RESUMEN
BACKGROUND: Several Rho GTPase-activating proteins (RhoGAPs) are implicated in tumor progression through their effects on Rho GTPase activity. ARHGAP21 is a RhoGAP with increased expression in head and neck squamous cell carcinoma and with a possible role in glioblastoma tumor progression, yet little is known about the function of ARHGAP21 in cancer cells. Here we studied the role of ARHGAP21 in two prostate adenocarcinoma cell lines, LNCaP and PC3, which respectively represent initial and advanced stages of prostate carcinogenesis. RESULTS: ARHGAP21 is located in the nucleus and cytoplasm of both cell lines and its depletion resulted in decreased proliferation and increased migration of PC3 cells but not LNCaP cells. In PC3 cells, ARHGAP21 presented GAP activity for RhoA and RhoC and induced changes in cell morphology. Moreover, its silencing altered the expression of genes involved in cell proliferation and cytoskeleton organization, as well as the endothelin-1 canonical pathway. CONCLUSIONS: Our results reveal new functions and signaling pathways regulated by ARHGAP21, and indicate that it could contribute to prostate cancer progression.
Asunto(s)
Adenocarcinoma/patología , Movimiento Celular , Proliferación Celular , Proteínas Activadoras de GTPasa/fisiología , Neoplasias de la Próstata/patología , Adenocarcinoma/metabolismo , Secuencia de Bases , Línea Celular Tumoral , Cartilla de ADN , Proteínas Activadoras de GTPasa/genética , Silenciador del Gen , Humanos , Etiquetado Corte-Fin in Situ , Masculino , Neoplasias de la Próstata/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Pyrimidine-5'-nucleotidase type I (P5'NI) deficiency is an autosomal recessive condition that causes nonspherocytic hemolytic anemia, characterized by marked basophilic stippling and pyrimidine nucleotide accumulation in erythrocytes. We herein present two African descendant patients, father and daughter, with P5'N deficiency, both born from first cousins. Investigation of the promoter polymorphism of the uridine diphospho glucuronosyl transferase 1A (UGT1A) gene revealed that the father was homozygous for the allele (TA7) and the daughter heterozygous (TA6/TA7). P5'NI gene (NT5C3) gene sequencing revealed a further change in homozygosity at amino acid position 56 (p.R56G), located in a highly conserved region. Both patients developed gallstones; however the father, who had undergone surgery for the removal of stones, had extremely severe intrahepatic cholestasis and, liver biopsy revealed fibrosis and siderosis grade III, leading us to believe that the homozygosity of the UGT1A polymorphism was responsible for the more severe clinical features in the father. Moreover, our results show how the clinical expression of hemolytic anemia is influenced by epistatic factors and we describe a new mutation in the P5'N gene associated with enzyme deficiency, iron overload, and severe gallstone formation. To our knowledge, this is the first description of P5'N deficiency in South Americans.
Asunto(s)
5'-Nucleotidasa/deficiencia , Anemia Hemolítica Congénita/genética , Colestasis/genética , Enfermedad de Gilbert/genética , Glicoproteínas/genética , Sobrecarga de Hierro/genética , Cirrosis Hepática/genética , 5'-Nucleotidasa/genética , Adulto , Alelos , Anemia Hemolítica Congénita/complicaciones , Anemia Hemolítica Congénita/enzimología , Anemia Hemolítica Congénita/patología , Niño , Colestasis/complicaciones , Colestasis/enzimología , Colestasis/patología , Consanguinidad , Epistasis Genética , Femenino , Enfermedad de Gilbert/complicaciones , Enfermedad de Gilbert/enzimología , Enfermedad de Gilbert/patología , Heterocigoto , Homocigoto , Humanos , Sobrecarga de Hierro/complicaciones , Sobrecarga de Hierro/enzimología , Sobrecarga de Hierro/patología , Hígado/enzimología , Hígado/patología , Cirrosis Hepática/complicaciones , Cirrosis Hepática/enzimología , Cirrosis Hepática/patología , Masculino , Regiones Promotoras Genéticas , Análisis de Secuencia de ADNRESUMEN
BACKGROUND: ß-carboline alkaloids exert a distinguished ability to impair cell growth and induce cell death in a variety of cancers and the evaluation of such new therapeutic candidates may denote new possibilities for leukemia treatment. In this present study, we screened 12 ß-carboline derivatives containing different substituents at 1- and 3-positions of ß-carboline nucleus for their antineoplastic activities in a panel of leukemia cell lines. METHODS: The cytotoxic effects of the ß-carboline derivatives were evaluated in different leukemia cell lines as well as reactive oxygen species (ROS) generation, autophagy, and important signaling pathways. RESULTS: Treatment with the ß-carboline derivatives resulted in a potent antineoplastic activity leading to a reduced cell viability that was associated with increased cell death in a concentration-dependent manner. Interestingly, the treatment of primary mononuclear cells isolated from the peripheral blood of healthy donors with the ß-carboline derivatives showed a minor change in cell survival. The antineoplastic activity occurs by blocking ROS production causing consequent interruption of the PI3K/AKT and MAPK/ERK signaling and modulating autophagy processes. Notably, in vivo, AML burden was diminished in peripheral blood and bone marrow of a xenograft mouse model. CONCLUSIONS: Our results indicated that ß-carboline derivatives have an on-target malignant cell-killing activity and may be promising candidates for treating leukemia cells by disrupting crucial events that promote leukemia expansion and chemotherapy resistance.
Asunto(s)
Antineoplásicos , Carbolinas , Supervivencia Celular , Leucemia Mieloide Aguda , Especies Reactivas de Oxígeno , Ensayos Antitumor por Modelo de Xenoinjerto , Carbolinas/farmacología , Humanos , Animales , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/patología , Ratones , Especies Reactivas de Oxígeno/metabolismo , Línea Celular Tumoral , Antineoplásicos/farmacología , Supervivencia Celular/efectos de los fármacos , Autofagia/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Relación Dosis-Respuesta a DrogaRESUMEN
Sickle cell disease (SCD) patients experience chronic inflammation and recurrent vaso-occlusive episodes during their entire lifetime. Inflammation in SCD occurs with the overexpression of several inflammatory mediators, including transforming growth factor beta-1 (TGF-ß1), a major immune regulator. In this study, we aimed to investigate the role played by TGF-ß1 in vascular inflammation and vaso-occlusion in an animal model of SCD. Using intravital microscopy, we found that a daily dose of recombinant TGF-ß1 administration for three consecutive days significantly reduced TNFα-induced leukocyte rolling, adhesion, and extravasation in the microcirculation of SCD mice. In contrast, immunological neutralization of TGF-ß, in the absence of inflammatory stimulus, considerably increased these parameters. Our results indicate, for the first time, that TGF-ß1 may play a significant ameliorative role in vascular SCD pathophysiology, modulating inflammation and vaso-occlusion. The mechanisms by which TGF-ß1 exerts its anti-inflammatory effects in SCD, however, remains unclear. Our in vitro adhesion assays with TNFα-stimulated human neutrophils suggest that TGF-ß1 can reduce the adhesive properties of these cells; however, direct effects of TGF-ß1 on the endothelium cannot be ruled out. Further investigation of the wide range of the complex biology of this cytokine in SCD pathophysiology and its potential therapeutical use is needed.
Asunto(s)
Anemia de Células Falciformes , Neutrófilos , Factor de Crecimiento Transformador beta1 , Enfermedades Vasculares , Anemia de Células Falciformes/complicaciones , Anemia de Células Falciformes/metabolismo , Animales , Humanos , Inflamación/metabolismo , Ratones , Neutrófilos/citología , Neutrófilos/efectos de los fármacos , Factor de Crecimiento Transformador beta1/farmacología , Factor de Necrosis Tumoral alfa/metabolismo , Enfermedades Vasculares/metabolismoRESUMEN
BACKGROUND: Sickle cell disease (SCD) incurs vaso-occlusive episodes and organ damage, including nephropathy. Despite displaying characteristics of vascular dysfunction, SCD patients tend to present relatively lower systemic blood pressure (BP), via an unknown mechanism. We investigated associations between BP and renin-angiotensin-system (RAS) components in SCD and determined whether an inhibitor of angiotensin converting enzyme (ACE; often used to slow SCD glomerulopathy) further modulates BP and RAS components in a murine model of SCD. METHODS: BP was compared in human subjects and mice with/without SCD. Plasma angiotensin II, ACE and renin were measured by immunoassay. BP was reevaluated after treating mice with enalapril (25 mg/kg, 5x/week) for 5 weeks; plasma and organs were stored for angiotensin II and ACE activity measurement, and quantitative real-time PCR. RESULTS: Diastolic BP and systolic BP were significantly lower in patients and mice with SCD, respectively, compared to controls. Reduced BP was associated with increased plasma renin and markers of kidney damage (mice) in SCD, as well as significantly decreased plasma ACE concentrations and ACE enzyme activity. As expected, enalapril administration lowered BP, plasma angiotensin II and organ ACE activity in control mice. In contrast, enalapril did not further reduce BP or organ ACE activity in SCD mice; however, plasma angiotensin II and renin levels were found to be significantly higher in enalapril-treated SCD mice than those of treated control mice. CONCLUSION: Relative hypotension was confirmed in a murine model of SCD, in association with decreased ACE concentrations in both human and murine disease. Given that ACE inhibition has an accepted role in decreasing BP, further studies should investigate mechanisms by which ACE depletion, via both Ang II-dependent and alternative pathways, could contribute to reduce BP in SCD and understand how ACE inhibition confers Ang II-independent benefits on kidney function in SCD.
Asunto(s)
Anemia de Células Falciformes/tratamiento farmacológico , Anemia de Células Falciformes/fisiopatología , Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Presión Sanguínea/efectos de los fármacos , Peptidil-Dipeptidasa A/biosíntesis , Adolescente , Adulto , Angiotensina II/metabolismo , Animales , Diástole , Modelos Animales de Enfermedad , Femenino , Humanos , Hidroxiurea/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Reacción en Cadena en Tiempo Real de la Polimerasa , Renina/sangre , Sistema Renina-Angiotensina , Sístole , Adulto JovenRESUMEN
The interaction between the bone marrow microenvironment and malignant hematopoietic cells can result in the protection of leukemia cells from chemotherapy in both myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). We, herein, characterized the changes in cytokine expression and the function of mesenchymal stromal cells (MSC) in patients with MDS, AML with myelodysplasia-related changes (MRC), a well-recognized clinical subtype of secondary AML, and de novo AML. We observed a significant inhibitory effect of MDS-MSC on T lymphocyte proliferation and no significant differences in any of the cytokines tested. AML-MSC inhibited T-cell proliferation only at a very low MSC/T cell ratio. When compared to the control, AML-MRCderived MSC presented a significant increase in IL6 expression, whereas de novo AML MSC presented a significant increase in the expression levels of VEGFA, CXCL12, RPGE2, IDO, IL1ß, IL6 and IL32, followed by a decrease in IL10 expression. Furthermore, data indicate that IL-32 regulates stromal cell proliferation, has a chemotactic potential and participates in stromal cell crosstalk with leukemia cells, which could result in chemoresistance. Our results suggest that the differences between AML-MRC and de novo AML also extend into the leukemic stem cell niche and that IL-32 can participate in the regulation of the bone marrow cytokine milieu.
Asunto(s)
Microambiente Celular , Leucemia Mieloide Aguda/etiología , Leucemia Mieloide Aguda/metabolismo , Síndromes Mielodisplásicos/etiología , Síndromes Mielodisplásicos/metabolismo , Microambiente Tumoral , Antimetabolitos Antineoplásicos/farmacología , Médula Ósea/patología , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/patología , Estudios de Casos y Controles , Línea Celular , Proliferación Celular , Microambiente Celular/efectos de los fármacos , Microambiente Celular/genética , Microambiente Celular/inmunología , Quimiotaxis/genética , Quimiotaxis/inmunología , Citarabina/farmacología , Citocinas/metabolismo , Femenino , Regulación de la Expresión Génica , Silenciador del Gen , Humanos , Inmunomodulación , Leucemia Mieloide Aguda/patología , Masculino , MicroARNs/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Síndromes Mielodisplásicos/patología , FN-kappa B/metabolismo , Interferencia de ARN , Transducción de Señal , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/genética , Microambiente Tumoral/inmunologíaRESUMEN
BACKGROUND: Telomeres, the ends of linear chromosomes, shorten during mitotic cell division and erosion may be aggravated by inflammation or proliferative and oxidative stress. As the bone marrow is under hyperproliferative pressure in sickle cell disease and several tissues are submitted to chronic inflammation, this study sought to determine the telomere length of patients with sickle cell disease. METHODS: The mean telomere length was measured in peripheral blood leukocytes by quantitative polymerase chain reaction. The age-adjusted telomere to single copy gene ratio was compared between 91 adult sickle cell disease patients and 188 controls. RESULTS: Sickle cell disease patients had significantly shorter telomeres than the controls (p-value<0.0001). Moreover, among sickle cell disease genotypes, Hb SS patients had significantly shorter telomeres compared to Hb SC and Hb Sß patients (p-value<0.0001). Patients on hydroxyurea also had shorter telomeres in comparison to those off the drug (p-value=0.02). A positive correlation was observed between telomere length and hemoglobin level (r=0.3; p-value=0.004), whereas negative correlations were detected between telomere length and lymphocyte count (r=-0.3; p-value=0.005) and interleukin-8 serum levels (r=-0.4; p-value=0.02). CONCLUSIONS: The findings of this study indicate that telomeres are short in sickle cell disease patients and that telomere erosion directly correlates with disease genotype, inflammation markers, and the use of hydroxyurea.
RESUMEN
Our hypothesis was to investigate the fatty acid potential as a bone induction factor. In vitro and in vivo studies were performed to evaluate this approach. Oleic acid was used in a 0.5 wt.% concentration. Polycaprolactone was used as the polymeric matrix by combining solvent-casting and particulate-leaching techniques, with a final porosity of 70 wt.%, investigated by SEM images. Contact angle measurements were produced to investigate the influence of oleic acid on polycaprolactone chains. Cell culture was performed using adipocyte-derived stem cells to evaluate biocompatibility and bioactivity properties. In addition, in vivo studies were performed to evaluate the induction potential of oleic acid addition. Adipocyte-derived stem cells were used to provide differentiation after 21 days of culture. Likewise, information were obtained with in vivo data and cellular invagination was observed on both scaffolds (polycaprolactone and polycaprolactone /oleic acid); interestingly, the scaffold with oleic acid addition demonstrated that cellular migrations are not related to the surrounding tissue, indicating bioactive potential. Our hypothesis is that fatty acid may be used as a potential induction factor for bone tissue engineering. The study's findings indicate oleic acid as a possible agent for bone induction, according to data on cell differentiation, proliferation, and migration. Impact statement The biomaterial combined in this study on bone regeneration is innovative and shows promising results in the treatment of bone lesions. Polycaprolactone (PCL) and oleic acid have been studied separately. In this research, we combined biomaterials to assess the stimulus and the speed of bone healing.
Asunto(s)
Huesos , Ácidos Grasos/metabolismo , Osteoblastos/efectos de los fármacos , Células Madre/efectos de los fármacos , Animales , Regeneración Ósea/efectos de los fármacos , Regeneración Ósea/fisiología , Huesos/efectos de los fármacos , Huesos/metabolismo , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Humanos , Masculino , Osteoblastos/citología , Poliésteres/farmacología , Ratas Wistar , Células Madre/citología , Ingeniería de Tejidos/métodosRESUMEN
ANKHD1 (Ankyrin repeat and KH domain-containing protein 1) is highly expressed and plays an important role in the proliferation and cell cycle progression of multiple myeloma (MM) cells. ANKHD1 downregulation modulates cell cycle gene expression and upregulates p21 irrespective of the TP53 mutational status of MM cell lines. The present study was aimed to investigate the role of ANKHD1 in MM in vitro clonogenicity and in vivo tumourigenicity, as well as the role of ANKHD1 in p21 transcriptional regulation. ANKHD1 silencing in MM cells resulted in significantly low no. of colonies formed and in slow migration as compared to control cells (p < 0.05). Furthermore, in xenograft MM mice models, tumour growth was visibly suppressed in mice injected with ANKHD1 silenced cells compared to the control group. There was a significant decrease in tumour volume (p = 0.006) as well as in weight (p = 0.02) in the group injected with silenced cells compared to those of the control group. Co-immunoprecipitation and chromatin immunoprecipitation (ChIP) assays confirmed the interaction between p21 and ANKHD1. Moreover, overexpression of ANKHD1 downregulated the activity of a p21 promoter in luciferase assays. Decrease in luciferase activity suggests a direct role of ANKHD1 in p21 transcriptional regulation. In addition confocal analysis after U266 cells were treated with Leptomycin B (LMB) for 24 h showed accumulation of ANKHD1 inside the nucleus as compared to untreated cells where ANKHD1 was found to be predominantly in cytoplasm. This suggests ANKHD1 might be shuttling between cytoplasm and nucleus. In conclusion, ANKHD1 promotes MM growth by repressing p21 a potent cell cycle regulator.
Asunto(s)
Proliferación Celular , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Mieloma Múltiple/genética , Regiones Promotoras Genéticas/genética , Proteínas de Unión al ARN/genética , Transporte Activo de Núcleo Celular/efectos de los fármacos , Animales , Western Blotting , Línea Celular Tumoral , Movimiento Celular/genética , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Ácidos Grasos Insaturados/farmacología , Femenino , Regulación Neoplásica de la Expresión Génica , Células HEK293 , Humanos , Masculino , Ratones Endogámicos NOD , Ratones SCID , Microscopía Confocal , Mieloma Múltiple/metabolismo , Mieloma Múltiple/patología , Unión Proteica , Interferencia de ARN , Proteínas de Unión al ARN/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Carga Tumoral/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Despite the detailed characterization of the inflammatory and endothelial changes observed in Sickle Cell Disease (SCD), the hierarchical relationship between elements involved in the pathogenesis of this complex disease is yet to be described. Meta-analyses of gene expression studies from public repositories represent a novel strategy, capable to identify key mediators in complex diseases. We performed several meta-analyses of gene expression studies involving SCD, including studies with patient samples, as well as in-vitro models of the disease. Meta-analyses were performed with the Inmex bioinformatics tool, based on the RankProd package, using raw gene expression data. Functional gene set analysis was performed using more than 60 gene-set libraries. Our results demonstrate that the well-characterized association between innate immunity, hemostasis, angiogenesis and heme metabolism with SCD is also consistently observed at the transcriptomic level, across independent studies. The enrichment of genes and pathways associated with innate immunity and damage repair-associated pathways supports the model of erythroid danger-associated molecular patterns (DAMPs) as key mediators of the pathogenesis of SCD. Our study also generated a novel database of candidate genes, pathways and transcription factors not previously associated with the pathogenesis of SCD that warrant further investigation in models and patients of SCD.
Asunto(s)
Anemia de Células Falciformes/etiología , Anemia de Células Falciformes/metabolismo , Regulación de la Expresión Génica , Inmunidad Innata , Transducción de Señal , Análisis por Conglomerados , Biología Computacional/métodos , Células Endoteliales/metabolismo , Perfilación de la Expresión Génica , Redes Reguladoras de Genes , Humanos , TranscriptomaRESUMEN
Leg ulcers represent a particularly disabling complication in patients with sickle cell disease (SCD). Platelet gel (PG) is a novel therapeutic strategy used for accelerating wound healing of a wide range of tissues through the continuous release of platelet growth factors. Here, we describe the use of PG preparation according to Anitua's PRGF (preparations rich in growth factors) protocol for treating chronic nonhealing ulcers in patients with SCD. A positive response occurred in 3 patients with an area reduction of 85.7% to 100%, which occurred within 7 to 10 weeks, and a 35.2% and 20.5% of area reduction in 2 other patients, who however, had large ulcers. After calcium chloride addition, the platelet-rich plasmas demonstrated enhanced platelet-derived growth factors-BB (P < .001), transforming growth factor-ß1 (P = .015), vascular endothelial growth factors (P = .03), and hepatocyte growth factors (nonsignificant) secretion. Furthermore, calcium chloride addition induced a significant decrease in platelet number (P = .0134) and there was no leukocyte detection in the PG product. These results demonstrate that PG treatment might impact the healing of leg ulcers in sickle cell disease, especially in patients with small ulcers.
RESUMEN
ABSTRACT Background: Telomeres, the ends of linear chromosomes, shorten during mitotic cell division and erosion may be aggravated by inflammation or proliferative and oxidative stress. As the bone marrow is under hyperproliferative pressure in sickle cell disease and several tissues are submitted to chronic inflammation, this study sought to determine the telomere length of patients with sickle cell disease. Methods: The mean telomere length was measured in peripheral blood leukocytes by quantitative polymerase chain reaction. The age-adjusted telomere to single copy gene ratio was compared between 91 adult sickle cell disease patients and 188 controls. Results: Sickle cell disease patients had significantly shorter telomeres than the controls (p-value < 0.0001). Moreover, among sickle cell disease genotypes, Hb SS patients had significantly shorter telomeres compared to Hb SC and Hb Sβ patients (p-value < 0.0001). Patients on hydroxyurea also had shorter telomeres in comparison to those off the drug (p-value = 0.02). A positive correlation was observed between telomere length and hemoglobin level (r = 0.3; p-value = 0.004), whereas negative correlations were detected between telomere length and lymphocyte count (r = -0.3; p-value = 0.005) and interleukin-8 serum levels (r = -0.4; p-value = 0.02). Conclusions: The findings of this study indicate that telomeres are short in sickle cell disease patients and that telomere erosion directly correlates with disease genotype, inflammation markers, and the use of hydroxyurea.
Asunto(s)
Humanos , Telómero , Homeostasis del Telómero , Inflamación , Anemia de Células FalciformesRESUMEN
ARHGAP21 is a 217 kDa RhoGAP protein shown to modulate cell migration through the control of Cdc42 and FAK activities. In the present work a 250 kDa-ARHGAP21 was identified by mass spectrometry. This modified form is differentially expressed among cell lines and human primary cells. Co-immunoprecipitations and in vitro SUMOylation confirmed ARHGAP21 specific modification by SUMO2/3 and mapped the SUMOylation site to ARHGAP21 lysine K1443. Immunofluorescence staining revealed that ARHGAP21 co-localizes with SUMO2/3 in the cytoplasm and membrane compartments. Interestingly, our results suggest that ARHGAP21 SUMOylation may be related to cell proliferation. Therefore, SUMOylation of ARHGAP21 may represent a way of guiding its function.
Asunto(s)
Proteínas Activadoras de GTPasa/metabolismo , Proteínas Modificadoras Pequeñas Relacionadas con Ubiquitina/metabolismo , Ubiquitinas/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Línea Celular Tumoral , Proliferación Celular , Proteínas Activadoras de GTPasa/química , Proteínas Activadoras de GTPasa/genética , Células HEK293 , Humanos , Lisina/química , Datos de Secuencia Molecular , Procesamiento Proteico-Postraduccional , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , SumoilaciónRESUMEN
The purpose of this study was to investigate the in vitro effects of thalidomide on long-term bone marrow cultures from patients with myelodysplastic syndrome. We demonstrated that thalidomide induced an increase in granulocyte-macrophage colony forming unit numbers and in IL-10 expression. Thalidomide also promoted a slight increase in IL-6, IL-1ß and TNF-α expression in the stromal layers. The numbers of erythroid burst forming units, the apoptosis rate of hematopoietic cells, and VEGF and TNF-α expression levels in culture supernatants were not modulated. Our results indicate a participation of thalidomide upon the hematopoietic microenvironment of patients with myelodysplastic syndromes, especially in the up regulation of IL-10.
Asunto(s)
Inhibidores de la Angiogénesis/uso terapéutico , Médula Ósea/efectos de los fármacos , Interleucina-10/metabolismo , Síndromes Mielodisplásicos/tratamiento farmacológico , Síndromes Mielodisplásicos/metabolismo , Células del Estroma/efectos de los fármacos , Talidomida/uso terapéutico , Adulto , Anciano , Técnicas de Cocultivo , Ensayo de Unidades Formadoras de Colonias , Ensayo de Inmunoadsorción Enzimática , Femenino , Células Madre Hematopoyéticas , Humanos , Interleucina-10/genética , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/genética , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células del Estroma/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
OBJECTIVES: To determine whether kidney disease and hemolysis are associated with bone mass density in a population of adult Brazilian patients with sickle cell disease. INTRODUCTION: Bone involvement is a frequent clinical manifestation of sickle cell disease, and it has multiple causes; however, there are few consistent clinical associations between bone involvement and sickle cell disease. METHODS: Patients over 20 years of age with sickle cell disease who were regularly followed at the Hematology and Hemotherapy Center of Campinas, Brazil, were sorted into three groups, including those with normal bone mass density, those with osteopenia, and those with osteoporosis, according to the World Health Organization criteria. The clinical data of the patients were compared using statistical analyses. RESULTS: In total, 65 patients were included in this study: 12 (18.5%) with normal bone mass density, 37 (57%) with osteopenia and 16 (24.5%) with osteoporosis. Overall, 53 patients (81.5%) had bone mass densities below normal standards. Osteopenia and osteoporosis patients had increased lactate dehydrogenase levels and reticulocyte counts compared to patients with normal bone mass density (p<0.05). Osteoporosis patients also had decreased hemoglobin levels (p<0.05). Hemolysis was significantly increased in patients with osteoporosis compared with patients with osteopenia, as indicated by increased lactate dehydrogenase levels and reticulocyte counts as well as decreased hemoglobin levels. Osteoporosis patients were older, with lower glomerular filtration rates than patients with osteopenia. There was no significant difference between the groups with regard to gender, body mass index, serum creatinine levels, estimated creatinine clearance, or microalbuminuria. CONCLUSION: A high prevalence of reduced bone mass density that was associated with hemolysis was found in this population, as indicated by the high lactate dehydrogenase levels, increased reticulocyte counts and low hemoglobin levels.
Asunto(s)
Anemia de Células Falciformes/complicaciones , Densidad Ósea/fisiología , Enfermedades Óseas Metabólicas/etiología , Hemólisis/fisiología , Absorciometría de Fotón , Adulto , Anemia de Células Falciformes/fisiopatología , Enfermedades Óseas Metabólicas/fisiopatología , Femenino , Tasa de Filtración Glomerular/fisiología , Humanos , L-Lactato Deshidrogenasa/sangre , Masculino , Persona de Mediana Edad , Osteoporosis/etiología , Osteoporosis/fisiopatología , Recuento de Reticulocitos , Adulto JovenRESUMEN
INTRODUCTION: Myelodysplastic syndromes encompass a heterogeneous group of clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis, refractory cytopenia and a tendency to progress toward acute myeloid leukemia. The accumulation of genetic alterations is closely associated with the progression of myelodysplastic syndromes toward acute myeloid leukemia. OBJECTIVE: To investigate the presence of mutations in the points most frequent for mutations (hotspot mutations) in phosphatidylinositol-3-kinase (PI3K), Janus kinase 2 (JAK2), FMS-like tyrosine kinase 3 (FLT3) and nucleophosmin (NPM1), which are involved in leukemia and other cancers, in a population of Brazilian MDS patients. METHODS: Fifty-one myelodysplastic syndromes patients were included in the study. According to French-American-British classification, the patients were distributed as follows: 31 with refractory anemia, 8 with refractory anemia with ringed sideroblasts, 7 with refractory anemia with excess blasts, 3 with refractory anemia with excess blasts in transformation and 2 with chronic myelomonocytic leukemia. Bone marrow samples were obtained and screened for the presence of hotspot mutations using analysis based on amplification with the polymerase chain reaction, sequencing, fragment size polymorphisms or restriction enzyme digestion. All patients were screened for mutations at the time of diagnosis, and 5 patients were also screened at the time of disease progression. RESULTS: These results show that hotspot mutations in the PI3K, JAK2, FLT3 and NPM1 genes are not common in MDS patients; nevertheless, JAK2 mutations may be present in myelodysplasia during disease progression. CONCLUSIONS: These results show that hotspot mutations in the PI3K, JAK2, FLT3 and NPM1 genes are not common in MDS patients; nevertheless, JAK2 mutations may be present in myelodysplasia during disease progression.
Asunto(s)
Janus Quinasa 2/genética , Mutación/genética , Síndromes Mielodisplásicos/genética , Proteínas Nucleares/genética , Fosfatidilinositol 3-Quinasa/genética , Tirosina Quinasa 3 Similar a fms/genética , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Pruebas Genéticas , Humanos , Masculino , Persona de Mediana Edad , NucleofosminaRESUMEN
Leukocytes are known to exacerbate inflammatory and vaso-occlusive processes in sickle cell disease (SCD). The aim of this study was to determine whether alterations in neutrophil maturity and/or cell-death modulating factors in the circulation contribute to the increased leukocyte counts and leukocyte survival observed in SCD. The maturity of circulating neutrophils from healthy control individuals (CON), SCD and SCD patients on hydroxyurea therapy (SCDHU) was determined immunophenotypically. Serum factors affecting neutrophil apoptosis (determined by annexin V-binding) were analyzed by culturing control neutrophils (CON neutrophils) with pooled serum from CON, SCD and SCDHU individuals. Immunophenotypic characterization of neutrophils suggested a slight, but significant, increase in the circulation of immature neutrophils in SCD. While SCD neutrophils cultured in the presence of CON serum presented delayed apoptosis, unexpectedly, the culture of CON neutrophils with SCD serum significantly augmented apoptosis and caspase-9 activity. Inhibition of the activity of serum interleukin-8, a neutrophil-apoptosis-inhibiting cytokine, significantly increased SCD serum-induced CON neutrophil apoptosis, indicating that SCD serum may have both apoptotic and antiapoptotic properties. The decreased maturity of SCD neutrophils observed is suggestive of an accelerated immigration of leukocytes from the bone marrow to the circulating pool that may contribute to an increase in cell survival, subject to modulation by a complex balance of both anti- and proapoptotic factors contained in the circulation of SCD individuals.
Asunto(s)
Anemia de Células Falciformes/patología , Senescencia Celular , Neutrófilos/patología , Adulto , Anciano , Apoptosis , Caspasa 9/metabolismo , Adhesión Celular , Técnicas de Cultivo de Célula , Supervivencia Celular , Femenino , Humanos , Interleucina-8/metabolismo , Recuento de Leucocitos , Masculino , Persona de Mediana Edad , SueroRESUMEN
OBJECTIVES: To determine whether kidney disease and hemolysis are associated with bone mass density in a population of adult Brazilian patients with sickle cell disease. INTRODUCTION: Bone involvement is a frequent clinical manifestation of sickle cell disease, and it has multiple causes; however, there are few consistent clinical associations between bone involvement and sickle cell disease. METHODS: Patients over 20 years of age with sickle cell disease who were regularly followed at the Hematology and Hemotherapy Center of Campinas, Brazil, were sorted into three groups, including those with normal bone mass density, those with osteopenia, and those with osteoporosis, according to the World Health Organization criteria. The clinical data of the patients were compared using statistical analyses. RESULTS: In total, 65 patients were included in this study: 12 (18.5 percent) with normal bone mass density, 37 (57 percent) with osteopenia and 16 (24.5 percent) with osteoporosis. Overall, 53 patients (81.5 percent) had bone mass densities below normal standards. Osteopenia and osteoporosis patients had increased lactate dehydrogenase levels and reticulocyte counts compared to patients with normal bone mass density (p<0.05). Osteoporosis patients also had decreased hemoglobin levels (p<0.05). Hemolysis was significantly increased in patients with osteoporosis compared with patients with osteopenia, as indicated by increased lactate dehydrogenase levels and reticulocyte counts as well as decreased hemoglobin levels. Osteoporosis patients were older, with lower glomerular filtration rates than patients with osteopenia. There was no significant difference between the groups with regard to gender, body mass index, serum creatinine levels, estimated creatinine clearance, or microalbuminuria. CONCLUSION: A high prevalence of reduced bone mass density that was associated with hemolysis was found in this population, as indicated by the high lactate dehydrogenase levels, increased reticulocyte counts and low hemoglobin levels.
Asunto(s)
Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven , Anemia de Células Falciformes/complicaciones , Densidad Ósea/fisiología , Enfermedades Óseas Metabólicas/etiología , Hemólisis/fisiología , Absorciometría de Fotón , Anemia de Células Falciformes/fisiopatología , Enfermedades Óseas Metabólicas/fisiopatología , Tasa de Filtración Glomerular/fisiología , L-Lactato Deshidrogenasa/sangre , Osteoporosis/etiología , Osteoporosis/fisiopatología , Recuento de ReticulocitosRESUMEN
INTRODUCTION: Myelodysplastic syndromes encompass a heterogeneous group of clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis, refractory cytopenia and a tendency to progress toward acute myeloid leukemia. The accumulation of genetic alterations is closely associated with the progression of myelodysplastic syndromes toward acute myeloid leukemia. OBJECTIVE: To investigate the presence of mutations in the points most frequent for mutations (hotspot mutations) in phosphatidylinositol-3-kinase (PI3K), Janus kinase 2 (JAK2), FMS-like tyrosine kinase 3 (FLT3) and nucleophosmin (NPM1), which are involved in leukemia and other cancers, in a population of Brazilian MDS patients. METHODS: Fifty-one myelodysplastic syndromes patients were included in the study. According to French-American-British classification, the patients were distributed as follows: 31 with refractory anemia, 8 with refractory anemia with ringed sideroblasts, 7 with refractory anemia with excess blasts, 3 with refractory anemia with excess blasts in transformation and 2 with chronic myelomonocytic leukemia. Bone marrow samples were obtained and screened for the presence of hotspot mutations using analysis based on amplification with the polymerase chain reaction, sequencing, fragment size polymorphisms or restriction enzyme digestion. All patients were screened for mutations at the time of diagnosis, and 5 patients were also screened at the time of disease progression. RESULTS: These results show that hotspot mutations in the PI3K, JAK2, FLT3 and NPM1 genes are not common in MDS patients; nevertheless, JAK2 mutations may be present in myelodysplasia during disease progression. CONCLUSIONS: These results show that hotspot mutations in the PI3K, JAK2, FLT3 and NPM1 genes are not common in MDS patients; nevertheless, JAK2 mutations may be present in myelodysplasia during disease progression.