Acute SIV Infection Research Articles

Nef gene evolution from a single transmitted strain in acute SIV infection

BackgroundThe acute phase of immunodeficiency virus infection plays a crucial role in determining steady-state virus load and subsequent progression of disease in both humans and nonhuman primates. The acute period is also the time when vaccine-mediated effects on host immunity are likely to exert their major effects on virus infection. Recently we developed a Monte-Carlo (MC) simulation with mathematical analysis of viral evolution during primary HIV-1 infection that enables classification of new HIV-1 infections originating from multiple versus single transmitted viral strains and the estimation of time elapsed following infection.ResultsA total of 322 SIV nef SIV sequences, collected during the first 3 weeks following experimental infection of two rhesus macaques with the SIVmac239 clone, were analyzed and found to display a comparable level of genetic diversity, 0.015% to 0.052%, with that of env sequences from acute HIV-1 infection, 0.005% to 0.127%. We confirmed that the acute HIV-1 infection model correctly identified the experimental SIV infections in rhesus macaques as "homogenous" infections, initiated by a single founder strain. The consensus sequence of the sampled strains corresponded to the transmitted sequence as the model predicted. However, measured sequential decrease in diversity at day 7, 11, and 18 post infection violated the model assumption, neutral evolution without any selection.ConclusionWhile nef gene evolution over the first 3 weeks of SIV infection originating from a single transmitted strain showed a comparable rate of sequence evolution to that observed during acute HIV-1 infection, a purifying selection for the founder nef gene was observed during the early phase of experimental infection of a nonhuman primate.

136 Increased IL-15 production is associated with higher infection of memory CD4 T cells during acute SIV infection

136 Increased IL-15 production is associated with higher infection of memory CD4 T cells during acute SIV infection

Critical Loss of the Balance between Th17 and T Regulatory Cell Populations in Pathogenic SIV Infection

Chronic immune activation and progression to AIDS are observed after SIV infection in macaques but not in natural host primate species. To better understand this dichotomy, we compared acute pathogenic SIV infection in pigtailed macaques (PTs) to non-pathogenic infection in African green monkeys (AGMs). SIVagm-infected PTs, but not SIVagm-infected AGMs, rapidly developed systemic immune activation, marked and selective depletion of IL-17-secreting (Th17) cells, and loss of the balance between Th17 and T regulatory (Treg) cells in blood, lymphoid organs, and mucosal tissue. The loss of Th17 cells was found to be predictive of systemic and sustained T cell activation. Collectively, these data indicate that loss of the Th17 to Treg balance is related to SIV disease progression.

Increased IL-15 Production Is Associated with Higher Susceptibility of Memory CD4 T Cells to Simian Immunodeficiency Virus during Acute Infection

Acute SIV infection is characterized by explosive infection of memory CD4 T cells in peripheral and mucosal tissues. Interestingly, relatively few memory CD4 T cells are infected until as late as days 7-8 after challenge. However, by day 10 postinfection, most of the memory CD4 T cells are infected and carry viral DNA. The rapidity with which infection expands within 2-3 days to encompass virtually the entire memory CD4 T cell compartment suggests significant alterations in the susceptibility of memory CD4 T cells to infection during this period. The mechanism(s) underlying this increased permissiveness to infection is not known. In this study, we show that IL-15 secretion significantly correlates with the up-regulated expression of CD4 on memory CD4 T cells that is associated with increased permissiveness to SIV infection. Activation and proliferation of memory CD8, but not memory CD4 T cells, preceded the amplification of viral infection. Although memory CD4 T cells did not express normal activation markers, they displayed a significant up-regulation in the density of CD4 but not CCR5 expression between days 7 and 10 postinfection that correlated with increased plasma IL-15 levels and infection in these cells. Culture of purified CD4 T cells with IL-15 and/or SIV was associated with a significant increase in the expression of CD4 and infection of these sorted cells. Our results demonstrate that IL-15 contributes to the increased susceptibility of memory CD4 T cells to SIV during the early phase of acute SIV infection.

Kinetics of T lymphocyte apoptosis and the cellular immune response in SIVmac239‐infected rhesus macaques

Although increased apoptosis is a central feature of AIDS, little is known about its kinetics or relationship to the early host response in acute HIV/SIV infection. Ex vivo apoptosis in freshly isolated peripheral blood and lymph node lymphocytes was monitored longitudinally in SIVmac239-infected rhesus macaques by flow-cytometric detection of active caspase-3, cleaved poly (ADP-ribose) polymerase, and fragmented DNA. Increased apoptosis of multiple lymphocyte subsets was observed in the first 2 weeks following SIV infection. Apoptosis of CD4+ T lymphocytes was of low magnitude but peaked earlier than other T lymphocyte subsets. A 10- to 36-fold increase in CD8+ T lymphocyte apoptosis coincided temporally with onset of the SIV-specific cellular immune response and enrichment of caspase-3-positive cells within recently proliferating, activated CD8+ T lymphocytes. The virus-specific T lymphocyte response to primary infection and generalized non-specific immune activation contribute to the pathogenesis of apoptosis in acute SIV infection.

Early Resolution of Acute Immune Activation and Induction of PD-1 in SIV-Infected Sooty Mangabeys Distinguishes Nonpathogenic from Pathogenic Infection in Rhesus Macaques

Primate lentiviruses are typically apathogenic in their evolutionarily coadapted host species but can be lethal when transferred to new host species. Why such infections are pathogenic in humans and rhesus macaques (RMs) but not in sooty mangabeys (SMs), a natural host, remains unclear. Studies of chronically infected animals point to the importance of diminished immune activation in response to the infection in SMs. In this study, we sought the causes and timing of the differences in immune activation in a comparative study of acute SIV infection in RMs and SMs. Surprisingly, we show that in acute infection immune activation is comparable in SMs and RMs but thereafter, SMs quickly resolve immune activation, whereas RMs did not. Early resolution of immune activation in SMs correlated with increased expression of PD-1 and with preservation of CD4(+) T cell counts and lymphatic tissue architecture. These findings point to early control of immune activation by host immunoregulatory mechanisms as a major determinant of the different disease outcomes in SIV infection of natural vs non-natural hosts.

Pathogenic mechanisms in simian immunodeficiency virus infection

Recent work in pathogenic simian immunodeficiency (SIV) infection of Asian macaques and in natural, nonpathogenic SIV infections of African nonhuman primate species has demonstrated that persistent activation has profound effects on CD4+ memory T-cell proliferation, differentiation and survival. Disease progression in pathogenic infection has been closely linked to these dynamics, reflecting a complex interplay of virus-mediated killing, the effects of systemic activation and host regenerative mechanisms. We review these recent advances. Massive depletion of CD4+ effector-memory T cells invariably occurs during acute CCR5-tropic SIV infection, but is initially stabilized by new production of these cells from spared central memory precursors above the threshold required to maintain clinical immune competence. In pathogenic (but not natural, apathogenic) infections, a persistent state of immune activation, characterized by multiple, recurrent bursts of lymphocyte proliferation, differentiation, migration, death and functional modification of 'resting' cells, is associated with progressive depletion of central memory CD4+ T cells, and ultimately, a collapse of effector site CD4+ memory populations that is closely associated with overt immune deficiency. The importance of maintaining the regenerative capacity of the central-memory compartment of CD4+ T cells is increasingly evident. Defining the physiologic and molecular mechanisms responsible for instability of the CD4+ central-memory T cell pool could enable new immunotherapeutic interventions.

Regulatory T cells in HIV infection: pathogenic or protective participants in the immune response?

Regulatory T cells in HIV infection: pathogenic or protective participants in the immune response?

An early and robust natural killer cell response during acute SIV infection in sooty mangabeys but not rhesus macaques is correlated with disease resistance

A need for defining optimal reagents and assays to identify the NK cell lineage in non‐human primates (NHP) is required because recent data clearly implicate an important role for NK cells in contributing to disease progression in HIV‐1 infected individuals. This cell lineage is particularly involved in the acute phase of infection which can optimally be studied in SIV infected NHP. A study of the role of NK cells in SIV infected disease resistant sooty mangabeys (SM) as compared with SIV infected disease susceptible rhesus macaques (RM) may provide important clues as to the contributing role of NK cells in the polarized clinical outcome in these two species. The use of previously identified reagents along with newly defined reagents and multi‐color flow cytometry has revealed that a gated population of CD3− CD8+ NKG2A+ cells identify 4 NK cell subsets with a predominant CD16+ CD56− subset primarily contributing to functional NK cytolytic activity. Cross sectional studies revealed a significant decline in this cytolytic subset in SIV‐infected RM but not in SIV‐infected SM. Importantly, longitudinal studies of individual animals pre and post SIV infection showed that while infection induces transient increases in NK cells in both species, there was a marked difference in the kinetics by which such increases occurred with SIV‐infected SM exhibiting a much earlier and more robust NK response than SIV‐infected RM.

Acute Loss of Intestinal CD4+ T Cells Is Not Predictive of Simian Immunodeficiency Virus Virulence

The predictive value of acute gut-associated lymphoid tissue (GALT) CD4+ T cell depletion in lentiviral infections was assessed by comparing three animal models illustrative of the outcomes of SIV infection: pathogenic infection (SIVsmm infection of rhesus macaques (Rh)), persistent nonprogressive infection (SIVagm infection of African green monkeys (AGM)), and transient, controlled infection (SIVagm infection of Rh). Massive acute depletion of GALT CD4+ T cells was a common feature of acute SIV infection in all three models. The outcome of this mucosal CD4+ T cell depletion, however, differed substantially between the three models: in SIVsmm-infected Rh, the acute GALT CD4+ T cell depletion was persistent and continued with disease progression; in SIVagm, intestinal CD4+ T cells were partially restored during chronic infection in the context of normal levels of apoptosis and immune activation and absence of damage to the mucosal immunologic barrier; in SIVagm-infected Rh, complete control of viral replication resulted in restoration of the mucosal barrier and immune restoration. Therefore, our data support a revised paradigm wherein severe GALT CD4+ T cell depletion during acute pathogenic HIV and SIV infections of humans and Rh is necessary but neither sufficient nor predictive of disease progression, with levels of immune activation, proliferation and apoptosis being key factors involved in determining progression to AIDS.

Estimating the Effectiveness of Simian Immunodeficiency Virus-Specific CD8+T Cells from the Dynamics of Viral Immune Escape

Antiviral CD8(+) T cells are thought to play a significant role in limiting the viremia of human and simian immunodeficiency virus (HIV and SIV, respectively) infections. However, it has not been possible to measure the in vivo effectiveness of cytotoxic T cells (CTLs), and hence their contribution to the death rate of CD4(+) T cells is unknown. Here, we estimated the ability of a prototypic antigen-specific CTL response against a well-characterized epitope to recognize and kill infected target cells by monitoring the immunodominant Mamu-A*01-restricted Tat SL8 epitope for escape from Tat-specific CTLs in SIVmac239-infected macaques. Fitting a mathematical model that incorporates the temporal kinetics of specific CTLs to the frequency of Tat SL8 escape mutants during acute SIV infection allowed us to estimate the in vivo killing rate constant per Tat SL8-specific CTL. Using this unique data set, we show that at least during acute SIV infection, certain antiviral CD8(+) T cells can have a significant impact on shortening the longevity of infected CD4(+) T cells and hence on suppressing virus replication. Unfortunately, due to viral escape from immune pressure and a dependency of the effectiveness of antiviral CD8(+) T-cell responses on the availability of sufficient CD4(+) T cells, the impressive early potency of the CTL response may wane in the transition to the chronic stage of the infection.

Perturbations in the CD4+CD25+ regulatory T cell population of the GALT during acute SIV infection

CD4+CD25+ regulatory T cells (Tregs) suppress the activation and proliferation of effector lymphocytes. Dysregulation of these cells in the peripheral blood and lymphoid tissue has been reported in HIV-1 infection. To elucidate the role of Tregs in HIV-1-induced GALT depletion of CD4+ T cells, we used the SIV/pigtailed macaque model of HIV-1 disease to determine the distribution of Tregs in a setting of acute infection, using FoxP3 as a marker for Tregs. CD4+ T cells from the ileum, lymph nodes, and peripheral blood were isolated on day 4 (n = 3), 14 (n = 3), and 114 (n = 6) post-inoculation from SIV-infected pigtailed macaques. Real-time RT-PCR was used to quantitate FOXP3 copy number in SIV-infected and uninfected control macaques (n = 5). Expression of FoxP3 in the ileum was decreased at all stages of infection when compared to the levels in uninfected macaques. In addition, functional analysis of ileal lamina propria CD4+ T cells from SIV-infected macaques revealed a lack of suppressive activity, suggesting the absence of Tregs in that compartment. These results indicate that Tregs are depleted in the GALT of SIV-infected macaques suggesting a role for the loss of Treg-mediated suppression in the pathogenesis of the disease.

Immunodomination in the Evolution of Dominant Epitope-Specific CD8+ T Lymphocyte Responses in Simian Immunodeficiency Virus-Infected Rhesus Monkeys

Because the control of HIV-1 replication is largely dependent on CD8+ T lymphocyte responses specific for immunodominant viral epitopes, vaccine strategies that increase the breadth of dominant epitope-specific responses should contribute to containing HIV-1 spread. Developing strategies to elicit such broad immune responses will require an understanding of the mechanisms responsible for focusing CD8+ T lymphocyte recognition on a limited number of epitopes. To explore this biology, we identified cohorts of rhesus monkeys that expressed the MHC class I molecules Mamu-A*01, Mamu-A*02, or both, and assessed the evolution of their dominant epitope-specific CD8+ T lymphocyte responses (Gag p11C- and Tat TL8-specific in the Mamu-A*01+ and Nef p199RY-specific in the Mamu-A*02+ monkeys) following acute SIV infection. The Mamu-A*02+ monkeys that also expressed Mamu-A*01 exhibited a significant delay in the evolution of the CD8+ T lymphocyte responses specific for the dominant Mamu-A*02-restricted SIV epitope, Nef p199RY. This delay in kinetics was not due to differences in viral load kinetics or magnitude or in viral escape mutations, but was associated with the evolution of the Mamu-A*01-restricted CD8+ T lymphocyte responses to the highly dominant SIV epitopes Gag p11C and Tat TL8. Thus, the evolution of dominant epitope-specific CD8+ T lymphocyte responses can be suppressed by other dominant epitope-specific responses, and this immunodomination is important in determining the kinetics of dominant epitope-specific responses.

Prolonged AIDS-free Survival for SIV-infected Macaques Treated With Anti-FasL

Acute SIV infection of macaques is a model for human AIDS typified by high levels of plasma viremia that decline with the onset of specific viral immunity. SIV infection limits the development of viral immunity and persists to allow for the establishment of chronic progressive disease. The destruction of CD4+ and CD4lymphocytes argues that both direct and indirect killing mechanisms contribute to the loss of cells. Although early intervention with antiretroviral drugs reduces viremia and disease, it is still unclear whether protection is due to diminished viral cytopathicity or due to blocking a host mechanism for cell killing that is triggered by the high level of viral replication. We tested the role of FasL killing of bystander lymphocytes by injecting monkeys with a humanized monoclonal anti-FasL. Treatment with antiFasL during acute infection reduced the level of apoptosis of circulating T and B cells, peak vRNA levels were unaffected but higher antibody and CTL responses to viral proteins lead to lower set-point viremia among treated macaques. Reduced bystander killing and increased viral immunity were associated with attenuated SIV disease and a significant increase in the life span of infected macaques after transient treatment with a monoclonal antibody against FasL. from 2005 International Meeting of The Institute of Human Virology Baltimore, USA, 29 August – 2 September 2005

Increased Expression of TLR3 in Lymph Nodes during Simian Immunodeficiency Virus Infection: Implications for Inflammation and Immunodeficiency

As pattern recognition receptors, TLRs signal and induce expression of multiple host defense genes including proinflammatory cytokines and chemokines. To investigate the mechanisms of up-regulation of proinflammatory cytokines and chemokines during SIV infection in rhesus macaques, we measured the relative levels of expression of TLRs 1-10 in lymphoid tissues during different stages of SIV infection. By real-time RT-PCR, TLR3 was determined to be up-regulated in macaque lymph nodes (LN) throughout the course of infection, whereas TLR9 was down-regulated during early stages of infection. CXCL9/Mig, CXCL10/IP-10, IFN-gamma, and IFN-alpha mRNAs were also increased during acute SIV infection and AIDS. Treatment of macaque spleen and LN cells with TLR3 and TLR9 ligands led to the induction of these same genes. TLR3 stimulation had disparate effects on viral transcription and viral replication, because poly(I:C), a model TLR3 ligand, stimulated the viral promoter but potently inhibited SIV replication in primary cultures of macaque spleen and LN cells. These findings identify roles for TLR3 inflammation in lymphoid tissues and in the immunopathogenesis of HIV-1/SIV, and suggest that TLR3 ligands could potentially be used to flush out latently infected cells that persist during antiretroviral therapies.

T-cell dynamics during acute SIV infection.

To delineate T-cell dynamics during acute SIV infection, particularly of phenotypically defined memory T cell subsets. T cells are a heterogeneous mix of naive and memory subsets delineated by simultaneously measuring CD4, CD8, CD45RA/RO, CD11a, CD28, and CD27. The effects of SIV infection on these subsets was measured to evaluate the impact of changes in functionally distinct cell types during pathogenesis. Peripheral blood was obtained from six SIV-infected macaques at multiple times before and after SIV infection and analyzed using 12-color flow cytometry. Acute infection was characterized by an initial lymphopenia caused by a decline in B cells. Total T-cell counts remained steady during the early acute phase; however, CD4 cell counts declined while CD8 T cells increased. The decline in CD4 T cells was a result of a decline in both naive and memory cells. CCR5+ or CD103+ subsets of CD4 T cells were depleted but only partially accounted for the decline of CD4 memory T cells, suggesting that acute infection was associated with a rapid redistribution of T cells from the periphery. Naive CD8 cell counts declined while memory CD8 cell counts increased. The increase coincided with declines in plasma viremia and was made up initially of CD27-CD28- (effector) cells; subsequently, the predominant phenotype became CD27+CD28-, akin to central memory cells. A complete understanding of the T-cell dynamics during acute SIV or HIV infection requires the simultaneous evaluation of a broad spectrum of T-cell subsets. Changes in homeostasis and associated immunopathogenesis can no longer be accurately described simply by measuring naive and memory T-cell subsets.

Changes in dendritic cell migration and activation during SIV infection suggest a role in initial viral spread and eventual immunosuppression.

Dendritic cells (DC) serve an essential function in linking the innate and acquired immune responses to antigen. Peripheral DC acquire antigen and migrate to draining lymph nodes, where they localize to the T cell-rich paracortex and function as potent antigen presenting cells. We examined the effects of human immunodeficiency virus (HIV) infection on DC function in vivo using the rhesus macaque/simian immunodeficiency virus (SIV) model. Our data show that during acute SIV infection, Langerhans cell density is reduced in skin and activated DC are increased in proportion in lymph nodes, whereas during AIDS, DC migration from skin and activation within lymph nodes are suppressed. These findings suggest that changes in DC function at different times during the course of infection may serve to promote virus dissemination and persistence: early during infection, DC mobilization may facilitate virus spread to susceptible lymph node T cell populations, whereas depressed DC function during advanced infection could promote generalized immunosuppression.

Enhanced Cellular Immune Response and Reduced CD8+ Lymphocyte Apoptosis in Acutely SIV-Infected Rhesus Macaques after Short-Term Antiretroviral Treatment

Losing the decisive virus-specific functions of both CD4+ and CD8+ T lymphocytes in the first weeks after immunodeficiency virus infection ultimately leads to AIDS. The SIV/rhesus monkey model for AIDS was used to demonstrate that a 4-week chemotherapeutic reduction of viral load during acute SIV infection of macaques allowed the development of a competent immune response able to control virus replication after discontinuation of treatment in two of five monkeys. Increasing SIV-specific CD4+ T-helper-cell proliferation was found in all macaques several weeks after treatment, independent of their viral load. However, only macaques with low viral loads showed persistent T-cell reactivity of lymph node cells. In contrast to animals with higher viral loads, T-helper-cell counts and memory T-helper cells did not decline in the two macaques controlling viral replication. Lymphocyte apoptosis was consistently low in all treated macaques. In contrast, high CD8+ lymphocyte death but only slightly increased CD4+ lymphocyte apoptosis were observed during the first weeks after infection in untreated control animals, indicating that early apoptotic death of virus-specific CTL could be an important factor for disease development. Antiretroviral treatment early after infection obviously retained virus-specific and competent T lymphocytes, whereby a virus-specific immune response could develop in two animals able to control the viral replication after cessation of treatment.

Mechanisms associated with thymocyte apoptosis induced by simian immunodeficiency virus.

Despite considerable research, the mechanisms by which HIV disrupts thymic function remain controversial. We have described the phenotypic changes that occur in the thymus of SIV-infected macaques during acute SIV infection. In this study, we analyzed the effects of SIV infection on apoptotic pathways in thymic tissue from newborn macaques infected with SIV. Thymocyte apoptosis was accompanied by a modest increase in surface Fas expression, a profound decrease in the frequency of bcl-2-positive cells, as well as the amount of bcl-2 per cell. With control of viral replication, levels of bcl-2 and Fas returned to baseline together with a return to basal levels of apoptosis. In the thymus, SIV infection resulted in depletion of CD4+CD8+ thymocytes, an increase in apoptosis of thymocytes, and a down-regulation of MHC class I molecules. These changes peaked 14-21 days after infection at or just after peak viremia. This data further suggests disruption of the antiapoptotic pathway regulated by bcl-2 plays a critical role in SIV-induced apoptosis of thymocytes.

Virus-induced cytokines regulate circulating lymphocyte levels during primary SIV infections.

Decline in blood CD4+ lymphocytes during primary symptomatic infections with HIV is usually attributed to viral killing, and has not been considered in terms of altered lymphocyte migration and sequestration. We therefore sought to examine whether CD4+ cell loss from blood of macaques undergoing an acute primary SIV infection might be due to increased synthesis of cytokines, known to profoundly affect lymphocyte trafficking, rather than to direct lymphocyte destruction by virus. The findings indicate that rapid lymphocyte depletion following acute infection is not selective for CD4+ cells, correlates precisely with increased plasma IFN-gamma and tumor necrosis factor-alpha levels, and is reversible. CD4/CD8 ratios in lymph nodes with high viral burdens remain relatively unchanged despite lymphocyte loss from blood. Levels of cytokine mRNA measured in lymphoid organs reflect neither cytokine plasma levels nor their potential to induce sequestration. These results support a model of cytokine-induced lymphocyte extravasation to account for the acute HIV/SIV-induced CD4+ cell lymphopenia and raise questions regarding the extent to which altered lymphocyte migration plays a role in the gradual CD4+ cell depletion throughout infection.