RESUMEN
One-lung anesthesia is a method of anesthesia performed by inserting the tip of a bronchial tube into either the right main bronchus or the left main bronchus. The right bronchial tube is a special structure. Since the distance of the carina to the right upper lobe bronchus is short, a side hole is made to prevent blockading of the right upper lobe bronchus, and the cuff is attached aslant to it. When inserting a bronchial tube into the right main bronchus, care is required to prevent the occurrence of atelectasis though a gap in the bronchial tube. We evaluated the structure of a trachea and a bronchus using the multidetector-row computed tomography (MD CT), and tried to select the right bronchial tube most suitable for each structure. There are individual differences in the structure of a trachea and a bronchus. By creating a 3-dimensional image of a trachea and a bronchus, the structure could be easily grasped, and therefore selection of the most appropriate bronchial tube according to the structure was possible.
Asunto(s)
Anestesia/métodos , Bronquios , Broncografía , Intubación Intratraqueal/instrumentación , Tomografía Computarizada por Rayos X , Anciano , Femenino , Humanos , Pulmón , Masculino , Persona de Mediana EdadAsunto(s)
Inmunoterapia/métodos , Interleucina-15/fisiología , Interleucina-2/fisiología , Linfocitos/citología , Linfocitos/inmunología , Animales , Anticuerpos Monoclonales/uso terapéutico , Apoptosis , Enfermedades Autoinmunes/terapia , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , Vacunas contra el Cáncer/uso terapéutico , Rechazo de Injerto/prevención & control , Humanos , Memoria Inmunológica , Interleucina-15/antagonistas & inhibidores , Interleucina-2/antagonistas & inhibidores , Leucemia/terapia , Activación de Linfocitos , Linfoma/terapia , Ratones , Modelos Biológicos , Neoplasias/terapiaRESUMEN
Activation of T cells usually requires two signals. Signal 1 is mediated via a peptide-MHC on the APC; signal 2 is mediated via a costimulatory molecule on the APC surface. We demonstrate here that naive CD4(+) T cells actually acquire the costimulatory molecule CD80 (B7-1) from syngeneic APCs after activation. This phenomenon was demonstrated showing acquisition of CD80 by T cells from CD80/CD86 (B7-2) knockout mice, and by treating T cells with cyclohexamide to further rule out endogenous expression of CD80 by T cells. Moreover, no CD80 mRNA could be detected in T cells that had acquired CD80. The amount of acquisition of CD80 by T cells was shown to be directly related to both the strength of signal 1 and the amount of CD80 on the APC. Specificity of this acquisition was also shown by the lack of acquisition by T cells from CD28 knockout mice (implicating CD28 in this process), the lack of acquisition of CD40 (another molecule on the APC surface) by T cells, and confocal microscopy studies. We demonstrate for the first time that 1) naive T cells, following acquisition of CD80 from APCs, were themselves shown to be capable of acting as APCs; and 2) memory T cells that have acquired CD80 from APCs undergo apoptosis in the presence of increased levels of signal 1. Thus we demonstrate both immunostimulatory and immunoregulatory functions as a result of CD80 acquisition by different T cell populations.
Asunto(s)
Antígeno B7-1/biosíntesis , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Animales , Presentación de Antígeno/genética , Células Presentadoras de Antígenos/inmunología , Células Presentadoras de Antígenos/metabolismo , Apoptosis/genética , Apoptosis/inmunología , Antígeno B7-1/genética , Antígeno B7-1/metabolismo , Antígenos CD28/genética , Antígenos CD28/metabolismo , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Células COS , Línea Celular , Células Cultivadas , Cicloheximida/farmacología , Relación Dosis-Respuesta Inmunológica , Memoria Inmunológica/genética , Interfase/genética , Interfase/inmunología , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/genética , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Reacción en Cadena de la Polimerasa , Unión Proteica/genética , Unión Proteica/inmunología , Inhibidores de la Síntesis de la Proteína/farmacología , ARN Mensajero/análisis , Transducción de Señal/genética , Transducción de Señal/inmunología , Especificidad de la Especie , Subgrupos de Linfocitos T/efectos de los fármacos , Células Tumorales Cultivadas , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genética , Regulación hacia Arriba/inmunologíaRESUMEN
A transgenic (Tg) mouse expressing human IL-15 was generated to define the role of IL-15 in the normal immune response. Overexpression of IL-15 resulted in an increase of NK, CD44(hi)CD8 memory T cells, and gammadelta T cells. Additionally, we observed the emergence of a novel type of NK-T cells with CD8alphaalpha' expression. Due to the expansion and activation of NK cells, the IL-15Tg mouse showed enhanced innate immunity. In adaptive T cell immunity, the roles of IL-15 contrasted with those of IL-2. IL-15 inhibited IL-2-induced T cell death, which plays a role in the maintenance of peripheral self-tolerance. IL-15 thus seems to contribute to enhanced immune memory by selectively propagating memory T cells and by blocking T cell death mediated by IL-2.
Asunto(s)
Muerte Celular/fisiología , Interleucina-15/fisiología , Interleucina-2/fisiología , Animales , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , Ciclo Celular , Citocinas/biosíntesis , Humanos , Interleucina-15/genética , Células Asesinas Naturales/citología , Células Asesinas Naturales/inmunología , Recuento de Linfocitos , Ratones , Ratones Transgénicos , TransgenesRESUMEN
We identified an interferon regulatory factor motif (IRF-E) upstream of an NF-kappaB binding site in the interleukin-15 (IL-15) promoter. Since these two motifs are part of the virus-inducible enhancer region of the beta interferon promoter, we speculated that there might be similar responses of these two genes to stimuli such as viruses. To test this hypothesis, L929 cells were infected with Newcastle disease virus (NDV), which led to the induction of IL-15 mRNA and protein expression. Using IL-15 promoter-reporter deletion constructs, a virus-inducible region, encompassing IRF-E, NF-kappaB, and a 13-nucleotide sequence flanked by these two motifs, was mapped to the -295-to--243 position relative to the transcription initiation site. Using cotransfection studies, it was demonstrated that all three motifs were essential to achieve the maximum promoter activity induced by IRF-1 and NF-kappaB expression plasmids. The presence of a virus-inducible region in the IL-15 promoter suggests a role for IL-15 as a component of host antiviral defense mechanisms.
Asunto(s)
Regulación de la Expresión Génica , Interleucina-15/genética , Virus de la Enfermedad de Newcastle/inmunología , Regiones Promotoras Genéticas/genética , Animales , Secuencia de Bases , Clonación Molecular , Proteínas de Unión al ADN/metabolismo , Fibroblastos , Factor 1 Regulador del Interferón , Interleucina-15/biosíntesis , Células L , Ratones , Datos de Secuencia Molecular , FN-kappa B/metabolismo , Fosfoproteínas/metabolismo , Análisis de Secuencia de ADNRESUMEN
Interleukin-2 (IL-2) and interleukin-15 (IL-15) are T-cell tropic factors that share beta and gammac subunits of their receptors on T/NK-cells. Although these two cytokines share receptor components, the IL-15Ralpha molecule is expressed constitutively by various tissue cells, whereas the IL-2Ralpha expression is mostly restricted to activated mononuclear cells. Consequently, we postulated that the biodistribution of IL-15 might be different from that of IL-2 and that individual alpha chains play an important role in this respect. This study investigated the differences between IL-2 and IL-15 in pharmacokinetics, biodistribution, and their tumor-targeting abilities. It found that only IL-2 showed specific binding to a protein, alpha2-macroglobulin, which may be the reason that IL-2 displays longer blood clearance than IL-15. Upon injection of these cytokines into mice, we observed that IL-15 accumulated significantly more than IL-2 in kidney, spleen, and bone. These are all tissues that express IL-15 receptor alpha but not IL-2 receptor alpha. To evaluate the tumor-targeting ability of each cytokine, we used nude mice xenografted with three A431 tumors, parental and cells transfected with alpha subunit of the receptor for either IL-2 or IL-15. When examined using radioiodinated IL-2 or IL-15, each cytokine accumulated on the target cells, expressing its respective alpha chain, suggesting that the expression of the alpha chains is sufficient to define specific biodistribution of IL-2 and IL-15, although these cytokines share the beta and yc molecules of their receptors. IL-15 displayed better target-specific accumulation and more rapid clearance from the circulation than did IL-2, and thus it can be considered to be a novel and unique therapeutic agent.
Asunto(s)
Carcinoma de Células Escamosas/metabolismo , Interleucina-15/farmacocinética , Interleucina-2/farmacocinética , Animales , Carcinoma de Células Escamosas/inmunología , Femenino , Humanos , Interleucina-15/sangre , Interleucina-2/sangre , Radioisótopos de Yodo , Tasa de Depuración Metabólica , Ratones , Ratones Desnudos , Proteínas Recombinantes/sangre , Proteínas Recombinantes/farmacocinética , Distribución Tisular , Transfección , Trasplante Heterólogo , alfa-Macroglobulinas/metabolismoRESUMEN
Two isoforms of interleukin (IL)-15 exist: one with a short and another with a long signal peptide (LSP). Experiments using combinations of the LSP and mature proteins IL-2, IL-15, and green fluorescent protein revealed complex pathways of intracellular trafficking. In one pathway, the LSP was unprocessed, and IL-15 was not glycosylated, remained in the cytoplasm, and was degraded. The second trafficking pathway involved endoplasmic reticulum entry, N-linked glycosylation, and alternative partial LSP processing. The third pathway involved endoplasmic reticulum entry, followed by glycosylation, complete processing, and ultimately secretion. The complex intracellular trafficking patterns of LSP-IL-15 with its impediments to secretion as well as impediments to translation may be required due to the potency of IL-15 as an inflammatory cytokine. In terms of a more positive role, we propose that intracellular infection may relieve the burdens on translation and intracellular trafficking to yield effective IL-15 expression.
Asunto(s)
Interleucina-15/metabolismo , Precursores de Proteínas/metabolismo , Procesamiento Proteico-Postraduccional , Señales de Clasificación de Proteína/metabolismo , Acetilcisteína/análogos & derivados , Acetilcisteína/farmacología , Secuencia de Aminoácidos , Animales , Bioensayo , Transporte Biológico , Células COS , Compartimento Celular , Inhibidores de Cisteína Proteinasa/farmacología , Retículo Endoplásmico/metabolismo , Glicosilación , Hexosaminidasas/farmacología , Interleucina-15/química , Datos de Secuencia Molecular , Isoformas de Proteínas , Precursores de Proteínas/química , Señales de Clasificación de Proteína/químicaRESUMEN
Interleukin-15 (IL-15) is a 14- to 15-kDa member of the 4 alpha-helix bundle family of cytokines. IL-15 expression is controlled at the levels of transcription, translation, and intracellular trafficking. In particular, IL-15 protein is posttranscriptionally regulated by multiple controlling elements that impede translation, including 12 upstream AUGs of the 5' UTR, 2 unusual signal peptides, and the C-terminus of the mature protein. IL-15 uses two distinct receptor and signaling pathways. In T and NK cells the IL-15 receptor includes IL-2/15R beta and gamma c subunits, which are shared with IL-2, and an IL-15-specific receptor subunit, IL-15R alpha. Mast cells respond to IL-15 with a receptor system that does not share elements with the IL-2 receptor but uses a novel 60- to 65-kDa IL-15RX subunit. In mast cells IL-15 signaling involves Jak2/STAT5 activation rather than the Jak1/Jak3 and STAT5/STAT3 system used in activated T cells. In addition to its other functional activities in immune and nonimmune cells, IL-15 plays a pivotal role in the development, survival, and function of NK cells. Abnormalities of IL-15 expression have been described in patients with rheumatoid arthritis or inflammatory bowel disease and in diseases associated with the retroviruses HIV and HTLV-I. New approaches directed toward IL-15, its receptor, or its signaling pathway may be of value in the therapy of these disorders.
Asunto(s)
Interleucina-15/genética , Interleucina-15/fisiología , Células Asesinas Naturales/inmunología , Animales , Enfermedades Autoinmunes/inmunología , Secuencia de Bases , Diferenciación Celular , ADN Complementario/genética , Regulación de la Expresión Génica , Humanos , Terapia de Inmunosupresión , Inmunoterapia , Inflamación/inmunología , Células Asesinas Naturales/citología , Neoplasias/inmunología , Receptores de Interleucina-15 , Receptores de Interleucina-2/fisiología , Infecciones por Retroviridae/inmunología , Transducción de SeñalRESUMEN
The authors have previously reported that the soluble serum form of the alpha subunit of the IL-2 receptor (sIL-2Ralpha), whose natural half-life is approximately 40 min, survived much longer in the circulation when bound by a specific antibody. In the present study, the authors evaluated the extent to which sIL-2Ralpha protected IL-2 in freshly collected serum using biochemical analyses, and a functional CTLL-2 assay. In particular, sIL-2Ralpha protected IL-2 from forming complexes with alpha(2)-macroglobulin and from inactivation in vitro. In addition, the authors demonstrated that the anti-IL-2Ralpha monoclonal antibody 7G7/B6, which does not inhibit the binding of IL-2 to its binding site on sIL-2Ralpha, protected IL-2 from degradation and inactivation in vivo in the presence of sIL-2Ralpha. Both(125)I-labelled and unlabelled IL-2 were injected into mice preinjected with humanized anti-Tac (hTac) or 7G7/B6 and sIL-2Ralpha, or sIL-2Ralpha alone. Using size-exclusion HPLC, ELISA, and CTLL-2 cell proliferation assays, we observed that the presence of 7G7/B6 led to formation of complexes with sIL-2Ralpha and increased the serum levels of IL-2 more than 3- to 40-fold those of groups receiving IL-2 alone, sIL-2Ralpha, or hTac. Taken as a whole, these results suggest that the complex of 7G7/B6 and sIL-2Ralpha not only prolongs the survival of IL-2 in vivo, but also maintains the bioactivity of IL-2. The use of antibodies against endogenous soluble receptors could increase the in vivo survival of cytokines, protect their bioactivity and thereby facilitate their clinical use in the treatment of various malignancies and AIDS.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Interleucina-2/sangre , Interleucina-2/inmunología , Receptores de Interleucina-2/sangre , Receptores de Interleucina-2/inmunología , Anticuerpos Monoclonales/sangre , Humanos , Unión Proteica , alfa-Macroglobulinas/inmunología , alfa-Macroglobulinas/metabolismoRESUMEN
Interleukin 15 (IL-15) mRNA is expressed in a wide variety of tissue types. However, with the exception of some T cell lines, IL-15 transcript expression has not been described in T cells. Herein we demonstrate that IL-15 mRNA can be detected in freshly isolated normal T cells and T cell lines. Furthermore, its expression is 3- to 4-fold higher in human T cell lymphotropic virus type I (HTLV-I)-infected T cells. By using reporter constructs bearing the 5' regulatory region of the IL-15 gene, we observed a positive correlation between HTLV-I Tax protein expression and IL-15 promoter activity in HTLV-I-infected T cells. Additionally, by using a Jurkat T cell transfectant that expresses Tax under an inducible promoter, we demonstrated that the expression of IL-15 mRNA increased 3-fold as Tax was expressed, suggesting that the Tax protein activates IL-15 transcription. An NF-kappaB consensus sequence is located at the -75 and -65 region of the IL-15 5' regulatory region. Mutations in the NF-kappaB motif or deletion of this sequence abrogated the promoter activity in both HTLV-I-positive and Jurkat Tax-transfectant cells. These data represent evidence for trans-activation of the IL-15 gene by the HTLV-I Tax protein through an NF-kappaB motif and suggest a potential role for IL-15 in HTLV-I-associated diseases such as adult T cell leukemia and HTLV-I-associated myopathy/tropical spastic paraparesis.
Asunto(s)
Productos del Gen tax/metabolismo , Virus Linfotrópico T Tipo 1 Humano/fisiología , Interleucina-15/biosíntesis , FN-kappa B/metabolismo , Regiones Promotoras Genéticas , Linfocitos T/inmunología , Transcripción Genética , Activación Transcripcional , Secuencia de Bases , Línea Celular , Células Cultivadas , Productos del Gen tax/biosíntesis , Genes Reporteros , Virus Linfotrópico T Tipo 1 Humano/inmunología , Virus Linfotrópico T Tipo 1 Humano/patogenicidad , Humanos , Interleucina-15/genética , Células Jurkat , Datos de Secuencia Molecular , FN-kappa B/biosíntesis , FN-kappa B/química , ARN Mensajero/biosíntesis , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/metabolismo , Secuencias Reguladoras de Ácidos Nucleicos , TransfecciónRESUMEN
Natural killer (NK) cells are critical for both innate and adaptive immunity. The development of NK cells requires interactions between their progenitors and the bone-marrow microenvironment; however, little is known about the molecular nature of such interactions. Mice that do not express the transcription factor interferon-regulatory factor-1 (IRF-1; such mice are IRF-1(-/-) mice) have been shown to exhibit a severe NK-cell deficiency. Here we demonstrate that the lack of IRF-1 affects the radiation-resistant cells that constitute the microenvironment required for NK-cell development, but not the NK-cell progenitors themselves. We also show that IRF-1(-/-) bone-marrow cells can generate functional NK cells when cultured with the cytokine interleukin-15 and that the interleukin-15 gene is transcriptionally regulated by IRF-1. These results reveal, for the first time, a molecular mechanism by which the bone-marrow microenvironment supports NK-cell development.
Asunto(s)
Proteínas de Unión al ADN/fisiología , Células Asesinas Naturales/citología , Fosfoproteínas/fisiología , Factores de Transcripción/fisiología , Animales , Células de la Médula Ósea/fisiología , Trasplante de Médula Ósea , Diferenciación Celular/fisiología , Células Cultivadas , Clonación Molecular , Regulación de la Expresión Génica/efectos de los fármacos , Factor 1 Regulador del Interferón , Interferón gamma/farmacología , Interleucina-15/genética , Interleucina-15/fisiología , Células Asesinas Naturales/trasplante , Lipopolisacáridos/farmacología , Ratones , Ratones Endogámicos C57BL , ARN Mensajero/metabolismo , Proteínas Recombinantes/farmacología , Secuencias Reguladoras de Ácidos Nucleicos , Quimera por TrasplanteRESUMEN
Both IL-15 and IL-2 are 14-15 kDa members of the four alpha-helical bundle family of cytokines that have T cell growth factor activity. In contrast to the pattern manifested by IL-2, IL-15 mRNA is produced by a wide variety of tissues other than T cells. We have demonstrated that IL-15 expression is posttranscriptionally regulated by multiple elements, including the ten upstream AUGs of the 5' UTR, a 48aa signal peptide and the carboxy-terminus of the mature protein. IL-15 utilizes two distinct receptor signaling pathways. In T cells the IL-15 receptor includes IL-2R beta and gamma c subunits shared with IL-2 as well as an IL-15 specific receptor, IL-15R alpha. However, mast cells respond to IL-15 using a receptor system that does not share elements with the IL-2R system but involves a novel 60-65 kDa IL-15RX subunit. In mast cells, IL-15 signaling involves JAK-2 and STAT-5 activation rather than the JAK-1 and JAK-3 as well as the STAT-3 and STAT-5 used by both IL-2 and IL-15 in activated T cells.
Asunto(s)
Interleucina-15/fisiología , Interleucina-2/fisiología , Receptores de Interleucina-2/fisiología , Receptores de Interleucina/fisiología , HumanosRESUMEN
Two isoforms of human interleukin 15 (IL-15) exist. One isoform has a shorter putative signal peptide (21 amino acids) and its transcript shows a tissue distribution pattern that is distinct from that of the alternative IL-15 isoform with a 48-aa signal peptide. The 21-aa signal isoform is preferentially expressed in tissues such as testis and thymus. Experiments using different combinations of signal peptides and mature proteins (IL-2, IL-15, and green fluorescent protein) showed that the short signal peptide regulates the fate of the mature protein by controlling the intracellular trafficking to nonendoplasmic reticulum sites, whereas the long signal peptide both regulates the rate of protein translation and functions as a secretory signal peptide. As a consequence, the IL-15 associated with the short signal peptide is not secreted, but rather is stored intracellularly, appearing in the nucleus and cytoplasmic components. Such production of an intracellular lymphokine is not typical of other soluble interleukin systems, suggesting a biological function for IL-15 as an intracellular molecule.
Asunto(s)
Interleucina-15/genética , Ingeniería de Proteínas , Animales , Secuencia de Bases , Células COS , Humanos , Interleucina-15/química , Interleucina-15/metabolismo , Datos de Secuencia Molecular , Péptidos/química , Péptidos/genética , Péptidos/metabolismoRESUMEN
Interleukin-15/T(IL-15) is a growth factor that utilizes IL-2 receptor (IL-2R) components in addition to its private binding protein IL-15R(alpha) in T-cells. Here, we report that IL-15 induces mast cell proliferation in the absence of IL-2R alpha and beta. Using transfectants of these cells with a cytoplasmic-truncated mutant of gamma(c), we demonstrated that IL-15 signaling in mast cells does not involve gamma(c). Cross-linking of mast cells with [(125)I]IL-15 revealed a 60-65 kDa IL-15 binding protein that is distinct from known components of T-cell IL-15 receptors. Mast cell IL-15 receptors recruit JAK-2 and STAT-5, instead of JAK1/3 and STAT3/5 that are activated in T-cells. Thus IL-15 is a mast cell growth factor that utilizes a novel receptor and distinct signaling pathway.
Asunto(s)
Interleucina-15/metabolismo , Mastocitos/metabolismo , Proteínas de la Leche , Proteínas Proto-Oncogénicas , Transducción de Señal , Animales , Células de la Médula Ósea , División Celular/efectos de los fármacos , Línea Celular , Proteínas de Unión al ADN/metabolismo , Humanos , Interleucina-15/aislamiento & purificación , Janus Quinasa 1 , Janus Quinasa 2 , Ratones , Fosforilación , Conformación Proteica , Proteínas Tirosina Quinasas/metabolismo , Proteínas/metabolismo , Receptores de Interleucina-2/química , Proteínas Recombinantes/aislamiento & purificación , Factor de Transcripción STAT3 , Factor de Transcripción STAT5 , Relación Estructura-Actividad , TYK2 Quinasa , Transactivadores/metabolismo , TransfecciónAsunto(s)
Interleucinas/genética , Interleucinas/fisiología , Animales , Secuencia de Bases , Línea Celular , ADN Complementario/genética , Regulación de la Expresión Génica , Virus Linfotrópico T Tipo 1 Humano/genética , Humanos , Interleucina-15 , Interleucinas/química , Células Asesinas Naturales/inmunología , Mastocitos/inmunología , Ratones , Datos de Secuencia Molecular , Estructura Molecular , ARN Mensajero/genética , Receptores de Interleucina-15 , Receptores de Interleucina-2/fisiología , Transducción de Señal , Linfocitos T/inmunologíaRESUMEN
We obtained human T-cell hybridomas that are specific for bee venom phospholipase A2 (PLA2) and constitutively secrete glycosylation inhibiting factor (GIF). Upon crosslinking of CD3, the hybridoma produced GIF having affinity for PLA2. When affinity-purified PLA2-binding GIF was used as an immunogen, monoclonal antibodies specific for the antigen-binding GIF were obtained. Monoclonal antibody 110BH3 bound the antigen-binding GIF but failed to bind the 13-kDa nonspecific GIF, as determined by both bioassay and ELISA. In contrast, 388F1, a monoclonal antibody against nonspecific GIF, gave ELISA signals with both the nonspecific GIF and the antigen-binding GIF. Gel filtration of affinity-purified antigen-binding GIF revealed the presence of a 72- to 80-kDa protein which gave ELISA signals with both 110BH3 and 388F1 and contained GIF bioactivity. Upon reduction and alkylation, the antigen-binding GIF dissociated into a 62- to 64-kDa protein which gave positive ELISA with antibody 110BH3 but no signal with antibody 388F1, and a 15-kDa protein, which gave ELISA signal with the 388F1 but not with 110BH3. Immunoblotting of a PLA2-binding GIF preparation revealed that under reducing conditions, the antigen-binding GIF dissociated a 13-kDa peptide which reacted with polyclonal antibodies against recombinant GIF. The results indicate that the 13-kDa nonspecific GIF is a subunit of antigen-binding GIF. The PLA2-binding GIF has affinity for an epitope, representing amino acid residues 19-28 in PLA2 which appears to be an external structure in the antigen.
Asunto(s)
Venenos de Abeja/enzimología , Linfocinas/metabolismo , Fosfolipasas A/inmunología , Proteínas de Secreción Prostática , Factores Supresores Inmunológicos/metabolismo , Linfocitos T/inmunología , Animales , Anticuerpos Monoclonales , Especificidad de Anticuerpos , Antígenos/inmunología , Ensayo de Inmunoadsorción Enzimática , Epítopos , Glicosilación , Humanos , Hibridomas , Linfocinas/genética , Linfocinas/inmunología , Linfocinas/aislamiento & purificación , Ratones , Fosfolipasas A/metabolismo , Fosfolipasas A2 , Proteínas Recombinantes/inmunología , Factores Supresores Inmunológicos/genética , Factores Supresores Inmunológicos/inmunología , Factores Supresores Inmunológicos/aislamiento & purificaciónRESUMEN
From the spleen cells of BALB/c mice primed with bee venom phospholipase A2 (PLA2), we established seven T cell hybridomas which constitutively secreted glycosylation inhibiting factor (GIF), expressed both CD3 and TCR alpha beta, and responded to antigen-pulsed antigen presenting cells (APC) for the formation of IgE-binding factor. Upon stimulation with antigen-pulsed APC, four of the seven hybridomas produced GIF having affinity for native PLA2. The antigen-binding GIF could suppress the anti-hapten antibody response of BALB/c mice to dinitrophenyl (DNP)-PLA2 conjugates in a carrier-specific manner and bound to immunosorbents coupled with either the mAb 14-12 or anti-TCR alpha chain, H28-710. Analysis of the epitope specificity of the TCR on the GIF-producing T hybridomas indicated that all of the hybridomas which could produce antigen-binding GIF upon antigenic stimulation recognized the synthetic peptide representing amino acid residues 19-34 in PLA2 molecules in the context of the product of the I-Ad subregion and the antigen-binding GIF formed by the cells had affinity for the peptide. The 3-D structure of bee venom PLA2 indicates that the sequence of amino acid 14-24 forms a loop in the PLA2 molecule and represents an external structure of the antigen, while peptide 25-37 forms an alpha helix. Evidence was obtained which suggests that the sequence of 25-34 contains amino acid residues interacting with Ia molecules, while peptide 19-24 contains residues involved in the interaction of p19-34-Ia complexes with TCR on the hybridomas. It was also found that not only the synthetic peptide 19-34, but also the peptides 13-28 and 19-30 inhibited the binding of antigen-binding GIF to PLA2-coupled Sepharose, while peptide 25-40 failed to do so. The results collectively indicate that the antigen-binding GIF and TCR on the cell source of the factor interact with a common epitope which is exposed on the surface of a nominal antigen.
Asunto(s)
Venenos de Abeja/inmunología , Epítopos , Linfocinas/biosíntesis , Fosfolipasas A/inmunología , Proteínas de Secreción Prostática , Factores Supresores Inmunológicos/biosíntesis , Linfocitos T Reguladores/inmunología , Secuencia de Aminoácidos , Animales , Antígenos/inmunología , Glicosilación , Hibridomas , Linfocinas/inmunología , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Fragmentos de Péptidos/inmunología , Fosfolipasas A2 , Receptores de Antígenos de Linfocitos T/inmunologíaRESUMEN
A high incidence of multiple primary neoplasms has been observed in our patients with ATL in comparison to persons with other forms of hematologic malignancy who we have observed during the past 23 years (1963-1985). Five of 15 patients with ATL (33.3 per cent) have had at least one other associated neoplasm in comparison to only 44 of 1156 patients with other forms of hematological malignancy (3.8 per cent). The incidence figures for secondary neoplasms associated with the other hematologic malignancies were 4.3 per cent (16/370) for acute non-lymphocytic leukemia (ANLL), 2.2 per cent (2/90) for acute lymphocytic leukemia (ALL), 4.8 per cent (1/21) for acute unclassifiable leukemia, 2.2 per cent (5/225) for chronic myelogenous leukemia, 4.7 per cent (2/43) for chronic lymphocytic leukemia, 5.9 per cent (8/136) for malignant monoclonal gammopathy and 3.7 per cent (10/271) for malignant lymphoma. The incidence of multiple neoplasms in patients with ATL in comparison to those with other hematological malignancies was statistically significant (p < 0.01 or p < 0.001). The neoplasms associated with ATL have been adenocarcinoma of the thyroid or stomach, and squamous cell carcinoma of the larynx, lip or lung. We identified ATL-derived factor (ADF) in the cytoplasm of the secondary neoplasms of the ATL patients by means of indirect immunofluoroscopy and immunohistochemical techniques utilizing anti-ADF antibody. We also identified ras p21 products in these neoplasms by means of p21 ras monoclonal antibody studies. The possibility that HTLV-I was the cause of the secondary neoplasms thus was investigated. HTLV-I provirus genome was not found in all the six cases of non-ATL leukemic cells of the patients with anti-HTLV-I antibodies as determined by means of Southern blot analysis utilizing pX DNA probe. These findings suggest that there is some association between ATL cells and pre-malignant cells through ADF or other unknown factors in the activation of ras oncogenes. Subsequent suppression of host immune defence mechanisms in ATL patients permits evolution of the secondary neoplasms.
Asunto(s)
Citocinas , Leucemia de Células T/patología , Neoplasias Primarias Múltiples/patología , Adenocarcinoma/patología , Adenocarcinoma/ultraestructura , Anciano , Carcinoma de Células Escamosas/patología , Carcinoma de Células Escamosas/ultraestructura , Neoplasias de Cabeza y Cuello/patología , Neoplasias de Cabeza y Cuello/ultraestructura , Virus Linfotrópico T Tipo 1 Humano/genética , Virus Linfotrópico T Tipo 1 Humano/inmunología , Humanos , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/análisis , Neoplasias Primarias Múltiples/ultraestructura , Proteínas Proto-Oncogénicas p21(ras)/análisis , Neoplasias Torácicas/patología , Neoplasias Torácicas/ultraestructuraRESUMEN
Adult T-cell leukemia-derived factor (ADF), originally defined as an interleukin-2 receptor inducer, is a human thioredoxin homologue. ADF is detected in many malignant tissues and has a growth-promoting effect on transformed cells. In this study, ADF expression was examined immunohistochemically in human liver cell lines and liver tissues, and its growth-promoting effect was tested on human hepatoma cells. On three liver cell line--PLC/PRF/5, HepG2, and Chang liver cells--ADF stained positively and also was detected by immunoblotting. ADF had strong staining in the fetal liver (n = 8), although it was faint in the normal adult liver (n = 6). In hepatocellular carcinoma (n = 25), ADF expression generally was enhanced and was very strong in 52% (13 of 25) of the cases, although it was moderate in cases of chronic hepatitis or cirrhosis. ADF augmented the growth of PLC/PRF/5 cells and showed an additive effect with epidermal growth factor. These results indicate possible involvement of ADF in cell activation and growth of hepatocytes, as is the case with lymphocytes.