RESUMEN
Malaria is one of the most significant tropical diseases and remains a major challenge due to the lack of a broadly effective vaccine and parasite resistance to current drugs. This means there is a need for new drug candidates with novel modes of action. Aromatic bisamidines, such as furamidine (DB75), were initially developed as anti-Trypanosoma agents however as clinical trials with furamidine highlighted potential side effects they were not pursued further in that setting. Despite apparent cytotoxicity liabilities the potency of furamidine against Plasmodium falciparum makes it a promising scaffold for the development of new anti-Plasmodium agents with improved selectivity. In this study a bisamidine compound series based on furamidine was synthesized by introducing modifications at the furan core structure and terminal amidine groups. The activity of the derived compounds was tested in vitro against drug sensitive and resistant P. falciparum lines and a human cell line (HEK293 cells) to generate anti-Plasmodium structure-activity relationships and to provide preliminary selectivity data.
Asunto(s)
Antimaláricos/síntesis química , Antimaláricos/farmacología , Furanos/síntesis química , Furanos/farmacología , Plasmodium falciparum/efectos de los fármacos , Antimaláricos/química , Antimaláricos/toxicidad , Células CACO-2 , Técnicas de Química Sintética , Diseño de Fármacos , Furanos/química , Furanos/toxicidad , Células HEK293 , Humanos , Relación Estructura-ActividadRESUMEN
Current treatments for acute myeloid leukemia (AML) are designed to target rapidly dividing blast populations with limited success in eradicating the functionally distinct leukemia stem cell (LSC) population, which is postulated to be responsible for disease resistance and relapse. We have previously reported high miR-126 expression levels to be associated with a LSC-gene expression profile. Therefore, we hypothesized that miR-126 contributes to 'stemness' and is a viable target for eliminating the LSC in AML. Here we first validate the clinical relevance of miR-126 expression in AML by showing that higher expression of this microRNA (miR) is associated with worse outcome in a large cohort of older (⩾60 years) cytogenetically normal AML patients treated with conventional chemotherapy. We then show that miR-126 overexpression characterizes AML LSC-enriched cell subpopulations and contributes to LSC long-term maintenance and self-renewal. Finally, we demonstrate the feasibility of therapeutic targeting of miR-126 in LSCs with novel targeting nanoparticles containing antagomiR-126 resulting in in vivo reduction of LSCs likely by depletion of the quiescent cell subpopulation. Our findings suggest that by targeting a single miR, that is, miR-126, it is possible to interfere with LSC activity, thereby opening potentially novel therapeutic approaches to treat AML patients.
Asunto(s)
Leucemia Mieloide Aguda/terapia , MicroARNs/antagonistas & inhibidores , Nanopartículas/administración & dosificación , Células Madre Neoplásicas/fisiología , Animales , Metilación de ADN , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/mortalidad , Antígenos Comunes de Leucocito/antagonistas & inhibidores , Ratones , Ratones Endogámicos C57BL , MicroARNs/fisiología , Células Madre Neoplásicas/efectos de los fármacosRESUMEN
High levels of microRNA-155 (miR-155) are associated with poor outcome in acute myeloid leukemia (AML). In AML, miR-155 is regulated by NF-κB, the activity of which is, in part, controlled by the NEDD8-dependent ubiquitin ligases. We demonstrate that MLN4924, an inhibitor of NEDD8-activating enzyme presently being evaluated in clinical trials, decreases binding of NF-κB to the miR-155 promoter and downregulates miR-155 in AML cells. This results in the upregulation of the miR-155 targets SHIP1, an inhibitor of the PI3K/Akt pathway, and PU.1, a transcription factor important for myeloid differentiation, leading to monocytic differentiation and apoptosis. Consistent with these results, overexpression of miR-155 diminishes MLN4924-induced antileukemic effects. In vivo, MLN4924 reduces miR-155 expression and prolongs the survival of mice engrafted with leukemic cells. Our study demonstrates the potential of miR-155 as a novel therapeutic target in AML via pharmacologic interference with NF-κB-dependent regulatory mechanisms. We show the targeting of this oncogenic microRNA with MLN4924, a compound presently being evaluated in clinical trials in AML. As high miR-155 levels have been consistently associated with aggressive clinical phenotypes, our work opens new avenues for microRNA-targeting therapeutic approaches to leukemia and cancer patients.
Asunto(s)
Ciclopentanos/farmacología , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , MicroARNs/genética , Pirimidinas/farmacología , Secuencias Repetidas en Tándem/genética , Ubiquitinas/antagonistas & inhibidores , Tirosina Quinasa 3 Similar a fms/genética , Animales , Apoptosis/efectos de los fármacos , Western Blotting , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Inmunoprecipitación de Cromatina , Resistencia a Antineoplásicos , Femenino , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Humanos , Leucemia Mieloide Aguda/patología , Ratones , Ratones Endogámicos NOD , Ratones SCID , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Monocitos/patología , Proteína NEDD8 , FN-kappa B/genética , FN-kappa B/metabolismo , Regiones Promotoras Genéticas , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal/efectos de los fármacos , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
OBJECTIVE: To evaluate the impact of operator experience in volumetric measurements on intra- and inter-observer variability of lung volume estimation by 3-dimensional (3D) ultrasound and magnetic resonance imaging (MRI) in fetuses with congenital diaphragmatic hernia (CDH). METHODS: We conducted a retrospective single-center study. Total fetal lung volume (TFLV) was measured twice using 52 stored 3D-ultrasound volumes and 52 corresponding MRI exams by a fetal medicine specialist with no experience in volumetric measurements and two operators experienced in the respective techniques. Bland-Altman analysis was performed to evaluate intra- and inter-observer variability. RESULTS: Measurements of TFLV by 3D-ultrasound showed higher intra-observer variability compared with MRI irrespective of operator experience, with narrower 95% limits of agreement on MRI as compared with ultrasound measurements. Similarly, 3D-ultrasound showed higher inter-observer variability as compared with MRI. Finally the 95% limits of agreement at ultrasound were quite comparable for intra-observer variability between the experienced and the inexperienced operator. CONCLUSION: Our study shows that operator experience has an impact on the variability of TFLV and, in the absence of experience in volumetric measurements, estimation of TFLV in fetuses with CDH by 3D-ultrasound shows higher variability than MRI measurements, but even in experienced hands variability is greater for 3D-ultrasound.
Asunto(s)
Competencia Clínica , Hernias Diafragmáticas Congénitas/diagnóstico por imagen , Curva de Aprendizaje , Pulmón/diagnóstico por imagen , Imagen por Resonancia Magnética , Ultrasonografía Prenatal , Femenino , Hernias Diafragmáticas Congénitas/embriología , Humanos , Imagenología Tridimensional , Pulmón/embriología , Mediciones del Volumen Pulmonar , Variaciones Dependientes del Observador , Embarazo , Análisis de Regresión , Reproducibilidad de los Resultados , Estudios Retrospectivos , Ultrasonografía Prenatal/métodosRESUMEN
Ferroquine (FQ, SSR97193) is currently the most advanced organo-metallic drug candidate and about to complete phase II clinical trials as a treatment for uncomplicated malaria. This ferrocene-containing compound is active against both chloroquine-susceptible and chloroquine-resistant Plasmodium falciparum and P. vivax strains and/or isolates. This article focuses on the discovery of FQ, its antimalarial activity, the hypothesis of its mode of action, the current absence of resistance in vitro and recent clinical trials.
Asunto(s)
Aminoquinolinas/farmacología , Aminoquinolinas/uso terapéutico , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Compuestos Ferrosos/farmacología , Compuestos Ferrosos/uso terapéutico , Malaria Falciparum/tratamiento farmacológico , Malaria Vivax/tratamiento farmacológico , Aminoquinolinas/química , Animales , Antimaláricos/química , Cloroquina/química , Cloroquina/farmacología , Ensayos Clínicos Fase II como Asunto , Resistencia a Medicamentos , Compuestos Ferrosos/química , Humanos , Metalocenos , Plasmodium falciparum/efectos de los fármacos , Plasmodium vivax/efectos de los fármacosRESUMEN
In previous studies of the infection of rats by P. berghei Anka, we have shown that primary blood stage infection induced the expansion of CD4+ T cells and CD8+ T cells in adult resistant rats while the number of CD4+CD25+ cells was found to be higher in young susceptible rats. In this work, the respective contribution of each cell population was determined in young and adult rats treated with monoclonal antibodies. Down-regulation of surface CD25 molecules, including those expressed by CD4+ cells did not significantly enhance the capacity of young rats to control the development of erythrocytic stages or modify the course of infection in adult infected rats. However, we observed a significant loss of protection when adult rats were treated with anti-CD4 mAb (W3/25) with higher blood parasitemia levels and approximately 50% of rats succumbed to infection. More importantly and in contrast to earlier studies performed in mice, we found a significant increase in blood parasite levels and a significant delay in parasite clearance in adult rats treated with anti-CD8 mAb OX8, known to deplete CD8+ cells. These results suggest that CD8+ cells play a critical role in the development of immune responses in rats to control the replication of blood stage parasites.
Asunto(s)
Antígenos CD4/inmunología , Antígenos CD8/inmunología , Subunidad alfa del Receptor de Interleucina-2/inmunología , Malaria/veterinaria , Plasmodium berghei/inmunología , Enfermedades de los Roedores/parasitología , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacología , Antígenos CD4/sangre , Linfocitos T CD4-Positivos/inmunología , Antígenos CD8/sangre , Linfocitos T CD8-positivos/inmunología , Predisposición Genética a la Enfermedad , Inmunidad Innata/efectos de los fármacos , Subunidad alfa del Receptor de Interleucina-2/sangre , Malaria/inmunología , Malaria/parasitología , Ratones , Ratas , Ratas Endogámicas F344 , Enfermedades de los Roedores/sangre , Enfermedades de los Roedores/genética , Enfermedades de los Roedores/inmunologíaAsunto(s)
Hepacivirus , Hepatitis C/epidemiología , Hepatitis C/virología , ARN Viral/sangre , Genotipo , Hepacivirus/clasificación , Hepacivirus/genética , Hepacivirus/aislamiento & purificación , Humanos , Líbano/epidemiología , Prevalencia , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Riesgo , Análisis de Secuencia de ADNRESUMEN
The effects of a novel nutrient formulation Epican Forte (EF) were evaluated on proliferation and induction of apoptosis using non-cytotoxic concentrations against HTLV-1 positive (HuT-102 & C91-PL) and negative (CEM & Jurkat) cells. EF showed anti-proliferative effect as determined by MTT assay and TGF mRNA protein expression using RT-PCR. EF resulted in the down-regulation of TGF-alpha and an up-regulation in TGF-beta2. EF caused a significant increase in apoptotic cells in the preG1 phase. These results were confirmed using Cell Death ELISA and Annexin V-FITC. Induction of apoptosis was caused by an up-regulation of p53, p21 and Bax protein levels and a down-regulation of Bcl-2alpha protein expression level.
Asunto(s)
Acetilcisteína/farmacología , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Arginina/farmacología , Ácido Ascórbico/farmacología , Catequina/análogos & derivados , Cobre/farmacología , Virus Linfotrópico T Tipo 1 Humano/efectos de los fármacos , Lisina/farmacología , Manganeso/farmacología , Prolina/farmacología , Selenio/farmacología , Linfocitos T/efectos de los fármacos , Catequina/farmacología , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , ADN/análisis , Suplementos Dietéticos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Ensayo de Inmunoadsorción Enzimática/métodos , Citometría de Flujo , Perfilación de la Expresión Génica , Virus Linfotrópico T Tipo 1 Humano/aislamiento & purificación , Humanos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Sensibilidad y Especificidad , Relación Estructura-Actividad , Linfocitos T/virología , Factores de Crecimiento Transformadores/genéticaRESUMEN
Although rats are widely used for the analysis of allergic reactions and parasitic infections where IL-5 is involved, nothing is currently known of the expression of IL-5 receptor in this species. In this study, the cDNA sequence, genomic structure and the transcriptional regulation of the rat IL-5Ralpha were analyzed. The rat IL-5Ralpha gene, which we localized to chromosome 4q34-q41, spans more than 25 kb and consists of 12 exons. Promoter activity was seen in different cell lines and analysis by deletion experiments allowed to identify two negative regulatory regions which did not differ when tested either with IL-5Ralpha-negative or positive cells. Finally, the investigation of the expression of IL-5Ralpha showed that it is expressed in lung, spleen, liver, and purified rat B cells from normal rat. This can provide an explanation for the role of rat IL-5 as B-cell growth factor and a relevant model in order to better understand the activity of IL-5 on human B cells.
Asunto(s)
Regiones no Traducidas 5'/análisis , Linfocitos B/fisiología , Expresión Génica , Regiones Promotoras Genéticas/genética , Receptores de Interleucina/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Mapeo Cromosómico , Cromosomas , Clonación Molecular , ADN/análisis , Dosificación de Gen , Genoma , Técnicas In Vitro , Datos de Secuencia Molecular , Ratas , Ratas Endogámicas F344 , Receptores de Interleucina/biosíntesis , Receptores de Interleucina-5 , Homología de Secuencia de Aminoácido , Transcripción GenéticaRESUMEN
IL-13 mediates its effects through a complex receptor system including IL-4Ralpha and a functional IL-13Ralpha1. IL-13 has been reported to have no effects on mouse B cells due to a lack of receptor expression. However, on human B cells a functional IL-13Ralpha1 has been described. Here, we identified the rat IL-13Ralpha1 in order to analyze its expression and function in rat B cells. The expression of IL-13Ralpha1 has been shown by the presence of mRNA and the corresponding protein in purified rat B cells and in rat hybridoma B cell line. Rat B cells are able to bind IL-13 and to proliferate when cultured with CD40 ligand and IL-13. In vivo experiments showed that administration of IL-13 did enhance IgE production. These results suggest a direct interaction of rat B cells with IL-13 through a functional receptor with an increase of IgE production and provide a relevant model to further study the activity of IL-13 and to better understand its role in human diseases.
Asunto(s)
Linfocitos B/metabolismo , Receptores de Interleucina/metabolismo , Secuencia de Aminoácidos , Animales , Clonación Molecular , Humanos , Inmunoglobulina E/metabolismo , Interleucina-13/metabolismo , Subunidad alfa1 del Receptor de Interleucina-13 , Interleucina-4/metabolismo , Datos de Secuencia Molecular , Unión Proteica , Ratas , Receptores de Interleucina/química , Receptores de Interleucina/genética , Receptores de Interleucina-13 , Proteínas Recombinantes/metabolismo , Alineación de SecuenciaRESUMEN
The discovery of a functional division in T helper cells on the basis of their cytokine secretion patterns has changed our vision of immunological responses. This dichotomy has equally shown the complexity of immune responses since there is a well orchestrated cross-regulation of cytokine production induced by viral, bacterial or parasitic pathogens. In the context of type 1-type 2 cytokine pattern, mice has been universally and extensively used to associate an infectious disease according to each category in order to better understand human infections. However, with respect to schistosomiasis, immunological observations in mice have not been confirmed in humans and particularly the nature of the protective immune response. This report will consider the relevance of extrapolating from immunological studies on schistosome in experimentally infected rats to studies on naturally infected humans.
Asunto(s)
Esquistosomiasis/inmunología , Animales , Citocinas/biosíntesis , Citocinas/metabolismo , Modelos Animales de Enfermedad , Infecciones/inmunología , Ratones , Ratas , Schistosoma/inmunología , Linfocitos T Colaboradores-Inductores/inmunologíaRESUMEN
Human schistosomiasis is associated with IgE and eosinophilia, feature of a type 2 response. In experimental investigations, murine model has been widely used in order to dissect the immune responses involved in the expression of protective immunity or disease in Schistosoma mansoni infection. Collectively, observations made in this model and in humans demonstrated a strong contrast since a Th2 response in infected mice is involved in the expression of pathology, however, in infected humans the same type of response is rather beneficial for the host. This review will consider the relevance of extrapolating studies of immune responses from experimentally infected rats a semi-permissive host, to studies on S. mansoni infected humans.
Asunto(s)
Modelos Animales de Enfermedad , Schistosoma mansoni/inmunología , Esquistosomiasis mansoni/inmunología , Animales , Humanos , Ratas , Esquistosomiasis mansoni/parasitologíaRESUMEN
We have previously demonstrated in rat experimental schistosomiasis an upregulation of IL-4 expression at the mRNA and protein levels which could explain, at least in part, the increased IgE production observed during infection. Using this model, we have investigated the expression of IL-13 which is also involved in the induction of the IgE response. In the present study, we have shown a significant increase in IL-13 mRNA expression in spleen, liver and lungs following primary and secondary infection. IL-13 protein was detected by intracellular staining in spleen cells from infected rats, and in the supernatants of antigen-stimulated spleen cells. Furthermore, circulating levels of IL-13 were increased in sera from infected rats as compared to those from non-infected control animals. These findings show that, similarly to IL-4, IL-13 is upregulated and secreted during rat schistosomiasis, suggesting an involvement of both cytokines in IgE induction. In the in vivo experiments, only rats cotreated with neutralizing anti-IL-4 and anti-IL-13 antibodies showed significant decrease in the IgE levels. Moreover, administration of IL-13 enhanced total IgE levels. These results demonstrate the implication of IL-4 and IL-13 in vivo in IgE production, and provide a relevant animal model for a better understanding of the role of IL-4 and IL-13 in humans.
Asunto(s)
Inmunoglobulina E/biosíntesis , Interleucina-13/biosíntesis , Esquistosomiasis mansoni/inmunología , Animales , Anticuerpos/farmacología , Secuencia de Bases , Cartilla de ADN/genética , Interleucina-13/antagonistas & inhibidores , Interleucina-13/genética , Interleucina-4/antagonistas & inhibidores , Masculino , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas Endogámicas F344 , Esquistosomiasis mansoni/genética , Esquistosomiasis mansoni/metabolismo , Regulación hacia ArribaRESUMEN
In contrast to most mouse strains, rats eliminate the primary schistosome burden around 4 weeks postinfection and subsequently develop protective immunity to reinfection. In rat schistosomiasis, we have shown predominant expression of a Th2-type cytokine response at the mRNA level after primary infection. In the present study, we showed a significant increase in interleukin-4 (IL-4) mRNA expression in inguinal lymph nodes early after a secondary infection. IL-5 mRNA expression showed a significant increase at days 2 and 4 postreinfection in the spleen and lymph nodes, respectively. We did not detect any gamma interferon (IFN-gamma) mRNA after a challenge infection. Analysis of cytokine secretion by stimulated spleen cells after a primary infection showed predominant expression of IL-4 with maximum production on day 21, accompanied by production of IL-5 from day 11 to day 67. A significant increase in IFN-gamma secretion was detected at day 21. Analysis of immunoglobulin G2b (IgG2b) and IgG2c (Th1-related isotypes) showed undetectable levels of IgG2b, but detectable levels of specific IgG2c antibodies were observed from day 42. The analysis of Th2-related isotypes showed high specific IgG1 and IgG2a antibody titers from day 29. After a secondary infection, only IL-4 and IL-5 secretion was sustained. This is supported by the increased production of Th2-related isotypes. These findings showed that S. mansoni infection can drive Th2 responses in rats in the absence of egg production which is required to induce a Th2 response in mice and are in favor of the role of Th2-type cytokines in protective immunity against reinfection.
Asunto(s)
Citocinas/metabolismo , Esquistosomiasis mansoni/inmunología , Células TH1/inmunología , Células Th2/inmunología , Animales , Citocinas/genética , Citocinas/inmunología , Isotipos de Inmunoglobulinas/clasificación , Interferón gamma/genética , Interferón gamma/inmunología , Interferón gamma/metabolismo , Interleucina-4/genética , Interleucina-4/inmunología , Interleucina-4/metabolismo , Interleucina-5/genética , Interleucina-5/inmunología , Interleucina-5/metabolismo , Cinética , Masculino , Ratas , Ratas Endogámicas F344 , Schistosoma mansoni/inmunología , Células TH1/metabolismo , Células Th2/metabolismoRESUMEN
The isolation of 2 genomic clones has allowed us to further characterize a Schistosoma mansoni serine protease designated SmSP1. The deduced amino acid sequence (248aa) considered as a 'light chain' encoding the active site, presents significant homologies with mouse plasma kallikrein and human factor I light chain. The secondary structure of SmSP1 'light chain' is correctly predicted and may be sufficient by itself to constitute an active enzyme. The biological function of SmSP1 is unknown, however, the homology with 2 serine proteases suggests that SmSP1 may play a role in the evasion of the host immune response. This is supported by the presence of the native protein corresponding to SmSP1 particularly in schistosomula released products (SRP) and in male dorsal spines. The expression of this enzyme is differentially regulated throughout the parasite life-cycle. However, infected animals with S. mansoni did not produce specific antibodies to recombinant SmSP1. The lack of such response could be advantageous to the parasite by protecting itself from host effector mechanisms.
Asunto(s)
Fibrinógeno/genética , Calicreínas/genética , Schistosoma mansoni/enzimología , Serina Endopeptidasas/genética , Serina Endopeptidasas/aislamiento & purificación , Secuencia de Aminoácidos , Animales , Anticuerpos Antihelmínticos/sangre , ADN de Helmintos/análisis , Ensayo de Inmunoadsorción Enzimática/métodos , Fibrinógeno/metabolismo , Regulación del Desarrollo de la Expresión Génica , Humanos , Calicreínas/metabolismo , Masculino , Ratones , Datos de Secuencia Molecular , Sistemas de Lectura Abierta/genética , Ratas , Ratas Endogámicas F344 , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Schistosoma mansoni/genética , Schistosoma mansoni/crecimiento & desarrollo , Schistosoma mansoni/inmunología , Esquistosomiasis mansoni/inmunología , Esquistosomiasis mansoni/parasitología , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Serina Endopeptidasas/química , Serina Endopeptidasas/metabolismoRESUMEN
The T-helper dependency of the IgA antibody response has been investigated in rats injected intravenously with Schistosoma mansoni eggs. This method, allowing the trapping of parasite eggs in the lung tissue, led to a strong anti-egg IgA antibody response in the bronchoalveolar lavage but not in the serum. To characterize the cytokine pattern associated with the IgA response, kinetic analysis of the cytokine mRNA expression in the lungs, periaortic nodes (PN) and spleen was undertaken. Under such conditions, significant levels of mRNA encoding IL-5 and IL-10 were recorded in spleen during the early period following egg injection, as well as a more prolonged expression of TGF-beta and IL-6 mRNAs. However, neither IFN-gamma nor IL-4 mRNA could be detected in these samples. Finally, in lungs and in PN, RT-PCR analysis revealed delayed production of cytokine mRNA. Taken together our data suggest that the rat mucosal IgA antibody response is predominantly linked to a Th2 response.
Asunto(s)
Citocinas/genética , Inmunoglobulina A/biosíntesis , Tejido Linfoide/metabolismo , ARN Mensajero/biosíntesis , Animales , Antígenos Helmínticos/administración & dosificación , Antígenos Helmínticos/inmunología , Citocinas/biosíntesis , Inmunidad Celular , Inyecciones Intravenosas , Pulmón/inmunología , Pulmón/metabolismo , Ganglios Linfáticos/inmunología , Ganglios Linfáticos/metabolismo , Tejido Linfoide/inmunología , Masculino , Ratones , Óvulo/inmunología , Ratas , Ratas Endogámicas F344 , Schistosoma mansoni/inmunología , Bazo/inmunología , Bazo/metabolismoRESUMEN
The nucleotide sequences containing the rat interleukin-12 p40 gene was determined. Sequencing revealed the presence of six exons and five introns. Analysis of the 5' non-coding region showed the presence of several possible sites involved in cytokine gene regulation at the transcriptional level. Comparison of the deduced amino acid sequence of rat IL-12 p40 with that of the mouse and of human p40, showed 92% and 65% identity respectively.
Asunto(s)
Genes/genética , Interleucina-12/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Sitios de Unión/genética , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , Exones/genética , Regulación de la Expresión Génica , Interleucina-12/química , Intrones/genética , Datos de Secuencia Molecular , Ratas , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Transcripción Genética/genéticaRESUMEN
The chemoattractant cytokine IL- 16 has been reported to suppress lymphocyte activation and to inhibit HIV-1 replication in acutely infected T cells. We have cloned and expressed human IL-16 in Escherichia coli and investigated whether the recombinant protein could regulate the level of lymphocyte apoptosis from HIV-1-infected subjects. After purification and refolding, only 2-10% of the recombinant cytokine was present in a biologically active homotetrameric form. This could explain the need for high concentrations of the bacterially derived IL- 16 to induce significant inhibition of HIV-1 replication. Addition of IL-16 to unstimulated peripheral blood mononuclear cell (PBMC) cultures from HIV-1-infected subjects did not modify the observed level of spontaneous lymphocyte apoptosis. In contrast, IL-16 added to PBMC cultures stimulated with anti-CD3, anti-CD95 or dexamethasone reduced significantly the percentage of lymphocytes undergoing AICD. This effect was found to correlate with the ability of the cytokine to decrease CD95 expression on activated CD4+ T cells. Comparative studies on PBMC from healthy individuals indicated that the regulation of apoptosis levels by IL-16 is a complex phenomenon and could depend on the nature of the activator used and/or the immune status of lymphocytes tested. The outcome of CD4 cross-linking on T cells by various ligands is discussed in the context of the observed beneficial activities of IL- 16 and its potential role in the treatment of HIV disease.
Asunto(s)
Apoptosis/efectos de los fármacos , Infecciones por VIH/sangre , VIH-1/efectos de los fármacos , VIH-1/fisiología , Interleucina-16/farmacología , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/virología , Replicación Viral/inmunología , Células Cultivadas , Escherichia coli , Infecciones por VIH/virología , Humanos , Proteínas Recombinantes/farmacologíaRESUMEN
As an animal model, rat schistosomiasis mansoni has provided considerable knowledge of immune mechanisms involved in the expulsion of worms and in a subsequent development of immunity to reinfection. Although it is clear that ADCC mechanisms participate in immunity to reinfection; the nature of the cytokines involved in immunity is unknown. To analyse the pattern of cytokines involved, the mRNA levels of different cytokines were assessed by RT-PCR as they occur within tissues during the course of infection. In spleens from infected rats, a significant elevation in IL-2 and IL-5 mRNA was observed during the early phase of infection (day 7). Analysis of pulmonary cytokine responses showed a dramatic increase in IL-4 and IL-5 on day 7. This was accompanied with a low but significant increase in IL-2 (day 11) and IL-12 (day 7) in the absence of augmented IFN-gamma expression. The cytokine expression patterns of draining lymph nodes (LN) from infected rats showed a significant increase of IL-2, IL-4 and IL-5 on day 21. Analysis of IL-10 expression showed exclusively a significant increase in LN on day 11, IFN-gamma mRNA was not detected in any tissue sample. Thus, rats develop a predominately Th2-type cytokine response during a primary infection which may be involved at least in part, in the expression of immunity against Schistosoma mansoni infection.
Asunto(s)
Interferón gamma/genética , Interleucinas/genética , ARN Mensajero/metabolismo , Esquistosomiasis mansoni/inmunología , Animales , Expresión Génica , Interferón gamma/metabolismo , Interleucinas/metabolismo , Pulmón/inmunología , Ganglios Linfáticos/inmunología , Reacción en Cadena de la Polimerasa , ARN Mensajero/genética , Ratas , Esquistosomiasis mansoni/parasitología , Bazo/inmunología , Células TH1/inmunología , Células Th2/inmunologíaRESUMEN
Genes encoding the heavy and light chains of an anti-idiotype antibody (AB2) mimicking a protective oligosaccharide of Schistosoma mansoni were cloned and expressed as a single-chain Fv fragment. The expression in a functional state was tested using the AB1. A specific binding between sFv and AB1 was observed. Immunization with the recombinant AB2 indicates its capacity to elicit anti-S. mansoni antibodies.