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Apoptosis has been proposed as a key mechanism responsible for CD4+ T cell depletion and immune dysfunction during HIV infection. We demonstrated that Q-VD-OPH, a caspase inhibitor, inhibits spontaneous and activation-induced death of T cells from SIV-infected rhesus macaques (RMs). When administered during the acute phase of infection, Q-VD-OPH was associated with (a) reduced levels of T cell death, (b) preservation of CD4+/CD8+ T cell ratio in lymphoid organs and in the gut, (c) maintenance of memory CD4+ T cells, and (d) increased specific CD4+ T cell response associated with the expression of cytotoxic molecules. Although therapy was limited to the acute phase of infection, Q-VD-OPH-treated RMs showed lower levels of both viral load and cell-associated SIV DNA as compared with control SIV-infected RMs throughout the chronic phase of infection, and prevented the development of AIDS. Overall, our data demonstrate that Q-VD-OPH injection in SIV-infected RMs may represent an adjunctive therapeutic agent to control HIV infection and delaying disease progression to AIDS.

The identification of human immunodeficiency virus type 1 (HIV-1) reservoir has been the center of extensive research for 25 years. In a recent work published in Cell Death and Differentiation, we show that mesenteric lymph nodes which drain intestine could represent the main reservoir for the virus. This concept has been established in a rhesus macaque model. Moreover, among the mechanisms associated with the lack of viral control, we suggest a major role of apoptosis in the death of CD8 T cells. This programmed cell death is associated with increased expression of immunosuppressive factors in lymph nodes such as TGF-b and two molecules regulating lymphocyte metabolism, IDO and PD-1. In this context, the virus benefits from the immune suppression which prevails within this "sanctuary", which offers optimal conditions for its persistence.

Simian immunodeficiency virus (SIV) infection of rhesus macaques (RMs) provides a reliable model to study the relationship between lentivirus replication, cellular immune responses, and CD4+ T-cell dynamics. Here we investigated, using SIVmac251-infected RMs of a Chinese genetic background (which experience a slower disease progression than Indian RMs), the dynamics of CD4+ CCR5+ T cells, as this subset of memory/activated CD4+ T cells is both a preferential target of virus replication and a marker of immune activation. As expected, we observed that the number of circulating CD4+ CCR5+ T cells decreases transiently at the time of peak viremia. However, at 60 days postinfection, i.e., when set-point viremia is established, the level of CD4+ CCR5+ T cells was increased compared to the baseline level. Interestingly, this increase correlated with faster disease progression, higher plasma viremia, and early loss of CD4+ T-cell function, as measured by CD4+ T-cell count, the fraction of memory CD4+ T cells, and the recall response to purified protein derivative. Taken together, these data show a key difference between the dynamics of the CD4+ CCR5+ T-cell pool (and its relationship with disease progression) in Chinese RMs and those described in previous reports for Indian SIVmac251-infected RMs. As the SIV-associated changes in the CD4+ CCR5+ T-cell pool reflect the opposing forces of SIV replication (which reduces this cellular pool) and immune activation (which increases it), our data suggest that in SIV-infected Chinese RMs the impact of immune activation is more prominent than that of virus replication in determining the size of the pool of CD4+ CCR5+ T cells in the periphery. As progression of HIV infection in humans also is associated with a relative expansion of the level of CD4+ CCR5+ T cells, we propose that SIV infection of Chinese RMs is a very valuable and important animal model for understanding the pathogenesis of human immunodeficiency virus infection.

SIV-infected macaques exhibit distinct rates of progression to AIDS and despite significant increases in CD8+ T cells, immune cells fail to control and eradicate SIV in vivo. Here, we investigated the interplay between viral reservoir sites, CD8+ T-cell activation/death and outcome. Our data provide strong evidence that mesenteric (Mes) lymph nodes represent major reservoirs not only for SIV-infected macaques progressing more rapidly toward AIDS but also in controllers. We demonstrate that macaques progressing faster display greater expression of TGF-beta and Indoleamine 2,3 dioxygenase in particular in intestinal tissues associated with a phosphorylation of the p53 protein on serine 15 in CD8+ T cells from Mes lymph nodes. These factors may act as a negative regulator of CD8+ T-cell function by inducing a Bax/Bak/Puma-dependent death pathway of effector/memory CD8+ T cells. Greater T-cell death and viral dissemination was associated with a low level of TIA-1+ expressing cells. Finally, we provide evidence that abrogation of TGF-beta in vitro enhances T-cell proliferation and reduces CD8+ T-cell death. Our data identify a mechanism of T-cell exhaustion in intestinal lymphoid organs and define a potentially effective immunological strategy for the modulation of progression to AIDS.

Most cell death stimuli trigger the mitochondrial release of cytochrome c and other cofactors that induce caspase activation and ensuing apoptosis. Apoptosis is also associated with massive mitochondrial fragmentation and cristae remodeling. Dynamin-related protein 1 (Drp1), a protein of the mitochondrial fission machinery, has been reported to participate in apoptotic mitochondrial fragmentation. Several theories explaining the mechanisms of cytochrome c release have been proposed. One suggests that it relies on the activation of Drp1-mediated mitochondrial fission. Here, we report that downregulation of Drp1 inhibits fragmentation of the mitochondrial network and partially prevents the release of cytochrome c but fails to prevent the release of other mitochondrial factors such as second mitochondria-derived activator of caspase/direct IAP-binding protein with low pI, Omi/HtrA2, adenylate kinase 2 and deafness dystonia peptide/TIMM8a. An explanation for the prevention of cytochrome c release is provided by our observation that inhibiting Drp1-mediated mitochondrial fission prevents the mitochondrial release of soluble OPA1 that was proposed to regulate cristae remodeling and complete cytochrome c release during apoptosis. Finally, we observed that downregulation of Drp1 delays but does not inhibit apoptosis, suggesting that mitochondrial fragmentation is not a prerequisite for apoptosis.

Mitochondrial outer membrane permeabilization (MOMP) is associated with most pro-apoptotic stimuli. MOMP results in the release of cytochrome c from the mitochondria into the cytosol, triggering caspase activation and subsequent apoptosis. Several theories explaining the mechanisms of MOMP have been proposed, one of which suggests that MOMP relies on the activation of the molecular machinery involved in fission, resulting in mitochondrial fragmentation. By contrast, several recent studies suggest that mitochondrial fragmentation occurs following MOMP. Moreover, under some conditions. MOMP occurs without mitochondrial fragmentation and, in fact, fragmentation even inhibits MOMP. Here, I discuss the apparently conflicting data and conclude that mitochondrial fragmentation is probably not a prerequisite for MOMP and cytochrome c release.

Immunological and virological events that occur during the earliest stages of SIV infection are now considered to have a major impact on subsequent disease progression. In the present study, we demonstrate a clear correlation between progression to AIDS and the rate of in vitro CD4+ (but not CD8+) T cell death in lymph nodes. The dying CD4+ T cells were effector memory T cells, which are critical for the immune response to pathogens. However, there was no correlation between the rate of the viral replication within lymph nodes and the extent of Fas ligand-mediated death, despite the increased sensitivity of CD4+ T cells to death in response to recombinant human Fas ligand. CD4+ T cell death was caspase and apoptosis-inducing factor independent but was clearly associated with mitochondrion damage. Interestingly, higher expression levels of the active form of Bak, a proapoptotic molecule involved in mitochondrial membrane permeabilization, were observed in SIV-infected macaques progressing more rapidly to AIDS. Finally, we demonstrated that the strain of SIV we used requires CCR5 and BOB/GRP15 molecules as coreceptors and caused death of unstimulated noncycling primary CD4+ T cells. Altogether, these results demonstrate that CD4+ T cell death occurring early after SIV infection is a crucial determinant of progression to AIDS and that it is mediated by the intrinsic death pathway.

In both Caenorhabditis elegans and mammals, Bcl-2 family members control apoptosis. In this issue of Molecular Cell, a paper by Delivani et al. (2006) sheds light on a new role of Bcl-2 family members as regulators of mitochondrial network morphology.

Pathogenic human immunodeficiency virus (HIV)/Simian immunodeficiency virus (SIV) infection is associated with increased T-cell apoptosis. In marked contrast to HIV infection in humans and SIV infection in macaques, the SIV infection of natural host species is typically nonpathogenic despite high levels of viral replication. In these nonpathogenic primate models, no observation of T-cell apoptosis was observed, suggesting that either SIV is less capable of directly inducing apoptosis in natural hosts (likely as a result of coevolution/coadaptation with the host) or, alternatively, that the indirect T-cell apoptosis plays the key role in determining the HIV-associated T-cell depletion and progression to acquired immune deficiency syndrome (AIDS). Understanding the molecular and cellular mechanisms responsible for the disease-free equilibrium in natural hosts for SIV infection, including those determining the absence of high levels of T-cell apoptosis, is likely to provide important clues regarding the mechanisms of AIDS pathogenesis in humans.

Immunological and virological events that occur during the earliest stages of HIV-1 infection are now considered to have a major impact on subsequent disease progression. We observed changes in the frequencies of CD8(bright) T cells expressing different chemokine receptors in the peripheral blood and lymph nodes of rhesus macaques during the acute phase of the pathogenic SIVmac251 infection; the frequency of CD8(bright) T cells expressing CXCR4 decreased, while the frequency of those expressing CCR5 increased. These reciprocal changes in chemokine receptor expression were associated with changes in the proportion of cycling (Ki67(+)) CD8(bright) T cells, and with the pattern of CD8(bright) T cell differentiation as defined by expression of CCR7 and CD45RA. In contrast, during the primary phase of the attenuated SIVmac251Deltanef infection, no major change was observed. Whereas during the acute phase of the infection with pathogenic SIV (2 wk postinfection) no correlate of disease protection was identified, once the viral load set points were established (2 mo postinfection), we found that the levels of cycling and of CCR5- and CXCR4-positive CD8(bright) T cells were correlated with the extent of viral replication and therefore with SIV-infection outcome. Our data reveal that, during primary SIV infection, despite intense CD8 T cell activation and an increase in CCR5 expression, which are considered as essential for optimal effector function of CD8(+) T cells, these changes are associated with a poor prognosis for disease progression to AIDS.

Highly active anti-retroviral therapy (HAART) has drastically altered the course of HIV-1 infection, resulting in a major decrease in morbidity and mortality. However, adverse drug reactions and long-term toxicities associated with HAART are now a concern. A major toxicity that has been highlighted by the increased use of HAART is related to mitochondrial side-effects. At the same time, analysis of the biochemical pathways involved in programmed cell death has revealed that mitochondria are main sensors in this process. In this article, the regulation of mitochondrial damage following the use of nucleoside analogue reverse transcriptase inhibitors (NRTIs) and protease inhibitors is discussed, with a particular focus on the putative molecular mechanisms involved.

Background: An attenuated immunodeficiency virus has been long considered innocuous. Nevertheless, converging data suggest that low levels of viral replication can still provoke AIDS. Pathogenesis of these attenuated infections is not understood., Objectives: To determine the pathogenicity of a long-term attenuated infection and to delineate T-cell dynamics during such an infection., Methods: This is a cross-sectional study of 12 rhesus macaques infected with SIV Delta nef for 8 years. We evaluated apoptosis (annexin V), activation (HLA-DR, Ki67), and newly generated T cells (TCR excision circle: TREC)., Results: Infection with SIV Delta nef induced pathological CD4 T-cell depletion after 8 years of infection. Virus replication and CD8 T-cell activation positively correlated with the rate of disease progression. The frequency of TREC within CD8+CD45RA+ cells increased in SIV Delta nef-infected animals compared to age-matched non-infected controls. Moreover, in the cohort of infected animals, TREC+CD45RA+CD4+ T-cell counts correlated strongly with non-progression to AIDS. The animal with the lowest rate of disease progression exhibited a 115-fold increase in TREC+CD45RA+CD4+ T-cell counts compared to age-matched non-infected controls. In contrast, the animal showing the fastest rate of progression to AIDS displayed 600-fold lower TREC+CD45RA+CD4+ T-cell counts compared to age-matched non-infected controls., Conclusions: Our results suggest that the thymus plays a major role in the pathogenesis of an attenuated SIV infection and that a sustained thymic output could maintain CD4 T-cell homeostasis in the context of low viral loads.

Among the many simian immunodeficiency virus (SIV) immunogens, only live attenuated viral vaccines have afforded strong protection to a natural pathogenic isolate. Since the promoter is crucial to the tempo of viral replication in general, it was reasoned that promoter exchange might confer a novel means of attenuating SIV. The core enhancer and promoter sequences of the SIV macaque 239nefstop strain (NF-kappaB/Sp1 region from -114 bp to mRNA start) have been exchanged for those of the human cytomegalovirus immediate-early promoter (CMV-IE; from -525 bp to mRNA start). During culture of the resulting virus, referred to as SIVmegalo, on CEMx174 or rhesus macaque peripheral blood mononuclear cells, deletions arose in distal regions of the CMV-IE sequences that stabilized after 1 or 2 months of culture. However, when the undeleted form of SIVmegalo was inoculated into rhesus macaques, animals showed highly controlled viremia during primary and persistent infection. Compared to parental virus infection in macaques, primary viremia was reduced by >1,000-fold to undetectable levels, with little sign of an increase of cycling cells in lymph nodes, CD4(+) depletion, or altered T-cell activation markers in peripheral blood. Moreover, in contrast to wild-type infection in most infected animals, the nef stop mutation did not revert to the wild-type codon, indicating yet again that replication was dramatically curtailed. Despite such drastic attenuation, antibody titers and enzyme-linked immunospot reactivity to SIV peptides, although slower to appear, were comparable to those seen in a parental virus infection. When animals were challenged intravenously at 4 or 6 months with the uncloned pathogenic SIVmac251 strain, viremia was curtailed by approximately 1,000-fold at peak height without any sign of hyperactivation in CD4(+)- or CD8(+)-T-cell compartment or increase in lymph node cell cycling. To date, there has been a general inverse correlation between attenuation and protection; however, these findings show that promoter exchange constitutes a novel means to highly attenuate SIV while retaining the capacity to protect against challenge virus.

Elevated CD4 T-cell turnover may lead to the exhaustion of the immune system during human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) infections. However, this hypothesis remains controversial. Most studies of this subject have concerned the blood, and information about the lymph nodes is rare and controversial. We used Ki67 expression to measure cycling T cells in the blood and lymph nodes of uninfected macaques and of macaques infected with a pathogenic SIVmac251 strain or with a nonpathogenic SIVmac251Deltanef clone. During the asymptomatic phase of infection, the number of cycling CD8(+) T cells progressively increased (two- to eightfold) both in the blood and in the lymph nodes of macaques infected with SIVmac251. This increase was correlated with viral replication and the progression to AIDS. In contrast, no increases in the numbers of cycling CD4(+) T cells were found in the blood or lymph nodes of macaques infected with the pathogenic SIVmac251 strain in comparison with SIVmac251Deltanef-infected or healthy macaques during this chronic phase. However, the lymph nodes of pre-AIDS stage SIVmac251-infected macaques contained more cycling CD4(+) T cells (low baseline CD4(+)-T-cell counts in the blood). Taken together, these results show that the profiles of CD4(+)- and CD8(+)-T-cell dynamics are distinct both in the lymph nodes and blood and suggest that higher CD4(+)-T-cell proliferation at the onset of AIDS may lead to the exhaustion of the immune system.

Infection with human immunodeficiency virus (HIV) is characterized by the gradual depletion of CD4+ T lymphocytes. The incorporation of the concept of apoptosis as a rationale to explain progressive T cell depletion has led to growing research in this field during the last 10 years. In parallel, the biochemical pathways implicated in programmed cell death have been extensively studied. Thus, the influence of mitochondrial control in the two major apoptotic pathways-the extrinsic and intrinsic pathways-is now well admitted. In this review, we summarized our current knowledge of the different pathways involved in the death of T cells in the course of HIV infection.

Objective: The acute phase of HIV and SIV infections leads to a host/virus equilibrium, and accumulating evidence suggests that this early phase dictates further progression towards AIDS. To gain insight into the early events that determine rapid disease progression, we performed a longitudinal study in the SIV rhesus macaque model, allowing an in-depth analysis of the primary stage of infection., Methods: We assessed viral replication (quantification of replicating and infected cells in lymph nodes, plasma viral load), immune response (cytotoxic T lymphocyte, antibody, proliferative responses), apoptosis and cycling cells (Ki-67 labelling) on lymph nodes and blood in nine rhesus macaques infected with the pathogenic SIVmac251 isolate., Results: Six primates remained asymptomatic during the one year follow-up period of the study, whereas three developed AIDS within 5-6 months. During the first 2 weeks of infection, peak numbers of apoptotic cells in the lymph node T-cell areas were significantly higher in the three future rapid progressors than in the six future slow progressors, and were correlated with subsequent viraemia levels measured 6 months after infection. The numbers of infected or cycling cells in the same lymph node T-cell areas, however, only became significantly different in future rapid and slow progressors 8 weeks after infection, at the end of the primary phase., Conclusion: Our findings identified extensive apoptosis induction in peripheral lymphoid organs as an early and predictive event that may play a crucial role in impairing the capacity of the immune system to control viral replication and progression towards disease.

We assessed the possible role in vivo CD4(+) CD8(+) T cells as a viral reservoir for simian immunodeficiency virus (SIV), in a macaque with 50% CD4(+) CD8(+) T cells in peripheral blood. During primary infection (day 14) of this rhesus macaque with the pathogenic SIVmac251 strain, proviruses were detected at similar frequencies in CD4(+) CD8(+) T cells (1/10) and CD4(+) T cells (1/10) and at a lower frequency in CD8(+) T cells (1/800). On day 235, no viral DNA was detected in CD8(+) cells, despite the persistent high viral load, indicating that CD8(+) cells do not constitute a reservoir during the chronic phase of SIV infection. Infection induced early lymphopenia of CD4(+), CD4(+) CD8(+), and CD8(+) cells; only the CD8(+) cell population returned to initial levels and expanded further. We found that CD4(+) CD8(+) T cells expressed the costimulatory CD28 molecule less and were more prone to die in vitro after phytohemagglutinin/interleukin 2 stimulation than were CD4(+) T cells. Taken together, massive death of CD4(+) CD8(+) T cells during acute stages of SIV infection may explain why CD8(+) T cells did not represent a major reservoir for SIV at the onset of infection.