RESUMEN
BACKGROUND: In K/BxN mice, anti-glucose-6-phosphate isomerase (GPI) antibodies (Abs) are arthritogenic, and their transfer into naive mice induces arthritis. Anti-GPI Abs develop in many human patients with RA and are associated with more severe forms of the disease. OBJECTIVE: To elucidate the serum and synovial fluid (SF) anti-GPI IgG profiles among different patient groups with a variety of arthritides. METHODS: Blood and SF obtained concomitantly from 91 patients with clinically well defined arthritis were tested for concentrations of total anti-GPI IgG, anti-GPI IgG subclasses, B lymphocyte stimulator (BLyS), and APRIL by ELISA. RESULTS: Anti-GPI IgG was detected in sera and SF of patients with many arthritic diseases, but was preferentially associated with inflammatory arthritis, in general, and RA, in particular. The anti-GPI IgG subclass usage was skewed and varied among the different arthritic disease groups. Inverse correlations between serum levels of BLyS and anti-GPI IgG and positive correlations between serum levels of APRIL and anti-GPI IgG were seen among immune based arthritic patients and patients with RA but not among non-immune based patients. No correlations were found in SF from any group of arthritic patients. CONCLUSION: Raised circulating anti-GPI Abs are not unique to patients with RA but are present in many patients with inflammatory arthritis. The difference in anti-GPI IgG subclass usage among disease groups may influence effector function and disease outcome. The inverse correlation between serum BLyS and anti-GPI IgG levels suggests that anti-GPI B cells may be regulated differently from other autoantibody producing B cells. Anti-GPI Abs may serve a pathogenic function in humans by promoting the maintenance of existing disease.
Asunto(s)
Artritis/inmunología , Autoanticuerpos/análisis , Glucosa-6-Fosfato Isomerasa/inmunología , Inmunoglobulina G/análisis , Líquido Sinovial/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Artritis/metabolismo , Artritis Reumatoide/inmunología , Artritis Reumatoide/metabolismo , Autoanticuerpos/sangre , Factor Activador de Células B , Ensayo de Inmunoadsorción Enzimática/métodos , Femenino , Glucosa-6-Fosfato Isomerasa/metabolismo , Humanos , Inmunoglobulina G/sangre , Masculino , Proteínas de la Membrana/análisis , Proteínas de la Membrana/sangre , Persona de Mediana Edad , Proteínas Nucleares/análisis , Proteínas Nucleares/sangre , Líquido Sinovial/metabolismo , Factor de Necrosis Tumoral alfa/análisisRESUMEN
OBJECTIVE: To assess longitudinal expression of a proliferation-inducing ligand (APRIL) in patients with systemic lupus erythematosus (SLE) and its correlation with B lymphocyte stimulator (BLyS) expression, serum anti-dsDNA titres, and clinical disease activity. METHODS: Sixty eight patients with SLE were longitudinally followed up for a median of 369 days. At each visit the physician assessed disease activity by SLEDAI, and blood was collected for determination of serum APRIL and BLyS levels and of blood APRIL and BLyS mRNA levels. Fifteen normal control subjects underwent similar laboratory evaluation. RESULTS: Dysregulation of APRIL was not as great as that of BLyS. Changes in serum levels of APRIL and BLyS over time were usually discordant, whereas blood levels of APRIL and BLyS mRNA strongly paralleled each other. Serum APRIL levels modestly, but significantly, inversely correlated with serum anti-dsDNA titres in anti-dsDNA positive patients analysed in aggregate. Moreover, serum APRIL levels modestly, but significantly, inversely correlated with clinical disease activity in all patients analysed in aggregate. CONCLUSION: Serum levels of APRIL and BLyS are differentially regulated. APRIL may serve as a down modulator of serological and/or clinical autoimmunity in patients with SLE. This may have important ramifications for BLyS targeted treatment, and it remains to be determined whether agents which neutralise only BLyS will be preferable to agents which neutralise both BLyS and APRIL.
Asunto(s)
Lupus Eritematoso Sistémico/sangre , Neuropéptidos/sangre , Proteínas Nucleares/sangre , Anticuerpos Antinucleares/sangre , Factor Activador de Células B , ADN/inmunología , Estudios de Seguimiento , Humanos , Proteínas de la Membrana/sangre , Proteínas de la Membrana/genética , Neuropéptidos/genética , Proteínas Nucleares/genética , ARN Mensajero/sangre , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
Striking cell losses occur during late B lymphocyte maturation, reflecting BcR-mediated selection coupled with requisites for viability promoting signals. How selection and survival cues are integrated remains unclear, but a key role for B lymphocyte stimulator (BLyS(TM); trademark of Human Genome Sciences, Inc.) is suggested by its marked effects on B cell numbers and autoantibody formation as well as the B lineage-specific expression of BLyS receptors. Our analyses of the B cell-deficient A/WySnJ mouse have established Bcmd as a gene controlling follicular B cell life span, and recent reports show Bcmd encodes a novel BLyS receptor. Here we show that A/WySnJ B cells are unresponsive to BLyS, affording interrogation of how Bcmd influences B cell homeostasis. Mixed marrow chimeras indicate A/WySnJ peripheral B cells compete poorly for peripheral survival. Moreover, in vivo BrdU labeling shows that (A/WySnJ x BALB/c)F(1) B cells have an intermediate but uniform life span, indicating viability requires continuous signaling via this pathway. Together, these findings establish the BLyS/Bcmd pathway as a dominant mediator of B cell survival, suggesting competition for BLyS/Bcmd signals regulates follicular B cell numbers.
Asunto(s)
Linfocitos B/citología , Recuento de Linfocitos , Proteínas de la Membrana , Receptores del Factor de Necrosis Tumoral/fisiología , Transducción de Señal/fisiología , Animales , Receptor del Factor Activador de Células B , Supervivencia Celular/fisiología , Heterocigoto , Ratones , Ratones Endogámicos BALB C , Receptores del Factor de Necrosis Tumoral/genéticaRESUMEN
OBJECTIVE: To determine whether serum levels of B lymphocyte stimulator (BLyS) are elevated in patients with systemic immune-based rheumatic diseases and correlate with serum Ig levels and/or autoantibody titers. METHODS: Sera from 185 patients with various systemic immune-based rheumatic diseases (95 with systemic lupus erythematosus [SLE], 67 with rheumatoid arthritis [RA], 23 with other diagnoses) were assayed for BLyS and Ig. In 7 patients who required arthrocentesis of a swollen knee, coincident serum and synovial fluid samples were assayed for BLyS. Medical charts were retrospectively reviewed for elevated autoantibody titers and proteinuria within a 1-month period before or after collection of sera for BLyS and Ig determination. Sera concurrently collected from 48 normal healthy subjects served as controls. RESULTS: Serum BLyS levels were elevated in 38 of 185 patients (21%) and correlated significantly with serum IgG levels. Serum BLyS levels did not correlate with the patients' age, sex, race, or medications, but correlated positively with anti-double-stranded DNA antibody titers among SLE patients and with rheumatoid factor titers among seropositive RA patients. In contrast, serum BLyS levels correlated inversely with nephrotic-range proteinuria among SLE patients. In every case tested, BLyS levels in clinically inflamed synovial fluids were greater than those in simultaneously obtained sera. CONCLUSION: BLyS may be an important factor in driving polyclonal hypergammaglobulinemia and elevated autoantibody titers in patients with systemic immune-based rheumatic diseases. Local production of BLyS in the joints may contribute to joint pathology. Patients with elevated serum BLyS levels may be ideal candidates for therapeutic targeting of BLyS.
Asunto(s)
Lupus Eritematoso Sistémico/sangre , Proteínas de la Membrana/sangre , Enfermedades Reumáticas/sangre , Adolescente , Adulto , Anciano , Animales , Anticuerpos Antinucleares/sangre , Factor Activador de Células B , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Inmunoglobulina G/sangre , Articulación de la Rodilla/metabolismo , Articulación de la Rodilla/patología , Lupus Eritematoso Sistémico/inmunología , Masculino , Proteínas de la Membrana/análisis , Persona de Mediana Edad , Estudios Retrospectivos , Enfermedades Reumáticas/inmunología , Factor Reumatoide/sangre , Líquido Sinovial/química , Líquido Sinovial/metabolismo , Factor de Necrosis Tumoral alfa/análisisRESUMEN
B-lymphocyte stimulator (BLyS) is a recently identified novel member of the tumor necrosis factor ligand superfamily shown to exist in a membrane-bound and soluble form. BLyS was found to be specifically expressed on cells of myeloid lineage and to selectively stimulate B-lymphocyte proliferation and immunoglobulin production. The expression of a cytokine involved in potentiation of humoral immune responses, such as BLyS, is expected to be strictly controlled. The goal of the present study was to examine regulation of BLyS levels in monocytic cells in response to cytokines and during their differentiation to macrophages and dendritic cells. The presence of BLyS on the cell surface and in the culture medium of both normal blood monocytes and on tumor cells of myelomonocytic origin was demonstrated. BLyS gene expression and levels of membrane-associated and soluble BLyS were found to be regulated by cytokines, in particular interferon (IFN)-gamma and to a lesser extent interleukin-10 (IL-10). The expression of BLyS on monocyte membranes was retained following differentiation into macrophages, but detection on the surface of monocyte-derived dendritic cells required stimulation with IFN-gamma. Both IFN-gamma and IL-10 enhanced the release of soluble BLyS that was active in B-cell proliferation assays. Cells transfected with BLyS complementary DNA mutated in a predicted cleavage site failed to release BLyS into the culture medium, thereby suggesting that soluble BLyS was derived from the membrane form. These results provide further support for an important role for BLyS expressed in myeloid cells in B-cell expansion and antibody responses.
Asunto(s)
Proteínas de la Membrana/genética , Células Mieloides/metabolismo , Factor de Necrosis Tumoral alfa/genética , Anticuerpos/metabolismo , Factor Activador de Células B , Linfocitos B/citología , División Celular/efectos de los fármacos , Citocinas/farmacología , Células Dendríticas/química , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Macrófagos/química , Proteínas de la Membrana/inmunología , Proteínas de la Membrana/metabolismo , Monocitos/metabolismo , Péptido Hidrolasas/metabolismo , ARN Mensajero/metabolismo , Solubilidad , Células Tumorales Cultivadas , Factor de Necrosis Tumoral alfa/inmunología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
B lymphocyte stimulator (BLyS) is a novel member of the TNF family of proteins expressed by myeloid cells as membrane-bound and soluble forms. BLyS was shown to act specifically on B cells, inducing proliferation and immunoglobulin production both in vitro and in vivo. The present study was undertaken to characterize binding of radiolabeled BLyS to its cognate receptor on human B lymphocytes and examine intracellular events initiated by BLyS binding. Similar to other TNF family members, BLyS is present in solution as a homotrimer as determined by gel filtration chromatography and light scattering analysis. BLyS binding to B cells is specific as other TNF family members tested did not compete for(125)I-BLyS binding. Analysis of equilibrium binding of(125)I-labeled BLyS to purified human tonsillar B cells demonstrated saturable binding. Scatchard analysis of the binding data revealed a single class of high-affinity binding on human B cells with approximately 2600 binding sites per cell and an apparent dissociation constant (K(D)) of about 0.1 nM. In addition we report that BLyS binding to B cells results in the activation of NF-kappaB and the Ets family transcription factor, ELF-1, and in the induction of mRNA for Polo-like kinase (PLK).
Asunto(s)
Linfocitos B/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas de la Membrana/metabolismo , FN-kappa B/metabolismo , Factores de Transcripción/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Factor Activador de Células B , Sitios de Unión , Reactivos de Enlaces Cruzados , Humanos , Técnicas In Vitro , Radioisótopos de Yodo , Cinética , Proteínas de la Membrana/química , Unión Proteica , Estructura Cuaternaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Transducción de Señal , Factor de Necrosis Tumoral alfa/químicaRESUMEN
A major problem with the use of umbilical cord/placental blood (UCB) is the limited blood volume that can be collected from a single donor. In this study, we evaluated a novel system for the collection of UCB and analyzed the kinetics of output of hematopoietic stem cells in the collected blood. Sequential UCB fractions were collected from 48 placentas by gravity following common procedures. When UCB flow was ended, collection was continued using the device. Nucleated cell (NC) density in each fraction was evaluated and the expression of CD34, CD38 and other hematopoietic markers was assessed by flow cytometry. The total collected volume was 60.9 +/- 26.2 ml (mean +/- SD, range 17-141.5). The device yield (volume collected using the device/total volume) was 26.5 +/- 15.1%. No significant difference was observed in NC count in sequential fractions. A significant increase in CD34(+) cell content in sequential fractions and a 2.07 +/- 1.18-fold increase in the percentage of CD34(+) cells in the last versus first fraction were observed. Furthermore, within the CD34(+) population, the percentage of CD38(-) pluripotent stem cells in the first fraction was 3.24 +/- 1.39, while in the last fraction it raised to 34.43 +/- 22.62. Thus, at the end of a collection performed following current procedures, further blood rich in the most primitive progenitor cells can be recovered. Therefore, the optimization and standardization of collection procedures are required to obtain maximal recovery from each placenta and increase the percentage of UCB units suitable for clinical use.
Asunto(s)
Antígenos CD , Separación Celular/métodos , Células Madre Hematopoyéticas/citología , ADP-Ribosil Ciclasa , ADP-Ribosil Ciclasa 1 , Antígenos CD34 , Antígenos de Diferenciación , Volumen Sanguíneo , Femenino , Sangre Fetal/citología , Citometría de Flujo/métodos , Células Madre Hematopoyéticas/inmunología , Humanos , Recuento de Leucocitos , Glicoproteínas de Membrana , NAD+ NucleosidasaRESUMEN
Herein, we report that Autographa californica nucleopolyhedrovirus, a member of the Baculoviridae family, is capable of stimulating antiviral activity in mammalian cells. Baculoviruses are not pathogenic to mammalian cells. Nevertheless, live baculovirus is shown here to induce interferons (IFN) from murine and human cell lines and induces in vivo protection of mice from encephalomyocarditis virus infection. Monoclonal antibodies specific for the baculovirus envelope gp67 neutralize baculovirus-dependent IFN production. Moreover, UV treatment of baculovirus eliminates both infectivity and IFN-inducing activity. In contrast, the IFN-inducing activity of the baculovirus was unaffected by DNase or RNase treatment. These data demonstrate that IFN production can be induced in mammalian cells by baculovirus even though the cells fail to serve as a natural host for an active viral infection. Baculoviruses, therefore, provide a novel model in which to study at least one alternative mechanism for IFN induction in mammalian cells.
Asunto(s)
Interferón-alfa/inmunología , Interferón beta/inmunología , Nucleopoliedrovirus/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Células COS , Infecciones por Cardiovirus/inmunología , Infecciones por Cardiovirus/prevención & control , ADN/inmunología , Femenino , Humanos , Interferón-alfa/biosíntesis , Interferón beta/biosíntesis , Lipopolisacáridos/inmunología , Mamíferos , Virus Maus Elberfeld/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Mariposas Nocturnas/virología , Pruebas de Neutralización , ARN Bicatenario/inmunologíaRESUMEN
This study describes the multilineage differentiation pattern of purified CD34+ stem cells obtained from human umbilical cord blood. CD34+ cells were collected from 49 umbilical cord blood samples. Following immunomagnetic purification, cells were double stained with anti CD34 and CD71, CD61, CD7, CD19, CD33, CD36 and triple stained with anti CD34, CD38 and HLA-DR. Analysis were performed using a FACScan flow cytometer. After purification, the mean CD34+ cells' purity was 85.49 +/- 7.08%. Several subpopulations of umbilical cord blood CD34+ cells were identified indicating different lineage commitment. The majority of CD34+ cells expressed both CD38 and HLA-DR (91.74 +/- 3.76%), while those lacking CD38 were 3.43 +/- 2.12% (CD38-DR+) and 1.81 +/- 1.54% (CD38-DR-). These data were compared to the expression of lineage commitment markers on purified CD34+ cells from 5 mobilized peripheral blood samples. The percentage of peripheral blood CD34+CD38-DR+) and CD34+CD38-DR- cells was significantly lower than umbilical cord blood, 0.24 +/- 0.18% and 0.04 +/- 0.03% respectively. The knowledge and standardized of umbilical cord blood CD34+ cells phenotype is critical since umbilical cord blood volume is limited. The homogeneity of CD34+ subpopulation phenotype suggests that monitoring of lineage differentiation antigens may not be relevant for clinical use of umbilical cord blood samples. However, the observed higher percentage of pluripotent CD34+38- stem cells in umbilical cord blood compared to peripheral blood, that might explain the successful clinical use of umbilical cord blood even when low number of cells are used, candidates these antigens as the predictive parameter for clinical use of umbilical cord blood samples.
Asunto(s)
Antígenos CD34/sangre , Sangre Fetal/citología , Antígenos CD/sangre , Antígenos CD34/inmunología , Separación Celular , Sangre Fetal/inmunología , Citometría de Flujo , Humanos , Separación Inmunomagnética/métodos , Leucaféresis , Leucocitos Mononucleares/inmunología , Fenotipo , Células Madre/citología , Células Madre/inmunologíaRESUMEN
The tumor necrosis factor (TNF) superfamily of cytokines includes both soluble and membrane-bound proteins that regulate immune responses. A member of the human TNF family, BLyS (B lymphocyte stimulator), was identified that induced B cell proliferation and immunoglobulin secretion. BLyS expression on human monocytes could be up-regulated by interferon-gamma. Soluble BLyS functioned as a potent B cell growth factor in costimulation assays. Administration of soluble recombinant BLyS to mice disrupted splenic B and T cell zones and resulted in elevated serum immunoglobulin concentrations. The B cell tropism of BLyS is consistent with its receptor expression on B-lineage cells. The biological profile of BLyS suggests it is involved in monocyte-driven B cell activation.
Asunto(s)
Linfocitos B/inmunología , Activación de Linfocitos , Proteínas de la Membrana/fisiología , Monocitos/inmunología , Factor de Necrosis Tumoral alfa/fisiología , Secuencia de Aminoácidos , Animales , Factor Activador de Células B , Receptor del Factor Activador de Células B , Subgrupos de Linfocitos B/inmunología , Línea Celular , Células Cultivadas , Humanos , Inmunoglobulinas/sangre , Interferón gamma/farmacología , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Proteínas de la Membrana/farmacología , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Receptores de Citocinas/metabolismo , Receptores del Factor de Necrosis Tumoral/metabolismo , Proteínas Recombinantes/farmacología , Alineación de Secuencia , Especificidad de la Especie , Factor de Necrosis Tumoral alfa/química , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/farmacología , Regulación hacia ArribaRESUMEN
After in vivo inoculation with abl/myc- and raf/myc-containing retroviruses, BALB/c mice predominantly develop late stage B cell tumors (plasmacytomas) and less frequently develop earlier B-lineage tumors while DBA/2 mice do not develop B-lineage tumors. We have investigated the in vitro tumorigenic potential of these viruses using cultured normal pre-B cell lymphocytes from both BALB/c and DBA/2 mice. Interestingly, both viruses infect cultured pre-B lymphocytes from both mouse strains. Following infection, IL-7 dependent pre-B cells become independent of normal in vitro growth requirements within 24 h and can rapidly form in vivo pre-B lymphomas in both mouse strains. Mechanisms mediating loss of IL-7 dependence are different depending on whether the raf or abl gene is present in myc-containing viruses. IL-7 JAK-STAT signaling is constitutively active in abl/myc induced pre-B cell tumors. In contrast, IL-7 JAK-STAT signaling is not constitutive in raf/myc induced pre-B cell tumors, demonstrating that subversion of this component of IL-7 signal transduction is not obligatory for pre-B cell transformation or loss of IL-7 dependence.
Asunto(s)
Linfocitos B , Transformación Celular Neoplásica , Transformación Celular Viral , Interleucina-7/metabolismo , Proteínas Proto-Oncogénicas c-abl/genética , Proteínas Proto-Oncogénicas c-raf/genética , Transducción de Señal , Animales , Células Cultivadas , Expresión Génica , Vectores Genéticos , Inmunidad Innata , Interleucina-7/farmacología , Linfoma de Células B/genética , Linfoma de Células B/metabolismo , Linfoma de Células B/virología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos DBA , Ratones SCID , Fenotipo , Proteínas Proto-Oncogénicas c-myc/genética , Provirus/genética , Retroviridae/fisiología , Células Madre , Integración ViralRESUMEN
Murine plasma cell tumors share a number of common features with human multiple myeloma, suggesting their possible use as a model for this disease. However, one major difference between the two is the peritoneal localization of murine tumors as opposed to bone marrow residence of malignant plasma cells in early stages of multiple myeloma. We have thus examined the ability of murine plasmacytoma to produce disseminated growth similar to that seen in myeloma or other lymphoid neoplasias. Of four murine cell lines evaluated, all were demonstrated to effect highly metastatic disease involving multiple organs, although variation was observed between lines. A temporal analysis was accordingly performed with the S107 line to assess the pattern of cellular localization. Both light microscopy and PCR analysis revealed that engraftment of plasma cells occurs first in the bone marrow, followed by dissemination to other sites including the spleen, lung, and liver. Cells passaged in vivo through the bone marrow display an entirely different metastatic pattern with no homing preference to bone marrow or any other organ, suggesting the occurrence of a phenotypic change. Microscopic osteolytic lesions were observed adjacent to plasma cell tumor masses in the bone marrow, indicating early stages of bone disease. These findings demonstrate previously unrecognized similarities between the murine and human diseases and suggest the use of this in vivo model for experimental approaches to the treatment of human disease.
Asunto(s)
Mieloma Múltiple/patología , Plasmacitoma/patología , Animales , Biomarcadores de Tumor/análisis , Médula Ósea/patología , Humanos , Cadenas Pesadas de Inmunoglobulina/análisis , Cadenas Pesadas de Inmunoglobulina/genética , Inyecciones Intraperitoneales , Ratones , Ratones Endogámicos BALB C , Proteínas de Mieloma/análisis , Proteínas de Mieloma/genética , Metástasis de la Neoplasia , Trasplante de Neoplasias , Especificidad de Órganos , Cavidad Peritoneal/patología , Células Plasmáticas/patología , Reacción en Cadena de la Polimerasa , Especificidad de la Especie , Células Tumorales CultivadasRESUMEN
Acute exposure of mammals to the environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) results in a diverse set of toxicologic and pathologic effects. The mechanism of some of these effects has been studied extensively in vitro and correlative studies have indicated the involvement of a transcription factor known as the aryl hydrocarbon receptor (AHR). However, a definitive association of the AHR with TCDD-mediated toxicity has been difficult to establish due to the diversity of effects and the ubiquitous expression of this receptor. In an effort to distinguish AHR-mediated TCDD toxicities from those resulting from alternative pathways, we have made use of the recently described AHR-deficient mouse that was generated by locus-specific homologous recombination in embryonic stem cells. Present studies demonstrate that AHR-deficient mice are relatively unaffected by doses of TCDD (2000 micrograms/kg) 10-fold higher than that found to induce severe toxic and pathologic effects in littermates expressing a functional AHR. Analyses of liver, thymus, heart, kidney, pancreas, spleen, lymph nodes, and uterus from AHR-deficient mice identified no significant TCDD-induced lesions. The resistance of AHR-deficient mice to TCDD-induced thymic atrophy appears restricted to processes involving AHR since the corticosteroid dexamethasone rapidly and efficiently induced cortical depletion in both AHR-deficient and normal littermate control mice. Taken together these results suggest that the pathological changes induced by TCDD in the liver and thymus are mediated entirely by the AHR. However, it is important to note that at high doses of TCDD, AHR-deficient mice displayed limited vasculitis and scattered single cell necrosis in their lungs and livers, respectively. The mechanism(s) responsible for these apparently receptor-independent processes remain unclear but may involve novel, alternative pathways for TCDD-induced toxicity.
Asunto(s)
Dibenzodioxinas Policloradas/toxicidad , Receptores de Hidrocarburo de Aril/deficiencia , Timo/efectos de los fármacos , Animales , Dexametasona/uso terapéutico , Femenino , Inyecciones Intraperitoneales , Hígado/efectos de los fármacos , Hígado/patología , Pulmón/efectos de los fármacos , Pulmón/patología , Ratones , Ratones Noqueados , Tamaño de los Órganos/efectos de los fármacos , Timo/patologíaRESUMEN
The development of murine plasma cell tumors induced by raf/myc containing retroviruses is facilitated by T cells and completely dependent on IL-6. To determine whether kinases with differing specificities reflect alternative biochemical pathways in B cell tumorigenesis, we have employed an abl/myc containing retrovirus to assess neoplastic development. In contrast with raf/myc, abl/myc disease is T cell and IL-6 independent. An examination of the IL-6 signal transduction pathway reveals that this pathway, as defined by activation of Stat3, is inducible by IL-6 in raf/myc tumors but constitutively activated in abl/myc tumors. These findings provide a mechanism for the derivation of cytokine-independent plasma cell tumors and suggest that both IL-6-dependent and independent tumors may arise in vivo depending on the particular mutational events incurred during tumorigenesis.
Asunto(s)
Genes abl/inmunología , Interleucina-6/inmunología , Plasmacitoma/genética , Infecciones por Retroviridae/genética , Transducción de Señal/genética , Transducción de Señal/inmunología , Infecciones Tumorales por Virus/genética , Animales , Secuencia de Bases , Interleucina-6/genética , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos DBA , Ratones Desnudos , Datos de Secuencia Molecular , Fenotipo , Plasmacitoma/inmunología , Infecciones por Retroviridae/inmunología , Infecciones Tumorales por Virus/inmunologíaRESUMEN
We show here that the mode of cell death in IL-6-starved T1165 and T1198 plasmacytoma cell lines is apoptosis, and that it can be suppressed by phorbol ester (PMA) treatment in a protein kinase C (PKC)-mediated process that involves alpha and/or delta isozymes. PMA-induced PKC activation, but not the depletion that follows it, participates in the suppression of apoptosis. Extended PKC activation is necessary but not sufficient for the apoptosis suppression. In addition, the cells must be in a "competent" state, which appears not to be determined by PKC. We observed two points of "competence" during the time between withdrawal of IL-6 and the start of massive cell death: one, immediately after withdrawal, and another, just before onset of apoptosis, at the time corresponding to maximal accumulation of cells in a G0/G1 block imposed by IL-6 withdrawal. Treatment with PMA and other PKC activators resulted in a shift of the cell population to S phase, lifting the G0/G1 block. We propose a model in which cells are rescued in a certain stage of the G1 phase of cell cycle. Death suppression occurs when a transient PMA-induced PKC activation occurs when a significant number of cells are in this part of G1, allowing them to pass the restriction point safely without initiating the cell death program.
Asunto(s)
Apoptosis/efectos de los fármacos , Ciclo Celular , Interleucina-6/farmacología , Proteína Quinasa C/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Alcaloides , Animales , Benzofenantridinas , Ciclo Celular/efectos de los fármacos , Línea Celular , Citosol/enzimología , Activación Enzimática , Inhibidores Enzimáticos/farmacología , Indoles/farmacología , Isoenzimas/antagonistas & inhibidores , Isoenzimas/metabolismo , Lactamas/farmacología , Maleimidas/farmacología , Ratones , Fenantridinas/farmacología , Ésteres del Forbol/farmacología , Plasmacitoma , Proteína Quinasa C/antagonistas & inhibidores , Acetato de Tetradecanoilforbol/análogos & derivados , Células Tumorales CultivadasRESUMEN
Peripheral lymphoid tissues contain a fibroblastic cell type referred to as stromal cells or reticulum cells which interact with lymphocytes as part of the lymphoid microenvironment. After isolation from human tonsils and expansion in vitro we analyzed the surface phenotype, extracellular matrix components, cytoskeletal products, cytokine production, binding and functional interaction with B lymphocytes of in vitro cultured stromal cells (HTSC) both in resting condition and after activation with tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma. Our results show that HTSC do not express specific myeloid, lymphoid, endothelial or epithelial markers. HTSC express CD54 (ICAM-1), CD49a (VLA-1), CD49b (VLA-2), CD49c (VLA-3), CD49e (VLA-5), CD49f (VLA-6), CD29, CD51, CD44 and produce vinculin, beta-tubulin, alpha-actin, vimentin, fibronectin, laminin and collagen types I, III and IV. Activation of HTSC up-regulated CD54 (ICAM-1) and induced HLA-DR and CD106 (VCAM-1). HTSC constitutively produce interleukin (IL)-6 which is enhanced upon activation with TNF-alpha. IL-8 and granulocyte/macrophage colony-stimulating factor are detected only in the supernatants of activated HTSC. Reverse transcriptase polymerase chain reaction analysis revealed that HTSC display mRNA for IL-1 alpha, leukemia inhibitory factor and IL-7. The adhesion of tonsillar B lymphocytes to activated HTSC is mediated by CD11a/CD18 and CD54. Furthermore, HTSC can induce maximal proliferation of IL-2-activated B lymphocytes cocultured in direct cell-cell contact with HTSC. These results clearly distinguish in vitro cultured HTSC from common fibroblasts and other non-lymphoid elements present in the lymphoid parenchyma, such as follicular dendritic cells, and show that HTSC actively participate in the lymphoid microenvironment. In vitro cultures of HTSC could therefore be a useful model system for detailed analysis of the interactions between stromal cells and lymphocytes under physiological and pathological conditions.
Asunto(s)
Linfocitos B/inmunología , Activación de Linfocitos , Tonsila Palatina/inmunología , Células del Estroma/clasificación , Células del Estroma/inmunología , Antígenos de Superficie/biosíntesis , Linfocitos B/citología , Secuencia de Bases , Adhesión Celular/inmunología , Células Cultivadas , Niño , Preescolar , Citocinas/biosíntesis , Proteínas del Citoesqueleto/análisis , Proteínas de la Matriz Extracelular/análisis , Humanos , Inmunofenotipificación , Datos de Secuencia Molecular , Tonsila Palatina/citología , Células del Estroma/citologíaRESUMEN
Interleukin (IL) 6 has been suggested to be the major cytokine responsible for proliferation of neoplastic plasma cells in both human myeloma and mouse plasmacytoma. Much of the evidence supporting this suggestion is derived from in vitro studies in which the survival or proliferation of some plasma cell tumors has been found to be IL-6 dependent. However, it remains unclear whether this dependency is the consequence of in vivo or in vitro selective pressures that preferentially expand IL-6-responsive tumor cells, or whether it reflects a critical in vivo role for IL-6 in plasma cell neoplasia. To address this question, we have attempted to induce plasma cell tumors in normal mice and in IL-6-deficient mice generated by introduction of a germline-encoded null mutation in the IL-6 gene. The results demonstrate that mice homozygous (+/+) or heterozygous (+/-) for the wild-type IL-6 allele yield the expected incidences of plasma cell tumors. In contrast, mice homozygous for the IL-6-null allele (-/-) are completely resistant to plasma cell tumor development. These studies define the essential role of IL-6 in the development of B lineage tumors in vivo and provide experimental support for continued efforts to modulate this cytokine in the treatment of appropriate human B cell malignancies.
Asunto(s)
Linfocitos B/fisiología , Interleucina-6/fisiología , Mieloma Múltiple/fisiopatología , Plasmacitoma/fisiopatología , Alelos , Animales , Secuencia de Bases , División Celular , Cocarcinogénesis , Cruzamientos Genéticos , Femenino , Citometría de Flujo , Gammaretrovirus/genética , Gammaretrovirus/fisiología , Predisposición Genética a la Enfermedad , Genotipo , Hibridación in Situ , Interleucina-6/deficiencia , Interleucina-6/genética , Linfoma de Células B/etiología , Linfoma de Células B/fisiopatología , Linfoma de Células T/etiología , Linfoma de Células T/fisiopatología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos , Ratones Noqueados , Datos de Secuencia Molecular , Mieloma Múltiple/etiología , Trasplante de Neoplasias , Oncogenes , Plasmacitoma/etiología , Reacción en Cadena de la Polimerasa , Terpenos/toxicidad , Infecciones Tumorales por Virus/virologíaRESUMEN
The aryl hydrocarbon (Ah) receptor (AHR) mediates many carcinogenic and teratogenic effects of environmentally toxic chemicals such as dioxin. An AHR-deficient (Ahr-/-) mouse line was constructed by homologous recombination in embryonic stem cells. Almost half of the mice died shortly after birth, whereas survivors reached maturity and were fertile. The Ahr-/- mice showed decreased accumulation of lymphocytes in the spleen and lymph nodes, but not in the thymus. The livers of Ahr-/- mice were reduced in size by 50 percent and showed bile duct fibrosis Ahr-/- mice were also nonresponsive with regard to dioxin-mediated induction of genes encoding enzymes that catalyze the metabolism of foreign compounds. Thus, the AHR plays an important role in the development of the liver and the immune system.
Asunto(s)
Inmunidad/fisiología , Hígado/fisiología , Receptores de Hidrocarburo de Aril/fisiología , Animales , Femenino , Regulación de la Expresión Génica/fisiología , Cirrosis Hepática Experimental/genética , Cirrosis Hepática Experimental/patología , Tejido Linfoide/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Hidrocarburo de Aril/genéticaRESUMEN
Major interest in the analysis of mature plasma cell neoplasias of mice and humans has focused on identification of precursor cells that give rise to mature malignant plasma cells. Although several laboratories have recently suggested that such cells are present in the granulomas of pristane-treated mice and the bone marrow of some multiple myeloma patients, the in vivo cellular interactions required for their differentiation into mature plasma cell tumors remains unclear. Given the extensive interactions of peripheral T cells and normal B cells, we assessed the potential role of T cells in plasma-cell tumor development, by using a myc, raf-containing retrovirus, J3V1, to induce plasmacytomas in normal BALB/c mice, T-cell-deficient nude mice, and T-cell-reconstituted nude mice. The B-lineage tumors arising in normal BALB/c mice were uniformly mature plasmacytomas, most of which secreted immunoglobulin. In contrast, nude mice yielded predominantly non-immunoglobulin-secreting B-cell lymphomas with a phenotype characteristic of peripheral B cells. T-cell reconstitution of nude mice prior to tumor induction resulted in a shift from B-cell lymphomas to plasmacytomas. These results imply that transformation can occur prior to terminal differentiation of B cells and that such transformed cells can be driven to terminal differentiation by peripheral T cells. These findings further suggest that, in human multiple myeloma, the ability of T cells to influence the differentiation state of transformed B cells may provide a mechanism by which malignant plasma cells found in the bone marrow could arise from clonotypically related less-mature B cells found in both the bone marrow and periphery.
Asunto(s)
Linfocitos B/patología , Transformación Celular Neoplásica , Plasmacitoma/patología , Linfocitos T/fisiología , Animales , Diferenciación Celular , Femenino , Humanos , Inmunocompetencia , Inmunoglobulinas/metabolismo , Linfoma de Células B/genética , Linfoma de Células B/patología , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Plasmacitoma/genética , Polimorfismo de Longitud del Fragmento de Restricción , RetroviridaeRESUMEN
Peyer's patch, peripheral lymph node, and mesenteric lymph node cells were transferred to immunodeficient SCID mice to assess the long-term (150-300 days) potential of these cells to repopulate the host's immune system. Results demonstrate that, irrespective of donor population, total serum Ig and isotype distribution appear normal within 4 weeks of reconstitution and remain at normal levels for up to one year following cell transfer. At the cellular level, each donor population reconstitutes splenic T and B cell compartments in a progressive and quantitatively indistinguishable manner. Immunohistological analyses of reconstituted mice indicate that, although some qualitative differences are evident, normal splenic composition and architecture are observed. In contrast, gut reconstitution varies significantly with donor population. Peyer's patch cells yield normal-appearing gut tissue with extensive infiltration of the lamina propria and intraepithelial compartments by T cells and IgA-secreting plasma cells. Peripheral lymph node cells give rise to T cells found almost exclusively in the lamina propria, while IgA secreting plasma cells are rarely detected. The course and extent of reconstitution further suggest that all donor populations contain long-lived T and B cells as well as self-renewing lymphocytes capable of extensive expansion. This latter observation has potentially important implications for both transplantation biology and gene therapy applications.