Induction of HTLV-III/LAV from a nonvirus-producing T-cell line: implications for latency

When the human T-cell line A3.01 is infected with HTLV-III/LAV, the virus associated with the acquired immune deficiency syndrome (AIDS), most of the cells are killed. However, a small number of cells that lack the Leu-3 surface marker survive. Under normal conditions these surviving cells do not produce virus, nor can they be infected by the virus, but they can be induced to produce virus by treatment with 5-iodo-2'-deoxyuridine. This response can be induced for as long as 3 months after the initial infection, suggesting that the cells may harbor a latent form of HTLV-III/LAV.     

Genetic variation in HTLV-III/LAV over time in patients with AIDS or at risk for AIDS

In a study of genetic variation in the AIDS virus, HTLV-III/LAV, sequential virus isolates from persistently infected individuals were examined by Southern blot genomic analysis, molecular cloning, and nucleotide sequencing. Four to six virus isolates were obtained from each of three individuals over a 1-year or 2-year period. Changes were detected throughout the viral genomes and consisted of isolated and clustered nucleotide point mutations as well as short deletions or insertions. Results from genomic restriction mapping and nucleotide sequence comparisons indicated that viruses isolated sequentially had evolved in parallel from a common progenitor virus. The rate of evolution of HTLV-III/LAV was estimated to be at least 10(-3) nucleotide substitutions per site per year for the env gene and 10(-4) for the gag gene, values a millionfold greater than for most DNA genomes. Despite this relatively rapid rate of sequence divergence, virus isolates from any one patient were all much more related to each other than to viruses from other individuals. In view of the substantial heterogeneity among most independent HTLV-III/LAV isolates, the repeated isolation from a given individual of only highly related viruses raises the possibility that some type of interference mechanism may prevent simultaneous infection by more than one major genotypic form of the virus.     

Isolation of a new human retrovirus from West African patients with AIDS

The etiological agent of AIDS, LAV/HTLV-III, is common in Central Africa but is not endemic in other areas of that continent. A novel human retrovirus, distinct from LAV/HTLV-III, has now been isolated from two AIDS patients from West Africa. Partial characterization of this virus revealed that it has biological and morphological properties very similar to LAV but that it differs in some of its antigenic components. Although the core antigens may share some common epitopes, the West African AIDS retrovirus and LAV differ substantially in their envelope glycoproteins. The envelope antigen of the West African virus can be recognized by serum from a macaque with simian AIDS infected by the simian retrovirus termed STLV-IIImac, suggesting that the West African AIDS virus may be more closely related to this simian virus than to LAV. Hybridization experiments with LAV subgenomic probes further established that this new retrovirus, here referred to as LAV-II, is distantly related to LAV and distinct from STLV-IIImac.     

Binding of HTLV-III/LAV to T4+ T cells by a complex of the 110K viral protein and the T4 molecule

Human T-lymphotropic virus type III (HTLV-III) or lymphadenopathy-associated virus (LAV) is tropic for human T cells with the helper-inducer phenotype, as defined by reactivity with monoclonal antibodies specific for the T4 molecule. Treatment of T4+ T cells with monoclonal antibodies to T4 antigen blocks HTLV-III/LAV binding, syncytia formation, and infectivity. Thus, it has been inferred that the T4 molecule itself is a virus receptor. In the present studies, the surfaces of T4+ T cells were labeled radioactively, and then the cells were exposed to virus. After the cells were lysed, HTLV-III/LAV antibodies were found to precipitate a surface protein with a molecular weight of 58,000 (58K). By blocking and absorption experiments, this 58K protein was identified as the T4 molecule. No cell-surface structures other than the T4 molecule were involved in the antibody-antigen complex formation. Two monoclonal antibodies, each reactive with a separate epitope of the T4 molecule, were tested for their binding capacities in the presence of HTLV-III/LAV. When HTLV-III/LAV was bound to T4+ T cells, the virus blocked the binding of one of the monoclonal antibodies, T4A (OKT4A), but not of the other, T4 (OKT4). When HTLV-III/LAV was internally radiolabeled and bound to T4+ T cells which were then lysed, a viral glycoprotein of 110K (gp110) coprecipitated with the T4 molecule. The binding of gp110 to the T4 molecule may thus be a major factor in HTLV-III/LAV tropism and may prove useful in developing therapeutic or preventive measures for the acquired immune deficiency syndrome.     

The epidemiology of AIDS: current status and future prospects

The reported incidence of acquired immune deficiency syndrome (AIDS) continues to increase in countries throughout the world. On the basis of a polynomial model for extrapolation, the cumulative number of cases diagnosed and reported since 1981 in the United States is expected to double during the next year with over 12,000 additional cases projected to be diagnosed by July 1986. The annual incidence rates for single (never-married) men in Manhattan and San Francisco, intravenous drug users in New York City and New Jersey, and persons with hemophilia A ranged from 261 to 350 per 100,000 population during 1984. For single men aged 25 to 44 years in Manhattan and San Francisco, AIDS was the leading cause of premature mortality in 1984 as measured by years of potential life lost. Infection with HTLV-III/LAV is considerably more common than reported AIDS in high-risk populations and can persist at least for several years, so the presence of specific antibody should be considered presumptive evidence of current infection. The screening of donated blood and plasma for antibody to HTLV-III/LAV and use of safer clotting factor concentrates should greatly reduce HTLV-III/LAV transmission through blood and blood products. Most HTLV-III/LAV infections occur through sexual transmission, use of contaminated needles, and as a result of infected mothers passing the virus to newborns. Continued research commitment is needed to develop an HTLV-III/LAV vaccine and therapy for this infection. In the interim, widespread community efforts are needed to minimize transmission.     

Cytokines alter production of HIV-1 from primary mononuclear phagocytes

Some strains of human immunodeficiency virus type 1 (HIV-1) can infect primary monocytes and monocyte-derived macrophages in vitro. In this report, the effect of cytokines on the production of one of these strains that shows a tropism for mononuclear phagocytes, designated HIV-1JR-FL, was studied. Primary peripheral blood mononuclear phagocytes infected with HIV-1JR-FL were treated with the hematopoietic factors: granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), macrophage colony-stimulating factor (M-CSF), and gamma-interferon (gamma-IFN). The M-CSF, GM-CSF, IL-3, and gamma-IFN were able to alter HIV-1 production under different conditions.     

New human T-lymphotropic retrovirus related to simian T-lymphotropic virus type III (STLV-IIIAGM)

This report describes serologic evidence for a virus similar to that known as simian T-lymphotropic virus type III of African Green monkeys (STLV-IIIAGM) infecting apparently healthy people in Senegal, West Africa, and the isolation of virus from these individuals. Serum samples from selected healthy West African people showed unusual serologic profiles when tested with antigens of HTLV-III/LAV, the etiologic agent of AIDS, and of STLV-IIIAGM. The samples reacted strongly with all of the major viral antigens of STLV-IIIAGM but showed variable or no reactivity with the major viral antigens of HTLV-III/LAV by radioimmunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A new human T-lymphotropic virus (HTLV-IV) isolated from these people was grown in vitro and shown to have retroviral type particles, growth characteristics, and major viral proteins similar to those of the STLV-III and HTLV-III/LAV group of retroviruses. The gp120/160, gp32, p64, p55, p53, p24, and p15 proteins precipitated were the same size as and reactive with STLV-IIIAGM proteins. The serologic data suggest that this virus shares more common epitopes with STLV-IIIAGM than with the prototype HTLV-III/LAV that infects people in the United States and Europe. Further study of this virus and of the origin of the HTLV-III/LAV group of viruses may expand our understanding of the human AIDS virus.     

Infection of HTLV-III/LAV in HTLV-I-carrying cells MT-2 and MT-4 and application in a plaque assay

The human T-cell lines MT-2 and MT-4 carry the human T-cell leukemia virus type I (HTLV-I). When MT-2 and MT-4 were infected with HTLV-III, the probable etiologic agent of the acquired immune deficiency syndrome (AIDS), rapid cytopathogenic effects and cytotoxicity were observed that made it possible to titrate the biologically active virus in a plaque-forming assay. The cytopathogenic effects were preceded by the rapid induction and increase of HTLV-III antigens as revealed by immunofluorescence and immunoprecipitation. Activities of HTLV-III were neutralized by the human antibodies against the virus when immunofluorescence and plaque assays were used. Essentially the same results were obtained with the lymphadenopathy-associated virus (LAV1).     

Isolation of T-lymphotropic retrovirus related to HTLV-III/LAV from wild-caught African green monkeys

Present evidence suggests that the acquired immune deficiency syndrome (AIDS) emerged in Central Africa as a new disease in recent decades. This disease has recently approached epidemic proportions in many parts of the world. The etiologic agent of AIDS is believed to be the virus HTLV-III/LAV, which has been proposed as having originated from a recent simian-human transmission in Africa. This report describes the isolation of a designated STLV-IIIAGM retrovirus closely related to HTLV-III/LAV from seven healthy wild-caught African Green monkeys (Cercopithecus aethiops) that showed the presence of antibodies designated STLV-IIIAGM. In vitro growth characteristics, ultrastructural morphology, and major proteins of 160,000 kilodaltons (kD), 120 kD, 55 kD, and 24 kD are similar to and cross-reactive with the analogous antigens of HTLV-III/LAV. The use of these serologic markers in the detection of STLV-IIIAGM-infected monkeys may be important in assuring the continued safety of a variety of biologic reagents that are derived from these primate species. The existence of a retrovirus closely related to HTLV-III/LAV that naturally infects an African nonhuman primate in the apparent absence of disease may provide a unique model for the study of human AIDS and the development of an effective vaccine.     

Hepatitis B virus DNA sequences in lymphoid cells from patients with AIDS and AIDS-related complex

A lymphotropic virus HTLV-III/LAV was recently identified as the etiologic agent of the acquired immune deficiency syndrome (AIDS). In a study of concomitant hepatitis B infections in patients with AIDS or the AIDS-related complex, DNA sequences of hepatitis B virus (HBV) were found in fresh and cultured lymphocytes from patients with AIDS even in the absence of conventional HBV serological markers. Furthermore, the restriction DNA pattern was consistent with the integration of the viral DNA. These results should prompt additional studies to reevaluate a possible role of HBV as a cofactor in AIDS in addition to the HTLV-III/LAV causal agent.     

Characterization of gp41 as the transmembrane protein coded by the HTLV-III/LAV envelope gene

Radiolabeled amino acid sequencing was used to characterize gp41, an antigen of HTLV-III/LAV, the virus believed to be the etiological agent of the acquired immune deficiency syndrome. This antigen is the one most commonly detected in immunoblot assays by sera of patients with AIDS or AIDS-related complex (ARC) and other individuals infected with HTLV-III/LAV. A mouse monoclonal antibody that was reactive with gp41 precipitated a 160-kilodalton protein (gp160) in addition to gp41, but did not precipitate a 120-kilodalton protein (gp120) from extracts of metabolically labeled cells producing HTLV-III. Extracts of infected cells that had been labeled with tritiated leucine or isoleucine were immunoprecipitated with the monoclonal antibody. The immunoprecipitates were fractionated by polyacrylamide gel electrophoresis and the p41 was eluted from the gel bands and subjected to amino-terminal radiolabeled amino acid sequencing by the semiautomated Edman degradation. Leucine residues occurred in cycles 7, 9, 12, 26, 33, and 34 among 40 cycles and isoleucine occurred in cycle 4 among 24 cycles analyzed. Comparison of the data with the deduced amino acid sequence of the env gene product of HTLV-III precisely placed gp41 in the COOH-terminal region of the env gene product. Gp160 is thus the primary env gene product and it is processed into gp120 and gp41.     

Specific tropism of HIV-1 for microglial cells in primary human brain cultures

Human immunodeficiency virus (HIV) frequently causes neurological dysfunction and is abundantly expressed in the central nervous system (CNS) of acquired immunodeficiency syndrome (AIDS) patients with HIV encephalitis or myelopathy. The virus is found mostly in cells of the monocyte-macrophage lineage within the CNS, but the possibility of infection of other glial cells has been raised. Therefore, the effects of different HIV-1 and HIV-2 strains were studied in primary cultures of adult human brain containing microglial cells, the resident CNS macrophages, and astrocytes. These cultures could be productively infected with macrophage-adapted HIV-1 isolates but not with T lymphocyte-adapted HIV-1 isolates or two HIV-2 isolates. As determined with a triple-label procedure, primary astrocytes did not express HIV gag antigens and remained normal throughout the 3-week course of infection. In contrast, virus replicated in neighboring microglial cells, often leading to their cell fusion and death. The death of microglial cells, which normally serve immune functions in the CNS, may be a key factor in the pathogenesis of AIDS encephalitis or myelopathy.     

Immunoregulatory lymphokines of T hybridomas from AIDS patients: constitutive and inducible suppressor factors

Supernatants derived from peripheral blood mononuclear cell cultures of certain patients with the acquired immunodeficiency syndrome (AIDS) or its prodromes have the capacity to block T cell-dependent immune reactivity in vitro. T cells derived from a patient positive for antibody to the lymphadenopathy associated virus ( LAV ), and elaborating high titers of these soluble suppressor factors, were fused to a mutagenized clone of the human T lymphoblastoid cell line KE37 . Molecules capable of profoundly depressing T cell-dependent polyclonal antibody production and DNA synthetic responses, either directly or after incubation with normal adherent cells, were isolated from stable hybrid clones.     

Secretion of neurotoxins by mononuclear phagocytes infected with HIV-1

Mononuclear phagocytes (microglia, macrophages, and macrophage-like giant cells) are the principal cellular targets for human immunodeficiency virus-1 (HIV-1) in the central nervous system (CNS). Since HIV-1 does not directly infect neurons, the causes for CNS dysfunction in acquired immunodeficiency syndrome (AIDS) remain uncertain. HIV-1-infected human monocytoid cells, but not infected human lymphoid cells, released toxic agents that destroy chick and rat neurons in culture. These neurotoxins were small, heat-stable, protease-resistant molecules that act by way of N-methyl-D-aspartate receptors. Macrophages and microglia infected with HIV-1 may produce neurologic disease through chronic secretion of neurotoxic factors.     

Long-term cultures of HTLV-III--infected T cells: a model of cytopathology of T-cell depletion in AIDS

Long-term cultures were established of HTLV-III-infected T4 cells from patients with the acquired immune deficiency syndrome (AIDS) and of T4 cells from normal donors after infection of the cells in vitro. By initially reducing the number of cells per milliliter of culture medium it was possible to grow the infected cells for 50 to 60 days. As with uninfected T cells, immunologic activation of the HTLV-III-infected cells with phytohemagglutinin led to patterns of gene expression typical of T-cell differentiation, such as production of interleukin-2 and expression of interleukin-2 receptors, but in the infected cells immunologic activation also led to expression of HTLV-III, which was followed by cell death. The results revealed a cytopathogenic mechanism that may account for T4 cell depletion in AIDS patients and suggest how repeated antigenic stimulation by infectious agents, such as malaria in Africa, or by allogeneic blood or semen, may be important determinants of the latency period in AIDS.     

Genomic heterogeneity of AIDS retroviral isolates from North America and Zaire

In an analysis of the genomic variation of AIDS retroviral isolates from patients living in New York, Alabama, and Zaire, restriction maps were constructed by using seven enzymes, each known to cleave the proviral DNA more than once, in conjunction with Southern blot analysis. The maps of LAV, HTLV-III, and ARV-2 as deduced from their published nucleotide sequences were included in this analysis. The results demonstrated that (i) several "signature" restriction sites were common to all isolates; (ii) with the exception of LAV and HTLV-III, the North American and European isolates were all different from one another and showed no geographical specificity; (iii) the African isolates as a group were more diverse than those from North America and Europe; and (iv) the genomic variability was concentrated within the env gene.