Evidence of thymic function in heavily antiretroviral-treated human immunodeficiency virus type 1-infected adults with long-term virologic treatment failure

Thymic function was evaluated in 32 heavily antiretroviral-treated human immunodeficiency virus type 1 (HIV-1)-infected adults with long-term virologic treatment failure by measuring thymic volume, by determining the absolute number of naive T cell phenotypes, and by determining the number of cells carrying T cell receptor excision circles (TRECs). There was a significant inverse correlation between age and thymic volume (r=-0.415; P=.018), and there was a significant direct correlation between thymic volume and total naive T cell counts (r=0.529; P=.002), naive CD4(+) cell counts (r=0.437; P=.012), naive CD8(+) cell counts (r=0.467; P=.007), and TREC levels (r=0.391; P=.027). In conclusion, this study found clear evidence that the thymus of heavily antiretroviral-treated HIV-1-infected adults with long-term virologic treatment failure is actively engaged in thymopoiesis, which generates new naive T cells for the peripheral lymphocyte pool.     

HIV-1 infection in children: a clinical and immunologic overview

Globally, HIV-1 is most often transmitted heterosexually so that nearly half of all infected adults are women of child-bearing age. Infants may acquire infection from vertical transmission. Without treatment most HIV-1 infected children in Africa die before their third birthday; as a result child mortality has increased overall by 35-50%, and by greater than 100% in areas of high seroprevalence. HIV-1 infection has a heterogeneous spectrum of clinical course. Compared to HIV-1-infected adults, survival times are considerably shorter for children who acquire the virus perinatally or during infancy. Factors contributing to accelerated disease progression in infants and children are poorly understood but may include relative immunological immaturity, thymic HIV-1-mediated destruction at a time of active thymopoiesis, and HLA class I sharing between mother and infant. This review will initially discuss clinical and biological determinants of mother-to-child transmission and disease progression in HIV-infected infants and children. Our current knowledge of the mechanisms of T cell depletion is summarised and the host immune response to HIV-1 (innate and adaptive) described in the context of Pediatric HIV-1 infection.     

Increased thymic mass and circulating naive CD4 T cells in HIV-1-infected adults treated with growth hormone

OBJECTIVE: To determine whether treatment with growth hormone (GH) enhances thymopoiesis in individuals infected with HIV-1. METHODS: Five HIV-1-infected adults were treated with GH for 6-12 months in a prospective open-label study. Immunological analyses were performed before GH treatment and repeated at 3 month intervals after GH initiation. Thymic mass was analysed using computed tomography with quantitative density and volume analysis. Analysis of circulating lymphocytes, including naive and memory T cell subsets, was performed using multiparameter flow cytometry. RESULTS: GH treatment was associated with a marked increase in thymic mass in all GH recipients. Circulating naive CD4 T cells also increased significantly in all patients during GH therapy, suggesting an enhancement of thymopoiesis. CONCLUSION: GH has significant effects on the human immune system, including the reversal of thymic atrophy in HIV-1-infected adults. De-novo T cell production may thus be inducible in immunodeficient adults.     

The thymus during HIV disease: role in pathogenesis and in immune recovery

The thymus is the primary lymphoid organ supplying new lymphocytes to the periphery. Clinical and morphologic studies of HIV-infected children and adults indicate that the thymus is affected by HIV. Thymic dysfunction and thymic involution occur during HIV disease and have been associated with rapid progression in infants infected perinatally with HIV. In vitro information of thymic organ culture, thymic epithelial cell culture, the SCID-hu mouse system and SHIV infection of primates have supported HIV-induced thymic damage. The mechanisms underlying this could be many, including direct thymocyte killing by the virus, apoptosis, or disruption of thymic stromal architecture. T cell receptor excision circles (TREC) have been developed as a marker of new thymic emigrants. Decreases in TREC concentrations have been found in both HIV-infected pediatric and adult patients. Mathematical models have suggested that thymic infection in children is more severe than in adults, particularly during infection with strains that use CXCR4 as coreceptor. Recent evidence suggests that thymic recovery may be achieved in some patients as a result of potent antiretroviral therapy. Extensive thymic damage may, however, hamper immune reconstitution, particularly in pediatric patients. This review attempts to summarize evidence for thymic involvement during HIV infection in children and in adults, the role of thymic infection in disease progression, and the contribution of the thymus to immune restoration following potent antiviral therapy. Immunologic interventions aiming at restoring thymic function in AIDS patients are also reviewed.     

Differential susceptibility of human thymic dendritic cell subsets to X4 and R5 HIV-1 infection

OBJECTIVES: Human thymus can be infected by HIV-1 with potential consequences on immune regeneration and homeostasis. We previously showed that CD4 thymocytes preferentially replicate CXCR4 tropic (X4) HIV-1 dependently on interleukin (IL)-7. Here we addressed the susceptibility of thymic dendritic cells (DC) to HIV-1 infection. METHODS: We investigated the replication ability of CXCR4 or CCR5 (R5) tropic HIV-1 in thymic micro-explants as well as in isolated thymic CD11clowCD14- DC, CD11chighCD14+ DC and plasmacytoid DC subsets. RESULTS: Thymic tissue was productively infected by both X4 and R5 viruses. However, X4 but not R5 HIV-1 replication was enhanced by IL-7 in thymic micro-explants, suggesting that R5 virus replication occurred in cells other than thymocytes. Indeed, we found that R5 HIV-1 replicated efficiently in DC isolated from thymic tissue. The replicative capacity of X4 and R5 viruses differed according to the different DC subsets. R5 but not X4 HIV-1 efficiently replicated in CD11chighCD14+ DC. In contrast, no HIV-1 replication was detected in CD11clowCD14- DC. Both X4 and R5 viruses efficiently replicated in plasmacytoid DC, which secreted interferon-alpha upon HIV-1 exposure. Productive HIV-1 infection also caused DC loss, consistent with different permissivity of each DC subset. CONCLUSIONS: Thymic DC sustain high levels of HIV-1 replication. DC might thus be the first target for R5 HIV-1 infection of thymus, acting as a Trojan horse for HIV-1 spread to thymocytes. Furthermore, DC death induced by HIV-1 infection may affect thymopoiesis.     

Porcine thymic grafts protect human thymocytes from HIV-1-induced destruction

Human immunodeficiency virus type 1 (HIV-1) infection depletes thymocytes and destroys thymic structure. Functional, tolerant human T cells develop in vivo in immunodeficient mice receiving porcine thymus and human fetal liver fragments under the kidney capsule. In this model, we evaluated the potential of porcine thymus to protect human thymocytes from the effects of HIV-1. Compared with that observed in control mice with human thymic grafts, porcine thymus attenuated human thymocyte depletion by the CCR5-tropic isolate JR-CSF without preventing thymocyte infection. Porcine thymus protected human thymocytes from infection and depletion by a CXCR4-tropic HIV-1 isolate without reducing peripheral blood viral loads or T cell infection. Human thymocytes from human but not porcine grafts showed decreased Bcl-2 expression and increased apoptosis after NL4.3 infection. Thus, porcine thymus protects human thymocytes from the cytopathic effect of HIV-1, suggesting a possible approach to achieving immune restoration in patients with acquired immunodeficiency syndrome who have incomplete responses to antiretroviral therapy. The model allows analysis of the mechanisms of HIV-mediated thymic dysfunction.     

In vitro studies of HIV-1 infection in thymic lymphocytes: a putative role of the thymus in AIDS pathogenesis.

To ascertain whether thymic lymphocytes represent suitable targets for HIV-1 infection, we infected thymic cell suspensions from normal donors with HIV-1 (HTLV-IIIB strain). We found that, in vitro, thymic lymphocytes are readily infected and highly permissive for HIV-1 replication. In addition, immature cells with the CD4+/CD8+ phenotype, most likely the precursors of mature circulating CD4+ and CD8+ lymphocytes, showed a marked susceptibility to viral infection and replication. These findings suggest that thymus infection may play a triggering role in the pathogenesis of AIDS, particularly in pediatric cases, and may partially explain the lack of restoration of peripheral CD4+ lymphocytes killed by HIV-1.     

Thymic volume, T-cell populations, and parameters of thymopoiesis in adolescent and adult survivors of HIV infection acquired in infancy

OBJECTIVES: Antiretroviral therapy has significantly prolonged the lifespan of children who acquire HIV infection in infancy, but the impact of HIV on thymus-mediated maintenance of T lymphocytes has not been studied. To examine the long-term effects of HIV infection in childhood on thymopoiesis, thymic volume and parameters of thymic function from clinically stable adolescents and young adults with HIV infection acquired in infancy were compared with those from uninfected controls. METHODS: Thymic volume was determined using three-dimensional reconstruction and volumetric analysis of non-contrast enhanced computed tomography images of the upper chest. The degree of fat involution was assessed using a semiquantitive scoring system. CD4 and CD8 T cell populations and T cell receptor recombination excision circles (TREC) concentrations in peripheral blood lymphocytes were measured in all subjects. RESULTS: Twenty youths (aged 17.6 +/- 2.5 years) with HIV infection acquired perinatally (n = 18) or by neonatal transfusion (n = 2) were enrolled whose HIV plasma viral load had been undetectable for a median of 3.1 years, along with 18 seronegative healthy young adults (aged 20.6 +/- 1.3 years). HIV infected subjects and controls had indistinguishable CD4 T cell counts, thymus volumes (20.5 versus 15.8 cm), thymic index scores, and TREC values. Thymic volume correlated with the number and percentage of CD4 T lymphocytes in the control group and with the number of TREC in CD4 lymphocytes in the HIV infected group. CONCLUSIONS: Long term survivors of pediatric HIV infection appear to have retained or recovered thymic volume and thymic activity approximating uninfected youths.     

Slow disease progression and robust therapy-mediated CD4+ T-cell recovery are associated with efficient thymopoiesis during HIV-1 infection

In chronic HIV infection, most untreated patients lose naive CD4+ and CD8+ T cells, whereas a minority preserve them despite persistent high viremia. Although antiretroviral therapy (ART)-mediated viral suppression generally results in a rise of naive and total CD4+ T cells, certain patients experience very little or no T-cell reconstitution. High peripheral T-cell activation has been linked to poor clinical outcomes, interfering with previous evaluations of thymic function in disease progression and therapy-mediated T-cell recovery. To circumvent this, we used the sj/betaTREC ratio, a robust index of thymopoiesis that is independent of peripheral T-cell proliferation, to evaluate the thymic contribution to the preservation and restoration of naive CD4+ T cells. We show that the loss of naive and total CD4+ T cells is the result of or is exacerbated by a sustained thymic defect, whereas efficient thymopoiesis supports naive and total CD4+ T-cell maintenance in slow progressor patients. In ART-treated patients, CD4+ T-cell recovery was associated with the normalization of thymopoiesis, whereas the thymic defect persisted in aviremic patients who failed to recover CD4+ T-cell counts. Overall, we demonstrate that efficient thymopoiesis is key in the natural maintenance and in therapy-mediated recovery of naive and total CD4+ T cells.     

Modelling thymic HIV-1 Nef effects

The nef gene is conserved among primate lentiviruses and is one of the first viral genes that is transcribed following infection. This suggests a critical role for Nef in the virus life cycle and in the pathogenesis of lentiviral infections. In vitro, several functions have been described, including down regulation of CD4 and MHC class I surface expression, altered T-cell signaling and activation, and enhanced viral infectivity. However, the impact of these individual functions on viral pathogenicity in general, and thymic T cell production in particular, remains elusive. Here, we review the observations from experimental models that have been used to study the pathogenic effect of HIV-1 Nef on the thymus. These in vitro and in vivo studies have led to a better understanding of Nef's mechanism of action, although there still exists discord as to the contribution of Nef-mediated disturbance of thymopoiesis in the pathogenesis of AIDS.     

HIV-1 replication and pathogenesis in the human thymus

How HIV replicates and causes destruction of the thymus, and how to restore thymic function, are among the most important questions of HIV-1 pathogenesis and therapy in adult as well as pediatric patients. The thymus appears to function, albeit at reduced levels, throughout the life of adults, to respond to T cell depletion induced by HIV and to be suppressed by HIV. In this review, we summarize recent findings concerning HIV replication and pathogenesis in the human thymus, focusing on mechanistic insights gleaned from studies in the SCID-hu Thy/Liv mouse and human fetal-thymus organ culture (HF-TOC) models. First, we discuss HIV viral determinants and host factors involved in the replication of HIV in the thymus. Second, we consider evidence that both viral factors and host factors contribute to HIV-induced thymocyte depletion. We thus propose that multiple mechanisms, including depletion and suppression of progenitor cells, paracrine and direct lytic depletion of thymocytes, and altered thymocyte selection are involved in HIV-induced pathology in the thymus. With the SCID-hu Thy/Liv mouse and HF-TOC models, it will be important in the coming years to further clarify the virological, cell biological, and immunological mechanisms of HIV replication and pathogenesis in human thymus, and to correlate their significance in HIV disease progression.     

Viral phenotype affects the thymic production of new T cells in HIV-1-infected children

OBJECTIVE: To determine whether viral phenotype has any effect on thymic production of new T cells in HIV-1-infected children. DESIGN: Differences in CD4+ T-cell counts and a marker of thymic output [T-cell antigen receptor (TCR) rearrangement excision circles (TRECs)], between HIV-1-infected children with non-syncytium-inducing (NSI) and syncytium-inducing (SI) viral strains were determined. PATIENTS AND METHODS: A cross-sectional study in 90 samples from vertically HIV-1-infected-children (median age 4.9 years) treated with combination therapy, and a longitudinal study in three children that underwent a change from NSI to SI phenotype were carried out. Viral load, viral phenotype, CD4+ T-cell counts, and quantification of TRECs values were determined. RESULTS: Children with SI virus showed significant lower levels of CD4+ T cells and a lower thymic production of new T cells than children with NSI. These reductions were independent of the treatment and the age of the children. However, there were no differences in viral load with the phenotype between those groups. In children with both NSI and SI viral phenotype, there was a significant correlation between CD4+ T-cell counts and TRECs values. CONCLUSION: The decrease of CD4+ T cells in presence of T-tropic viruses would be mainly due to a lower production of new CD4+ T cells as consequence of the inhibitory effect of these T-tropic strains on thymic function. This effect is not due either to the amount of circulating virus or to the replication kinetics of those strains, but rather depends on the ability of T-tropic viruses to infect T-cell precursors using CXCR4 receptors, which are highly expressed in immature thymocytes.     

CD4 T cell surface CCR5 density as a host factor in HIV-1 disease progression

OBJECTIVE AND DESIGN: We have recently shown that the number of CCR5 molecules at the surface of peripheral blood CD4 T cells (CCR5 density) correlates with the viral RNA plasma level in HIV-1-infected individuals. As viral load is a strong predictor of outcome in HIV infection, the present study examines the correlation between CCR5 density and HIV-1 disease progression. METHODS: Using a quantitative flow cytometry assay, we measured CCR5 density in HIV-1-infected adults and control healthy volunteers. The CCR5 genotype (presence of a Delta 32 allele) was also determined. RESULTS: CCR5 density was stable over time on non-activated, HLA-DR(-)CD4 T cells of infected individuals. In a study cohort of 25 patients, asymptomatic and non-treated, we observed a correlation between CCR5 density on HLA-DR(-)CD4 T cells and the CD4 T cell slope (P = 0.026), which was independent of the presence or absence of the Delta 32CCR5 deletion. In particular, slow progressors expressed lower CCR5 densities than non-slow progressors (P = 0.004) and non-infected control subjects (P = 0.002). CONCLUSION: These results are compatible with the hypothesis that CCR5 density, which is a key factor of HIV-1 infectability, determines in-vivo HIV production, and thereby the rate of CD4 cell decline. Consequently, CCR5 density quantitation could be a new valuable prognostic tool in HIV-1 infection. Moreover, these data emphasize the therapeutic potential of treatments that reduce functional CCR5 density.     

Tropism-restricted neutralization by secretory IgA from parotid saliva of HIV type 1-infected individuals

In this study we examined secretory IgA, isolated from the parotid saliva of 10 HIV-1-infected subjects, for its ability to influence HIV-1 infection of peripheral blood mononuclear cells with two R5 and two X4 primary isolates. Salivary IgA from four subjects was found to inhibit both R5 viruses but not the X4 viruses. In another subject, salivary IgA inhibited both X4 viruses but not the R5 viruses. The specificity of these antibodies seemed to be directed against, but not restricted to, gp160 and gp120. Compared with subjects whose salivary IgA did not inhibit HIV-1 infection, subjects who displayed neutralizing activity were in relatively early stages of disease and had CD4(+) T cell counts greater than 200 cells/microl. Our data indicate the presence of tropism-specific (more frequently R5-specific) neutralizing antibodies in HIV-1-infected subjects. Because mucosal transmission of HIV-1 occurs exclusively in R5 viruses, and X4 viruses often emerge in established infection and account for viral persistence later in disease, our data suggest a potential role for secretory IgA in preventing viral transmission, but a less likely effect on chronic infection.     

Human papillomavirus type 16 intratypic variant infection and risk for cervical neoplasia in southern China

A mutation of the stromal cell-derived factor 1 gene (SDF-1 3'A) was shown to protect adults exposed to human immunodeficiency virus type 1 (HIV-1) from infection and to affect HIV disease progression in adults. The presence of this mutation in HIV-1-infected Kenyan children did not predict mother-to-child virus transmission. The SDF-1 3'A polymorphism was studied in 256 HIV-1-infected, 118 HIV-1-exposed but uninfected, and 170 unexposed and uninfected children of Italian origin, and the frequency of SDF-1 3'A heterozygosity and homozygosity in each of the 3 groups was similar. Of the 256 HIV-1-infected children, 194 were regularly followed up and were assigned to groups according to disease progression. The frequency of the SDF-1 3'A allele was substantially lower among children with long-term nonprogression than among children with rapid (P =.0329) or delayed (P =.0375) progression. We show that the presence of the SDF-1 3'A gene correlates with accelerated disease progression in HIV-1-infected children born to seropositive mothers but does not protect against mother-to-child HIV-1 transmission.     

De novo T-cell generation in patients at different ages and stages of HIV-1 disease

We assessed de novo T-cell generation by determining T-cell receptor-rearrangement excision circles (TRECs) based on patient age and on stage of HIV-1 infection. TRECs were measured in purified CD4 and CD8 T cells of a large cohort of HIV-1-infected subjects (n = 297) with chronic infection but no previous antiretroviral treatment and of a control group of HIV-negative subjects (n = 120). HIV-1-infected subjects were stratified on the basis of CD4 T-cell counts in 3 groups, early-stage disease (more than 500 CD4 T cells), intermediate-stage disease (200-500 CD4 T cells), and late-stage disease (fewer than 200 CD4 T cells). Compared with the control group, CD8 TREC contents were severely reduced (P <.001) in HIV-1-infected subjects regardless of the stage of HIV disease. In contrast, CD4 TREC contents were significantly increased (P =.003) in HIV-1-infected subjects during early-stage disease, similar at intermediate-stage disease, and severely reduced only at late-stage disease. We show that the increase in CD4 TRECs was mostly limited to younger (younger than 45 years) patients at early-stage disease. Our results demonstrate a dichotomy between TREC contents in CD4 and CD8 T-cell populations in HIV-1 infection and indicate that thymus function in younger subjects is preserved at early and intermediate stages of HIV infection.     

Causal relationships between HIV-1 coreceptor utilization, tropism, and pathogenesis in human thymus

The pathogenic differences between CXCR4 (X4)- and CCR5 (R5)-utilizing strains of HIV-1 may be predominantly due to differences in viral tropism, which in turn may be due to differential coreceptor utilization. We tested this hypothesis in the human thymus organ of the SCID-hu Thy/Liv mouse, using recombinants of NL4-3 that were isogenic except for Env coreceptor-binding determinants of the V1-V3 loops. Conversion of NL4-3 from an X4 to an R5 isolate was associated with altered tropism for cell subpopulations within the Thy/Liv organ (with a higher frequency of infection of thymic stromal cells, including macrophages), a slower rate of replication, and a lower level of cytopathicity. These observations underscore the causal relationships between tropism, coreceptor use, and cytopathicity in the human thymus in vivo.     

Gut mucosal T cell responses and gene expression correlate with protection against disease in long-term HIV-1-infected nonprogressors

Limited information is available on the molecular mechanisms by which long-term HIV-1-infected nonprogressors suppress HIV-1 infection and maintain immune functions. The intestinal mucosal immune system is an early target for HIV-1 infection and severe CD4+ T cell depletion. We evaluated mucosal T lymphocyte subsets, virus-specific cellular responses, gene expression profiles, and viral loads in intestinal mucosal biopsies of long-term nonprogressor (LTNP) patients as compared to chronically HIV-1-infected patients with high viral loads (HVLs) and CD4+ T cell loss, as well as HIV-seronegative healthy individuals. This study aims to identify the mucosal correlates of HIV disease progression and to determine the molecular changes associated with immune and intestinal dysfunction. LTNP patients had undetectable viral loads, normal CD4+ T cell levels, and virus-specific cellular responses in peripheral blood and mucosal compartments. Microarray analysis revealed a significant increase in gene expression regulating immune activation, cell trafficking, and inflammatory response in intestinal mucosa of HVL patients as compared to LTNP patients. Genes associated with cell cycle regulation, lipid metabolism, and epithelial cell barrier and digestive functions were down-regulated in both HVL and LTNP patients. This may adversely influence nutrient adsorption and digestive functions, with the potential to impact the efficacy of antiretroviral therapy. We demonstrate that the maintenance of mucosal T cells, virus-specific responses, and distinct gene expression profiles correlate with clinical outcome in LTNP patients. However, the intestinal mucosal immune system remains an important target of HIV-1 infection in LTNP, and these effects may ultimately contribute toward disease progression.     

Fetal human immunodeficiency virus type 1 infection of different organs in the second trimester

Human immunodeficiency virus type 1 (HIV-1) infection in utero was examined by isolating the virus and detecting the HIV-1 DNA sequence from different fetal tissues. The brain, thymus, lung, liver, spleen, and placenta tissues from fetuses (10-23 weeks of gestation) born to HIV-1-infected asymptomatic mothers were examined. HIV-1 was isolated from 2 of 7, 1 of 7, and 1 of 7 cocultures of splenic, thymic, and trypsin-resistant cells from the liver and placenta, respectively, with peripheral blood mononuclear cells; 20-30% and 40-60% of splenic and of thymic cells were CD4+ lymphoid cells and 40-80% of trypsin-resistant cells were mononuclear phagocytes. The HIV-1 DNA sequence was detected in 4 of 7, 3 of 7, 1 of 7, 1 of 7, 2 of 7, and 2 of 6 samples from the spleen, thymus, brain, lung, liver, and placenta, respectively, using the polymerase chain reaction. In one case, the intensity of the HIV-1 DNA sequence appeared to be correlated with the success of viral isolation. We indicate that fetal HIV-1 infection may frequently occur in the second trimester and the cells responsible for the infection may be CD4+ lymphoid cells and mononuclear phagocytes.     

Interferon gamma and interleukin 6 modulate the susceptibility of macrophages to human immunodeficiency virus type 1 infection

The effect of interferon gamma (IFN-gamma) and interleukin 6 (IL-6) on infection of macrophages with human immunodeficiency virus type 1 (HIV-1) was investigated. By using a polymerase chain reaction-based viral entry assay and viral infectivity assay, it was demonstrated that IL-6 and IFN-gamma augmented susceptibility of monocyte-derived macrophages (MDMs) to infection with T-cell tropic CXCR4-utilizing (X4) HIV-1 strains. Consistent with this finding, IFN-gamma and IL-6 augmented fusion of MDMs with T-tropic envelope-expressing cells. The enhanced fusion of cytokine-treated MDMs with T-tropic envelopes was inhibited by the CXCR4 ligand, SDF-1, and by T22 peptide. IFN-gamma and IL-6 did not affect expression of surface CXCR4 or SDF-1-induced Ca(++) flux in MDMs. In contrast to the effect of IFN-gamma on the infection of MDMs with X4 strains, IFN-gamma inhibited viral entry and productive infection of MDMs with macrophage-tropic (M-tropic) HIV-1. Consistent with this finding, IFN-gamma induced a decrease in fusion with M-tropic envelopes that correlated with a modest reduction in surface CCR5 and CD4 on MDMs. It was further demonstrated that macrophage inflammatory protein (MIP)-1alpha and MIP-beta secreted by cytokine-treated MDMs augmented their fusion with T-tropic-expressing cells and inhibited their fusion with M-tropic envelope-expressing cells. These data indicate that proinflammatory cytokines, which are produced during opportunistic infections or sexually transmitted diseases, may predispose macrophages to infection with X4 strains that, in turn, could accelerate disease progression.