A cellular trafficking signal in the SIV envelope protein cytoplasmic domain is strongly selected for in pathogenic infection.

The HIV/SIV envelope glycoprotein (Env) cytoplasmic domain contains a highly conserved Tyr-based trafficking signal that mediates both clathrin-dependent endocytosis and polarized sorting. Despite extensive analysis, the role of these functions in viral infection and pathogenesis is unclear. An SIV molecular clone (SIVmac239) in which this signal is inactivated by deletion of Gly-720 and Tyr-721 (SIVmac239ΔGY), replicates acutely to high levels in pigtail macaques (PTM) but is rapidly controlled. However, we previously reported that rhesus macaques and PTM can progress to AIDS following SIVmac239ΔGY infection in association with novel amino acid changes in the Env cytoplasmic domain. These included an R722G flanking the ΔGY deletion and a nine nucleotide deletion encoding amino acids 734-736 (ΔQTH) that overlaps the rev and tat open reading frames. We show that molecular clones containing these mutations reconstitute signals for both endocytosis and polarized sorting. In one PTM, a novel genotype was selected that generated a new signal for polarized sorting but not endocytosis. This genotype, together with the ΔGY mutation, was conserved in association with high viral loads for several months when introduced into naïve PTMs. For the first time, our findings reveal strong selection pressure for Env endocytosis and particularly for polarized sorting during pathogenic SIV infection in vivo.

Depletion of Bone Marrow Hematopoietic Cells in Ebolavirus-Infected Rhesus Macaques: A Possible Cause of Hematologic Abnormalities in Ebolavirus Disease.

The pathophysiology of long-recognized hematologic abnormalities in Ebolavirus (EBOV) disease (EVD) is unknown. From limited human sampling (of peripheral blood), it has been postulated that emergency hematopoiesis plays a role in severe EVD, but the systematic characterization of the bone marrow (BM) has not occurred in human disease or in nonhuman primate models. In a lethal rhesus macaque model of EVD, 18 sternal BM samples exposed to the Kikwit strain of EBOV were compared to those from uninfected controls (n = 3). Immunohistochemistry, RNAscope in situ hybridization, transmission electron microscopy, and confocal microscopy showed that EBOV infects BM monocytes/macrophages and megakaryocytes. EBOV exposure was associated with severe BM hypocellularity, including depletion of myeloid, erythroid, and megakaryocyte hematopoietic cells. These depletions were negatively correlated with cell proliferation (Ki67 expression) and were not associated with BM apoptosis during disease progression. In EBOV-infected rhesus macaques with terminal disease, BM showed marked hemophagocytosis, megakaryocyte emperipolesis, and the release of immature hematopoietic cells into the sinusoids. Collectively, these data demonstrate not only direct EBOV infection of BM monocytes/macrophages and megakaryocytes but also that disease progression is associated with hematopoietic failure, notably in peripheral cytopenia. These findings inform current pathophysiologic unknowns and suggest a crucial role for BM dysfunction and/or failure, including emergency hematopoiesis, as part of the natural history of severe human disease.

Ebola Virus Disease Features Hemophagocytic Lymphohistiocytosis/Macrophage Activation Syndrome in the Rhesus Macaque Model.

BACKGROUND: Ebola virus (EBOV) disease (EVD) is one of the most severe and fatal viral hemorrhagic fevers and appears to mimic many clinical and laboratory manifestations of hemophagocytic lymphohistiocytosis syndrome (HLS), also known as macrophage activation syndrome. However, a clear association is yet to be firmly established for effective host-targeted, immunomodulatory therapeutic approaches to improve outcomes in patients with severe EVD. METHODS: Twenty-four rhesus monkeys were exposed intramuscularly to the EBOV Kikwit isolate and euthanized at prescheduled time points or when they reached the end-stage disease criteria. Three additional monkeys were mock-exposed and used as uninfected controls. RESULTS: EBOV-exposed monkeys presented with clinicopathologic features of HLS, including fever, multiple organomegaly, pancytopenia, hemophagocytosis, hyperfibrinogenemia with disseminated intravascular coagulation, hypertriglyceridemia, hypercytokinemia, increased concentrations of soluble CD163 and CD25 in serum, and the loss of activated natural killer cells. CONCLUSIONS: Our data suggest that EVD in the rhesus macaque model mimics pathophysiologic features of HLS/macrophage activation syndrome. Hence, regulating inflammation and immune function might provide an effective treatment for controlling the pathogenesis of acute EVD.

Immunological Responses to the Relapsing Fever Spirochete Borrelia turicatae in Infected Rhesus Macaques: Implications for Pathogenesis and Diagnosis.

The global public health impact of relapsing fever (RF) spirochetosis is significant, since the pathogens exist on five of seven continents. The hallmark sign of infection is episodic fever and the greatest threat is to the unborn. With the goal of better understanding the specificity of B-cell responses and the role of immune responses in pathogenicity, we infected rhesus macaques with Borrelia turicatae (a new world RF spirochete species) by tick bite and monitored the immune responses generated in response to the pathogen. Specifically, we evaluated inflammatory mediator induction by the pathogen, host antibody responses to specific antigens, and peripheral lymphocyte population dynamics. Our results indicate that B. turicatae elicits from peripheral blood cells key inflammatory response mediators (interleukin-1ß and tumor necrosis factor alpha), which are associated with preterm abortion. Moreover, a global decline in peripheral B-cell populations was observed in all animals at 14 days postinfection. Serological responses were also evaluated to assess the antigenicity of three surface proteins: BipA, BrpA, and Bta112. Interestingly, a distinction was observed between antibodies generated in nonhuman primates and mice. Our results provide support for the nonhuman primate model not only in studies of prenatal pathogenesis but also for diagnostic and vaccine antigen identification and testing.

Lack of T-cell-mediated IL-2 and TNFα production is linked to decreased CD58 expression in intestinal tissue during acute simian immunodeficiency virus infection.

For an effective T-cell activation and response, co-stimulation is required in addition to the antigen-specific signal from their antigen receptors. The CD2/CD58 interaction is considered as one of the most important T-cell co-stimulatory pathways for T-cell activation and proliferation, and its role in regulating intestinal T-cell function in acute and chronic SIV -infected macaques is poorly documented. Here, we demonstrated a significant reduction of CD58 expression in both T- and B-cell populations during acute SIV infection along with high plasma viral load and a loss of intestinal CD4+ T cells compared to SIV-uninfected control macaques. The reduction of CD58 expression in T cells was correlated with the reduced expression of T-cell-mediated IL-2 and TNFα production. Together, these results indicate that reduction in the CD2/CD58 interaction pathway in mucosal lymphocytes might play a crucial role in mucosal T-cell dysfunction during acute SIV/HIV infection.

An HSV-2 Trivalent Vaccine Is Immunogenic in Rhesus Macaques and Highly Efficacious in Guinea Pigs.

A genital herpes vaccine is urgently needed to prevent pain and suffering, reduce the incidence of neonatal herpes, and decrease the risk of HIV acquisition and transmission that accompanies genital infection. We evaluated a trivalent HSV-2 subunit antigen vaccine administered with CpG and alum in rhesus macaques and guinea pigs. The vaccine contains glycoproteins C, D and E (gC2, gD2, gE2) to block virus entry by gD2 and immune evasion by gC2 and gE2. In rhesus macaques, the trivalent vaccine induced plasma and mucosa neutralizing antibodies, antibodies that block gC2 and gE2 immune evasion activities, and stimulated CD4 T cell responses. After intravaginal challenge, a self-limited vaginal infection of brief duration was detected by histopathology and immunohistochemistry in naïve, but not in trivalent immunized macaques. Vaccine efficacy was evaluated in female guinea pigs. Animals were mock immunized, or immunized with gD2, the trivalent vaccine or the trivalent vaccine followed by a booster dose of gD2 (trivalent + gD2). The trivalent and trivalent + gD2 groups were 97% and 99% efficacious, respectively in preventing genital lesions and both outperformed gD2 alone. As a marker of transmission risk, vaginal swabs were evaluated daily for HSV-2 DNA and replication competent virus between five and seven weeks after challenge. HSV-2 DNA shedding was reduced in all groups compared with mock. Shedding of replication competent virus occurred on fewer days in the trivalent than gD2 immunized animals while the trivalent + gD2 group had no shedding of replication competent virus. Overall, the trivalent group had genital lesions on < 1% days and shedding of replication competent virus on 0.2% days. The vaccine has outstanding potential for prevention of genital herpes in humans.

A significant productive in vivo infection of resting cells with simian immunodeficiency virus in a macaque with AIDS.

Identifying the cells that can be infected with HIV in vivo and thus potentially persist as latent reservoirs is of high priority. Here, we report the major infected cells in a chronically simian immunodeficiency virus (SIV)-infected macaque that developed AIDS and encephalitis. A majority of the infected cells were detected as non-proliferating resting cells. SIV-infected non-proliferating resting cells were found to be playing an important role in viral pathogenesis, persistence, or reservoir formation.

The Role of Defensins in HIV Pathogenesis.

Profound loss of CD4+ T cells, progressive impairment of the immune system, inflammation, and sustained immune activation are the characteristics of human immunodeficiency virus-1 (HIV-1) infection. Innate immune responses respond immediately from the day of HIV infection, and a thorough understanding of the interaction between several innate immune cells and HIV-1 is essential to determine to what extent those cells play a crucial role in controlling HIV-1 in vivo. Defensins, divided into the three subfamilies α-, ß-, and θ-defensins based on structure and disulfide linkages, comprise a critical component of the innate immune response and exhibit anti-HIV-1 activities and immunomodulatory capabilities. In humans, only α- and ß-defensins are expressed in various tissues and have broad impacts on HIV-1 transmission, replication, and disease progression. θ-defensins have been identified as functional peptides in Old World monkeys, but not in humans. Instead, θ-defensins exist only as pseudogenes in humans, chimpanzees, and gorillas. The use of the synthetic θ-defensin peptide "retrocyclin" as an antiviral therapy was shown to be promising, and further research into the development of defensin-based HIV-1 therapeutics is needed. This review focuses on the role of defensins in HIV-1 pathogenesis and highlights future research efforts that warrant investigation.

Elite Control, Gut CD4 T Cell Sparing, and Enhanced Mucosal T Cell Responses in Macaca nemestrina Infected by a Simian Immunodeficiency Virus Lacking a gp41 Trafficking Motif.

UNLABELLED: Deletion of Gly-720 and Tyr-721 from a highly conserved GYxxØ trafficking signal in the SIVmac239 envelope glycoprotein cytoplasmic domain, producing a virus termed ΔGY, leads to a striking perturbation in pathogenesis in rhesus macaques (Macaca mulatta). Infected macaques develop immune activation and progress to AIDS, but with only limited and transient infection of intestinal CD4(+) T cells and an absence of microbial translocation. Here we evaluated ΔGY in pig-tailed macaques (Macaca nemestrina), a species in which SIVmac239 infection typically leads to increased immune activation and more rapid progression to AIDS than in rhesus macaques. In pig-tailed macaques, ΔGY also replicated acutely to high peak plasma RNA levels identical to those for SIVmac239 and caused only transient infection of CD4(+) T cells in the gut lamina propria and no microbial translocation. However, in marked contrast to rhesus macaques, 19 of 21 pig-tailed macaques controlled ΔGY replication with plasma viral loads of <15 to 50 RNA copies/ml. CD4(+) T cells were preserved in blood and gut for up to 100 weeks with no immune activation or disease progression. Robust antiviral CD4(+) T cell responses were seen, particularly in the gut. Anti-CD8 antibody depletion demonstrated CD8(+) cellular control of viral replication. Two pig-tailed macaques progressed to disease with persisting viremia and possible compensatory mutations in the cytoplasmic tail. These studies demonstrate a marked perturbation in pathogenesis caused by ΔGY's ablation of the GYxxØ trafficking motif and reveal, paradoxically, that viral control is enhanced in a macaque species typically predisposed to more pathogenic manifestations of simian immunodeficiency virus (SIV) infection. IMPORTANCE: The pathogenesis of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) reflects a balance between viral replication, host innate and adaptive antiviral immune responses, and sustained immune activation that in humans and Asian macaques is associated with persistent viremia, immune escape, and AIDS. Among nonhuman primates, pig-tailed macaques following SIV infection are predisposed to more rapid disease progression than are rhesus macaques. Here, we show that disruption of a conserved tyrosine-based cellular trafficking motif in the viral transmembrane envelope glycoprotein cytoplasmic tail leads in pig-tailed macaques to a unique phenotype in which high levels of acute viral replication are followed by elite control, robust cellular responses in mucosal tissues, and no disease. Paradoxically, control of this virus in rhesus macaques is only partial, and progression to AIDS occurs. This novel model should provide a powerful tool to help identify host-specific determinants for viral control with potential relevance for vaccine development.

Breadth and magnitude of antigen-specific antibody responses in the control of plasma viremia in simian immunodeficiency virus infected macaques.

BACKGROUND: Increasing evidence suggests an unexpected potential for non-neutralizing antibodies to prevent HIV infection. Consequently, identification of functional linear B-cell epitopes for HIV are important for developing preventative and therapeutic strategies. We therefore explored the role of antigen-specific immune responses in controlling plasma viremia in SIV infected rhesus macaques. METHODS: Thirteen rhesus macaques were inoculated either intravaginally or intrarectally with SIVMAC251. Peripheral blood CD4+ T-cells were quantified. Plasma was examined for viremia, antigen specific IgG, IgA and IgM binding responses and neutralizing antibodies. Regions containing binding epitopes for antigen-specific IgG, IgM and IgA responses were determined, and the minimum size of linear Envelope epitope responsible for binding antibodies was identified. RESULTS: The presence of neutralizing antibodies did not correlate the outcome of the disease. In a few SIV-infected macaques, antigen-specific IgG and IgM responses in plasma correlated with decreased plasma viremia. Early induction and the breadth of antigen-specific IgG responses were found to be significantly correlated with the control of plasma viral load. Immunoglobulin classes share similar functional linear B-cell epitopes. SIV-specific linear envelope B-cell epitopes were found to be 12 amino-acids in length. CONCLUSIONS: Early induction of combination of peptide-specific IgG responses were found to be responsible for the control of plasma viral load and indicative of disease outcome in SIV-infected rhesus macaques and might be important for the development of therapeutic strategies for control or prevention of HIV/AIDS.

Septicemia in an Indian Rhesus Macaque (Macaca mulatta) associated with Providencia stuartii.

Providencia stuartii (P. stuartii) is an opportunistic pathogen and major concern in urinary catheter-related infections in human medicine. Here we report P. stuartii-induced septicemia in an eighteen-year-old, female India-origin Rhesus macaque with multiple traumatic wounds. The animal had neutrophilic leukocytosis, necrosuppurative meningoencephalitis, hypophysitis and bronchopneumonia with vasculitis, thrombosis, and clusters of extracellular Gram-negative bacilli. P. stuartii was isolated from the lesions of the brain and lung and confirmed by PCR and sequencing. To the authors' knowledge, this is the first case of septicemia associated with P. stuartii in a non-human primate.

Transforming growth factor-ß1 regulated phosphorylated AKT and interferon gamma expressions are associated with epithelial cell survival in rhesus macaque colon explants.

Transforming growth factor-ß1 (TGF-ß1) is an important immunoregulatory cytokine that plays an obligate role in regulating T-cell functions. Here, we demonstrated the role of TGF-ß1 in regulating the survival of intestinal epithelial cells (ECs) in rhesus colon explant cultures using either anti-TGF-ß1 antibody or recombinant TGF-ß1 proteins. Neutralization of endogenous TGF-ß1 using anti-TGF-ß1 antibodies induced apoptosis of both intestinal ECs and lamina propria (LP) cells. Additionally, endogenous TGF-ß1 blocking significantly increased expression of IFNγ, TNFα, CD107a and Perforin in LP cells compared to media and isotype controls. A significant decrease in pAKT expression was detected in anti-TGF-ß1 MAbs treated explants compared to isotype and rTGF-ß1 protein treated explants. Our results demonstrated TGF-ß1 regulated pAKT and IFNγ expressions were associated with epithelial cell survival in rhesus macaque colon explants and suggest a potential role of mucosal TGF-ß1 in regulating intestinal homeostasis and EC integrity.

Cytokine/Chemokine responses in activated CD4+ and CD8+ T cells isolated from peripheral blood, bone marrow, and axillary lymph nodes during acute simian immunodeficiency virus infection.

UNLABELLED: Understanding the cytokine/chemokine networks in CD4(+) and CD8(+) T cells during the acute phase of infection is crucial to design therapies for the control of early human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) replication. Here, we measured early changes in CD4(+) and CD8(+) T cells in the peripheral blood (PB), bone marrow (BM), and axillary lymph node (ALN) tissue of rhesus macaques infected with SIVMAC251. At 21 days after infection, all tissues showed a statistically significant loss of CD4(+) T cells along with immune activation of CD8(+) T cells in PB and ALN tissue. Twenty-eight different cytokines/chemokines were quantified in either anti-CD3/28 antibody- or staphylococcal enterotoxin B-stimulated single-positive CD4(+) and CD8(+) T cells. PB CD4(+) T cells produced predominantly interleukin-2 (IL-2), whereas CD4(+) and CD8(+) T-cell subsets in tissues produced ß-chemokines both before and 21 days after SIV infection. Tissues generally exhibited massive upregulation of many cytokines/chemokines following infection, possibly in an attempt to mitigate the loss of CD4(+) T cells. There was no evidence of a T-helper 1 (TH1)-to-TH2 shift in CD4(+) T cells or a T-cytotoxic 1 (TC1)-to-TC2 cytokine shift in CD8(+) T cells in PB, BM, and ALN T-cell subsets during the acute phase of SIV infection. Despite the upregulation of several important effector cytokines/chemokines (IL-2, IL-12, IL-17, gamma interferon, granulocyte-macrophage colony-stimulating factor) by CD4(+) and CD8(+) T cells, upregulation of ß-chemokines (CCL2 and CCL22), basic fibroblast growth factor (FGF-basic), hepatocyte growth factor (HGF), and migration inhibition factor (MIF) may provide a poor prognosis either by inducing increased virus replication or by other unknown mechanisms. Therefore, drugs targeting ß-chemokines (CCL2 and CCL22), FGF-basic, HGF, or MIF might be important for developing effective vaccines and therapeutics against HIV. IMPORTANCE: Human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) infection results in early depletion of CD4(+) T cells and dysregulation of protective immune responses. Therefore, understanding the cytokine/chemokine networks in CD4(+) and CD8(+) T cells in different tissues during the acute phase of infection is crucial to the design of therapies for the control of early viral replication. Here, we measured early changes in CD4(+) and CD8(+) T cells in peripheral blood (PB), bone marrow (BM), and axillary lymph node (ALN) tissue of rhesus macaques infected with SIVMAC251. There was no evidence of a T-helper 1 (TH1)-to-TH2 shift in CD4(+) T cells or a T-cytotoxic 1 (TC1)-to-TC2 cytokine shift in CD8(+) T cells in PB, BM, and ALN T-cell subsets during the acute phase of SIV infection. Despite the upregulation of several important effector cytokines/chemokines by CD4(+) and CD8(+) T cells, upregulation of ß-chemokines, fibroblast growth factor-basic, hepatocyte growth factor, and migration inhibition factor may provide a poor prognosis.

Lack of interleukin-10-mediated anti-inflammatory signals and upregulated interferon gamma production are linked to increased intestinal epithelial cell apoptosis in pathogenic simian immunodeficiency virus infection.

UNLABELLED: Interleukin-10 (IL-10) is an immunomodulatory cytokine that is important for maintenance of epithelial cell (EC) survival and anti-inflammatory responses (AIR). The majority of HIV infections occur through the mucosal route despite mucosal epithelium acting as a barrier to human immunodeficiency virus (HIV). Therefore, understanding the role of IL-10 in maintenance of intestinal homeostasis during HIV infection is of interest for better characterization of the pathogenesis of HIV-mediated enteropathy. We demonstrated here changes in mucosal IL-10 signaling during simian immunodeficiency virus (SIV) infection in rhesus macaques. Disruption of the epithelial barrier was manifested by EC apoptosis and loss of the tight-junction protein ZO-1. Multiple cell types, including a limited number of ECs, produced IL-10. SIV infection resulted in increased levels of IL-10; however, this was associated with increased production of mucosal gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α), suggesting that IL-10 was not able to regulate AIR. This observation was supported by the downregulation of STAT3, which is necessary to inhibit production of IFN-γ and TNF-α, and the upregulation of SOCS1 and SOCS3, which are important regulatory molecules in the IL-10-mediated AIR. We also observed internalization of the IL-10 receptor (IL-10R) in mucosal lymphocytes, which could limit cellular availability of IL-10 for signaling and contribute to the loss of a functional AIR. Collectively, these findings demonstrate that internalization of IL-10R with the resultant impact on IL-10 signaling and dysregulation of the IL-10-mediated AIR might play a crucial role in EC damage and subsequent SIV/HIV pathogenesis. IMPORTANCE: Interleukin-10 (IL-10), an important immunomodulatory cytokine plays a key role to control inflammatory function and homeostasis of the gastrointestinal mucosal immune system. Despite recent advancements in the study of IL-10 and its role in HIV infection, the role of mucosal IL-10 in SIV/HIV infection in inducing enteropathy is not well understood. We demonstrated changes in mucosal IL-10 signaling during SIV infection in rhesus macaques. Disruption of the intestinal epithelial barrier was evident along with the increased levels of mucosal IL-10 production. Increased production of mucosal IFN-γ and TNF-α during SIV infection suggested that the increased level of mucosal IL-10 was not able to regulate anti-inflammatory responses. Our findings demonstrate that internalization of IL-10R with the resultant impact on IL-10 signaling and dysregulation of the IL-10-mediated anti-inflammatory responses might play a crucial role in epithelial cell damage and subsequent SIV/HIV pathogenesis.

Chronic Binge Alcohol Administration Increases Intestinal T-Cell Proliferation and Turnover in Rhesus Macaques.

BACKGROUND: Alcohol use results in changes in intestinal epithelial cell turnover and microbial translocation, yet less is known about the consequences on intestinal lymphocytes in the gut. Here, we compared T-cell subsets in the intestine of macaques before and after 3 months of chronic alcohol administration to examine the effects of alcohol on intestinal T-cell subsets. METHODS: Rhesus macaques received either alcohol or isocaloric sucrose as a control treatment daily over a 3-month period via indwelling gastric catheters. Intestinal lymphocyte subsets were identified in biopsy samples by flow cytometry. Twenty-four hours prior to sampling, animals were inoculated with bromo-deoxyuridine (BrdU) to assess lymphocyte proliferation. Immunohistochemistry was performed on tissue samples to quantitate CD3+ cells. RESULTS: Animals receiving alcohol had increased rates of intestinal T-cell turnover of both CD4+ and CD8+ T cells as reflected by increased BrdU incorporation. However, absolute numbers of T cells were decreased in intestinal tissues as evidenced by immunohistochemistry for total CD3 expression per mm(2) intestinal lamina propria in tissue sections. Combining immunohistochemistry and flow cytometry data showed that the absolute numbers of CD8+ T cells were significantly decreased, whereas absolute numbers of total CD4+ T cells were minimally decreased. CONCLUSIONS: Collectively, these data indicate that alcohol exposure to the small intestine results in marked loss of CD3+ T cells, accompanied by marked increases in CD4+ and CD8+ T-cell proliferation and turnover, which we speculate is an attempt to maintain stable numbers of T cells in tissues. This suggests that alcohol results in accelerated T-cell turnover in the gut, which may contribute to premature T-cell senescence. Further, these data indicate that chronic alcohol administration results in increased levels of HIV target cells (proliferating CD4+ T cells) that may support higher levels of HIV replication in intestinal tissues.

Divergent kinetics of proliferating T cell subsets in simian immunodeficiency virus (SIV) infection: SIV eliminates the "first responder" CD4+ T cells in primary infection.

Although increased lymphocyte turnover in chronic human immunodeficiency virus and simian immunodeficiency virus (SIV) infection has been reported in blood, there is little information on cell turnover in tissues, particularly in primary SIV infection. Here we examined the levels of proliferating T cell subsets in mucosal and peripheral lymphoid tissues of adult macaques throughout SIV infection. To specifically label cells in S-phase division, all animals were inoculated with bromodeoxyuridine 24 h prior to sampling. In healthy macaques, the highest levels of proliferating CD4(+) and CD8(+) T cells were in blood and, to a lesser extent, in spleen. Substantial percentages of proliferating cells were also found in intestinal tissues, including the jejunum, ileum, and colon, but very few proliferating cells were detected in lymph nodes (axillary and mesenteric). Moreover, essentially all proliferating T cells in uninfected animals coexpressed CD95 and many coexpressed CCR5 in the tissues examined. Confocal microscopy also demonstrated that proliferating cells were substantial viral target cells for SIV infection and viral replication. After acute SIV infection, percentages of proliferating CD4(+) and CD8(+) T cells were significantly higher in tissues of chronically infected macaques and macaques with AIDS than in those of the controls. Surprisingly, however, we found that proliferating CD4(+) T cells were selectively decreased in very early infection (8 to 10 days postinoculation [dpi]). In contrast, levels of proliferating CD8(+) T cells rapidly increased after SIV infection, peaked by 13 to 21 dpi, and thereafter remained significantly higher than those in the controls. Taken together, these findings suggest that SIV selectively infects and destroys dividing, nonspecific CD4(+) T cells in acute infection, resulting in homeostatic changes and perhaps continuing loss of replication capacity to respond to nonspecific and, later, SIV-specific antigens.

Dynamics of cytokine/chemokine responses in intestinal CD4+ and CD8+ T Cells during Acute Simian Immunodeficiency Virus Infection.

Loss of intestinal CD4(+) T cells was associated with decreased production of several T-helper 1 (TH1) and TH2 cytokines and increased production of interleukin 17 (IL-17), gamma interferon (IFN-γ), CCL4, and granulocyte-macrophage colony-stimulating factor (GM-CSF) by CD8(+) T cells 21 days after simian immunodeficiency virus (SIV) infection in rhesus macaques. Shifting of mucosal TH1 to TH2 or T-cytotoxic 1 (TC1) to TC2 cytokine profiles was not evident. Additionally, both CD4(+) and CD8(+) T cells showed upregulation of macrophage migration inhibition factor (MIF) and basic fibroblast growth factor (FGF-basic) cytokines that have been linked to HIV disease progression.

A single amino acid mutation in the envelope cytoplasmic tail restores the ability of an attenuated simian immunodeficiency virus mutant to deplete mucosal CD4+ T cells.

Disruption of the conserved motif GYxxØ in the simian immunodeficiency virus (SIV) SIVmac239 envelope (Env) cytoplasmic tail resulted in a virus (ΔGY) that exhibited a high plasma peak but uniquely failed to acutely deplete mucosal CD4(+) T cells. Here, we show that ΔGY containing a flanking S727P mutation that was acquired in ΔGY-infected macaques reacquired the ability to rapidly deplete CD4(+) T cells in lamina propria. This suggests that the GYxxØ motif and S727P each contribute to SIV's targeting to mucosal tissues.

Loss of a tyrosine-dependent trafficking motif in the simian immunodeficiency virus envelope cytoplasmic tail spares mucosal CD4 cells but does not prevent disease progression.

A hallmark of pathogenic simian immunodeficiency virus (SIV) and human immunodeficiency virus (HIV) infections is the rapid and near-complete depletion of mucosal CD4(+) T lymphocytes from the gastrointestinal tract. Loss of these cells and disruption of epithelial barrier function are associated with microbial translocation, which has been proposed to drive chronic systemic immune activation and disease progression. Here, we evaluate in rhesus macaques a novel attenuated variant of pathogenic SIVmac239, termed ΔGY, which contains a deletion of a Tyr and a proximal Gly from a highly conserved YxxØ trafficking motif in the envelope cytoplasmic tail. Compared to SIVmac239, ΔGY established a comparable acute peak of viremia but only transiently infected lamina propria and caused little or no acute depletion of mucosal CD4(+) T cells and no detectable microbial translocation. Nonetheless, these animals developed T-cell activation and declining peripheral blood CD4(+) T cells and ultimately progressed with clinical or pathological features of AIDS. ΔGY-infected animals also showed no infection of macrophages or central nervous system tissues even in late-stage disease. Although the ΔGY mutation persisted, novel mutations evolved, including the formation of new YxxØ motifs in two of four animals. These findings indicate that disruption of this trafficking motif by the ΔGY mutation leads to a striking alteration in anatomic distribution of virus with sparing of lamina propria and a lack of microbial translocation. Because these animals exhibited wild-type levels of acute viremia and immune activation, our findings indicate that these pathological events are dissociable and that immune activation unrelated to gut damage can be sufficient for the development of AIDS.

Interleukin-10 prevents epithelial cell apoptosis by regulating IFNγ and TNFα expression in rhesus macaque colon explants.

Interleukin-10 (IL-10) is an important immunomodulatory cytokine that plays an obligate role in regulating inflammatory responses. Here we demonstrated the role of IL-10 in regulating crypts length and breadth as well as maintaining the survival of epithelial cells using rhesus colon explant cultures. Anti-IL-10 antibody treatment of colon explant cultures induced increased production of inflammatory cytokines/molecules like IFNγ, TNFα, CD107a and perforin as well as increased epithelial cell apoptosis compared to media controls tested. Our results suggest that IL-10 plays a crucial role in maintaining mucosal homeostasis by regulating mucosal IFNγ and TNFα cytokine production.