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Investigación Biomédica/organización & administración , Investigación Biomédica/normas , Infecciones por Coronavirus/epidemiología , Neumonía Viral/epidemiología , Centros Médicos Académicos , Betacoronavirus , Bancos de Muestras Biológicas/normas , Bancos de Muestras Biológicas/estadística & datos numéricos , Investigación Biomédica/estadística & datos numéricos , COVID-19 , Humanos , Comunicación Interdisciplinaria , Italia/epidemiología , Pandemias , Sistema de Registros/normas , Sistema de Registros/estadística & datos numéricos , SARS-CoV-2RESUMEN
The role of progenitor/stem cells in pituitary tumorigenesis, resistance to pharmacological treatments and tumor recurrence is still unclear. This study investigated the presence of progenitor/stem cells in non-functioning pituitary tumors (NFPTs) and tested the efficacy of dopamine receptor type 2 (DRD2) and somatostatin receptor type 2 (SSTR2) agonists to inhibit in vitro proliferation. They found that 70% of 46 NFPTs formed spheres co-expressing stem cell markers, transcription factors (DAX1, SF1, ERG1) and gonadotropins. Analysis of tumor behavior showed that spheres formation was associated with tumor invasiveness (OR = 3,96; IC: 1.05-14.88, p = 0.036). The in vitro reduction of cell proliferation by DRD2 and SSTR2 agonists (31 ± 17% and 35 ± 13% inhibition, respectively, p < 0.01 vs. basal) occurring in about a half of NFPTs cells was conserved in the corresponding spheres. Accordingly, these drugs increased cyclin-dependent kinase inhibitor p27 and decreased cyclin D3 expression in spheres. In conclusion, they provided further evidence for the existence of cells with a progenitor/stem cells-like phenotype in the majority of NFPTs, particularly in those with invasive behavior, and demonstrated that the antiproliferative effects of dopaminergic and somatostatinergic drugs were maintained in progenitor/stem-like cells.
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Carcinogénesis/genética , Recurrencia Local de Neoplasia/tratamiento farmacológico , Neoplasias Hipofisarias/tratamiento farmacológico , Receptores de Dopamina D2/genética , Receptores de Somatostatina/genética , Adulto , Carcinogénesis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Ciclina D3/biosíntesis , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/biosíntesis , Receptor Nuclear Huérfano DAX-1/biosíntesis , Dopaminérgicos/administración & dosificación , Resistencia a Antineoplásicos/genética , Canal de Potasio ERG1/biosíntesis , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Gonadotropinas/biosíntesis , Humanos , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/genética , Recurrencia Local de Neoplasia/patología , Células Madre Neoplásicas/efectos de los fármacos , Neoplasias Hipofisarias/genética , Neoplasias Hipofisarias/patología , Factores de Empalme de ARN/biosíntesis , Receptores de Dopamina D2/agonistas , Receptores de Somatostatina/agonistas , Esferoides Celulares/efectos de los fármacos , Esferoides Celulares/patologíaRESUMEN
Non-functioning pituitary tumors (NFPTs) frequently present local invasiveness. Dopamine receptor 2 (DRD2) agonists are the only medical therapy that induces tumor shrinkage in some patients. Invasion requires cytoskeleton rearrangements that are tightly regulated by cofilin pathway, whose alterations correlate with invasion in different tumors. We investigated the effect of DR2D agonist on NFPT cells migration/invasion and the molecular mechanisms involved. We demonstrated that DRD2 agonist reduced migration (-44 ± 25%, p < 0.01) and invasion (-34 ± 6%, p < 0.001) and increased about 4-fold Ser3-phosphorylated inactive cofilin (P-cofilin) in NFPT cells. These effects were abolished by inhibiting ROCK, a kinase that phosphorylates cofilin. The overexpression of wild-type or phosphodeficient S3A-cofilin increased HP75 cells migration (+49 ± 6% and +57 ± 9% vs empty vector, respectively, p < 0.05), while phosphomimetic mutant had no effect. Interestingly, P-cofilin levels were lower in invasive vs non-invasive tumors by both western blot (mean P-cofilin/total cofilin ratio 0.77 and 1.93, respectively, p < 0.05) and immunohistochemistry (mean percentage of P-cofilin positive cells 17.6 and 45.7, respectively, p < 0.05). In conclusion, we showed that the invasiveness of pituitary tumors is promoted by the activation of cofilin, which can be regulated by DRD2 and might represent a novel biomarker for pituitary tumors' invasive behavior.
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Factores Despolimerizantes de la Actina/metabolismo , Movimiento Celular , Neoplasias Hipofisarias/enzimología , Receptores de Dopamina D2/metabolismo , Quinasas Asociadas a rho/metabolismo , Factores Despolimerizantes de la Actina/genética , Adulto , Anciano , Antineoplásicos/farmacología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Agonistas de Dopamina/farmacología , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Masculino , Persona de Mediana Edad , Invasividad Neoplásica , Fosforilación , Neoplasias Hipofisarias/tratamiento farmacológico , Neoplasias Hipofisarias/genética , Neoplasias Hipofisarias/patología , Mutación Puntual , Inhibidores de Proteínas Quinasas/farmacología , Receptores de Dopamina D2/agonistas , Transducción de Señal , Transfección , Células Tumorales Cultivadas , Quinasas Asociadas a rho/antagonistas & inhibidoresRESUMEN
PURPOSE: Hypoxia-inducible factor 1α (HIF-1α) activity is one of the major players in hypoxia-mediated glioma progression and resistance to therapies, and therefore the focus of this study was the evaluation of HIF-1α modulation in relation to tumour response with the purpose of identifying imaging biomarkers able to document tumour response to treatment in a murine glioma model. METHODS: U251-HRE-mCherry cells expressing Luciferase under the control of a hypoxia responsive element (HRE) and mCherry under the control of a constitutive promoter were used to assess HIF-1α activity and cell survival after treatment, both in vitro and in vivo, by optical, MRI and positron emission tomography imaging. RESULTS: This cell model can be used to monitor HIF-1α activity after treatment with different drugs modulating transduction pathways involved in its regulation. After temozolomide (TMZ) treatment, HIF-1α activity is early reduced, preceding cell cytotoxicity. Optical imaging allowed monitoring of this process in vivo, and carbonic anhydrase IX (CAIX) expression was identified as a translatable non-invasive biomarker with potential clinical significance. A preliminary in vitro evaluation showed that reduction of HIF-1α activity after TMZ treatment was comparable to the effect of an Hsp90 inhibitor, opening the way for further elucidation of its mechanism of action. CONCLUSION: The results of this study suggest that the U251-HRE-mCherry cell model can be used for the monitoring of HIF-1α activity through luciferase and CAIX expression. These cells can become a useful tool for the assessment and improvement of new targeted tracers for potential theranostic procedures.
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Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/metabolismo , Neoplasias Encefálicas/tratamiento farmacológico , Dacarbazina/análogos & derivados , Glioma/tratamiento farmacológico , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Animales , Biomarcadores de Tumor/genética , Anhidrasas Carbónicas/genética , Anhidrasas Carbónicas/metabolismo , Línea Celular Tumoral , Dacarbazina/uso terapéutico , Evaluación Preclínica de Medicamentos/métodos , Femenino , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Ratones , Ratones Desnudos , Imagen Óptica , TemozolomidaRESUMEN
PURPOSE: The aim of this study was to characterize a cell-based model for the molecular study of hypoxia-inducible factor (HIF)-1α activity, in the context of hypoxia, by means of different imaging techniques. PROCEDURES: Engineered U251-HRE glioma cells were used to analyze the molecular mechanisms underlying HIF-1α activity in vitro in relation to luciferase expression. The same cells were orthotopically implanted in mice to evaluate tumor progression and hypoxia induction by bioluminescence imaging, fluorescence imaging, positron emission tomography (PET), and magnetic resonance imaging (MRI). RESULTS: In vitro analyses highlighted the relationship between HIF-1α and luciferase activity in hypoxic conditions and after pharmacological treatments in U251-HRE cells. Through in vivo studies, it was possible to assess hypoxia establishment in relation to tumor growth by optical imaging, PET and MRI. CONCLUSIONS: The findings of this study indicate that the U251-HRE orthotopic murine model can be used to reliably evaluate processes modulating HIF-1α activity, using both molecular and preclinical non-invasive imaging techniques.
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Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Modelos Biológicos , Imagen Multimodal/métodos , Animales , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Hipoxia de la Célula/efectos de los fármacos , Línea Celular Tumoral , Forma de la Célula/efectos de los fármacos , Deferoxamina/farmacología , Glioma/diagnóstico , Glioma/metabolismo , Glioma/patología , Humanos , Inmunohistoquímica , Luciferasas/metabolismo , Imagen por Resonancia Magnética , Ratones , Imagen Óptica , Tomografía de Emisión de Positrones , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Renal cell carcinoma (RCC) accounts for about 3% of all human malignancies and its incidence is increasing. There are no standard biomarkers currently used in the clinical management of patients with renal cell carcinoma. A promising strategy for new biomarker detection is comparative proteomics of urinary exosomes (UE), nanovesicles released by every epithelial cell facing the urinary space, enriched in renal proteins and excluding high-abundance plasmatic proteins, such as albumin. Aim of the work is to establish the protein profile of exosomes isolated from urines of RCC patient compared with control subjects. We enrolled 29 clear cell RCC patients and 23 control healthy subjects (CTRL), age and sex-matched, for urine collection and vesicle isolation by differential centrifugation. Such vesicles were morphologically and biochemically characterized and proved to share exosome properties. Proteomic analysis, performed on 9 urinary exosome (UE) pooled samples by gel based digestion followed by LC-MS/MS, led to the identification of 261 proteins from CTRL subject UE and 186 from RCC patient UE, and demonstrated that most of the identified proteins are membrane associated or cytoplasmic. Moreover, about a half of identified proteins are not shared between RCC and control UE. Starting from these observations, and from the literature, we selected a panel of 10 proteins, whose UE differential content was subjected to immunoblotting validation. Results show for the first time that RCC UE protein content is substantially and reproducibly different from control UE, and that these differences may provide clues for new RCC biomarker discovery.
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Carcinoma de Células Renales/metabolismo , Exosomas/metabolismo , Neoplasias Renales/metabolismo , Proteoma/análisis , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/patología , Femenino , Humanos , Riñón/metabolismo , Neoplasias Renales/genética , Neoplasias Renales/patología , Masculino , Persona de Mediana Edad , Análisis por Matrices de Proteínas , Proteínas/análisis , ProteómicaRESUMEN
The expression of small, non-coding RNA or microRNAs (miR), is frequently deregulated in human cancer, but how these pathways affect disease progression is still largely elusive. Here, we report on a miR, miR-296, which is progressively lost during tumor progression and correlates with metastatic disease in colorectal, breast, lung, gastric, parathyroid, liver and bile ducts cancers. Functionally, miR-296 controls a global cell motility gene signature in epithelial cells by transcriptionally repressing the cell polarity-cell plasticity module, Scribble (Scrib). In turn, loss of miR-296 causes aberrantly increased and mislocalized Scrib in human tumors, resulting in exaggerated random cell migration and tumor cell invasiveness. Re-expression of miR-296 in MDA-MB231 cells inhibits tumor growth in vivo. Finally, miR-296 or Scrib levels predict tumor relapse in hepatocellular carcinoma patients. These data identify miR-296 as a global repressor of tumorigenicity and uncover a previously unexplored exploitation of Scrib in tumor progression in humans.
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Polaridad Celular , MicroARNs/fisiología , Neoplasias/etiología , Animales , Movimiento Celular , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica , Humanos , Proteínas de la Membrana/genética , Ratones , Neoplasias/genética , Neoplasias/patología , Proteínas Supresoras de Tumor/genéticaRESUMEN
BACKGROUND: Survivin is involved in modulation of cell death and cell division processes. Survivin expression in normal adult tissues has not been fully understood, although it is markedly lower than in cancer, where it is over-expressed. OBJECTIVE: To investigate survivin expression in normal, potentially malignant and cancerous oral mucosa. METHODS: We measured survivin mRNA levels by real-time RT-PCR in specimens of oral mucosa (15 from normal mucosa, 17 from potentially malignant lesions, 17 from neoplasms). Scores were compared using Kruskal-Wallis test and post hoc according to Conover. Chi-squared test was used for dichotomous data. RESULTS: The median relative levels of survivin mRNA resulted six for normal mucosa, eight for potentially malignant lesions, 13 for cancers: differences among these three groups were statistically significant, as between cancer and potentially malignant lesions. Expression in normal mucosa and potentially lesions group showed no significant difference. Low, but not marginal expression of survivin in normal mucosa is a new finding, and it could be explained with the higher sensibility of our methods. CONCLUSIONS: Survivin expression in oral potentially malignant lesions might indicate a progressive deregulation of expression paralleling oncogenesis, particularly during the first stages of process, suggesting a putative predictive role for survivin.
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Carcinoma de Células Escamosas/metabolismo , Leucoplasia Bucal/metabolismo , Proteínas Asociadas a Microtúbulos/biosíntesis , Mucosa Bucal/metabolismo , Neoplasias de la Boca/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Carcinoma de Células Escamosas/química , Estudios de Casos y Controles , Distribución de Chi-Cuadrado , Femenino , Humanos , Proteínas Inhibidoras de la Apoptosis , Leucoplasia Bucal/química , Masculino , Persona de Mediana Edad , Mucosa Bucal/química , Neoplasias de la Boca/química , Pronóstico , ARN Mensajero/análisis , ARN Neoplásico/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Fumar/metabolismo , Estadísticas no Paramétricas , Survivin , Adulto JovenRESUMEN
Parathyroid carcinoma (PaC) is a rare cause of primary hyperparathyroidism. Though the loss of the oncosuppressor CDC73/HRPT2 gene product, parafibromin, has been involved in the hyperparathyroidism-jaw tumor syndrome and in a consistent set of sporadic PaCs, parathyroid carcinogenesis remains obscure. MicroRNAs are a new class of small, non-coding RNAs implicated in development of cancer, since their deregulation can induce aberrant expression of several target genes. The aim of the present study was to identify differentially expressed microRNAs in parathyroid cancers compared with normal tissues. We performed a TaqMan low-density array profiling of four parathyroid cancers harboring CDC73 inactivating mutations and negative for parafibromin immunostaining. Their microRNA profiling was compared with that of two normal parathyroid biopsies. Out of 362 human microRNAs assayed, 279 (77%) were successfully amplified. Fourteen and three microRNAs were significantly down- and over-expressed in parathyroid cancers respectively. Of these, miR-296 and miR-139 were down-regulated, and miR-503 and miR-222 were over-expressed with a null false discovery rate. Carcinomas could be discriminated from parathyroid adenomas by a computed score based on the expression levels of miR-296, miR-222, and miR-503 as miR-139 was similarly down-regulated in both cancers and adenomas. Finally, miR-296 and miR-222 levels negatively correlated with mRNA levels of the hepatocyte growth factor receptor-regulated tyrosine kinase substrate and p27/kip1 levels respectively. These results suggest the existence of an altered microRNA expression pattern in PaCs together with a potential role of miR-296 as novel oncosuppressor gene in these neoplasia.
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Adenoma/genética , Carcinoma/genética , MicroARNs/genética , Glándulas Paratiroides/metabolismo , Neoplasias de las Paratiroides/genética , Adenoma/diagnóstico , Adulto , Carcinoma/diagnóstico , Análisis por Conglomerados , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Genes Supresores de Tumor/fisiología , Salud , Humanos , MicroARNs/metabolismo , MicroARNs/fisiología , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , Neoplasias de las Paratiroides/diagnóstico , PronósticoRESUMEN
OBJECTIVES: Aberrant cAMP signaling is involved in the pathogenesis of somatotropinomas. The aim of the study was to screen acromegalic patients for the presence of variants of phosphodiesterase type 11A (PDE11A) gene, which have been recently identified in adrenocortical and testicular tumors. SUBJECTS AND METHODS: We sequenced the PDE11A gene-coding region in 78 acromegalic patients and 110 controls. Immunohistochemistry for PDE11A was performed in a subgroup of adenomas and normal pituitary samples. RESULTS: We found 15 nonsynonymous germline substitutions in 13 acromegalic patients (17%), i.e. 14 missense variants (Y727C in six, R804H in one, R867G in four, and M878V in three) and one truncating mutation (FS41X), with a prevalence only slightly higher than that observed in controls (14%). Immunohistochemistry revealed PDE11A expression higher in somatotropinomas than in normal somatotrophs, without significant difference between tumors with or without PDE11A variants, with the exception of two tumors (one with loss of heterozygosity (LOH) at the PDE11A locus and one with FS41X mutation) showing markedly reduced PDE11A staining. No significant differences in hormonal and clinical parameters between patients with or without PDE11A variants were observed, although patients with PDE11A changes showed a tendency to have a more aggressive tumor compared with patients with wild-type sequence (extrasellar extension in 69 vs 45%). CONCLUSIONS: This study first demonstrated the presence of PDE11A variants in a subset of acromegalic patients, which was only slightly more frequent than in controls. The normal expression of the enzyme in the majority of tumor tissues together with the lack of significant clinical phenotype suggests that these variants might only marginally contribute to the development of somatotropinomas.
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Acromegalia/enzimología , Adenoma/enzimología , Hidrolasas Diéster Fosfóricas/genética , Neoplasias Hipofisarias/enzimología , 3',5'-GMP Cíclico Fosfodiesterasas , Acromegalia/sangre , Acromegalia/genética , Adenoma/sangre , Adenoma/genética , Adulto , Secuencia de Bases , ADN de Neoplasias/química , ADN de Neoplasias/genética , Femenino , Variación Genética , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Neoplasias Hipofisarias/sangre , Neoplasias Hipofisarias/genética , Reacción en Cadena de la Polimerasa , Análisis de RegresiónRESUMEN
The cAMP-protein kinase A (PKA) pathway is the major signal transduction pathway involved in melanocyte-stimulating hormone receptor-mediated signaling and melanin production, whereas its role in the control of melanocyte proliferation is still controversial. In this study, we evaluated the effects of selective activation of the different PKA regulatory subunits type 1A (R1A) and type 2B (R2B) on melanocyte proliferation. Immunohistochemistry demonstrated that normal melanocytes lacked R1A protein whereas this subunit was highly expressed in all human melanomas studied (N=20) and in six human melanoma cell lines. Pharmacological activation of the R2 subunits by the cAMP analogue 8-Cl-cAMP inhibited proliferation and increased caspase-3 activity by 68.77+/-10.5 and 72+/-9% respectively, in all cell lines with the exception of the only p53-mutated one. Similar effects were obtained by activating R2 subunits with other analogues and by silencing R1A expression. The antiproliferative and proapoptotic effects of 8-Cl-cAMP were comparable to those observed with commonly used antitumoral drugs. Moreover, 8-Cl-cAMP potentiated the effects of these drugs on both cell proliferation and caspase-3 activity. In conclusion, this study first reports that human melanomas are characterized by a high R1/R2 ratio and that pharmacological and genetic manipulations able to revert this unbalanced expression cause significant antiproliferative and proapoptotic effects in melanoma cells.
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Proliferación Celular , Subunidad RIIbeta de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Melanoma/metabolismo , Adulto , Anciano , Apoptosis , Western Blotting , Caspasa 3/metabolismo , AMP Cíclico/metabolismo , Humanos , Técnicas para Inmunoenzimas , Melanocitos/metabolismo , Melanoma/patología , Persona de Mediana Edad , Células Tumorales CultivadasRESUMEN
The four regulatory subunits (R1A, R1B, R2A, R2B) of protein kinase A (PKA) are differentially expressed in several cancer cell lines and exert distinct roles in growth control. Mutations of the R1A gene have been found in patients with Carney complex and in a minority of sporadic primary pigmented nodular adrenocortical disease (PPNAD). The aim of this study was to evaluate the expression of PKA regulatory subunits in non-PPNAD adrenocortical tumors causing ACTH-independent Cushing's syndrome and to test the impact of differential expression of these subunits on cell growth. Immunohistochemistry demonstrated a defective expression of R2B in all cortisol-secreting adenomas (n=16) compared with the normal counterpart, while both R1A and R2A were expressed at high levels in the same tissues. Conversely, carcinomas (n=5) showed high levels of all subunits. Sequencing of R1A and R2B genes revealed a wild type sequence in all tissues. The effect of R1/R2 ratio on proliferation was assessed in mouse adrenocortical Y-1 cells. The R2-selective cAMP analogue 8-Cl-cAMP dose-dependently inhibited Y-1 cell proliferation and induced apoptosis, while the R1-selective cAMP analogue 8-HA-cAMP stimulated cell proliferation. Finally, R2B gene silencing induced up-regulation of R1A protein, associated with an increase in cell proliferation. In conclusion, we propose that a high R1/R2 ratio favors the proliferation of well differentiated and hormone producing adrenocortical cells, while unbalanced expression of these subunits is not required for malignant transformation.
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Neoplasias de la Corteza Suprarrenal/enzimología , Neoplasias de la Corteza Suprarrenal/genética , Adenoma Corticosuprarrenal/enzimología , Adenoma Corticosuprarrenal/genética , Proliferación Celular/efectos de los fármacos , Proteínas Quinasas Dependientes de AMP Cíclico/genética , Hidrocortisona/metabolismo , 8-Bromo Monofosfato de Adenosina Cíclica/análogos & derivados , 8-Bromo Monofosfato de Adenosina Cíclica/farmacología , Neoplasias de la Corteza Suprarrenal/metabolismo , Adenoma Corticosuprarrenal/metabolismo , Secuencia de Bases/genética , Carcinoma/enzimología , Carcinoma/genética , Carcinoma/metabolismo , Línea Celular Tumoral , Transformación Celular Neoplásica/genética , Subunidad RIIbeta de la Proteína Quinasa Dependiente de AMP Cíclico/genética , Subunidad RIIbeta de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/genética , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Relación Dosis-Respuesta a Droga , Regulación Enzimológica de la Expresión Génica/genética , Regulación Neoplásica de la Expresión Génica/genética , Silenciador del Gen/fisiología , Humanos , Regulación hacia Arriba/genéticaRESUMEN
Gastric carcinoma is one of the major causes of cancer mortality worldwide. Early detection results in excellent prognosis for patients with early cancer (EGC), whereas the prognosis of advanced cancer (AGC) patients remains poor. It is not clear whether EGC and AGC are molecularly distinct, and whether they represent progressive stages of the same tumor or different entities ab initio. Gene expression profiles of EGC and AGC were determined by Affymetrix technology and quantitative polymerase chain reaction. Representative regulated genes were further analysed by in situ hybridization (ISH) on tissue microarrays. Expression analysis allowed the identification of a signature that differentiates AGC from EGC. In addition, comparison with normal gastric mucosa indicated that the majority of alterations associated with EGC are retained in AGC, and that further expression changes mark the transition from EGC to AGC. Finally, ISH analysis showed that representative genes, differentially expressed in the invasive areas of EGC and AGC, are not differentially expressed in the non-invasive areas of the same tumors. Our data are more directly compatible with a progression model of gastric carcinogenesis, whereby EGC and AGC may represent different molecular stages of the same tumor. Finally, the identification of an AGC-specific signature might help devising novel therapeutic strategies for advanced gastric cancer.
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Biomarcadores de Tumor/biosíntesis , Biomarcadores de Tumor/genética , Perfilación de la Expresión Génica , Neoplasias Gástricas/genética , Neoplasias Gástricas/patología , Diferenciación Celular/genética , Proliferación Celular , Progresión de la Enfermedad , Estudios de Seguimiento , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Índice de Severidad de la Enfermedad , Neoplasias Gástricas/clasificación , Neoplasias Gástricas/metabolismoRESUMEN
BACKGROUND: An increased risk of testicular cancer in men with infertility and poor semen quality has been reported. Our aim was to investigate the prevalence of testicular nodules and cancer in azoospermic subjects with different spermatogenetic patterns. METHODS: A total of 1443 consecutive infertile men were investigated, out of which 145 (10.0%) were found to be azoospermic. By using clinical examination and testicular ultrasound, 11 out of the 145 patients showed testicular nodules (2.8-26 mm). To obtain spermatozoa for assisted reproduction, 97 subjects required testicular sperm extraction (TESE) and biopsy, including the 11 patients with nodules. They were divided into two groups according to biopsy results: Group A (n = 38) with complete Sertoli cell-only syndrome (SCOS) and Group B (n = 59) with varying spermatogenetic patterns. Ten nodules were found in Group A and one in Group B. RESULTS: In azoospermic men, the overall prevalence of nodules was 7.5%. In complete SCOS, the prevalence of nodules and cancer was 10/38 (26.3%) and 4/38 (10.5%), respectively. Amongst the cancers, one embryonal carcinoma, one seminoma and two in-situ carcinomas were found. CONCLUSION: The prevalence of testicular nodules and cancer in azoospermic men with complete SCOS is very high. In these subjects, the role of clinical evaluation, ultrasound and biopsy should be emphasized.
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Azoospermia/epidemiología , Infertilidad Masculina/diagnóstico , Espermatogénesis , Neoplasias Testiculares/epidemiología , Adulto , Biopsia , Humanos , Infertilidad Masculina/epidemiología , Células Intersticiales del Testículo/metabolismo , Masculino , Prevalencia , Células de Sertoli/metabolismo , Espermatozoides/metabolismo , Síndrome , Testículo/diagnóstico por imagen , Testículo/patología , Ultrasonografía/métodosRESUMEN
Survivin is a dual regulator of cell proliferation and cell viability overexpressed in most human tumors. Although strategies to lower survivin levels have been pursued for rational cancer therapy, the molecular circuitries controlling survivin expression in tumors have not been completely elucidated. Here, we show that stimulation with insulin-like growth factor-1 (IGF-1) results in increased survivin expression in prostate cancer cells. This response is independent of de novo gene transcription, changes in mRNA expression or modifications of survivin protein stability. Instead, IGF-1 induced persistence and translation of a pool of survivin mRNA, in a reaction abolished by the mTOR (mammalian target of rapamycin) inhibitor, rapamycin. Forced expression of the mTOR target p70S6K1 reproduced the increase in survivin expression in prostate cancer cells, whereas acute ablation of endogenous p70S6K1 by small interfering RNA downregulated survivin levels. Rapamycin, alone or in combination with suboptimal concentrations of taxol reduced survivin protein levels, and decreased viability of prostate cancer cells. Therefore, IGF-1/mTOR signaling elevates survivin in prostate cancer cells via rapid changes in mRNA translation. Antagonists of this pathway may be beneficial to lower an antiapoptotic threshold maintained by survivin in prostate cancer.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Factor I del Crecimiento Similar a la Insulina/farmacología , Proteínas Asociadas a Microtúbulos/genética , Proteínas de Neoplasias/genética , Neoplasias de la Próstata/metabolismo , Proteínas Quinasas/metabolismo , Transducción de Señal , Animales , Apoptosis , Ciclo Celular , Proliferación Celular , Regulación hacia Abajo , Células HeLa , Humanos , Proteínas Inhibidoras de la Apoptosis , Masculino , Ratones , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/antagonistas & inhibidores , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/metabolismo , Regiones Promotoras Genéticas , Neoplasias de la Próstata/genética , Proteínas Quinasas/genética , Estabilidad del ARN , ARN Interferente Pequeño/farmacología , Receptor IGF Tipo 1/genética , Receptor IGF Tipo 1/fisiología , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , Sirolimus , Survivin , Serina-Treonina Quinasas TOR , Transcripción Genética , Células Tumorales Cultivadas/efectos de los fármacosRESUMEN
In view of a potential clinical use we aimed this study to assess the selective homing to the injured myocardium and the definitive fate of peripherally injected labeled and previously cryopreserved Bone Marrow Mononuclear cells (BMMNCs). The myocardial damage (cryoinjury) was produced in 59 rats (45 treated, 14 controls). From 51 donor rats 4.4 x 10(9) BMMNCs were isolated and cryopreserved (slow-cooling protocols); the number of CD34+ and the viability of pooled cells was assessed by flow-cytometry analysis before and after cryopreservation and simulated delivery through a 23G needle. Seven days after injury, BMMNCs were thawed, labeled with PKH26 dye and peripherally injected (20 x 10(6) cells in 500 microl) in recipient rats. Two weeks after experimental injury, the heart, lungs, liver, kidneys, spleen and thymus were harvested to track transplanted cells. Except a small amount in the spleen, PKH26+ cells were found only in the infarcted myocardium of the treated animals. Typical vascular structures CD34+ were found in the infarcted areas of all animals; treated rats showed a significantly higher number of these structures if compared with untreated. Morphological ultra-structural examination of infarcted areas confirmed in treated rats the presence of early-stage PKH26+ vascular structures derived from injected BMMNCs. The estimated mean CD34+ cells loss due to the cryopreservation procedure and to the system of delivery was 0.24% and 0.1%, respectively, confirming the feasibility of the procedure. This study supports the possible therapeutic use of cryopreserved peripherally injecetd BMMNCs as a source of CD34+ independent vascular structures following myocardial damage.
Asunto(s)
Células de la Médula Ósea/fisiología , Criopreservación , Trasplante de Células Madre Hematopoyéticas , Leucocitos Mononucleares/fisiología , Infarto del Miocardio/terapia , Neovascularización Fisiológica , Animales , Antígenos CD34/análisis , Movimiento Celular , Masculino , Infarto del Miocardio/fisiopatología , Ratas , Ratas Endogámicas F344RESUMEN
Somatostatin analogs currently used in the treatment of acromegaly and other neuroendocrine tumors inhibit hormone secretion and cell proliferation by binding to somatostatin receptor type (SST) 2 and 5. The antiproliferative pathways coupled to these receptors have been only partially characterized. The aim of this study was to evaluate the effect of octreotide and super selective SST2 (BIM23120) and SST5 (BIM23206) analogs on apoptotic activity and apoptotic gene expression in human somatotroph tumor cells. Eight somatotroph tumors expressing similar levels of SST2 and SST5 evaluated by real-time PCR and western blot analyses were included in the study. In cultured cells obtained from these tumors, octreotide induced a dose-dependent increase of caspase-3 activity (160+/-20% vs basal at 10 nM) and cleaved cytokeratin 18 levels (172+/-25% vs basal) at concentrations higher than 0.1 nM. This effect was due to SST2 activation since BIM23120 elicited comparable responses, while BIM23206 was ineffective. BIM23120-stimulated apoptosis was dependent on phosphatases, since it was abrogated by the inhibitor orthovanadate, and independent from the induction of apoptosis-related genes, such as p53, p63, p73, Bcl-2, Bax, BID, BIK, TNFSF8, and FADD. In somatotroph tumors, both BIM23120 and BIM2306 caused growth arrest as indicated by the increase in p27 and decrease in cyclin D1 expression. In conclusion, the present study showed that octreotide-induced apoptosis in human somatotroph tumor cells by activating SST2. This effect, together with the cytostatic action exerted by both SST2 and SST5 analogs, might account for the tumor shrinkage observed in acromegalic patients treated with long-acting somatostatin analogs.
Asunto(s)
Apoptosis/efectos de los fármacos , Octreótido/farmacología , Neoplasias Hipofisarias/patología , Receptores de Somatostatina/fisiología , Acromegalia , Caspasa 3/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Neoplasias Hipofisarias/genética , ARN Mensajero/genética , Receptores de Somatostatina/efectos de los fármacos , Receptores de Somatostatina/genética , Células Tumorales CultivadasRESUMEN
AIM: To evaluate the clinical implications of c-kit (CD117) expression in plasma cell myeloma (PCM). METHODS AND RESULTS: We first evaluated the reliability of immunohistochemistry in assessing c-kit expression by comparing the results with those obtained by flow cytometry and gene expression arrays in 22 PCM and in 10 PCM cell lines. Immunohistochemical results showed a perfect concordance with those of flow cytometry; likewise, immunohistochemical and gene expression data were also concordant in all but one PCM and cell lines analysed. Then, we investigated the clinical implications of c-kit immunoreactivity in bone marrow biopsies of 85 PCM patients with a mean follow-up of 41 months. C-kit immunoreactivity was detected in 24 (28.2%) of the 85 cases and it was significantly associated with a high microvessel density, but not with traditional clinicopathological characteristics or with survival. CONCLUSIONS: Our findings suggest that immunohistochemistry is a reliable indicator of c-kit gene expression and reinforce the notion that approximately one-third of PCM express high levels of c-kit. The lack of association with traditional clinicopathological parameters and patient survival suggests that c-kit expression may not be an adjunct in predicting the clinical course of the disease.
Asunto(s)
Mieloma Múltiple/patología , Proteínas Proto-Oncogénicas c-kit/biosíntesis , Adulto , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Femenino , Citometría de Flujo , Regulación Neoplásica de la Expresión Génica , Humanos , Inmunohistoquímica , Masculino , Glicoproteínas de Membrana/análisis , Persona de Mediana Edad , Mieloma Múltiple/genética , Mieloma Múltiple/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteoglicanos/análisis , Proteínas Proto-Oncogénicas c-kit/genética , Análisis de Supervivencia , SindecanosRESUMEN
BACKGROUND: Loss of TP53 function through gene mutation is a critical event in the development and progression of many tumour types including colorectal cancer (CRC). In vitro studies have found considerable heterogeneity amongst different TP53 mutants in terms of their transactivating abilities. The aim of this work was to evaluate whether TP53 mutations classified as functionally inactive (< or=20% of wildtype transactivation ability) had different prognostic and predictive values in CRC compared with mutations that retained significant activity. MATERIALS AND METHODS: TP53 mutations within a large, international database of CRC (n = 3583) were classified according to functional status for transactivation. RESULTS: Inactive TP53 mutations were found in 29% of all CRCs and were more frequent in rectal (32%) than proximal colon (22%) tumours (P < 0.001). Higher frequencies of inactive TP53 mutations were also seen in advanced stage tumours (P = 0.0003) and in tumours with the poor prognostic features of vascular (P = 0.006) and lymphatic invasion (P = 0.002). Inactive TP53 mutations were associated with significantly worse outcome only in patients with Dukes' stage D tumours (RR = 1.71, 95%CI 1.25-2.33, P < 0.001). Patients with Dukes' C stage tumours appeared to gain a survival benefit from 5-fluorouracil-based chemotherapy regardless of TP53 functional status for transactivation ability. CONCLUSIONS: Mutations that inactivate the transactivational ability of TP53 are more frequent in advanced CRC and are associated with worse prognosis in this stage of disease.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias Colorrectales/genética , Mutación , Proteína p53 Supresora de Tumor/genética , Adenocarcinoma/tratamiento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/patología , Anciano , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/patología , Análisis Mutacional de ADN , Exones , Femenino , Estudios de Seguimiento , Humanos , Agencias Internacionales , Masculino , Persona de Mediana Edad , Invasividad Neoplásica/genética , Invasividad Neoplásica/patología , Estadificación de Neoplasias , Tasa de SupervivenciaRESUMEN
Hesx1 is a paired-like homeobox gene first expressed during mouse embryogenesis in the anterior midline visceral endoderm. As gastrulation proceeds, Hesx1 is expressed in the ventral prosencephalon and, subsequently, at E9.0 appears in the ventral diencephalon and in the thickened layer of oral ectoderm that give rise to Rathke's pouch, the primordium of the anterior pituitary gland. Hesx1 continues to be expressed in the developing anterior pituitary until E11.5 when its transcripts disappear in a spatiotemporal sequence corresponding to progressive pituitary cell differentiation, becoming undetectable by E15.5. In the present study, we investigated whether HESX1 is expressed during adult life in human normal pituitaries and in different types of human pituitary adenomas. We analysed, using quantitative RT-PCR method, three normal pituitaries, seven GH-, two TSH-, two PRL-, one ACTH-secreting adenomas, and seven nonfunctioning pituitary tumors. HESX1 mRNA was found to be expressed in normal pituitaries and in all the pituitary tumors that we have analysed. These results suggest that in humans HESX1 is not turned-off during the adult life as it occurs in mice. Thus, HESX1 in humans might play a role in the maintenance of the anterior pituitary cell types and function, as well as in the differentiation of pituitary adenomas, whose pathogenetic mechanisms remain to be further investigated. This is the first study on HESX1 expression in humans during adult life.