Human Vgamma2Vdelta2 T cells contain cytoplasmic RANTES.

The adult human Vgamma2Vdelta2 T cell repertoire is a product of chronic selection in the periphery. Endogenous antigens drive the expansion of cells expressing the Vgamma2Vdelta2 TCR. Thus, we would expect the majority of circulating Vgamma2Vdelta2 T cells to be antigen experienced and to have memory phenotype, in contrast to the alpha/beta TCR+ subsets that include a substantial fraction of naive cells. We sought to characterize functional aspects of Vgamma2Vdelta2 T cells that might show whether circulating cells are memory or naive. For these studies, we focus on the expression of the CC chemokine regulated upon activation normal T cell expressed and secreted (RANTES). In naive alphabeta T cells, an initial stimulus triggers the onset of RANTES transcription followed later by protein expression. In memory CD8+ alphabeta T cells, RANTES mRNA is already present in unstimulated cells and protein expression is triggered immediately by TCR signaling; some cells may also contain RANTES protein in cytoplasmic stores. We show here that the vast majority of circulating human T cells contain RANTES protein in cytoplasmic stores and the chemokine is secreted rapidly after TCR signaling. Primary Vgamma2Vdelta2 T cell lines obtained after in vitro stimulation with phosphoantigens behaved similarly to circulating Vgamma2Vdelta2 T cells and contained both RANTES mRNA and protein, but only very low levels of mRNA or protein for macrophage inflammatory protein (MIP)-1alpha or MIP-1beta. The presence of stored RANTES shows that circulating Vgamma2Vdelta2 T cells are mostly memory phenotype and capable of rapid chemokine responses to phosphoantigen stimulation. Considering that one of 40 circulating CD3+ lymphocytes is Vgamma2Vdelta2+, they comprise the largest circulating memory population against a single antigen, and phosphoantigen stimulation will trigger a rapid activation with immediate release of RANTES.

Innate T-cell immunity in HIV infection: the role of Vgamma9Vdelta2 T lymphocytes.

There is growing interest in the use of innate immune reactions in the therapy and prophylaxis of various diseases. Natural T (NT) lymphocytes that recognize infected cells or microbial compounds without the classical genetic restriction by polymorphic MHC molecules are crucial components of innate immunity. NT cells bearing the Vgamma9Vdelta2 T-cell receptor (TCR) are broadly reactive against intracellular pathogens, can lyse human immunodeficiency virus (HIV) infected cells, and release cytokines capable of regulating HIV replication. The potent antiviral activities of Vgamma9Vdelta2 T cells may help to contain viral spread during acute HIV infection and/or to prevent the establishment of viral persistence. Substantial changes in the composition and function of circulating gammadelta T-cell pools occur in HIV-infected patients. These changes a) may contribute to the etiopathogenesis of opportunistic infections and neoplasms, and b) are partly reversed by highly active anti-retroviral therapy (HAART). In addition to direct antiviral activities, activated gammadelta T cells influence dendritic cell maturation and the adaptive alphabeta T-cell response. Vgamma9Vdelta2 T cells can be stimulated in vivo and in vitro by various nonpeptidic antigens (NpAgs) and recent animal experimental data suggest that activated Vgamma9Vdelta2 T cells may help to control SIV replication. Currently, NpAgs are being assessed as potential therapeutic agents in AIDS, tuberculosis and certain cancers susceptible to Vgamma9Vdelta2 T-cell effector mechanisms.

Macaque models for early immunosuppression and T cell loss during acute immunodeficiency virus infections.

The interval of acute infection with immunodeficiency viruses is critically important for determining the long-term rate of disease progression. The steps of initial infection, systemic dissemination, and explosive replication of pathogenic SIV or SHIV in macaques are being mapped to show the mechanisms responsible for remodeling host immunity, for establishing the persistent infection, and for promoting disease progression. Here, we describe recent studies on two ways in which CD4+ T cell populations are depleted during acute infection. Initially, we discuss recent work on the mechanisms for CD4+ T cell-mediated, MHC-unrestricted cytolysis. This mechanism shows how even soluble viral antigens such as the envelope glycoprotein, can prime CD4+ lymphocytes to be both effector and target cells in an unrestricted cytolysis mechanism. The consequence of unrestricted cytolysis is a more rapid destruction of the CD4+ T cell population. Secondly, we discuss the broader issue of T cell hyperactivation during acute infection. Inappropriate activation of this lymphocyte population renders cells susceptible to activation induced cell death and also increases the rate of virus replication. Macaque immunization studies have shown a clear role for extracellular Tat in hyperactivation. These two mechanisms, unrestricted cytolysis and T cell hyperactivation, are components of the acute infection that remodel host immunity and dictate the rate of progression to AIDS.

Staphylococcal superantigens induce lymphotactin production by human CD4+ and CD8+ T cells.

Lymphotactin is a potent chemotactic cytokine (chemokine) that is produced by and also attracts T and natural killer (NK) cells. We are studying whether chemokines that affect mainly T cells might also regulate immune responses by preferentially recruiting individual subsets or by affecting cytokine or other chemokine responses. In order to pursue these questions, we need to learn more about the mechanisms regulating lymphotactin production and the cell types capable of releasing this factor. We used new monoclonal antibodies against human lymphotactin to develop a sensitive antigen-capture enzyme linked immunoabsorbent assay (ELISA) that measures chemokine levels in culture fluids. Using this capture ELISA, we showed that lymphotactin could be produced by CD4+ and CD8+ T cells, but only after T cell-receptor-dependent stimulation using bacterial superantigens and not after treatment by inflammatory cytokines or lipopolysaccharide (LPS). Our data show that lymphotactin production responds mainly to T cell-receptor signals in CD4+ and CD8+ T cells, and suggests a mechanism whereby this chemokine could help to regulate T cell immune responses.

Monomeric solution structure of the prototypical 'C' chemokine lymphotactin.

Lymphotactin, the sole identified member of the C class of chemokines, specifically attracts T lymphocytes and natural killer cells. This 93-residue protein lacks 2 of the 4 conserved cysteine residues characteristic of the other 3 classes of chemokines and possesses an extended carboxyl terminus, which is required for chemotactic activity. We have determined the three-dimensional solution structure of recombinant human lymphotactin by NMR spectroscopy. Under the conditions used for the structure determination, lymphotactin was predominantly monomeric; however, pulsed field gradient NMR self-diffusion measurements and analytical ultracentrifugation revealed evidence of dimer formation. Sequence-specific chemical shift assignments were determined through analysis of two- and three-dimensional NMR spectra of (15)N- and (13)C/(15)N-enriched protein samples. Input for the torsion angle dynamics calculations used in determining the structure included 1258 unique NOE-derived distance constraints and 60 dihedral angle constraints obtained from chemical-shift-based searching of a protein conformational database. The ensemble of 20 structures chosen to represent the structure had backbone and heavy atom rms deviations of 0.46 +/- 0.11 and 1.02 +/- 0.14 A, respectively. The results revealed that human lymphotactin adopts the conserved chemokine fold, which is characterized by a three-stranded antiparallel beta-sheet and a C-terminal alpha-helix. Two regions are dynamically disordered as evidenced by (1)H and (13)C chemical shifts and [(15)N]-(1)H NOEs: residues 1-9 of the amino terminus and residues 69-93 of the C-terminal extension. A functional role for the C-terminal extension, which is unique to lymphotactin, remains to be elucidated.

Down-regulation of CXCR1 and CXCR2 expression on human neutrophils upon activation of whole blood by S. aureus is mediated by TNF-alpha.

It was suggested that bacterial products can inhibit the expression of leucocyte chemokine receptors during sepsis and affect leucocyte functions in septic syndrome. Superantigens and toxins produced by Staphylococcus aureus are capable of activating leucocytes via binding to MHC-II antigens on monocytes and T-cell receptor molecules on T lymphocytes. It was recently shown that staphylococcal enterotoxins directly down-regulate the expression of CC chemokine receptors on monocytes through binding to MHC class II molecules. We studied the effects of killed S. aureus on the expression of interleukin-8 receptors, CXCR1 and CXCR2, on polymorphonuclear leucocytes (PMN), which are known to lack the expression of MHC-II antigens. It was shown that S. aureus down-regulated the cell-surface expression of CXCR1 and CXCR2 on PMN in the whole blood and total blood leucocyte fraction containing PMN and monocytes, but did not modulate IL-8 receptor expression in purified PMN suspension. Antibody to TNF-alpha abrogated down-regulation of IL-8 receptors induced by S. aureus. In contrast, LPS reduced CXCR1 and CXCR2 expression in purified PMN and whole blood in a TNF-alpha-independent manner. We further showed that TNF-alpha-induced decrease of CXCR1 and CXCR2 expression was associated with lower IL-8 binding and lower CXCR1 and CXCR2 mRNA levels, and was abrogated by protease inhibitors. We suggest that during septicemia, S. aureus may inhibit neutrophil responsiveness to IL-8 and other CXC chemokines via TNF-alpha- mediated down-regulation of CXCR1 and CXCR2.

In vitro stimulation with a non-peptidic alkylphosphate expands cells expressing Vgamma2-Jgamma1.2/Vdelta2 T-cell receptors.

The majority of peripheral blood gammadelta T cells in healthy adult humans express the Vgamma2/Vdelta2 T-cell receptor (TCR) and generate TCR-mediated, major histocompatibility complex (MHC)-unrestricted proliferative responses to low molecular weight alkylphosphates. Vgamma2/Vdelta2 populations after antigen proliferation maintained diversity in the CDR3s of Vgamma2 mRNA, indicating that the response was polyclonal or oligoclonal, and were enriched for Vgamma2 TCR chains containing the Jgamma1.2 segment. Alkylphosphate stimulation further skewed an already biased peripheral blood gammadelta T-cell population and increased the abundance of Vgamma2-Jgamma1.2/Vdelta2 T cell receptors, suggesting similarities between the alkylphosphate response and peripheral selection mechanisms shaping this repertoire in human beings.

Blood-brain barrier disruption in simian immunodeficiency virus encephalitis.

Infected monocyte-derived macrophages (MDM) are thought by some investigators to play a central role in the neuropathogenesis of human immunodeficiency virus encephalitis (HIVE). It was recently proposed that these cells gain access to the central nervous system (CNS) through disruptions in blood-brain barrier (BBB) tight junctions, which occur in HIVE in association with accumulation of activated, HIV-1-infected, perivascular macrophages and serum protein extravasation (Am J Pathol 1999, 155: 1915-27). The present study tested this hypothesis in basal ganglia tissue from simian immunodeficiency virus (SIV)-infected macaques with encephalitis by examining vessels for immunohistochemical alterations in the tight junction-associated proteins, occludin and zonula occludens-1 (ZO-1). Compared to non-infected macaques and SIV-infected macaques without encephalitis, cerebral vessels from macaques with SIVE showed fragmentation and decreased immunoreactivity for both tight junction proteins. These alterations were associated with accumulation of perivascular macrophages and aberrant occludin and ZO-1 immunoreactivity within these cells. In addition, perivascular extravasation of fibrinogen, a plasma protein, and a change from a strong linear staining pattern to a more irregular pattern of glucose transporter isoform-1 (GLUT-1), a metabolic BBB marker, were observed in regions with vascular tight junction protein alterations. These findings demonstrate that tight junction disruption occurs in SIVE in association with perivascular macrophage accumulation. While it cannot be ascertained from these studies whether such changes precede macrophage infiltration, or are secondary to the chronic presence of macrophages around cerebral vessels, disruptions in BBB integrity could serve as portals for additional accumulation of perivascular macrophages in SIVE.

Whole body positron emission tomography imaging of activated lymphoid tissues during acute simian-human immunodeficiency virus 89.6PD infection in rhesus macaques.

Mechanisms of acute retroviral pathogenesis have been examined during primary infection of rhesus macaques with simian-human immunodeficiency virus 89.6PD (SHIV(89.6PD)). During acute infection, between initial exposure and establishment of antigen-specific immune responses that stabilize the virus burden, rapid immune system changes influence the viral set-point and dictate subsequent steps in disease progression. In a previous study, we described specific patterns of lymphocyte activation during acute SHIV(89.6PD) infection. We now extend these studies to describe lymphoid tissue activation, using whole body positron emission tomography (PET) and the radioactive tracer 2-[(18)F]fluorodeoxyglucose (FDG). Within a few days after primary infection by intravenous, intrarectal, or intravaginal routes, PET-FDG imaging revealed a distinct pattern of lymphoid tissue activation centered on axillary, cervical, and mediastinum lymph nodes. Increased tissue FDG uptake preceded fulminant virus replication at these sites, suggesting that a diffusible factor of host or viral origin was responsible for lymphoid tissue changes. These data show that activation of lymphoid tissues in the upper body is an early response to virus infection and that diffusible mediators of activation might be important targets for vaccine or therapeutic intervention strategies.

Definition of five new simian immunodeficiency virus cytotoxic T-lymphocyte epitopes and their restricting major histocompatibility complex class I molecules: evidence for an influence on disease progression.

Simian immunodeficiency virus (SIV) infection of the rhesus macaque is currently the best animal model for AIDS vaccine development. One limitation of this model, however, has been the small number of cytotoxic T-lymphocyte (CTL) epitopes and restricting major histocompatibility complex (MHC) class I molecules available for investigating virus-specific CTL responses. To identify new MHC class I-restricted CTL epitopes, we infected five members of a family of MHC-defined rhesus macaques intravenously with SIV. Five new CTL epitopes bound by four different MHC class I molecules were defined. These included two Env epitopes bound by Mamu-A*11 and -B*03 and three Nef epitopes bound by Mamu-B*03, -B*04, and -B*17. All four restricting MHC class I molecules were encoded on only two haplotypes (b or c). Interestingly, resistance to disease progression within this family appeared to be associated with the inheritance of one or both of these MHC class I haplotypes. Two individuals that inherited haplotypes b and c separately survived for 299 and 511 days, respectively, while another individual that inherited both haplotypes survived for 889 days. In contrast, two MHC class I-identical individuals that did not inherit either haplotype rapidly progressed to disease (survived <80 days). Since all five offspring were identical at their Mamu-DRB loci, MHC class II differences are unlikely to account for their patterns of disease progression. These results double the number of SIV CTL epitopes defined in rhesus macaques and provide evidence that allelic differences at the MHC class I loci may influence rates of disease progression among AIDS virus-infected individuals.

Importance of the CD3 marker for evaluating changes in rhesus macaque CD4/CD8 T-cell ratios.

BACKGROUND: Until recently, there were no CD3 antibodies that crossreacted with rhesus macaque T cells. Consequently, studies relying on CD8 counts or CD4/CD8 ratios enumerated this subpopulation on the basis of CD8+ or CD8bright+ staining. We used a rhesus-specific, anti-CD3 antibody to better define the CD8+ T-cell population, and to show the effects of better measurements on CD4/CD8 ratios and changes in T cells as macaques age. METHODS: We used three-color flow cytometry to measure CD4 and CD8 populations with and without CD3 costaining. Venous blood samples were obtained from 52 colony-bred macaques between 2 months and 9 years of age. RESULTS: The CD8+ T cells defined by CD3 and CD8 double staining were approximately 60% of all cells that were stained by CD8 alone. Improved detection of this lymphocyte subset showed that CD4/CD8 ratios were close to the range of 1.5-2.0. Declining CD4/CD8 ratios during aging are predominantly due to decreasing CD4+ T-cell counts. CONCLUSIONS: Better quantitation of the CD8+ T-cell population showed that the CD4/CD8 ratio was not inverted as had been reported, but is actually very similar to the values observed in human beings. Although the two species differ in the pattern of CD8 expression, the general immune system characteristics are very similar.

Vaccination with tat toxoid attenuates disease in simian/HIV-challenged macaques.

The Tat protein is essential for HIV type 1 (HIV-1) replication and may be an important virulence factor in vivo. We studied the role of Tat in viral pathogenesis by immunizing rhesus macaques with chemically inactivated Tat toxoid and challenging these animals by intrarectal inoculation with the simian/human immunodeficiency virus 89.6PD. Immune animals had significantly attenuated disease with lowered viral RNA, interferon-alpha, and chemokine receptor expression (CXCR4 and CCR5) on CD4(+) T cells; these features of infection have been linked to in vitro effects of Tat and respond similarly to extracellular Tat protein produced during infection. Immunization with Tat toxoid inhibits key steps in viral pathogenesis and should be included in therapeutic or preventive HIV-1 vaccines.

Murine immune responses to mucosally delivered Salmonella expressing Lassa fever virus nucleoprotein.

Arenaviruses are emerging pathogens known to infect via the mucosa, however no formal attempts to make mucosal vaccines have been undertaken. Here we describe a recombinant aroA attenuated Salmonella typhimurium that expresses the nucleoprotein (NP) gene of Lassa fever virus (LAS). The complete NP gene was cloned downstream of the bacterial groEL promotor and integrated into the aroA locus of S. typhimurium. Lassa NP protein was detected in whole cell extracts from the recombinant Salmonella by immunoblot analysis with serum from Lassa-infected people. Mice were inoculated by intragastric intubation with 5 x 10(9) S. typhimurium and boosted with the same recombinant Salmonella 21 days after the primary inoculation. Both local mucosal IgA and serum immunoglobulins against Lassa NP were observed. Splenic cytotoxic T-lymphocyte responses to LAS NP were detected after the boost and they cross-reacted with target cells infected with the related arenavirus, lymphocytic choriomeningitis virus. Recombinant Salmonella elicits humoral and cell mediated immune responses against Lassa fever virus in mice and should be considered as a potential vaccine strategy in man.

Gammadelta T cell receptor repertoire in blood and colonic mucosa of rhesus macaques.

Although their precise roles are not well defined, gammadelta T lymphocytes are recognized as regular components of immune responses. These cells express a limited T cell receptor repertoire and they can be stimulated by soluble ligands without conventional processing and presentation by major histocompatibility antigens. Progress in this area has been limited by the substantial differences between murine and human gammadelta T cells and the lack of knowledge about these cells in nonhuman primates. We used molecular analysis of T cell receptor diversity to characterize gammadelta T cell populations from peripheral blood and colon of rhesus macaques (Macaca mulatta). The gammadelta T cell receptor diversity was limited and distinct for these tissue compartments, particularly in the TCRGV2 family. Furthermore, the TCRDV1 + subset of peripheral blood gammadelta T cells showed signs of progressive oligoclonalization as a function of age. Similar observations have been reported for human tissue samples and our results validate rhesus macaques as an appropriate animal model for studying primate gammadelta T cell populations.

Characterization of primary isolate-like variants of simian-human immunodeficiency virus.

Several different strains of simian-human immunodeficiency virus (SHIV) that contain the envelope glycoproteins of either T-cell-line-adapted (TCLA) strains or primary isolates of human immunodeficiency virus type 1 (HIV-1) are now available. One of the advantages of these chimeric viruses is their application to studies of HIV-1-specific neutralizing antibodies in preclinical AIDS vaccine studies in nonhuman primates. In this regard, an important consideration is the spectrum of antigenic properties exhibited by the different envelope glycoproteins used for SHIV construction. The antigenic properties of six SHIV variants were characterized here in neutralization assays with recombinant soluble CD4 (rsCD4), monoclonal antibodies, and serum samples from SHIV-infected macaques and HIV-1-infected individuals. Neutralization of SHIV variants HXBc2, KU2, 89.6, and 89.6P by autologous and heterologous sera from SHIV-infected macaques was restricted to an extent that these viruses may be considered heterologous to one another in their major neutralization determinants. Little or no variation was seen in the neutralization determinants on SHIV variants 89.6P, 89.6PD, and SHIV-KB9. Neutralization of SHIV HXBc2 by sera from HXBc2-infected macaques could be blocked with autologous V3-loop peptide; this was less true in the case of SHIV 89.6 and sera from SHIV 89.6-infected macaques. The poorly immunogenic but highly conserved epitope for monoclonal antibody IgG1b12 was a target for neutralization on SHIV variants HXBc2, KU2, and 89.6 but not on 89.6P and KB9. The 2G12 epitope was a target for neutralization on all five SHIV variants. SHIV variants KU2, 89.6, 89.6P, 89.6PD, and KB9 exhibited antigenic properties characteristic of primary isolates by being relatively insensitive to neutralization in peripheral blood mononuclear cells with serum samples from HIV-1-infected individuals and 12-fold to 38-fold less sensitive to inhibition with recombinant soluble CD4 than TCLA strains of HIV-1. The utility of nonhuman primate models in AIDS vaccine development is strengthened by the availability of SHIV variants that are heterologous in their neutralization determinants and exhibit antigenic properties shared with primary isolates.

Lymphocyte activation during acute simian/human immunodeficiency virus SHIV(89.6PD) infection in macaques.

Host-virus interactions control disease progression in human immunodeficiency virus-infected human beings and in nonhuman primates infected with simian or simian/human immunodeficiency viruses (SHIV). These interactions evolve rapidly during acute infection and are key to the mechanisms of viral persistence and AIDS. SHIV(89.6PD) infection in rhesus macaques can deplete CD4(+) T cells from the peripheral blood, spleen, and lymph nodes within 2 weeks after exposure and is a model for virulent, acute infection. Lymphocytes isolated from blood and tissues during the interval of acute SHIV(89.6PD) infection have lost the capacity to proliferate in response to phytohemagglutinin (PHA). T-cell unresponsiveness to mitogen occurred within 1 week after mucosal inoculation yet prior to massive CD4(+) T-cell depletion and extensive virus dissemination. The lack of mitogen response was due to apoptosis in vitro, and increased activation marker expression on circulating T cells in vivo coincided with the appearance of PHA-induced apoptosis in vitro. Inappropriately high immune stimulation associated with rapid loss of mature CD4(+) T cells suggested that activation-induced cell death is a mechanism for helper T-cell depletion in the brief period before widespread virus dissemination. Elevated levels of lymphocyte activation likely enhance SHIV(89.6PD) replication, thus increasing the loss of CD4(+) T cells and diminishing the levels of virus-specific immunity that remain after acute infection. The level of surviving immunity may dictate the capacity to control virus replication and disease progression. We describe this level of immune competence as the host set point to show its pivotal role in AIDS pathogenesis.

Virus-specific cytotoxic T-lymphocyte responses select for amino-acid variation in simian immunodeficiency virus Env and Nef.

Cytotoxic T-lymphocyte (CTL) responses to human immunodeficiency virus arise early after infection, but ultimately fail to prevent progression to AIDS. Human immunodeficiency virus may evade the CTL response by accumulating amino-acid replacements within CTL epitopes. We studied 10 CTL epitopes during the course of simian immunodeficiency virus disease progression in three related macaques. All 10 of these CTL epitopes accumulated amino-acid replacements and showed evidence of positive selection by the time the macaques died. Many of the amino-acid replacements in these epitopes reduced or eliminated major histocompatibility complex class I binding and/or CTL recognition. These findings strongly support the CTL 'escape' hypothesis.

High major histocompatibility complex-unrestricted lysis of simian immunodeficiency virus envelope-expressing cells predisposes macaques to rapid AIDS progression.

Before the development of virus-specific immune responses, peripheral blood mononuclear cells (PBMC) from uninfected rhesus monkeys and human beings have the capacity to lyse target cells expressing simian immunodeficiency virus (SIV) or human immunodeficiency virus-1 (HIV) envelope (gp130 and gp120) antigens. Lysis by naive effector cells does not require major histocompatibility complex (MHC)-restricted antigen presentation, is equally effective for allogeneic and xenogeneic targets, and is designated MHC-unrestricted (UR) lysis. UR lysis is not sensitive to EGTA and does not require de novo RNA or protein synthesis. Several kinds of envelope-expressing targets, including cells that poorly express MHC class I antigens, can be lysed. CD4(+) effectors are responsible for most of the lytic activity. High lysis is correlated with high expression of HIV or SIV envelope, specifically, the central one-third of the gp130 molecule, and lysis is completely inhibited by a monoclonal antibody against envelope. Our work extends observations of human lymphocytes expressing HIV gp120 to the SIV/rhesus monkey model for AIDS. Additionally, we address the relevance of UR lysis in vivo. A survey of PBMC from 56 uninfected rhesus monkeys indicates that 59% of the individuals had peak UR lytic activity above 15% specific lysis. Eleven of these monkeys were subsequently infected with SIV. Animals with UR lytic activity above 15% specific lysis were predisposed to more rapid disease progression than animals with low UR lytic activity, suggesting a strong correlation between this form of innate immunity and disease progression to AIDS.

Rapid and slow progressors differ by a single MHC class I haplotype in a family of MHC-defined rhesus macaques infected with SIV.

Highly polymorphic HLA class I molecules may influence rates of disease progression of HIV-infected individuals. Recent evidence suggests that individuals who mount vigorous CTL responses to multiple HIV-1 epitopes have reduced viral loads, and survive longer than individuals that make a less robust or less diverse CTL response. It has been difficult, however, to define associations between particular HLA class I alleles and rates of disease progression. This may be due, in part, to the uncontrolled variables associated with naturally acquired HIV infections. Studies using MHC-defined, non-human primates infected with well characterized viral stocks should help to clarify this relationship. To explore the possibility that MHC class I polymorphism can influence disease progression, we infected four Mamu-DRB-identical individuals from a family of MHC-defined rhesus macaques intravenously with 40 TCID50SIVmac239. Two of these macaques developed severe wasting and were euthanized within 80 days of infection, while the other two survived for more than 400 days without showing any symptoms of disease. Since all four of these macaques were Mamu-DRB-identical, we were able to exclude the MHC class II DRB loci as determinant of disease progression. Interestingly, both of the slow progressors made CTL responses to the same three SIV CTL epitopes, which were restricted by two molecules (Mamu-B*03 and B*04) encoded by their common maternal haplotype. The two rapid progressors did not share this haplotype with the slow progressors, and we were unable to detect CTL responses in these two siblings. These observations implicate products of the Mamu-B*03 and B*04 alleles in resistance to disease progression in this family of SIV-infected macaques, and provide additional evidence that certain MHC class I-restricted CTL responses may play a significant role in delaying the onset of AIDS.

Recombinant, attenuated Salmonella typhimurium stimulate lymphoproliferative responses to SIV capsid antigen in rhesus macaques.

Recombinant bacteria are useful vectors for delivering foreign antigens to mucosal surfaces and may elicit immune protection against sexually-transmitted pathogens. Recombinant, attenuated Salmonella typhimurium expressing the Simian Immunodeficiency Virus capsid protein (p27) were given to rhesus macaques by intragastric intubation. This route of immunization was compared with intramuscular injection of soluble p27 in adjuvant, and with immunization protocols that combined intragastric and intramuscular antigen exposures. Recombinant Salmonella stimulated p27-specific lymphoproliferative responses that were present transiently in peripheral blood, and were recalled easily by booster immunizations. Intramuscular p27 injection elicited strong serum antibody responses, but only low level capsid-specific proliferative responses. Recombinant Salmonella immunization elicited low levels of p27-specific antibodies in serum and did not suppress subsequent responses to parenteral immunization. Intragastric immunization of macaques with recombinant Salmonella typhimurium was safe and induced immune responses specific for the expressed, foreign antigen.