RESUMEN
There is an ongoing question regarding the structure forming capabilities of water at ambient temperatures. To probe for different structures, we studied effects in pure water following magnetic field exposures corresponding to the ion cyclotron resonance of H3O(+). Included were measurements of conductivity and pH. We find that under ion cyclotron resonance (ICR) stimulation, water undergoes a transition to a form that is hydroxonium-like, with the subsequent emission of a transient 48.5 Hz magnetic signal, in the absence of any other measurable field. Our results indicate that hydronium resonance stimulation alters the structure of water, enhancing the concentration of EZ-water. These results are not only consistent with Del Giudice's model of electromagnetically coherent domains, but they can also be interpreted to show that these domains exist in quantized spin states.
Asunto(s)
Ciclotrones , Compuestos Onio/química , Agua/química , Campos Electromagnéticos , Hidrógeno/química , Concentración de Iones de Hidrógeno , Iones , Temperatura , Factores de Tiempo , VibraciónRESUMEN
We have analyzed the V beta usage by CD4+ and CD8+ T cells from human immunodeficiency virus (HIV)-infected individuals in response to an in vitro stimulation with the superantigenic erythrogenic toxin A (ETA) of Streptococcus pyogenes. ETA amplifies specifically CD4+ and CD8+ T cells from control donors expressing the V beta 8 and the V beta 12 elements. When peripheral T cells from asymptomatic HIV-infected individuals were stimulated with ETA, there was a complete lack of activation of the V beta 8+ T cell subset, whereas the V beta 12+ T cell subset responded normally to the superantigen. This V beta-specific anergy, which was also observed in response to staphylococcal enterotoxin E (SEE), affected both CD4+ and CD8+ T cells and represented an intrinsic functional defect rather than a specific lack of response to bacterial superantigens since it was also observed after a stimulation with V beta 8 monoclonal antibodies. The V beta 8 anergic T cells did not express interleukin 2 receptors (IL-2Rs) and failed to proliferate in response to exogenous IL-2 or IL-4, suggesting that this anergy was not a reversible process, at least by the use of these cytokines. The unresponsiveness of the V beta 8 T cell subset is frequent since it was found in 56% of the patients studied, and comparison of the clinical status of responder vs. anergic patients indicated that the only known common factor between them was HIV infection. In addition, it is noteworthy that the anergy of the V beta 8 subset may be a very early phenomenon since it was found in a patient at Centers for Disease Control stage I of the disease. These data provide evidence that a dominant superantigen may be involved in the course of HIV infection and that the contribution of HIV has to be considered.
Asunto(s)
Proteínas Bacterianas , Linfocitos T CD4-Positivos/inmunología , Anergia Clonal , Infecciones por VIH/inmunología , Proteínas de la Membrana , Linfocitos T Reguladores/inmunología , Adulto , Anticuerpos Monoclonales/inmunología , Antígenos Bacterianos/inmunología , Antígenos CD4 , Antígenos CD8 , Exotoxinas/inmunología , Femenino , Humanos , Masculino , Receptores de Interleucina-2/biosíntesis , Streptococcus pyogenes/inmunología , Superantígenos/inmunologíaRESUMEN
gp120 and CD4 are two glycoproteins that are considered to interact together to allow the binding of HIV to CD4+ cells. We have utilized enzymatic digestion by endoglycosidases in order to analyze N-linked carbohydrate chains of these proteins and their possible role in the interaction of gp120 or gp160 with CD4. SDS denaturation was not necessary to obtain optimal deglycosylation of either molecule, but deglycosylation of CD4, nonetheless, depended on the presence of 1% Triton X-100. Endo H and Endo F that cleave high mannose type and biantennary glycans diminish the molecular mass of the glycoproteins from 120 or 160 Kd to 90 or 130 Kd, respectively; but these enzymes had no action on CD4 glycans. Endo F N-glycanase mixture, which acts on all glycan species, including triantennary chains, led to complete deglycosylation of gp120/160 and of CD4. Therefore, probably half of the glycan moieties of gp120/160 are composed of high mannose and biantennary chains, the other half being triantennary species. The carbohydrate structures of CD4 seems to be triantennary chains. To analyze the binding of gp120/160 to CD4, we used a molecular assay in which an mAb (110-4) coupled to Sepharose CL4B allowed the attachment of soluble gp120/160 to the beads; 125I-sCD4 was then added to measure the binding of CD4 to different amounts of gp120/160. Binding to gp160 was not modified when using completely deglycosylated 125I-sCD4, while deglycosylation of gp120 or of gp160 resulted in the decrease of the binding to native CD4 by two- and fivefold, respectively. Native and deglycosylated gp120/160 bound to CD4+ cells with comparable affinities. In addition, deglycosylated gp120 displaced 125I-gp160 binding to CD4+ cells and inhibited fusion of fresh Molt-T4 cells with CEM HIV1- or HIV2-infected cells to the same extent. Taken together, these results indicate that carbohydrates of CD4 and of gp120/160 do not play a significant role in the in vitro interaction between these two molecules.
Asunto(s)
Antígenos de Diferenciación de Linfocitos T , VIH , Polisacáridos/fisiología , Receptores Virales/metabolismo , Proteínas de los Retroviridae/metabolismo , Proteínas del Envoltorio Viral/metabolismo , Conformación de Carbohidratos , Línea Celular , Electroforesis en Gel de Poliacrilamida , Glicósido Hidrolasas/metabolismo , Glicosilación , Proteína gp120 de Envoltorio del VIH , Proteínas gp160 de Envoltorio del VIH , Hexosaminidasas/metabolismo , Manosa/análisis , Manosil-Glicoproteína Endo-beta-N-Acetilglucosaminidasa , Peso Molecular , Receptores del VIH , Relación Estructura-Actividad , Linfocitos T/metabolismoRESUMEN
Using mAbs and genomic probe to the CD4 molecule, the HIV receptor, we demonstrated that HIV replication induces the disappearance of its functional receptor from the cell surface by two distinct mechanisms. First, after being expressed onto the cell surface, HIV envelope gp110 will complex CD4, efficiently masking the CD4 epitope used by the virus to bind its receptor. This phenomenon occurs on the surface of each infected cell and is not due to the release of soluble gp110; infection with recombinant HIV/vaccinia viruses expressing a mutated HIV env gene designed to prevent gp110 release from the cell surface induces a similar gp/CD4 complexes formation. Second, virus replication induces a dramatic and rapid loss of CD4 mRNA transcripts, preventing new CD4 molecules from being synthesized. These two mechanisms of receptor modulation could have been developed to avoid reinfection of cells replicating the virus as well as to produce more infectious particles. These results suggest that the classical virus interference documented for other retroviruses might not only be due to receptor/envelope interaction, but might also depend on receptor gene expression.
Asunto(s)
Membrana Celular/metabolismo , VIH/crecimiento & desarrollo , ARN Mensajero/metabolismo , Receptores Virales/metabolismo , Células Cultivadas , Regulación de la Expresión Génica , VIH/genética , Antígenos VIH/metabolismo , Receptores del VIH , Receptores Virales/genética , Recombinación Genética , Virus Vaccinia/genética , Proteínas del Envoltorio Viral/metabolismo , Replicación ViralRESUMEN
Interferon-alpha 1 from Escherichia coli transformed with a hybrid plasmid containing a human leukocyte complementary DNA insert, induces resistance to virus in appropriate target cells. It also shares the following properties with natural leukocyte interferon (IFN). (i) It enhances natural killing activity of human lymphocytes, (ii) it enhances antibody-dependent cell-mediated cytotoxicity, (iii) it suppresses antigen- and mitogen-induced leukocyte migration inhibition, (iv) it inhibits growth of IFN-sensitive Burkitt lymphoma cells. Since these activities are exhibited by a cloned protein species, they are due to IFN itself and not to other human proteins.
Asunto(s)
ADN Recombinante , Interferones/farmacología , Citotoxicidad Celular Dependiente de Anticuerpos/efectos de los fármacos , División Celular/efectos de los fármacos , Inhibición de Migración Celular , Clonación Molecular , Escherichia coli , Humanos , Hipersensibilidad Tardía/inmunología , Inmunidad Celular/efectos de los fármacos , Interferones/genética , Relación Estructura-ActividadRESUMEN
A retrovirus belonging to the family of recently discovered human T-cell leukemia viruses (HTLV), but clearly distinct from each previous isolate, has been isolated from a Caucasian patient with signs and symptoms that often precede the acquired immune deficiency syndrome (AIDS). This virus is a typical type-C RNA tumor virus, buds from the cell membrane, prefers magnesium for reverse transcriptase activity, and has an internal antigen (p25) similar to HTLV p24. Antibodies from serum of this patient react with proteins from viruses of the HTLV-I subgroup, but type-specific antisera to HTLV-I do not precipitate proteins of the new isolate. The virus from this patient has been transmitted into cord blood lymphocytes, and the virus produced by these cells is similar to the original isolate. From these studies it is concluded that this virus as well as the previous HTLV isolates belong to a general family of T-lymphotropic retroviruses that are horizontally transmitted in humans and may be involved in several pathological syndromes, including AIDS.
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida/microbiología , Retroviridae/aislamiento & purificación , Infecciones Tumorales por Virus/microbiología , Adulto , Animales , Anticuerpos Antivirales/inmunología , Células Cultivadas , Homólogo de la Proteína Chromobox 5 , Humanos , Masculino , Microscopía Electrónica , ADN Polimerasa Dirigida por ARN/metabolismo , Linfocitos T/microbiologíaRESUMEN
The presence of antibodies to lymphadenopathy-associated retrovirus (LAV) was determined by a radioimmunoprecipitation assay and by an enzyme-linked immunosorbent solid assay of sera from Zairian patients with the acquired immune deficiency syndrome (AIDS) in 1983. Thirty-five of 37 patients (94 percent) and 32 of 36 patients (88 percent), respectively, were seropositive by the two tests. In a control group of 26 patients, six (23 percent) showed positive results in these tests. Of these six control patients, five had clinically demonstrable infectious diseases and a low ratio of T4 to T8 lymphocytes. In addition, sera collected from a control group of Zairian mothers in 1980 were positive for LAV in 5 of 100 cases. Other serologic data suggest that LAV was present as early as 1977 in Zaire.
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida/inmunología , Anticuerpos Antivirales/análisis , Enfermedades Linfáticas/microbiología , Retroviridae/inmunología , República Democrática del Congo , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Masculino , Radioinmunoensayo , Linfocitos T Colaboradores-Inductores/citología , Linfocitos T Reguladores/citologíaRESUMEN
A strain of lymphadenopathy associated retrovirus ( LAV ) passaged in vitro was used to infect a lymphoblastoid cell line obtained by transformation with Epstein-Barr virus of B lymphocytes from a healthy donor. The virus produced from this line (B- LAV ) was also able to grow at a high rate in some other lymphoblastoid lines and in a Burkitt lymphoma line. This adapted strain retained the biochemical, ultrastructural, and antigenic characteristics of the original strain, as well as its tropism for normal T4+ lymphocytes. It is thus possible to grow LAV in large quantities that can be used for the preparation of diagnostic reagents. The interaction between such a human retrovirus and Epstein-Barr virus, a DNA virus, may have some implication for the pathology of the acquired immunodeficiency syndrome and related diseases.
Asunto(s)
Linfocitos B/microbiología , Herpesvirus Humano 4/metabolismo , Retroviridae/crecimiento & desarrollo , Replicación Viral , Síndrome de Inmunodeficiencia Adquirida/microbiología , Anticuerpos Monoclonales/inmunología , Línea Celular , Transformación Celular Viral , Deltaretrovirus/metabolismo , Humanos , Linfocitos T/microbiologíaRESUMEN
Lymphadenopathy-associated virus ( LAV ), a human T- lymphotrophic retrovirus isolated from a homosexual man with lymphadenopathy, has been causally associated with acquired immunodeficiency syndrome (AIDS). A sensitive and specific radioimmunoprecipitation test was developed for the detection of antibodies to the major core protein of LAV , p25 (molecular weight 25,000). Antibody to LAV p25 was found in the serum of 51 of 125 AIDS patients, 81 of 113 patients with lymphadenopathy syndrome, 0 of 70 workers at the Centers for Disease Control (some of whom had handled specimens from AIDS patients), and 0 of 189 random blood donors. Of a group of 100 homosexual men from San Francisco whose serum was obtained in 1978, only one had antibody to LAV p25; in contrast, of a group of 50 homosexual men in the same community whose serum was obtained in 1984, 12 had antibodies to LAV p25.
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida/microbiología , Anticuerpos Antivirales/inmunología , Infecciones por Retroviridae/inmunología , Retroviridae/inmunología , Proteínas Virales/inmunología , Síndrome de Inmunodeficiencia Adquirida/inmunología , Donantes de Sangre , Deltaretrovirus/inmunología , Homosexualidad , Humanos , MasculinoRESUMEN
The increasing incidence of a variety of cancers after the Second World War confronts scientists with the question of their origin. In Western countries, expansion and ageing of the population as well as progress in cancer detection using new diagnostic and screening tests cannot fully account for the observed growing incidence of cancer. Our hypothesis is that environmental factors play a more important role in cancer genesis than it is usually agreed. (1) Over the last 2-3 decades, alcohol consumption and tobacco smoking in men have significantly decreased in Western Europe and North America. (2) Obesity is increasing in many countries, but the growing incidence of cancer also concerns cancers not related to obesity nor to other known lifestyle-related factors. (3) There is evidence that the environment has changed over the time period preceding the recent rise in cancer incidence, and that this change, still continuing, included the accumulation of many new carcinogenic factors in the environment. (4) Genetic susceptibility to cancer due to genetic polymorphism cannot have changed over one generation and actually favours the role of exogenous factors through gene-environment interactions. (5) Age is not the unique factor to be considered since the rising incidence of cancers is seen across all age categories, including children, and adolescents. (6) The fetus is specifically vulnerable to exogenous factors. A fetal exposure during a critical time window may explain why current epidemiological studies may still be negative in adults. We therefore propose that the involuntary exposure to many carcinogens in the environment, including microorganisms (viruses, bacteria and parasites), radiations (radioactivity, UV and pulsed electromagnetic fields) and many xenochemicals, may account for the recent growing incidence of cancer and therefore that the risk attributable to environmental carcinogen may be far higher than it is usually agreed. Of major concern are: outdoor air pollution by carbon particles associated with polycyclic aromatic hydrocarbons; indoor air pollution by environmental tobacco smoke, formaldehyde and volatile organic compounds such as benzene and 1,3 butadiene, which may particularly affect children and food contamination by food additives and by carcinogenic contaminants such as nitrates, pesticides, dioxins and other organochlorines. In addition, carcinogenic metals and metalloids, pharmaceutical medicines and some ingredients and contaminants in cosmetics may be involved. Although the risk fraction attributable to environmental factors is still unknown, this long list of carcinogenic and especially mutagenic factors supports our working hypothesis according to which numerous cancers may in fact be caused by the recent modification of our environment.
Asunto(s)
Contaminantes Ambientales/toxicidad , Estilo de Vida , Neoplasias/inducido químicamente , Envejecimiento/fisiología , Contaminantes Atmosféricos/toxicidad , Consumo de Bebidas Alcohólicas/efectos adversos , Niño , Dieta , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos , Ejercicio Físico/fisiología , Contaminación de Alimentos , Humanos , Leucemia/epidemiología , Esperanza de Vida , Neoplasias/epidemiología , Neoplasias/genética , Obesidad/complicaciones , Enfermedades Profesionales/epidemiología , Virus Oncogénicos , Sobrepeso/complicaciones , Fumar/efectos adversosRESUMEN
An epithelioid variant, which forms characteristic colonies in soft agar medium, has been isolated from the BHK21/13 line of hamster fibroblasts. After infection with a high multiplicity of polyoma virus, a fraction of the cells is morphologically transformed. Transformation may be demonstrated in two ways: 1) When infected cells are grown on glass, dense colonies of transformed cells develop from the confluent monolayer of untransformed cells in which multiplication has stopped because of their contiguity. 2) In agar, colonies derived from transformed cells are larger and morphologically distinct from those formed by the untransformed cells. The variant cells produce tumors in hamsters. Their transplantability is increased by polyoma transformation. Transformed cells possess a new polyoma-specific cell antigen.
Asunto(s)
Antígenos Virales , Efecto Citopatogénico Viral , Células Epitelioides/patología , Células Epitelioides/virología , Poliomavirus , Animales , Línea Celular , Transformación Celular Neoplásica , Cricetinae , Células Epitelioides/inmunología , Fibroblastos/inmunología , Fibroblastos/patología , Fibroblastos/virología , Infecciones por Polyomavirus/patología , Infecciones Tumorales por Virus/patologíaRESUMEN
HIV infection is associated with immune activation, which in turn stimulates HIV replication. Certain other co-infections cause immune activation and may contribute to an increased viral load. The impact of co-infections by microorganisms such as Mycobacterium tuberculosis can be important for patient survival, particularly those at high risk of exposure to infection and with poor access to medical care.
Asunto(s)
Infecciones Oportunistas Relacionadas con el SIDA/inmunología , Infecciones por VIH/etiología , Infecciones Oportunistas Relacionadas con el SIDA/virología , Animales , Fármacos Anti-VIH/farmacología , Antígenos/administración & dosificación , VIH/inmunología , VIH/patogenicidad , Infecciones por VIH/inmunología , Infecciones por VIH/virología , Humanos , Sistema Inmunológico/efectos de los fármacos , Tuberculosis/complicaciones , Tuberculosis/inmunología , Tuberculosis/virología , Replicación ViralRESUMEN
We have recently described in BHK cells a plasma membrane protein of molecular weight 177,000, which is significantly increased in Hamster Sarcoma Virus-transformed cells (Lage-Davila, A. and Montagnier, L. (1977) Biochem. Biophys. Res. Commun. 79, 577--584). We present now a study of proteins from purified plasma membrane fractions in the same pair of clones. Solubilization conditions, cross-linking experiments, metabolic labelling and enzymatic radioiodination allow to characterize this 177,000 transformation-increased protein as an integral membrane glycoprotein partially exposed at the outer cell surface. Additional information on other membrane proteins in this system is also given.
Asunto(s)
Transformación Celular Viral , Proteínas de la Membrana/metabolismo , Virus Oncogénicos , Animales , Línea Celular , Membrana Celular/metabolismo , Cobre , Cricetinae , Glucosamina/metabolismo , Yoduros/metabolismo , Mercaptoetanol , Metionina/metabolismo , Peso Molecular , Fenantrolinas , PolietilenglicolesAsunto(s)
Enfermedad Crítica/terapia , Estrés Oxidativo , Especies Reactivas de Oxígeno/metabolismo , Lesión Renal Aguda/complicaciones , Antioxidantes/metabolismo , Antioxidantes/uso terapéutico , Biomarcadores/sangre , Quemaduras/complicaciones , Cuidados Críticos , Humanos , Hiperglucemia/complicaciones , Macrófagos/metabolismo , Enfermedades Mitocondriales/prevención & control , Neutrófilos/metabolismo , Síndrome de Dificultad Respiratoria/complicaciones , Sepsis/complicacionesRESUMEN
The V2 region of simian immunodeficiency virus (SIV) and V3 region of human immunodeficiency virus type 1 (HIV-1) have been reported to be neutralization epitopes. We analysed the corresponding regions in HIV-2. Synthetic peptides modeling the V2 (aa 149-168) and V3 (CV3: aa 298-315 and NV3: aa 306-324) regions of the HIV-2 external envelope glycoprotein were coupled to KLH and used as immunogens in rabbits. We characterized the resulting antiV2 and antiV3 antibodies for their ability to recognize native and deglycosylated HIV-2 envelope glycoprotein, to block gp-CD4 interaction and to inhibit syncytium formation in vitro. The three synthetic peptides induced antibodies able to recognize specifically the native HIV-2 envelope glycoprotein with a significant avidity (K0.5 between 6 x 10(-7) and 8 x 10(-9) M). Interestingly, the reactivity of antibodies produced against the V2 peptide, which contains two potential sites of N-glycosylation, was higher against the fully deglycosylated than glycosylated HIV-2 external envelope glycoprotein (gp105). The antipeptide antibodies were used to investigate the topography of these regions in the preformed gp-CD4 complex in indirect immunofluorescence assays. The V2 and V3 regions in the complex remained accessible to their respective antibodies. Moreover, preincubation of gp105 with anti V2 or anti V3 antibodies did not prevent gp-CD4 interaction. Thus the V2 and V3 regions are not directly involved in the gp105 binding site for the CD4 receptor. Finally, in contrast with results obtained with antibodies produced against the V3 region of HIV-1 gp120 and monoclonal antibodies produced against the V3 of SIV, antibodies produced against V2 and V3 of HIV-2 were unable to inhibit syncytium formation induced by HIV-2 in vitro.
Asunto(s)
Especificidad de Anticuerpos , Productos del Gen env/inmunología , Anticuerpos Anti-VIH/inmunología , Proteína gp120 de Envoltorio del VIH/inmunología , VIH-2/inmunología , Fragmentos de Péptidos/inmunología , Precursores de Proteínas/inmunología , Secuencia de Aminoácidos , Animales , Antígenos CD4/inmunología , Datos de Secuencia Molecular , Conejos , Virus de la Inmunodeficiencia de los Simios/inmunología , Productos del Gen env del Virus de la Inmunodeficiencia HumanaRESUMEN
HIV-1 infected Molt4-T4 cells provide an efficient system for the production of cellular precursor gp160 of HIV envelope glycoproteins, gp120 and gp41. The precursor gp160 was purified on an immuno-affinity column containing antibodies from sera of HIV-1-seropositive patients. The precursor gp160 was then isolated by preparative polyacrylamide gel electrophoresis. Two out of four Balb/c mice, immunized with these purified preparations of gp160, developed specific circulating antibodies. A hybridoma cell line was subsequently isolated producing monoclonal antibody KL49/19 (IgG1, K) specific for gp160. This monoclonal antibody can specifically immunoprecipitate gp160, existing in HIV-1-infected cells. In an immunoblotting assay, it identifies mainly gp160 and shows a slight affinity for the mature glycoprotein, gp120. The monoclonal antibody is probably directed against an epitope in the polypeptide residue of gp160 since it can recognize a deglycosylated polypeptide of molecular weight 90,000, a product of gp160 digestion by endoglycosidase H (Endo H). It does not cross-react with any protein of HIV-2 by immunoblot or immunoprecipitation assays. By virtue of its specificity, the monoclonal antibody KL49/19 might provide a powerful probe with which to detect gp160 in cells which might partially express the HIV-1 genes.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Anticuerpos Antivirales/inmunología , VIH/inmunología , Proteínas de los Retroviridae/inmunología , Animales , Especificidad de Anticuerpos , Línea Celular , Anticuerpos Anti-VIH , Proteína gp120 de Envoltorio del VIH , Humanos , Precursores de Proteínas/inmunologíaRESUMEN
With a view to obtaining human monoclonal antibodies against HIV-1 antigens we used the Epstein-Barr virus immortalization technique to induce lymphoblastoid cell lines from peripheral blood lymphocytes of 10 people who were seropositive for HIV-1 and had no clinical symptoms. A number of polyclonal lines were obtained which synthesized antibodies against most of the major proteins and glycoproteins of HIV-1. Three stable clones were characterized for class, secretion characteristics and specificity. Two of these clones produce antibodies which react with gp41, and the third reacts with p25. One of the anti-gp41 antibodies was found to have neutralizing activity.
Asunto(s)
Anticuerpos Monoclonales/biosíntesis , Anticuerpos Anti-VIH/biosíntesis , Proteína gp41 de Envoltorio del VIH/inmunología , Seropositividad para VIH/inmunología , VIH-1/inmunología , Proteínas del Núcleo Viral/inmunología , Secuencia de Aminoácidos , Especificidad de Anticuerpos , Linfocitos B/inmunología , Western Blotting , Línea Celular Transformada , Homólogo de la Proteína Chromobox 5 , Ensayo de Inmunoadsorción Enzimática , Técnica del Anticuerpo Fluorescente , Anticuerpos Anti-VIH/análisis , Anticuerpos Anti-VIH/inmunología , Antígenos VIH/inmunología , Herpesvirus Humano 4/fisiología , Humanos , Inmunoglobulina G/biosíntesis , Inmunoglobulina G/inmunología , Datos de Secuencia Molecular , Ensayo de RadioinmunoprecipitaciónRESUMEN
We expressed peptides from the HIV-1 envelope protein at the surface of Escherichia coli by genetic insertions into an exposed loop of the outer membrane protein LamB. Recombinant bacteria expressing eight peptides from gp110 (pep1-pep8), conserved between HIV-1 and HIV-2, were used as live immunogens in rabbits by the intravenous route. The eight constructions elicited anti-LamB antibodies, showing that the hybrid proteins were immunogenic. One of them, LamB-pep8, gave rise to antibodies able to react with gp160 and to neutralize HIV-1 in vitro. We also show that this type of recombinant E. coli can provide a convenient reagent to monitor and characterize specific antibodies. Recombinant clones were used to test sera of seropositive individuals, as well as to narrow down the monoclonal antibody 110-1 recognition site to a cluster of eight residues at the carboxy-terminal end of gp110.
Asunto(s)
Escherichia coli/genética , Antígenos VIH/inmunología , VIH-1/inmunología , Proteínas del Envoltorio Viral/inmunología , Secuencia de Aminoácidos , Animales , Anticuerpos Monoclonales/inmunología , Proteínas de la Membrana Bacteriana Externa , Western Blotting , Productos del Gen env/genética , Productos del Gen env/inmunología , Antígenos VIH/genética , Proteínas gp160 de Envoltorio del VIH , Datos de Secuencia Molecular , Pruebas de Neutralización , Porinas , Precursores de Proteínas/genética , Precursores de Proteínas/inmunología , Conejos , Receptores Virales/inmunología , Recombinación Genética , Proteínas del Envoltorio Viral/genéticaRESUMEN
We have investigated the possible involvement in the interaction between HIV gp110 and its CD4 receptor of epitopes different from the currently known binding site(s) of the molecule. Four monoclonal antibodies (MAbs) to gp110 were used (Genetic Systems Corporation, Seattle, Washington, USA): one (110-1) recognized a peptide corresponding to the C-terminal part of gp110 (494-517); the other three (110-3, 110-4, 110-5) recognized the same peptide located at position 308-328. HIV or purified gp110 obtained from a vaccinia recombinant (Transgene S.A., Strasbourg, France) were pre-incubated with the MAb prior to addition to CD4+ cells. Specific binding of viral particles or of the soluble molecule was then determined by flow cytometer analysis, compared with that of control preparations where the MAb was added after HIV or gp 110 had been allowed to bind CD4+ cells. Significant inhibition of HIV binding was noted with the three MAbs to peptide (308-328), but not with 110-1. At the molecular level, these same MAbs decreased the affinity of interaction between CD4 and soluble gp110, although they could still label the latter molecule after it had bound to CD4+ cells. Therefore, steric hindrance may account for neutralization of HIV binding by antibodies that are actually directed to epitopes topographically distinct from the site of binding of gp110 to CD4.
Asunto(s)
Anticuerpos Monoclonales/inmunología , VIH/metabolismo , Glicoproteínas de Membrana/metabolismo , Receptores Virales/metabolismo , Epítopos , Humanos , Glicoproteínas de Membrana/inmunología , Receptores del VIHRESUMEN
The cellular receptor for HIV-1 is the leucocyte differentiation antigen, CD4. Blocking of HIV-1 infectivity can be achieved with monoclonal antibodies (MAbs) to some, but not all epitopes of this antigen. We demonstrate here, by inhibition of virus infection, blocking of syncytium formation and inhibition of pseudotype infection with a panel of CD4 MAbs, that HIV-1, HIV-2 and simian immunodeficiency virus (SIV) isolates share the same cellular receptor, the CD4 glycoprotein. It is also shown that very similar epitopes of this molecule are involved in virus binding. We infer from these data that the binding sites on these viruses are highly conserved regions, and may therefore make good targets for potential vaccines. In addition, we show that cell surface expression of CD4 is similarly modulated after infection of cell lines by all the viruses.