Immunodeficiency Virus-Related Immune Activation and Inflammation in Gut Mucosal Tissues.

UNLABELLED: The role of the adenosine (ADO) pathway in human immunodeficiency virus type 1/simian immunodeficiency virus (HIV-1/SIV) infection remains unclear. We compared SIVsab-induced changes of markers related to ADO production (CD39 and CD73) and breakdown (CD26 and adenosine deaminase) on T cells from blood, lymph nodes, and intestine collected from pigtailed macaques (PTMs) and African green monkeys (AGMs) that experience different SIVsab infection outcomes. We also measured ADO and inosine (INO) levels in tissues by mass spectrometry. Finally, we assessed the suppressive effect of ADO on proinflammatory cytokine production after T cell receptor stimulation. The baseline level of both CD39 and CD73 coexpression on regulatory T cells and ADO levels were higher in AGMs than in PTMs. Conversely, high INO levels associated with dramatic increases in CD26 expression and adenosine deaminase activity were observed in PTMs during chronic SIV infection. Immune activation and inflammation markers in the gut and periphery inversely correlated with ADO and directly correlated with INO. Ex vivo administration of ADO significantly suppressed proinflammatory cytokine production by T cells in both species. In conclusion, the opposite dynamics of ADO pathway-related markers and contrasting ADO/INO levels in species with divergent proinflammatory responses to SIV infection support a key role of ADO in controlling immune activation/inflammation in nonprogressive SIV infections. Changes in ADO levels predominately occurred in the gut, suggesting that the ADO pathway may be involved in sparing natural hosts of SIVs from developing SIV-related gut dysfunction. Focusing studies of the ADO pathway on mucosal sites of viral replication is warranted. IMPORTANCE: The mechanisms responsible for the severe gut dysfunction characteristic of progressive HIV and SIV infection in humans and macaques are not completely elucidated. We report that ADO may play a key role in controlling immune activation/inflammation in nonprogressive SIV infections by limiting SIV-related gut inflammation. Conversely, in progressive SIV infection, significant degradation of ADO occurs, possibly due to an early increase of ADO deaminase complexing protein 2 (CD26) and adenosine deaminase. Our study supports therapeutic interventions to offset alterations of this pathway during progressive HIV/SIV infections. These potential approaches to control chronic immune activation and inflammation during pathogenic SIV infection may prevent HIV disease progression.

Kinetics of myeloid dendritic cell trafficking and activation: impact on progressive, nonprogressive and controlled SIV infections.

We assessed the role of myeloid dendritic cells (mDCs) in the outcome of SIV infection by comparing and contrasting their frequency, mobilization, phenotype, cytokine production and apoptosis in pathogenic (pigtailed macaques, PTMs), nonpathogenic (African green monkeys, AGMs) and controlled (rhesus macaques, RMs) SIVagmSab infection. Through the identification of recently replicating cells, we demonstrated that mDC mobilization from the bone marrow occurred in all species postinfection, being most prominent in RMs. Circulating mDCs were depleted with disease progression in PTMs, recovered to baseline values after the viral peak in AGMs, and significantly increased at the time of virus control in RMs. Rapid disease progression in PTMs was associated with low baseline levels and incomplete recovery of circulating mDCs during chronic infection. mDC recruitment to the intestine occurred in all pathogenic scenarios, but loss of mucosal mDCs was associated only with progressive infection. Sustained mDC immune activation occurred throughout infection in PTMs and was associated with increased bystander apoptosis in blood and intestine. Conversely, mDC activation occurred only during acute infection in nonprogressive and controlled infections. Postinfection, circulating mDCs rapidly became unresponsive to TLR7/8 stimulation in all species. Yet, stimulation with LPS, a bacterial product translocated in circulation only in SIV-infected PTMs, induced mDC hyperactivation, apoptosis and excessive production of proinflammatory cytokines. After infection, spontaneous production of proinflammatory cytokines by mucosal mDCs increased only in progressor PTMs. We thus propose that mDCs promote tolerance to SIV in the biological systems that lack intestinal dysfunction. In progressive infections, mDC loss and excessive activation of residual mDCs by SIV and additional stimuli, such as translocated microbial products, enhance generalized immune activation and inflammation. Our results thus provide a mechanistic basis for the role of mDCs in the pathogenesis of AIDS and elucidate the causes of mDC loss during progressive HIV/SIV infections.