RESUMEN
The adenovirus death protein (ADP) is expressed at late times during a lytic infection of species C adenoviruses. ADP promotes the release of progeny virus by accelerating the lysis and death of the host cell. Since some human lymphocytes survive while maintaining a persistent infection with species C adenovirus, we compared ADP expression in these cells with ADP expression in lymphocytes that proceed with a lytic infection. Levels of ADP were low in KE37 and BJAB cells, which support a persistent infection. In contrast, levels of ADP mRNA and protein were higher in Jurkat cells, which proceed with a lytic infection. Epithelial cells infected with an ADP-overexpressing virus died more quickly than epithelial cells infected with an ADP-deleted virus. However, KE37, and BJAB cells remained viable after infection with the ADP-overexpressing virus. Although the levels of ADP mRNA increased in KE37 and BJAB cells infected with the ADP-overexpressing virus, the fraction of cells with detectable ADP was unchanged, suggesting that the control of ADP expression differs between epithelial and lymphocytic cells. When infected with an ADP-deleted adenovirus, Jurkat cells survived and maintained viral DNA for greater than 1 month. These findings are consistent with the notion that the level of ADP expression determines whether lymphocytic cells proceed with a lytic or a persistent adenovirus infection.
Asunto(s)
Infecciones por Adenoviridae/virología , Proteínas E3 de Adenovirus/metabolismo , Adenovirus Humanos/metabolismo , Linfocitos/virología , Proteínas E3 de Adenovirus/genética , Adenovirus Humanos/genética , Línea Celular , Humanos , Liberación del Virus , Replicación ViralRESUMEN
Rabbit antisera to a mouse testicular teratoma, absorbed with normal mouse tissues, react by immunofluorescence with plasma membrane antigens of a variety of transplantable mouse tumor cells and transformed fibroblast cell lines including Clone 1D, SV-40-3T3, and 3T12. Trypsin treatment of cells of "normal" lines, 3T3 and FR-SV-3T3, uncovers reactivity on these as well. Early passage mouse embryo fibroblast cell cultures do not react even after trypsinization. By cross-absorbtion studies, the anti-teratoma serum appears to react with an antigen common to most tumor cells investigated thus far. When this antigen on Clone 1D cells is "capped," H-2 antigens collect with the teratoma antigens in the cap indicating a physical association between the molecules. Molecules specified by both the H-2D and H-2K regions are bound to the teratoma antigens in the Clone 1D plasma membrane. This antigen is also found in soluble tumor cell fractions where it is believed to be free of H-2. A second cell surface antigen defined by anti-teratoma serum is expressed only by hepatoma and teratoma itself. This second antigen is apparently a secretory product of teratoma cells. A third surface antigen defined by anti-teratoma serum appears to be specific for the teratoma.
Asunto(s)
Antígenos/análisis , Membrana Celular/inmunología , Antígenos de Histocompatibilidad/análisis , Teratoma/inmunología , Neoplasias Testiculares/inmunología , Animales , Reacciones Antígeno-Anticuerpo , Antígenos de Neoplasias , Antígenos Virales , Células Cultivadas , Técnica del Anticuerpo Fluorescente , Sueros Inmunes , Masculino , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratones Endogámicos , Neoplasias Experimentales/inmunología , Conejos , Virus 40 de los Simios , Tripsina/farmacologíaRESUMEN
BACKGROUND: In search of a proposed viral aetiology of childhood acute lymphoblastic leukaemia (ALL), the common species C adenoviruses were analysed in Guthrie cards. METHODS: Guthrie cards from 243 children who later developed ALL and from 486 matched controls were collected and analysed by nested polymerase chain reaction for the presence of adenovirus DNA. RESULTS: Adenovirus DNA was reliably detected from only two subjects, both of whom developed ALL. CONCLUSION: Adenovirus DNA is detected in Guthrie card samples at too low a frequency to reveal an association between adenovirus and the development of leukaemia.
Asunto(s)
Infecciones por Adenoviridae/virología , Adenoviridae/genética , ADN Viral/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/virología , Infecciones por Adenoviridae/diagnóstico , Infecciones por Adenoviridae/genética , Adolescente , Estudios de Casos y Controles , Niño , Preescolar , Femenino , Humanos , Lactante , Masculino , Reacción en Cadena de la Polimerasa , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , PronósticoRESUMEN
Although species C human adenoviruses establish persistent infections, the molecular details of this lifestyle remain poorly understood. We previously reported that adenovirus DNA is found in human mucosal T lymphocytes in a noninfectious form (C. T. Garnett, D. Erdman, W. Xu, and L. R. Gooding, J. Virol. 76:10608-10616, 2002). In this study, human tonsil and adenoid tissues were analyzed to determine the dynamics of infection, the rate of clearance of viral DNA, and the possibility of reactivation of virus from these tissues. The presence of viral DNA peaked at 4 years of age and declined thereafter. The average number of viral genomes declined with the age of the donor. The frequency of virus-bearing cells ranged from 3 x 10(-7) to 3.4 x 10(-4), while the amount of viral DNA per cell varied less, with an average of 280 copies per cell. All species C serotypes were represented in these tissues, although adenovirus type 6 was notably rare. Infectious virus was detected infrequently (13 of 94 of donors tested), even among donors with the highest levels of adenoviral DNA. Adenovirus transcripts were rarely detected in uncultured lymphocytes (2 of 12 donors) but appeared following stimulation and culture (11 of 13 donors). Viral DNA replication could be stimulated in most donor samples by lymphocyte stimulation in culture. New infectious virus was detected in 13 of 15 donors following in vitro stimulation. These data suggest that species C adenoviruses can establish latent infections in mucosal lymphocytes and that stimulation of these cells can cause viral reactivation resulting in RNA transcription, DNA replication, and infectious virus production.
Asunto(s)
Adenoviridae/aislamiento & purificación , Infecciones por Adenovirus Humanos/virología , Portador Sano/virología , ADN Viral/aislamiento & purificación , Tonsila Palatina/virología , Latencia del Virus , Adenoviridae/clasificación , Adenoviridae/fisiología , Adolescente , Factores de Edad , Células Cultivadas , Niño , Preescolar , Humanos , Linfocitos/virología , Serotipificación , Activación ViralRESUMEN
A quantitative assay was used to measure the rate of collection of a population of embryonic neural retina cells to the surface of cell aggregates. The rate of collection of freshly trysinized cells was limited in the initial stages by the rate of replacement of trypsin-sensitive cell- surface components. When cells were preincubated, or "recovered," and then added to cell aggregates, collection occurred at a linear rate and was independent of protein and glycoprotein synthesis. The adhesion of recovered cells was temperature and energy dependent, and was reversibly inhibited by cytochalasin B. Colchicine had little effect on collection of recovered cells. Antiserum directed against recovered cell membranes was shown to bind to recovered cells by indirect immunofluorescence. The antiserum also was shown to inhibit collection of recovered cells to aggregates, suggesting that at least some of the antigens identified might be involved in the adhesion process. The inhibitory effect of the antiserum was dose dependent . Freshly trypsinized cells absorbed neither the immunofluorescence activity nor the adhesion-inhibiting activity. Recovered cells absorbed away both activities. In specificity studies, dorsal neural retina cells adhered to aggregates of ventral optic tectum in preference to aggregates of dorsal optic tectum. The adhesive specificity of the dorsal retina cells was less sensitive to trypsin than the adhesive specificity of ventral retina cells which adhered preferentially to dorsal tectal aggregates only after a period of recovery.
Asunto(s)
Adhesión Celular , Membrana Celular/fisiología , Retina/citología , Animales , Antígenos , Adhesión Celular/efectos de los fármacos , Membrana Celular/efectos de los fármacos , Membrana Celular/inmunología , Células Cultivadas , Embrión de Pollo , Colchicina/farmacología , Cianuros/farmacología , Cicloheximida/farmacología , Citocalasina B/farmacología , Diazooxonorleucina/farmacología , Dinitrofenoles/farmacología , Colículos Superiores/citología , Tripsina/farmacologíaRESUMEN
The adenovirus E1A and E1B proteins are required for transformation of primary rodent cells. When expressed in the absence of the 19,000-dalton (19K) E1B protein, however, the E1A proteins are acutely cytotoxic and induce host cell chromosomal DNA fragmentation and cytolysis, analogous to cells undergoing programmed cell death (apoptosis). E1A alone can efficiently initiate the formation of foci which subsequently undergo abortive transformation whereby stimulation of cell growth is counteracted by continual cell death. Cell lines with an immortalized growth potential eventually arise with low frequency. Coexpression of the E1B 19K protein with E1A is sufficient to overcome abortive transformation to produce high-frequency transformation. Like E1A, the tumoricidal cytokine tumor necrosis factor alpha (TNF-alpha) evokes a programmed cell death response in many tumor cell lines by inducing DNA fragmentation and cytolysis. Expression of the E1B 19K protein by viral infection, by transient expression, or in transformed cells completely and specifically blocks this TNF-alpha-induced DNA fragmentation and cell death. Cosegregation of 19K protein transforming activity with protection from TNF-alpha-mediated cytolysis demonstrates that both activities are likely the consequence of the same function of the protein. Therefore, we propose that by suppressing an intrinsic cell death mechanism activated by TNF-alpha or E1A, the E1B 19K protein enhances the transforming activity of E1A and enables adenovirus to evade TNF-alpha-dependent immune surveillance.
Asunto(s)
Adenovirus Humanos/fisiología , Muerte Celular , Transformación Celular Viral , Proteínas Oncogénicas Virales/fisiología , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Proteínas Precoces de Adenovirus , Animales , Cricetinae , Efecto Citopatogénico Viral , Regulación hacia Abajo , Células HeLa , Humanos , Técnicas In Vitro , Receptores de Superficie Celular/metabolismo , Receptores del Factor de Necrosis TumoralRESUMEN
We have used immunoprecipitation of radioactivity from labeled cell membrane preparations to study further the association between the histocompatibility-2 (H-2) peptides and a tumor-associated embryonic antigen (Ag I) on the surface of L-cells. Since earlier studies had shown that Ag I and H-2 co-cap on L-cells, the present study was initiated to test the hypothesis that Ag I presents an altered H-2 peptide expressed on transformed cells. Indeed, sodium dodecyl sulfate-gel electrophoresis of Ag I shows two major peaks at approximately 110,000 and 40,000 daltons, the latter co-migrating with the major H-2 peptide. However, further analysis using sequential immunoprecipitation indicates that the molecules reacting with antisera to H-2 and to Ag I are separate entities, precipitating independently of one another. This has led us to postulate that Ag I and H-2 participate in a unique kind of association in the cell membrane, that is, that the bonds between them which allow them to co-cap may be in the lipid bilayer itself, and that these bonds are distrupted upon solubilization of the membrane.
Asunto(s)
Antígenos de Neoplasias , Feto/inmunología , Células L/inmunología , Animales , Membrana Celular/inmunología , Transformación Celular Neoplásica , Antígenos de Histocompatibilidad , Ratones , Teratoma/inmunologíaRESUMEN
Mucosal lymphocytes support latent infections of species C adenoviruses. Because infected lymphocytes resist re-infection with adenovirus, we sought to identify changes in cellular gene expression that could inhibit the infectious process. The expression of over 30,000 genes was evaluated by microarray in persistently infected B-and T-lymphocytic cells. BBS9, BNIP3, BTG3, CXADR, SLFN11 and SPARCL1 were the only genes differentially expressed between mock and infected B cells. Most of these genes are associated with oncogenesis or cancer progression. Histone deacetylase and DNA methyltransferase inhibitors released the repression of some of these genes. Cellular and viral gene expression was compared among leukemic cell lines following adenovirus infection. Childhood leukemic B-cell lines resist adenovirus infection and also show reduced expression of CXADR and SPARCL. Thus adenovirus induces limited changes to infected B-cell lines that are similar to changes observed in childhood leukemic cell lines.
Asunto(s)
Infecciones por Adenovirus Humanos/complicaciones , Infecciones por Adenovirus Humanos/virología , Adenovirus Humanos/fisiología , Regulación Leucémica de la Expresión Génica , Leucemia/etiología , Latencia del Virus , Adulto , Factores de Edad , Linfocitos B/metabolismo , Linfocitos B/patología , Linfocitos B/virología , Línea Celular Tumoral , Niño , Análisis por Conglomerados , Perfilación de la Expresión Génica , Humanos , Leucemia de Células B/etiologíaRESUMEN
One potential limitation of adenovirus (Ad)-based vectors for the gene therapy of cystic fibrosis (CF) and other genetic diseases is the transience of expression observed in most in vivo systems. In this study, the influence of various factors on persistence of transgene expression in the lung was investigated. In the absence of immune pressure, such as in the nude mouse, the genomic structure of the vector was found to be predominant in determining the persistence of expression; Ad vector constructs with an E1-E3+E4ORF6+ backbone encoding beta-galactosidase (beta-Gal) or the cystic fibrosis transmembrane conductance regulator (CFTR) produced declining levels of expression while an Ad/CMV beta Gal vector with an E1-E3+E4+ backbone gave rise to sustained, long-term reporter gene expression. The ability of the latter vector to persist was in turn limited in part by the presence of cytotoxic T lymphocytes (CTLs). Adoptive transfer experiments indicated that CTLs directed against either viral proteins or the beta-Gal reporter gene product were able to reduce expression in nude C57BL/6 mice stably expressing beta-Gal from the E4+ vector. Finally, the specificity and strength of the CTL response elicited by Ad vector was found to vary considerably depending on mouse strain haplotype. These results indicate that persistence of transgene expression in a given system is determined by the interplay between several factors including genomic structure of the vector, host background, and immune response.
Asunto(s)
Adenoviridae/genética , Regulación de la Expresión Génica/genética , Técnicas de Transferencia de Gen , Animales , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , ADN Recombinante , ADN Viral/genética , ADN Viral/metabolismo , Genes Reporteros/genética , Terapia Genética , Vectores Genéticos/genética , Vectores Genéticos/metabolismo , Haplotipos/genética , Pulmón/metabolismo , Ratones , Ratones Endogámicos , Ratones Desnudos , Linfocitos T Citotóxicos/inmunología , Linfocitos T Citotóxicos/metabolismo , Proteínas Virales/inmunología , Proteínas Virales/metabolismo , beta-Galactosidasa/genética , beta-Galactosidasa/metabolismoRESUMEN
Using digitonin permeabilization to assay mitochondrial electron transfer, we have found that respiratory activity (succinoxidase and cytochrome oxidase) in three mouse fibroblast lines is completely eliminated by incubation with human recombinant tumor necrosis factor-alpha (hrTNF). As with cytotoxicity, hrTNF-induced mitochondrial dysfunction occurs in resistant cells upon inhibition of protein synthesis, whereas sensitive cells exhibit spontaneous respiratory inhibition. In C3HA cells, inhibition is detectable 1.5-2 h after hrTNF addition, preceding cell lysis by at least 5 h (as measured by dye exclusion), and is approximately coincidental with morphological changes we have previously reported for this cell line. LM cells also exhibit inhibition of electron transfer, coincidental with morphological changes. These results suggest that bioenergetic dysfunction may be involved in the cytotoxic mechanism of TNF.
Asunto(s)
Mitocondrias/efectos de los fármacos , Factor de Necrosis Tumoral alfa/farmacología , Animales , Línea Celular , Permeabilidad de la Membrana Celular/efectos de los fármacos , Transporte de Electrón/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Humanos , Ratones , Mitocondrias/fisiología , Consumo de Oxígeno/efectos de los fármacos , Biosíntesis de ProteínasAsunto(s)
Proteínas E1A de Adenovirus/fisiología , Adenovirus Humanos/inmunología , Apoptosis/inmunología , Adenovirus Humanos/química , Caspasa 2 , Caspasas/fisiología , Humanos , Receptores del Factor de Necrosis Tumoral/metabolismo , Transducción de Señal/inmunología , Linfocitos T Citotóxicos/inmunología , Factor de Necrosis Tumoral alfa/fisiología , Receptor fasAsunto(s)
Adenovirus Humanos/inmunología , Presentación de Antígeno/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Proteínas E1A de Adenovirus/inmunología , Proteínas E3 de Adenovirus/química , Proteínas E3 de Adenovirus/inmunología , Animales , Regulación hacia Abajo , HumanosAsunto(s)
Adenoviridae/inmunología , Adenoviridae/genética , Adenoviridae/patogenicidad , Proteínas Precoces de Adenovirus , Animales , Transporte Biológico , Transformación Celular Viral , Citotoxicidad Inmunológica/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Humanos , Interferones/fisiología , Proteínas Oncogénicas Virales/inmunología , ARN Viral/fisiología , Factor de Necrosis Tumoral alfa/fisiologíaAsunto(s)
Antígenos de Histocompatibilidad/análisis , Inmunidad Celular , Inmunidad , Linfocitos T/inmunología , Animales , Reacciones Antígeno-Anticuerpo , Unión Competitiva , Pruebas Inmunológicas de Citotoxicidad , Sueros Inmunes , Leucemia Experimental/inmunología , Ratones , Ratones Endogámicos BALB CRESUMEN
Epidemiological evidence suggests that childhood acute lymphoblastic leukaemia (ALL) may be initiated by an in infection in utero. Adenovirus DNA was detected in 13 of 49 neonatal blood spots from ALL patients but only in 3 of 47 controls (P=0.012) suggesting a correlation between prenatal adenovirus infection and the development of ALL.
Asunto(s)
Infecciones por Adenoviridae/complicaciones , Adenoviridae/genética , ADN Viral/aislamiento & purificación , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras/etiología , Adenoviridae/aislamiento & purificación , Adenoviridae/patogenicidad , Estudios de Casos y Controles , Niño , HumanosRESUMEN
Human adenoviruses are among a growing number of human, animal, and even plant viruses that encode gene products that interfere with the defense mechanisms of the host. The host mechanisms most often affected by these viral products are the innate and inflammatory responses such as interferon and the proinflammatory cytokines interleukin-1 (IL-1) and tumor necrosis factor (TNF). This review discusses examples of viral anti-immune mechanisms from many different viruses and then focuses on the molecular interactions between adenoviruses and TNF and the effect of these interactions on the ensuing pathogenesis of virus infection.
Asunto(s)
Infecciones por Adenovirus Humanos/inmunología , Adenovirus Humanos/inmunología , Inflamación/virología , Factor de Necrosis Tumoral alfa/inmunología , Adenovirus Humanos/genética , Animales , Muerte Celular , ADN Viral/genética , Regulación Viral de la Expresión Génica , Genoma Viral , Humanos , Inflamación/inmunología , Interleucina-1/inmunologíaRESUMEN
Cell-mediated immunity to SV40-transformed C3H and C3H-SW cell lines was measured by using both 51Cr and 125IUdR release assays. Killing by cytotoxic cells generated on in vitro sensitization of immune spleen cells with syngeneic SV40 cells by either assay is specific for syngeneic SV40 transformants. Cytolysis mediated by in vitro sensitized cells is ablated by treatment of the effector cells with anti-theta serum and complement. Intraperitoneal immunization with syngeneic SV40 cells yields two distinct killer-cell populations in the peritoneal exudate when assayed by 125IUdR release. The first, nylon wool nonadherent and sensitive to anti-theta and complement, is indistinguishable from the killers generated in vitro. The second population, present in larger numbers and more efficient on a per-cell basis in killing of SV40 targets than the first, is nylon adherent and is not removed by treatment with anti-theta and complement. This second population will kill any SV40 transformed target, whether syngeneic or allogeneic. The possible roles of T cell and non-T cell effectors in rejection of syngeneic SV40 tumors are discussed.
Asunto(s)
Transformación Celular Neoplásica , Virus 40 de los Simios/inmunología , Linfocitos T/inmunología , Animales , Suero Antilinfocítico/farmacología , Líquido Ascítico/citología , Línea Celular , Proteínas del Sistema Complemento , Pruebas Inmunológicas de Citotoxicidad , Inmunidad Celular , Linfocitos/inmunología , Ratones , Ratones Endogámicos C3H , Bazo/inmunologíaRESUMEN
Antibody blockade of cell-mediated lysis was used to probe the cell surface structures recognized by T lymphocytes generated to syngeneic SV40 virus-transformed mouse cells. Alloantisera to H-2 antigens were highly effective in inhibiting lysis. Anti-H2 antibody blockade of lysis was haplotype specific even on transformed F1 target cells. Inhibition occurred only when antibody was bound to the target cell. Antibody binding to the effector did not inhibit lysis. Inhibition required that antibody be bound to the H-2 molecule itself; antibody to molecules associated with H-2, such as beta2-microglobulin, had no effect. Attempts to block lysis by using antisera to known virus-coded molecules were uniformly unsuccessful. These results are discussed in light of current controversy concerning the nature of the SV40 virus-specific transplantation antigen.
Asunto(s)
Anticuerpos , Citotoxicidad Inmunológica , Virus 40 de los Simios/inmunología , Linfocitos T/inmunología , Animales , Especificidad de Anticuerpos , Antígenos de Superficie/inmunología , Suero Antilinfocítico/farmacología , Unión Competitiva , Antígenos H-2 , Sueros Inmunes/farmacología , Isoanticuerpos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Endogámicos DBA , Conejos , Microglobulina beta-2/inmunologíaRESUMEN
T lymphocyte effectors to syngeneic SV40-transformed cells, generated by secondary in vitro sensitization of immune spleen cells, lyse SV40 transformed targets that are syngeneic at the H-2 locus. In this study we have employed recombinants within the H-2 region to examine in detail this H-2 specificity. H-2b effectors were found to lyse SV40-transformed targets from recombinants bearing either H-2Kb or H-2Db.H-2k effectors recognized only SV40-transformed H-2Kk, and not H-2Dk target cells. By using the same protocol for sensitization, no effector cells could be detected in H-2d mice. Effectors generated in H-2 recombinant mice showed that the response capacity resides with K and D. For example, HTG, which is H-2d except at the D locus (H-2Db), produced effector cells specific for SV40-transformed H-2Db targets. Thus, the secondary in vitro response to SV40 transformants was found to depend only on the K and D alleles and not to be modified by the I region to any measurable extent.
Asunto(s)
Citotoxicidad Inmunológica , Antígenos H-2/genética , Recombinación Genética , Virus 40 de los Simios/inmunología , Linfocitos T/inmunología , Animales , Línea Celular , Transformación Celular Viral , Epítopos , Ratones , Ratones Endogámicos A , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Endogámicos C57BLRESUMEN
Cultured SV40-transformed fibroblasts from C3H mice (SV-C3H) were "adapted" to in vivo growth by serial passage through sublethally irradiated, syngeneic recipients. After four in vivo passages, a population of cells was obtained (V4) that was weakly oncogenic in nonirradiated mice. Cells isolated from large V4 tumors (V5) were found to be highly oncogenic, producing lethal tumors at doses of less than 10(3) cells. V5 is insensitive to SV40-specific transplantation immunity in syngeneic animals but can be rejected completely by H-2 allogeneic mice. In vitro studies revealed that although V4 and the parent SV-C3H cells can induce SV40-specific cytotoxic T cells (CTL) in vitro and are lysed by these CTL, V5 does neither. The failure of V5 to interact with CTL was traced to the loss of H-2Kk antigen expression on these cells. The correlation between H-2Kk loss and immunoresistance in vivo suggests a central role for the cytotoxic T cell in in vivo tumor elimination in this system.