Origin of the HIV-susceptible human CD4+ cell line H9.
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CD4-Positive T-Lymphocytes↗
Biomedical subjects
R C Gallo
Publications and source records attributed to R C Gallo.
Monoclonal antibodies to HTLV-III451 gp41: delineation of an immunoreactive conserved epitope in the transmembrane region of divergent isolates of HIV-1.
We report on the development of monoclonal antibodies directed against the transmembrane portion of the envelope of HTLV-III451 gp41. One of these monoclonal antibodies, designated M71/2B4, was found to cross-react with transmembrane proteins from other independent isolates of HIV-1, namely IIIB, MN, and RF. Thus, this monoclonal antibody identifies an epitope located in a region of gp41 that is conserved among all these isolates. To identify this conserved region a series of E. coli recombinant proteins were screened in immunoblot with M71/2B4. From these results the epitope recognized by this antibody appears to map at the amino terminus of gp41, in the region indicated between the cleavage site with gp120 (aa 508) and the HindIII site (aa647).
Animals↗
Purification and partial characterization of human immunodeficiency virus type 2 reverse transcriptase.
We have raised a rabbit monospecific antibody against a synthetic peptide derived from a sequence within the COOH-terminal portion of the reverse transcriptase (RT) of HIV-1. This sequence was also found to be conserved in the predicted amino acid sequence of HIV-2. The antibody, designated C2003, cross-reacted with HIV-2 RT on immunoblots of HIV-2 virus extract and directly inhibited HIV-2 RT activity. Fractionation of HIV-2 RT by immunoaffinity chromatography with C2003 antibody yielded a pair of viral proteins of 68 and 55 kD associated with both RT and RNAse H activities. Both proteins were found to be highly immunogenic, recognized by 11 of 11 human sera that previously tested positive for antibodies to HIV-2 antigens in immunoblot assays.
Amino Acid Sequence↗
A recombinant clone of HIV-1 preferentially transmitted in normal peripheral blood mononuclear cells.
To study biologic properties associated with specific regions of HIV-1, a chimera, pHX-JY1, was constructed by exchanging the vif-env region of a Zairian molecular clone (JY1) with that of pHXB2gpt, a full-length biologically active proviral clone of North American origin. Virus was produced by transfection of permissive cells with parental and recombinant clones, and the biologic and molecular properties of these viruses were compared. Virus derived from pHXB2gpt infected phytohemagglutinin (PHA)-activated normal peripheral blood mononuclear cells (PBMC) and CD4+ leukemic T cell lines equally well. In contrast, virus derived from pHX-JY1 was transmitted slowly to both PBMC and cell lines, and the infectivity of pHX-JY1 virus was two orders of magnitude greater for PBMC than for T cell lines. All essential viral genes in the exchanged JY1 vif-env region were intact and functioned comparably to those of the parent clone in transfected COS-1 cells. The findings suggest differences in these regions of the HIV-1 genome may play an important role in differential cell tropism.
CD4-Positive T-Lymphocytes↗
Polymorphism of human herpesvirus 6 DNA from five Japanese patients with exanthem subitum.
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DNA, Viral↗
Human retroviruses: their role in neoplasia and immunodeficiency.
Human retroviruses (HTLVs and HIVs) infect the cells of the immune system and cause mild-to-severe immune dysfunction. They are directly or indirectly responsible for associated neoplasia and central nervous system disorders. The study of these viruses is of great importance, not only because they cause grave illnesses like AIDS, neoplasias, and CNS disease, but also because they have the ability to exert such fine levels of gene regulatory control in their replication and expression. These studies will ultimately shed light on fundamental mechanisms of genetic control in human cells in their normal state and the alterations of these controls in neoplastic or immunologically aberrant states.
Acquired Immunodeficiency Syndrome↗
Challenge of chimpanzees (Pan troglodytes) immunized with human immunodeficiency virus envelope glycoprotein gp120.
The human immunodeficiency virus type 1 (HIV-1), the causative agent of acquired immunodeficiency syndrome, infects humans and chimpanzees. To determine the efficacy of immunization for preventing infection, chimpanzees were immunized with gp120 purified from human T-cell lymphotrophic virus type IIIB (HTLV-IIIB)-infected cell membranes and challenged with the homologous virus, HTLV-IIIB. A challenge stock of HTLV-IIIB was prepared by using unconcentrated HTLV-IIIB produced in H9 cells. The titer of the virus from this stock on human and chimpanzee peripheral blood mononuclear cells and in human lymphoid cell lines was determined; a cell culture infectivity of 10(4) was assigned. All chimpanzees inoculated intravenously with 40 cell culture infectious units or more became infected, as demonstrated by virus isolation and seroconversion. One of two chimpanzees inoculated with 4 cell culture infectious units became infected. Chimpanzees immunized with gp120 formulated in alum developed antibodies which precipitated gp120 and neutralized HTLV-IIIB. Peripheral blood mononuclear cells from gp120-vaccinated and HIV-infected animals showed a significantly greater response in proliferation assays with HIV proteins than did peripheral blood mononuclear cells from nonvaccinated and non-HIV-infected chimpanzees. Two of the gp120-alum-immunized chimpanzees were challenged with virus from the HTLV-IIIB stock. One animal received 400 cell culture infectious units, and one received 40 infectious units. Both animals became infected with HIV, indicating that the immune response elicited by immunization with gp120 formulated in alum was not effective in preventing infection with HIV-1.
Animals↗
Identification of simian immunodeficiency virus SIVMAC env gene products.
A monoclonal antibody recognizing an antigenic determinant on the env transmembrane protein, gp32 of simian immunodeficiency virus SIVMAC has been developed and designated SF8/5E11. The reactivity of this antibody was found to be type specific, since it did not cross-react with either SIVSMM or SIVMNe transmembrane proteins. The availability of both this antibody and the complete nucleotide sequence of SIVMAC allowed us to define the organization of the env gene products of this virus. Radiolabel sequencing of the amino termini of both gp160 and gp32 confirmed the positions of both cleavage sites predicted by alignment of the inferred amino acid sequences of the SIVMAC and human immunodeficiency virus type 1 env genes. The cleavage site between the signal peptide and the external env glycoprotein resides between the cysteine residue at position 21 and the threonine residue at position 22, starting from the first residue after the env gene initiator methionine. The env precursor polyprotein gp160 is cleaved between arginine 526 and glycine 527 to give rise to the external glycoprotein and the transmembrane of SIVMAC.
Animals↗
Proliferative T-cell response to HIV envelope glycoprotein in immunized and infected primates and human beings.
Human immunodeficiency virus (HIV)-specific helper T-cell response was studied in human subjects and nonhuman primates either infected with HIV or immunized with different HIV protein preparations. A strong group-specific T-cell response involving T-cell proliferation and lymphokine secretion was observed in immunized chimpanzees and rhesus monkeys as well as HIV-infected chimpanzees and gibbons. HIV-infected people demonstrated a low or no HIV-specific T-cell response. In contrast, five of 14 HIV antibody-negative sexual partners of HIV-infected men recognized one or more T-cell epitopes in the envelope glycoprotein of HIV.
Acquired Immunodeficiency Syndrome↗
AIDS-associated Kaposi's sarcoma: a molecular model for its pathogenesis.
Kaposi's sarcoma is a mesenchymal tumor that is frequently observed in AIDS patients. Although the tumor is clearly associated with HIV-1 infection, little is known about the molecular events underlying its pathogenesis. Here we consider the potential roles of specific cytokines and the virally encoded tat product in inducing the tumor.
Acquired Immunodeficiency Syndrome↗
Utilization of human hematopoietic cell lines for the propagation and characterization of HBLV (human herpesvirus 6).
In 1986, we reported the discovery and isolation of a novel human herpesvirus (HBLV) from AIDS and other lymphoproliferative disorders. Because HBLV is distinct from other members of the herpesvirus family and can infect B- and T-lymphocytes and other human cells (megakaryocytes and glioblastoma cells), we suggested human herpesvirus-6 (HHV-6) as the taxonomic designation for this virus. In cultures from patients' peripheral blood, the evidence of HBLV can be recognized from the appearance of short-lived giant cells (2-10%), which are large, refractile, and are often mono- and binucleated. As these cells degenerate, extracellular virus particles are found in the culture medium. HBLV can infect fresh mononuclear cells, established B- and T-lymphoblastoid cell lines, megakaryocytes and glioblastoma cell lines. HBLV infection can be detected by: a. morphological changes; b. indirect immunofluorescence assay, in situ hybridization, southern blot analysis, polymerase chain reaction amplification; and c. electron microscopy. Because of its wide cell tropism, HBLV DNA sequences have been detected in B-cell lymphomas and short term cultured cells from Sjogren's patients. Expression of HBLV RNA was also detected in sarcoidosis. The etiological role of HBLV in human tumors is unclear. While in vitro data may not necessarily apply to in vivo conditions, the infection of various cell lines from tumors and fresh mononuclear cells suggests HBLV involvement in a variety of diseases.
Cell Line↗
Use of Old World monkeys for acquired immunodeficiency syndrome research.
Mangabeys, macaques, and baboons persistently infected with human immunodeficiency virus (HIV)-2 NIH-DZ demonstrated no signs of immunodeficiency disease after 6-11 months following seroconversion. Thus Old World monkeys provide an animal model to investigate the effects of passive immunization (anti-HIV-2 antibodies) on HIV infection in primates.
Acquired Immunodeficiency Syndrome↗
Human T-lymphotropic virus I-infected T cells constitutively express lymphotoxin in vitro.
We have studied the pattern of expression of the lymphokines tumor necrosis factor (TNF alpha) and lymphotoxin (TNF beta) in T-cell lines established by transformation with human T-lymphotropic virus, type I (HTLV-I), the etiologic agent of adult T-cell leukemia (ATL). We report here that nine of nine HTLV-I-infected T-cell lines, established by in vitro infection with HTLV-I, including those with CD4+ or CD8+ as well as CD4-/CD8- phenotypes, constitutively produce high levels of TNF alpha and -beta mRNA and secrete biologically active TNF beta into the culture medium. Similar patterns of expression are seen in six of six HTLV-I-infected T-cell lines directly established from ATL patients. In contrast, several T-cell lines, either uninfected or infected with human immunodeficiency virus I, did not produce comparable levels of the TNF beta. Comparisons of a normal functional T-cell clone before and after infection with HTLV-I show that expression of TNF beta mRNA is induced in the infected cells. The high level expression in HTLV-I-infected cell lines dose not seem to involve perturbation of the TNF alpha/beta genetic loci by proviral integration. A cell line (81-66/45) nonproductively transformed with HTLV-I that produces tat-1 in the absence of viral structural proteins, produces both TNF alpha and -beta mRNA. This suggests that expression of these cytokines could be mediated in trans by the tat-1 gene product.
Blotting, Southern↗
HIV encephalopathy and lymphadenopathy: cells associated with viral antigens.
HIV associated subacute, micronodular encephalitis and lymphadenopathy were compared with regard to demonstrable HIV antigens and characterization of HIV antigen expressing cells. In the brain, both gag- and env-coded antigens were confined to cells of micronodular lesions with immunophenotype of monocyte/macrophages (KiM6+, 9.4+, CD4+/-). The micronodular lesions were also infiltrated by some lymphoid cells, predominantly CD8+. In lymphadenopathic nodes, gag-encoded antigens were demonstrable almost exclusively in follicles/germinal centres in association with follicular dendritic cells, whereas env-antigens usually were not found. Follicular involution was related to destruction of follicular dendritic cells and infiltration of CD8+ lymphocytes within the germinal centres. During follicular involution, HIV-gag antigens diminished in parallel with destruction of the network of dendritic reticulum cells. These findings indicate important tissue related differences in HIV expression. In addition, a possible participation of CD8+ cells in the pathogenesis of HIV induced tissue lesions is suggested.
AIDS-Related Complex↗
Utilization of human hematopoietic cell lines for the propagation and characterization of HBLV (human herpesvirus 6).
Details of the productive infection of established human cell lines of diverse origin by HBLV (also designated Human Herpesvirus 6) are described in this report. The infection and replication of HBLV in several T and B lymphoid and other cell lines was observed by electron microscopic examination, by the detection of viral antigen expression by indirect immunofluorescence assay (IFA) and by the presence of HBLV DNA by Southern blot hybridization. Several of these cell lines produced large amounts of virus. For this reason and because of the absence of other human herpesviruses, these lines have provided a valuable resource for the preparation of reagents and the development of assays for the detection and characterization of HBLV. The isolation and characterization of new HBLV isolates from patients with chronic fatigue syndrome were also facilitated by using some of the cell lines reported here. The host range of HBLV in established cell lines, therefore, does not appear to be limited to the B lymphocytes, as initially suggested by in vivo studies. The infection of T and B lymphocytes, megakaryocytes and neuronal cells in vitro suggests a need for the evaluation of diverse hematological and neurological disorders to shed light on a possible HBLV involvement.
Blotting, Southern↗
Angiogenic properties of Kaposi's sarcoma-derived cells after long-term culture in vitro.
Cells derived from lung biopsies and pleural effusions from AIDS patients with Kaposi's sarcoma (KS) of the lungs were established in long-term culture with the aid of conditioned medium from HTLV-II-transformed T cells (HTLV-II CM). These AIDS-KS cells were similar to the so-called spindle cells in KS lesions and had some of their features. They produced factors that supported their own growth (autocrine) and the growth of other cells (paracrine), including umbilical vein endothelium and fibroblasts. That the AIDS-KS cells also expressed potent angiogenic activity was demonstrated by the chorioallantoic membrane assay and by subcutaneous inoculation of AIDS-KS cells into nude mice, which resulted in the development of angiogenic lesions composed of mouse cells and showing histological features similar to those of human KS lesions. These data suggest that AIDS-associated KS and possibly other types of KS may be initiated by signals that induce the growth of particular cells (spindle cells of lymphatic or vascular origin) and the expression of autocrine and paracrine activities.
Acquired Immunodeficiency Syndrome↗
Kaposi's sarcoma cells: long-term culture with growth factor from retrovirus-infected CD4+ T cells.
Studies of the biology and pathogenesis of Kaposi's sarcoma (KS) have been hampered by the inability to maintain long-term cultures of KS cells in vitro. In this study AIDS-KS-derived cells with characteristic spindle-like morphology were cultured with a growth factor (or factors) released by CD4+ T lymphocytes infected with human T-lymphotropic virus type I or II (HTLV-I or HTLV-II) or with human immunodeficiency virus type 1 or 2 (HIV-1 or HIV-2). Medium conditioned by HTLV-II-infected, transformed lines of T cells (HTLV-II CM) contained large amounts of this growth activity and also supported the temporary growth of normal vascular endothelial cells, but not fibroblasts. Interleukin-1 and tumor necrosis factor-alpha stimulated the growth of the KS-derived cells, but the growth was only transient and these could be distinguished from that in HTLV-II CM. Other known endothelial cell growth promoting factors, such as acidic and basic fibroblast growth factors and epidermal growth factor, did not support the long-term growth of the AIDS-KS cells. The factor released by CD4+ T cells infected with human retroviruses should prove useful in studies of the pathogenesis of KS.
Acquired Immunodeficiency Syndrome↗
Cell-mediated immune response toward viral envelope and core antigens in gibbon apes (Hylobates lar) chronically infected with human immunodeficiency virus-1.
The specific cellular immune response toward envelope and core proteins of human immunodeficiency virus-1 (HIV-1) was investigated in gibbon apes chronically infected with the HTLV-IIIB isolate. After in vitro stimulation of PBMC from infected and control animals with HIV-1 Ag, DNA synthesis, IL-2R expression and IL-2 release were assayed. Cells from infected gibbon apes demonstrated a group-specific response toward whole virus preparations from three divergent HIV-1 isolates (HTLV-IIIB, HTLV-IIIRF, HTLV-IIIMN). Consistent responses were also detected against purified HIV-1 Ag, i.e., native gp120 envelope glycoprotein, recombinant gp160 glycoprotein, a synthetic peptide (peptide 7) representing a highly conserved region of gp120, and purified native core protein p24. In addition, lymphocytes from infected gibbon apes displayed a specific, MHC-restricted, cytotoxic activity against autologous cells expressing HIV-1 envelope or gag proteins. The specific T cell reactivity toward HIV-1 proteins observed in infected gibbons contrasts with findings in HIV-1 infected humans, and may help to explain the apparent discrepancy in the natural history of the infection between the two species.
Acquired Immunodeficiency Syndrome↗