RESUMEN

Chronic lymphocytic leukemia (CLL) is a clonal disease of B lymphocytes manifesting as an absolute lymphocytosis in the blood. However, not all lymphocytoses are leukemic. In addition, first-degree relatives of CLL patients have an ~15 % chance of developing a precursor condition to CLL termed monoclonal B cell lymphocytosis (MBL), and distinguishing CLL and MBL B lymphocytes from normal B cell expansions can be a challenge. Therefore, we selected FMOD, CKAP4, PIK3C2B, LEF1, PFTK1, BCL-2, and GPM6a from a set of genes significantly differentially expressed in microarray analyses that compared CLL cells with normal B lymphocytes and used these to determine whether we could discriminate CLL and MBL cells from B cells of healthy controls. Analysis with receiver operating characteristics and Bayesian relevance determination demonstrated good concordance with all panel genes. Using a random forest classifier, the seven-gene panel reliably distinguished normal polyclonal B cell populations from expression patterns occurring in pre-CLL and CLL B cell populations with an error rate of 2 %. Using Bayesian learning, the expression levels of only two genes, FMOD and PIK3C2B, correctly distinguished 100 % of CLL and MBL cases from normal polyclonal and mono/oligoclonal B lymphocytes. Thus, this study sets forth effective computational approaches that distinguish MBL/CLL from normal B lymphocytes. The findings also support the concept that MBL is a CLL precursor.

Asunto(s)

Linfocitos B/fisiología , Fosfatidilinositol 3-Quinasas Clase II/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Leucemia Linfocítica Crónica de Células B/diagnóstico , Linfocitosis/diagnóstico , Lesiones Precancerosas/diagnóstico , Valor Predictivo de las Pruebas , Proteoglicanos/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Fosfatidilinositol 3-Quinasas Clase II/genética , Biología Computacional , Diagnóstico Diferencial , Proteínas de la Matriz Extracelular/genética , Fibromodulina , Regulación Neoplásica de la Expresión Génica , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Activación de Linfocitos/genética , Linfocitosis/genética , Análisis por Micromatrices , Lesiones Precancerosas/genética , Pronóstico , Proteoglicanos/genética , Transcriptoma

RESUMEN

Retinoic acid (RA) can induce growth arrest and neuronal differentiation of neuroblastoma cells and has been used in clinic for treatment of neuroblastoma. It has been reported that RA induces the expression of several HOXD genes in human neuroblastoma cell lines, but their roles in RA action are largely unknown. The HOXD cluster contains nine genes (HOXD1, HOXD3, HOXD4, and HOXD8-13) that are positioned sequentially from 3' to 5', with HOXD1 at the 3' end and HOXD13 the 5' end. Here we show that all HOXD genes are induced by RA in the human neuroblastoma BE(2)-C cells, with the genes located at the 3' end being activated generally earlier than those positioned more 5' within the cluster. Individual induction of HOXD8, HOXD9, HOXD10 or HOXD12 is sufficient to induce both growth arrest and neuronal differentiation, which is associated with downregulation of cell cycle-promoting genes and upregulation of neuronal differentiation genes. However, induction of other HOXD genes either has no effect (HOXD1) or has partial effects (HOXD3, HOXD4, HOXD11 and HOXD13) on BE(2)-C cell proliferation or differentiation. We further show that knockdown of HOXD8 expression, but not that of HOXD9 expression, significantly inhibits the differentiation-inducing activity of RA. HOXD8 directly activates the transcription of HOXC9, a key effector of RA action in neuroblastoma cells. These findings highlight the distinct functions of HOXD genes in RA induction of neuroblastoma cell differentiation.

Asunto(s)

Diferenciación Celular/genética , Regulación Neoplásica de la Expresión Génica , Proteínas de Homeodominio/genética , Familia de Multigenes/genética , Neuroblastoma/genética , Neuroblastoma/patología , Ciclo Celular/efectos de los fármacos , Ciclo Celular/genética , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Quinasas Ciclina-Dependientes/genética , Quinasas Ciclina-Dependientes/metabolismo , Ciclinas/genética , Ciclinas/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genes Relacionados con las Neoplasias , Proteínas de Homeodominio/metabolismo , Humanos , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Tretinoina/farmacología

RESUMEN

BACKGROUND: Abnormal NF-κB2 activation has been implicated in the pathogenesis of multiple myeloma, a cancer of plasma cells. However, a causal role for aberrant NF-κB2 signaling in the development of plasma cell tumors has not been established. Also unclear is the molecular mechanism that drives the tumorigenic process. We investigated these questions by using a transgenic mouse model with lymphocyte-targeted expression of p80HT, a lymphoma-associated NF-κB2 mutant, and human multiple myeloma cell lines. METHODS: We conducted a detailed histopathological characterization of lymphomas developed in p80HT transgenic mice and microarray gene expression profiling of p80HT B cells with the goal of identifying genes that drive plasma cell tumor development. We further verified the significance of our findings in human multiple myeloma cell lines. RESULTS: Approximately 40% of p80HT mice showed elevated levels of monoclonal immunoglobulin (M-protein) in the serum and developed plasma cell tumors. Some of these mice displayed key features of human multiple myeloma with accumulation of plasma cells in the bone marrow, osteolytic bone lesions and/or diffuse osteoporosis. Gene expression profiling of B cells from M-protein-positive p80HT mice revealed aberrant expression of genes known to be important in the pathogenesis of multiple myeloma, including cyclin D1, cyclin D2, Blimp1, survivin, IL-10 and IL-15. In vitro assays demonstrated a critical role of Stat3, a key downstream component of IL-10 signaling, in the survival of human multiple myeloma cells. CONCLUSIONS: These findings provide a mouse model for human multiple myeloma with aberrant NF-κB2 activation and suggest a molecular mechanism for NF-κB2 signaling in the pathogenesis of plasma cell tumors by coordinated regulation of plasma cell generation, proliferation and survival.

Asunto(s)

Diferenciación Celular/genética , Mutación , Subunidad p52 de NF-kappa B/genética , Plasmacitoma/genética , Transducción de Señal , Animales , Proteínas Sanguíneas/metabolismo , Línea Celular Tumoral , Proliferación Celular , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Ratones , Ratones Transgénicos , Subunidad p52 de NF-kappa B/metabolismo , Plasmacitoma/metabolismo , Plasmacitoma/patología , Factor de Transcripción STAT3/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo

RESUMEN

Differentiation status in neuroblastoma strongly affects clinical outcomes and inducing differentiation is a treatment strategy in this disease. However, the molecular mechanisms that control neuroblastoma differentiation are not well understood. Here, we show that high-level HOXC9 expression is associated with neuroblastoma differentiation and is prognostic for better survival in neuroblastoma patients. HOXC9 induces growth arrest and neuronal differentiation in neuroblastoma cells by directly targeting both cell-cycle-promoting and neuronal differentiation genes. HOXC9 expression is upregulated by retinoic acid (RA), and knockdown of HOXC9 expression confers resistance to RA-induced growth arrest and differentiation. Moreover, HOXC9 expression is epigenetically silenced in RA-resistant neuroblastoma cells, and forced HOXC9 expression is sufficient to inhibit their proliferation and tumorigenecity. These findings identified HOXC9 as a key regulator of neuroblastoma differentiation and suggested a therapeutic strategy for RA-resistant neuroblastomas through epigenetic activation of HOXC9 expression.

Asunto(s)

Diferenciación Celular , Proteínas de Homeodominio/fisiología , Neuroblastoma/patología , Neuronas/citología , Línea Celular Tumoral , Proliferación Celular , Epigénesis Genética , Fase G1 , Proteínas de Homeodominio/genética , Humanos , Neuroblastoma/mortalidad , Pronóstico , Regiones Promotoras Genéticas , Tretinoina/farmacología

RESUMEN

Recent studies from our laboratory have shown that intravenous administration of human umbilical cord blood (HUCB) mononuclear cells to mice improved blood glucose levels, survival, atherosclerosis and prostate cancer. In this study, we examined the effect of HUCB cells on the production of IL-10 levels in IL-10 knockout mice. It has been proposed that administration of IL-10 may be beneficial in the treatment of inflammatory bowl disease. The results show that mice treated with HUCB cells (100 x 10(6)) produce IL-10, as demonstrated by both qualitative and quantitative analyses, and that the levels of this cytokine persisted until the mice were sacrificed (5.5 months after administration). Immunohistochemical staining of the intestine using HuNu antibody cocktail demonstrated the presence of HUCB cells in the knockout mouse. Although the mice did not receive any immunosuppression, there was no evidence of graft-versus-host disease. Our data suggest that HUCB cells are capable of producing IL-10, and the use of these cells or HUCB may be indicated in the treatment of certain human diseases.

Asunto(s)

Trasplante de Células Madre de Sangre del Cordón Umbilical , Sangre Fetal/metabolismo , Enfermedades Inflamatorias del Intestino/inmunología , Interleucina-10/metabolismo , Mucosa Intestinal/metabolismo , Leucocitos Mononucleares/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Sangre Fetal/citología , Sangre Fetal/inmunología , Sangre Fetal/trasplante , Humanos , Terapia de Inmunosupresión , Enfermedades Inflamatorias del Intestino/sangre , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/terapia , Interleucina-10/sangre , Interleucina-10/genética , Intestinos/inmunología , Intestinos/patología , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/trasplante , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Embarazo , Quimera por Trasplante

RESUMEN

Since its discovery in follicular lymphoma cells at the breakpoint t(14;18), Bcl-2 has been studied extensively in many basic and clinical science settings. Bcl-2 can locate as an integral mitochondrial membrane component, where its primary role is to block apoptosis by maintaining membrane integrity. Here we show that Bcl-2 also can position on the outer cell surface membrane of B cells from patients with chronic lymphocytic leukemia (B-CLL) and certain other leukemias that do not classically possess the chromosomal breakpoint t(14;18). Although low levels of Bcl-2 can be detected on the surface membrane of apparently healthy leukemic and normal B cells, expression of Bcl-2 correlates best with spontaneous or induced apoptosis. Notably, upon induction of apoptosis, B-CLL cells were much more efficient in upregulating surface Bcl-2 than normal B cells. It is not clear if this surface membrane expression is a passive consequence of the apoptotic process or an active attempt by the B cell to abort cell death by stabilizing the plasma membrane.

Asunto(s)

Linfocitos B/metabolismo , Membrana Celular/metabolismo , Leucemia de Células B/patología , Leucemia Linfocítica Crónica de Células B/patología , Linfoma de Células B/patología , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Apoptosis , Linfocitos B/patología , Membrana Celular/ultraestructura , Células Cultivadas , Perfilación de la Expresión Génica , Regulación Leucémica de la Expresión Génica , Humanos , Leucemia de Células B/metabolismo , Leucemia Linfocítica Crónica de Células B/metabolismo , Linfoma de Células B/metabolismo , Microscopía Confocal , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Proto-Oncogénicas c-bcl-2/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas

RESUMEN

We investigated the possibility that manifestations of Lyme disease in certain hosts, such as arthritis and carditis, may be autoimmunity mediated due to molecular mimicry between the bacterium Borrelia burgdorferi and self-components. We first compared amino acid sequences of Streptococcus pyogenes M protein, a known inducer of antibodies that are cross-reactive with myosin, and B. burgdorferi and found significant homologies with OspA protein. We found that S. pyogenes M5-specific antibodies and sera from B. burgdorferi-infected mice reacted with both myosin and B. burgdorferi proteins by Western blots and enzyme-linked immunosorbent assay. To investigate the relationship between self-reactivity and the response to B. burgdorferi, NZB mice, models of autoimmunity, were infected. NZB mice infected with B. burgdorferi developed higher degrees of joint swelling and higher anti-B. burgdorferi immunoglobulin M cross-reactive responses than other strains with identical major histocompatibility complex (DBA/2 and BALB/c). These studies reveal immunological cross-reactivity and suggest that B. burgdorferi may share common epitopes which mimic self-proteins. These implications could be important for certain autoimmunity-susceptible individuals or animals who become infected with B. burgdorferi.

Asunto(s)

Artritis Infecciosa/etiología , Enfermedades Autoinmunes/complicaciones , Borrelia burgdorferi/inmunología , Enfermedad de Lyme/complicaciones , Enfermedad de Lyme/inmunología , Miocarditis/etiología , Animales , Anticuerpos Antibacterianos/sangre , Anticuerpos Antibacterianos/inmunología , Antígenos de Superficie/genética , Antígenos de Superficie/inmunología , Artritis Infecciosa/inmunología , Artritis Infecciosa/fisiopatología , Enfermedades Autoinmunes/inmunología , Enfermedades Autoinmunes/microbiología , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas de la Membrana Bacteriana Externa/inmunología , Vacunas Bacterianas , Reacciones Cruzadas , Humanos , Inmunoglobulina M/sangre , Inmunoglobulina M/inmunología , Lipoproteínas/genética , Lipoproteínas/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos DBA , Ratones Endogámicos NZB , Miocarditis/inmunología

RESUMEN

RNA interference, or RNAi, is designed to work by Watson-Crick base pairing and to result in a posttranscriptional block in protein synthesis. Antiapoptotic proteins are a major focus of cancer therapy and make attractive targets for RNAi. An IL-10 RNAi sequence was designed in accordance with Tuschl rules and was modeled to a hairpin configuration. In chronic lymphocytic leukemia (CLL), the most common leukemia in the Western world, the failure to undergo apoptosis may be responsible for the accumulation of malignant B-1 cells. Interleukin-10, despite controversy, has been shown to have antiapoptotic properties, and increased endogenous IL-10 production has been found in CLL by several labs. A malignant B-1 cell line, LNC, derived from an NZB mouse (a murine model for CLL) was utilized as a target for IL-10 RNAi. Our earlier studies of antisense IL-10 resulted in antiproliferative and proapoptotic effects. The cytotoxic effects of IL-10 RNAi were dose- and time-dependent, with an optimal dose 10-fold lower than that of antisense IL-10. IL-10 RNAi lowered IL-10 protein as measured by ELISA. 2 micro M IL-10 RNAi initiated a G2/M block and a decrease in the message for cdc25C, the M-phase inducer phosphatase. IL-10 RNAi efficiently induced apoptosis. Bcl7C, a member of the antiapoptotic Bcl family, was significantly down-regulated. IL-10 modulating Bcl7C expression represents a novel mechanism in the evasion of apoptosis. This approach, by itself or in conjunction with current therapies, merits consideration in similar B-cell malignancies.

Asunto(s)

Apoptosis/genética , Linfocitos B/patología , Interleucina-10/antagonistas & inhibidores , Interleucina-10/genética , Leucemia Linfocítica Crónica de Células B/terapia , Interferencia de ARN , ARN Interferente Pequeño/farmacología , Animales , Ciclo Celular/genética , Ensayo de Inmunoadsorción Enzimática , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/patología , Ratones , Ratones Endogámicos NZB , Modelos Moleculares , Conformación de Ácido Nucleico , ARN Bicatenario/genética , ARN Interferente Pequeño/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa

RESUMEN

BACKGROUND: The B-1 malignancy, CLL has been associated with a failure to undergo apoptosis and increased endogenous IL-10 production. This study was undertaken to identify IL-10 effects in the NZB murine model of CLL. MATERIALS AND METHODS: Antisense IL-10 was employed in vitro and in vivo to decrease IL-10 protein. Following treatment, cells were analyzed for alterations in cell cycle and RNA was studied for alterations in gene expression. Additional in vivo studies employed NZB mice in which the IL-10 gene was deleted. RESULTS: IL-10 (-/-) knockout NZB mice overwhelmingly failed to develop leukemia. In vitro antisense IL-10 treatment resulted in a G2/M block and apoptosis and in vivo treatment with antisense IL-10 increased the survival of mice. Microarray analysis indicated a significant role for IL-10 in cell cycle regulation via cdc25C up-regulation and decreased p47phox redox activity. CONCLUSION: In summary, IL-10 is a critical survival factor for malignant B cells via anti-apoptotic and cell cycle effects.