Characterization of CD4 and CD8 T cell responses in MuSK myasthenia gravis.

Muscle specific tyrosine kinase myasthenia gravis (MuSK MG) is a form of autoimmune MG that predominantly affects women and has unique clinical features, including prominent bulbar weakness, muscle atrophy, and excellent response to therapeutic plasma exchange. Patients with MuSK MG have predominantly IgG4 autoantibodies directed against MuSK on the postsynaptic muscle membrane. Lymphocyte functionality has not been reported in this condition. The goal of this study was to characterize T cell responses in patients with MuSK MG. Intracellular production of IFN-gamma, TNF-alpha, IL-2, IL-17, and IL-21 by CD4+ and CD8+ T cells was measured by polychromatic flow cytometry in peripheral blood samples from 11 Musk MG patients and 10 healthy controls. Only one MuSK MG patient was not receiving immunosuppressive therapy. Regulatory T cells (Treg) were also included in our analysis to determine if changes in T cell function were due to altered Treg frequencies. CD8+ T cells from MuSK MG patients had higher frequencies of polyfunctional responses than controls, and CD4+ T cells had higher IL-2, TNF-alpha, and IL-17. MuSK MG patients had a higher percentage of CD4+ T cells producing combinations of IFN-gamma/IL-2/TNF-gamma, TNF-alpha/IL-2, and IFN-gamma/TNF-alpha. Interestingly, Treg numbers and CD39 expression were not different from control values. MuSK MG patients had increased frequencies of Th1 and Th17 cytokines and were primed for polyfunctional proinflammatory responses that cannot be explained by a defect in CD39 expression or Treg number.

HIV-1 gp41 envelope IgA is frequently elicited after transmission but has an initial short response half-life.

Prevention of HIV-1 transmission at mucosal surfaces will likely require durable pre-existing mucosal anti-HIV-1 antibodies (Abs). Defining the ontogeny, specificities and potentially protective nature of the initial mucosal virus-specific B-cell response will be critical for understanding how to induce protective Ab responses by vaccination. Genital fluids from patients within the earliest stages of acute HIV-1 infection (Fiebig I-VI) were examined for multiple anti-HIV specificities. Gp41 (but not gp120) Env immunoglobulin (Ig)A Abs were frequently elicited in both plasma and mucosal fluids within the first weeks of transmission. However, shortly after induction, these initial mucosal gp41 Env IgA Abs rapidly declined with a t(½) of ∼2.7 days. B-cell-activating factor belonging to the TNF family (BAFF) was elevated immediately preceding the appearance of gp41 Abs, likely contributing to an initial T-independent Ab response. HIV-1 transmission frequently elicits mucosal HIV-1 envelope-specific IgA responses targeted to gp41 that have a short half-life.

Polyfunctional cytomegalovirus-specific immunity in lung transplant recipients receiving valganciclovir prophylaxis.

Cytomegalovirus (CMV) is a common opportunistic infection after lung transplant. Despite effective antiviral medications to treat CMV, invasive CMV disease contributes to lung allograft dysfunction and worse survival. Efforts to prevent CMV have led to the use of valganciclovir prophylaxis for increasingly longer periods after transplant. A pivotal concern with long-term antiviral prophylaxis is that it may prevent or delay the development of CMV-specific immunity and increase the subsequent risk of late onset disease. To address this issue, we conducted a pilot study to determine if CMV-specific immunity was detectable in lung transplant recipients at risk for CMV while on antiviral prophylaxis. Utilizing polychromatic flow cytometry panels, CMV-specific immunity was determined by peripheral blood CD4 and CD8 T cell expression of cytokines in response to the HLA restricted CMV peptides pp65 and IE-1. We determined CMV seropositive lung transplant recipients on valganciclovir for a median of 6 months from transplant have a detectable polyfunctional CMV-specific T cell response which is comparable to seropositive recipients not on antiviral medications and to healthy seropositive nontransplant controls. Thus, valganciclovir prophylaxis does not appear to impair the development of CMV-specific immunity in lung transplantation.

Excellent safety and tolerability of the human immunodeficiency virus type 1 pGA2/JS2 plasmid DNA priming vector vaccine in HIV type 1 uninfected adults.

A vaccine consisting of DNA priming followed by recombinant modified vaccinia Ankara (rMVA) boosting has achieved long-term control of a pathogenic challenge with a chimera of simian and human immunodeficiency viruses (SHIV-89.6P) in rhesus macaques. Based on these results, clade B HIV-1 DNA and rMVA immunogens have been developed for trials in humans. We conducted a first-time in humans phase I safety trial using the pGA2/JS2 (JS2) HIV-1 DNA priming vector expressing Gag, Pol, Env, Tat, Rev, and Vpu. Thirty HIV-uninfected adults were vaccinated with 0.3 or 3 mg of JS2 DNA, or a saline placebo, by intramuscular injection at months 0 and 2. Both doses of DNA were safe and well-tolerated with no differences between the control, 0.3 mg, or 3 mg groups (n = 6, 12, and 12, respectively) through 12 months of postvaccination follow- up. A chromium-release assay using fresh peripheral blood mononuclear cells (PBMCs) and a validated IFN-gamma ELISpot assay with frozen PBMCs failed to detect CD4(+) or CD8(+) HIV-1-specific T cell responses. HIV-specific neutralizing antibodies were also not detected. The vaccine is being further developed as a priming vector for a combined DNA plus rMVA prime/boost HIV vaccination regimen.

Immunologic and virologic analyses of an acutely HIV type 1-infected patient with extremely rapid disease progression.

The immunologic and virologic factors that impact on the rate of disease progression after acute infection with human immunodeficiency virus (HIV) type 1 are poorly understood. A patient with an extraordinarily rapid disease course leading to AIDS-associated death within 6 months of infection was studied intensively for the presence of anti-HIV immune reactivities as well as changes in the genetic and biologic properties of virus isolates. Although altered humoral responses were evident, the most distinctive immunologic feature was a nearly complete absence of detectable HIV-specific CTL responses. In addition to a rapid decline in CD3+CD4+ cells, elevated percentages of CD8+CD45RA+ and CD8+CD57+ cells and diminished CD8+CD45R0+ and CD8+CD28+ cells were evident. Primary viral isolates recovered throughout the course of infection exhibited limited sequence diversity. Cloned viral envelopes were found to have unusually broad patterns of coreceptor usage for cell-cell fusion, although infectivity studies yielded no evidence of infection via these alternative receptors. The infectivity studies demonstrated that these isolates and their envelopes maintained an R5 phenotype throughout the course of disease. The absence of demonstrable anti-HIV CTL reactivities, coupled with a protracted course of seroconversion, highlights the importance of robust HIV-specific immune responses in the control of disease progression.

CD8 CTL responses in vaccines: emerging patterns of HLA restriction and epitope recognition.

We evaluated MHC-class I-restricted CTL responses induced by HIV-1 clade B-based vaccines in nine HIV-1 seronegative vaccine recipients with regard to their patterns of HLA restriction and epitope recognition. We found that seven of nine volunteers developed detectable CTL reactivities against novel epitopes within the HIV-1 Env and Gag proteins. Although four of nine subjects were HLA-A*0201, none of the cellular responses was restricted in the context of this allele. The type of responses observed in this sampling of vaccines appeared similar to those reported during primary infection and among long term non-progressors, with three out of nine subjects recognizing HLA-B27 or HLA-B17(57)-restricted epitopes. Although the majority of CTL responses were directed against novel epitopes, these effectors were still able to mediate cross-clade reactivities.

Immunologic profile of human immunodeficiency virus-infected patients during viral remission and relapse on antiretroviral therapy.

A dissociation between plasma human immunodeficiency virus (HIV) RNA levels and CD4(+) cell counts has been reported in patients experiencing viral relapse while receiving antiretroviral therapy. This study compared patients with stable CD4(+) lymphocytes during viral relapse while receiving treatment with patients who had sustained virus suppression. Plasma HIV RNA levels, lymphocyte immunophenotyping, and T cell receptor excision circle (TREC) levels were measured. Naive CD4(+) lymphocyte phenotype and TREC levels were not significantly different in patients with virus suppression or in those who had relapsed. However, CD8(+) lymphocyte activation, including the number and percentage of activated cells and CD38 antibody-binding capacity, was significantly elevated during viral relapse, compared with that in suppressed patients. By multivariable regression analyses, CD8(+) and CD4(+) lymphocyte activation were associated significantly with increasing plasma HIV RNA levels.

Thymopoiesis in HIV-infected adults after highly active antiretroviral therapy.

The thymus of HIV-seropositive patients can enlarge as CD4+ T cell counts increase on highly active anti-retroviral therapy (HAART). This may indicate development of new T cells or represent mature peripheral T cells recirculating to the thymus. To define the etiology of the enlargement, the thymuses of two HIV-infected individuals on HAART were biopsied. For more than 3 years before initiation of HAART, both patients (38 and 41 years of age) had documented CD4+ T lymphopenia. Peripheral blood samples were obtained to assess circulating CD4+ CD45RA+ CD62L+ T cells, which were thought to have recently developed in the thymus. Peripheral blood T cells from both patients and thymocytes from the second patient were also tested for levels of DNA episomes formed during T cell receptor gene rearrangement (T cell receptor rearrangement excision circles, TRECs). With HAART, peripheral blood CD4+ T cell counts increased from approximately 60/mm(3) to 552/mm(3) and 750/mm(3) for patients 1 and 2, respectively. Thymic biopsies from both patients showed normal thymus histology with active thymopoiesis. Percentages of peripheral blood CD4+ CD45RA+ CD62L+ T cells and quantitation of T cell TRECs also reflected active thymopoiesis in both patients. Thus, in these two HIV-seropositive adults examined after initiation of HAART, thymic enlargement represented active thymopoiesis. Thymopoiesis in adult AIDS patients may contribute to immune reconstitution even after prolonged CD4+ T lymphopenia.

Identification of highly conserved and broadly cross-reactive HIV type 1 cytotoxic T lymphocyte epitopes as candidate immunogens for inclusion in Mycobacterium bovis BCG-vectored HIV vaccines.

One of the fundamental goals of current strategies to develop an efficacious vaccine for AIDS is the elicitation of cytotoxic T lymphocyte (CTL) reactivities capable of recognizing cells infected with different subtypes of the human immunodeficiency virus type 1 (HIV-1). In efforts to explore new vaccine candidates by the UNAIDS/WHO Vaccine Committee, we review the most recent data concerning CTL epitopes that are conserved among the different HIV-1 subtypes. Moreover, we examine HLA allelic frequencies in several different populations, to determine those that could contribute to the goal of a cumulative phenotype frequency (CP) of at least 80%. By analyzing conserved epitopes in the context of HLA restricting alleles, we define a set of HIV-1 gene regions that may have the greatest potential to induce cross-clade reactive CTLs. The absence of well-defined correlates of immune protection that link CTL epitopes to delayed disease progression and/or prevention of infection does not permit an assignment of rank order of the most relevant component of a candidate vaccine. Thus far, most of the studies conducted in clade B-infected patients to define conserved and immunodominant epitopes indicate gag and pol gene products to be the most conserved among the HIV-1 subtypes. Moreover, anti-Pol and -Gag CTL responses appear to correlate inversely with disease progression, suggesting that they should be among the first choice of antigens to be included in a candidate vaccine construct aimed at induction of broad CTL responses. The impact of a clade B-based vaccine as a worldwide candidate capable of inducing protective immune responses can be determined only after "in vivo" studies. Meanwhile, extensive parallel studies in populations infected with non-clade B HIV-1 subtypes should define the patterns of immunodominant epitopes and HLA for comparison with the data already collected in clade B-infected subjects.

A distinctive clade B HIV type 1 is heterosexually transmitted in Trinidad and Tobago.

HIV-1 transmission worldwide is predominantly associated with heterosexual activity, and non-clade B viruses account for the most spread. The HIV-1 epidemic in Trinidad/Tobago and the Caribbean shares many features with such heterosexual epidemics, including a prominent role for coincident sexually transmitted diseases. This study evaluates the molecular epidemiology of HIV-1 in Trinidad/Tobago during a period when abrupt transition from homosexual to heterosexual transmission occurred in the absence of injecting drug use, concomitant with a rapid rise in HIV-1 prevalence in the heterosexual population. Of 31 viral isolates studied during 1987-1995, all cluster with subtype B reference strains. In the analysis of full env genes from 22 early seroconverters, the Trinidad isolates constitute a significant subcluster within the B subtype. The Trinidad V3 consensus sequence differs by a single amino acid from the prototype B V3 consensus and demonstrates stability over the decade of this study. In the majority of isolates, the V3 loop of env contains a signature threonine deletion that marks the lineage of the Trinidad HIV-1 clade B epidemic from pre-1984. No phenotypic features, including syncitium induction, neutralization profiles, and chemokine receptor usage, distinguish this virus population from other subtype B viruses. Thus, although the subtype B HIV-1 viruses being transmitted in Trinidad are genetically distinguishable from other subtype B viruses, this is probably the result of a strong founder effect in a geographically circumscribed population rather than genetic selection for heterosexual transmission. These results demonstrate that canonical clade B HIV-1 can generate a typical heterosexual epidemic.

Effect of highly active antiretroviral therapy and thymic transplantation on immunoreconstitution in HIV infection.

The purpose of this study was to determine whether thymic transplantation in addition to highly active antiretroviral therapy (HAART) will restore T cell function in HIV infection. Eight treatment-naive HIV-infected patients with CD4+ T cell counts of 200-500/mm3 were randomized into thymic transplantation and control arms. All patients received HAART (zidovudine, lamivudine, and ritonavir) for 6 weeks prior to transplantation. Thymic transplantation was done without immunosuppression, using postnatal HLA-unmatched cultured allogeneic thymus tissue. Patients were immunized every 6 months with the neoantigen keyhole limpet hemocyanin (KLH) and the recall antigen tetanus toxoid (TT). T cell phenotype and function and T cell receptor rearrangement excision circles (TRECs) were assessed. Thymic allografts were biopsied at 2 months. Six HIV-infected patients completed the study. Four patients received cultured allogeneic postnatal thymic grafts, two others were controls. CD4+ T cell counts increased and T cell-proliferative responses to Candida antigen and TT normalized in all patients. Proliferative responses to KLH developed in three of four transplant recipients and one of two controls. Patients responding to KLH after secondary immunization had greater TREC increases compared with the patients who did not respond. All thymic allografts were rejected within 2 months. In summary, four of six patients developed T cell-proliferative responses to the neoantigen KLH over the first 2 years of HAART. The transplanted thymus tissue, however, was rejected. There was no clear difference in restoration of T cell function in the transplant recipients compared with the controls. Increases in TRECs after initiation of HAART may correlate with improved immune function.

CD8+ T cell-mediated suppressive activity inhibits HIV-1 after virus entry with kinetics indicating effects on virus gene expression.

Individuals infected with HIV-1 have varying rates of progression to AIDS. Cellular immune responses, comprised of cytolytic and noncytolytic CD8(+) T cell effector functions, are considered important for controlling viremia and maintaining the clinically asymptomatic state. Although there is general agreement regarding CD8(+) T lymphocyte cytotoxic functions, considerable controversy exists over the nature of the noncytolytic antiviral activity of CD8(+) cells. The discovery that RANTES (regulated on activation, normal T cell expressed and secreted), MIP-1alpha, and MIP-1beta (macrophage inflammatory protein 1 alpha and beta) could inhibit HIV-1 replication by blocking viral entry processes led to the notion that these molecules are responsible for the CD8(+) cell suppressive activity. However, T tropic HIV isolates requiring the CXCR4 coreceptor for entry are insensitive to the antiviral effects of these beta-chemokines. Using a CXCR4-dependent virus, we determined that the mechanism of CD8(+) T cell-mediated activity did act after viral entry into the host cell. We also define the kinetics of the HIV life cycle in primary activated human CD4(+)-enriched T cells by using an HIV-1 reporter virus system pseudotyped with the CXCR4-dependent HIV-1 envelope gene of NL4-3. Analysis of these kinetic data indicates that CD8(+) T cell-mediated suppressive activity acts at a stage in the viral life cycle after entry and independently of the HIV envelope. Additionally, we show that the antiviral activity targets stages of the virus life cycle correlating with transcription and early proviral gene expression. These findings not only provide a range of possible targets for the CD8(+) T cell-mediated activity but also support the notion that this antiviral activity is multifactorial in nature.

HIV vaccine development at Duke University Medical Center.

With the AIDS epidemic continuing to spread throughout the world, development of a safe, practical, and effective HIV vaccine is a national priority. HIV vaccine research efforts are currently targeted towards design of HIV immunogens that induce both cellular and humoral immunity. This brief review summarizes ongoing work at the Duke University School of Medicine on HIV vaccine development.

Immune responses in asymptomatic HIV-1-infected patients after HIV-DNA immunization followed by highly active antiretroviral treatment.

Intensive chemotherapy is capable of reducing the viral load in HIV-1-infected individuals while infected cells are still present. A special property of DNA immunization is to induce both new CTL and Ab responses. We evaluated the possibility of inducing new immune responses in already infected individuals by means of DNA constructs encoding the nef, rev, or tat regulatory HIV-1 genes. Significant changes in viral loads and CD4+ counts were observed in four patients who started highly active antiretroviral treatment (HAART) during the immunization study. The DNA immunization induced Ag-specific T cell proliferation, which persisted up to 9 mo after the last DNA injection, and cytolytic activities but did not, by itself, reduce viral load. Increased levels of CTL precursor cells were induced in all nine DNA-immunized patients. The profile of IFN-gamma secretion observed when human PBMC were transfected with the nef, rev, and tat DNA resembled that found in the CTL activity (nef > tat > rev). Ab responses that occurred after immunizations were of a low magnitude. In accordance with the high IL-6 production induced by the nef DNA plasmid, IgG titers were highest in patients immunized with nef DNA. The initiation of HAART appears to contribute to the induction of new HIV-specific CTL responses, but by itself did not cause obvious re-induction of these activities.

A canarypox vaccine expressing multiple human immunodeficiency virus type 1 genes given alone or with rgp120 elicits broad and durable CD8+ cytotoxic T lymphocyte responses in seronegative volunteers.

Induction of CD8+ cytotoxic T cells is considered one of the important correlates for the protective efficacy of candidate human immunodeficiency virus type 1 (HIV-1) vaccines. To induce CD8+ cytotoxic T lymphocytes (CTLs) along with neutralizing antibody and CD4+ T cell help, a live canarypox virus construct expressing gp120, transmembrane gp41, the gag and protease genes, and sequences containing CTL epitopes in nef and pol was given simultaneously with, or followed by, rgp120 SF2. CD8+ CTLs were detected in 61% of volunteers at some time during the trial. Three to 6 months after the last immunization, the gene-specific responses were gag, 26/81; env, 17/77; nef, 12/77; and pol, 3/16. Simultaneous immunization with the canarypox vector and the subunit, beginning with the initial immunization, resulted in earlier antibody responses. In summary, a strategy of immunization with a canarypox vector expressing multiple genes of HIV-1 given with gp120 results in durable CD8+ CTL responses to a broad range of epitopes.

Analysis of the adult thymus in reconstitution of T lymphocytes in HIV-1 infection.

A key question in understanding the status of the immune system in HIV-1 infection is whether the adult thymus contributes to reconstitution of peripheral T lymphocytes. We analyzed the thymus in adult patients who died of HIV-1 infection. In addition, we studied the clinical course of HIV-1 infection in three patients thymectomized for myasthenia gravis and determined the effect of antiretroviral therapy on CD4(+) T cells. We found that five of seven patients had thymus tissue at autopsy and that all thymuses identified had inflammatory infiltrates surrounding lymphodepleted thymic epithelium. Two of seven patients also had areas of thymopoiesis; one of these patients had peripheral blood CD4(+) T-cell levels of <50/mm3 for 51 months prior to death. Of three thymectomized patients, one rapidly progressed to AIDS, one progressed to AIDS over seven years (normal progressor), whereas the third remains asymptomatic at least seven years after seroconversion. Both latter patients had rises in peripheral blood CD4(+) T cells after antiretroviral therapy. Most patients who died of complications of HIV-1 infection did not have functional thymus tissue, and when present, thymopoiesis did not prevent prolonged lymphopenia. Thymectomy before HIV-1 infection did not preclude either peripheral CD4(+) T-cell rises or clinical responses after antiretroviral therapy.

Coadministration of zidovudine and interleukin-2 increases absolute CD4 cells in subjects with Walter Reed stage 2 human immunodeficiency virus infection: results of ACTG protocol 042.

Interleukin-2 (IL-2) can increase numbers of absolute CD4 cells in persons infected with the human immunodeficiency virus who are receiving antiretroviral therapy. Twenty-five subjects with > 400/mm3 absolute CD4 cells received zidovudine and low-dose intravenous or subcutaneous IL-2 (< or = 10(6) U/m2). Absolute CD4 cells increased significantly during IL-2 treatment, and 56% of the subjects achieved a maximal increase of > or = 500 cells/mm3. A dose-response relationship favored increasing IL-2 doses, and subcutaneous delivery offered greater increases than intravenous administration. Fifteen subjects had persistent increases of > or = 100 cells/mm3 6 weeks after IL-2 was discontinued. No changes occurred in delayed-type hypersensitivity or helper T cell responses to recall antigens. Cell-mediated cytotoxicities increased against Daudi cells. IL-2 was well tolerated and only 1 subject required dose reduction. Relatively low-dose IL-2 delivered by subcutaneous or intravenous routes may provide an important complement to antiretroviral therapy to increase absolute CD4 cells with the potential for less toxicity than with higher IL-2 doses.

Lymphokine-activated killer (LAK) cell anti-HIV-1 ADCC reactivity: a potential strategy for reduction of virus-infected cellular reservoirs.

Lymphocytes from HIV-1-seropositive and -seronegative individuals were examined to determine whether HIV-1 infection interfered with the ability to generate a lymphokine-activated killer (LAK) cell response. Following a 3-day ex vivo incubation in the presence of 1000 U/ml of recombinant interleukin-2, lymphocytes from seropositive individuals exhibited a LAK cell response which was equivalent to or greater than that of seronegative controls as measured against Daudi cell targets. LAK cells from seropositive and seronegative donors showed no specific cytolytic activity against gp120-coated or HIV-1-infected targets. However, in the presence of patient sera, significant levels of virus-specific LAK cell-mediated antibody-dependent cellular cytotoxicity (ADCC) were observed. The level of this specific LAK cell-mediated ADCC was greater than that mediated under similar conditions by freshly isolated peripheral blood mononuclear cells. The greatest improvement in ADCC over baseline activity was seen with lymphocytes from AIDS patients after the 3-day ex vivo activation, suggesting that this patient population might benefit the most from adaptive LAK cell therapy.

Safety and immunogenicity of an HLA-based HIV envelope polyvalent synthetic peptide immunogen. DATRI 010 Study Group. Division of AIDS Treatment Research Initiative.

OBJECTIVE: To evaluate the safety and immunogenicity of a polyvalent (PV) HIV envelope synthetic peptide immunogen, C4-V3. The immunogen comprised four peptides containing T-helper epitopes from the fourth constant region (C4) of gp120 of HIV-1MN, and T-helper, cytotoxic T-lymphocyte HLA-B7-restricted, and B-cell neutralizing epitopes from the gp120 third variable region (V3) of four clade B HIV-1 isolates, HIV-1MN, HIV-1RF, HIV-1EV91, and HIV-1Can0A. DESIGN: A pilot, Phase I controlled trial [Division of AIDS Treatment Research Initiative (DATRI) 010] conducted at a single center. METHODS: Ten HIV-infected, HLA-B7-positive patients with CD4 cells > 500 x 10(6)/l were enrolled. Eight patients received the C4-V3 PV immunogen emulsified in incomplete Freund's adjuvant in five intramuscular injections over 24 weeks, and two controls received incomplete Freund's adjuvant alone. All subjects were followed for 52 weeks. RESULTS: Four out of eight C4-V3 PV recipients generated at least fourfold rise in serum antibody titers to at least three immunogen peptides in contrast to none of the control subjects. Four out of eight C4-V3 PV recipients and none of the controls had an at least fourfold rise in neutralizing antibodies to either HIV-1MN, HIV-1RF, or HIV-1(4489-5) laboratory-adapted HIV isolates. 3H-Thymidine incorporation assays of peripheral blood mononuclear cells increased at least fivefold over the baseline stimulation index to at least one of the immunogen peptides in two consecutive post-immunization timepoints in five out of eight C4-V3 PV recipients versus none of the controls. CD4 cell counts and plasma HIV RNA levels did not change in patients who received either C4-V3 PV or adjuvant alone. Adverse events consisted primarily of grade 1 injection site reactions in six subjects (four C4-V3 recipients, two controls). CONCLUSIONS: C4-V3 PV synthetic peptides demonstrated both immunogenicity and safety in HIV-infected patients.

Herpesvirus saimiri transformation of HIV type 1 suppressive CD8+ lymphocytes from an HIV type 1-infected asymptomatic individual.

CD8+ T lymphocytes from HIV+ individuals can potently suppress HIV-1 replication in a noncytolytic manner. This suppression appears to be multifactorial and the molecules contributing have not been fully elucidated. As an approach to this question we used herpesvirus saimiri (HVS) to transform CD8+ T lymphocytes from an HIV+ asymptomatic donor to a continuously growing, activation-independent, IL-2-dependent phenotype. The transformed cell population, termed CD8(HVS), had an activated phenotype, contained HVS sequences, did not shed infectious HVS virus, and was polyclonal. The CD8(HVS) cells, despite the absence of detectable CTL activity, potently suppressed HIV-1 production by both autologous and heterologous CD4+ cells from infected donors. The CD8(HVS) cells in coculture also suppressed virus production from PBMCs acutely infected with syncytium-inducing (SI) strains or NSI primary isolates of HIV-1. The supernatants from the CD8(HVS) cells and their concentrates derived from these supernatants were suppressive to NSI primary isolates of HIV-1 but not to SI strains. Fractionation of these concentrates showed that the suppressive activity was associated with low molecular mass (6500- to 19,300-Da) protein species. Western blotting and ELISA indicated that the CC chemokines MIP-1alpha, MIP-1beta, and RANTES were present in these fractions. Antibody-blocking studies with antibodies to the CC chemokines indicated that a significant portion of the soluble HIV-suppressive activity was due to these molecules. However, these experiments also suggested the inhibitory activity of the CD8(HVS) cells in coculture is not due exclusively to the CC chemokines. The HVS-transformed cells provide a useful tool for the study of noncytolytic CD8+ T lymphocyte-mediated suppression of HIV-1.