RESUMEN

The epidermal growth factor receptor (EGFR) is involved in the regulation of several cellular processes and in the development of many human cancers. Somatic mutations of EGFR at tyrosine kinase domain have been associated with clinical response to tyrosine kinase inhibitors (TKIs) in lung cancer patients. In this study, we evaluated the frequency of point mutations in EGFR for future use of TKI in clinical treatment of bladder cancer. A total, 50 Moroccan patient specimens with bladder cancer and 48 healthy controls were analysed for EGFR mutations in the region delimiting exons 18-21 by PCR amplification and direct sequencing. Our results showed the absence of mutations in the EGFR kinase domain in these exons in all analysed specimens. However, sequence analysis of the EGFR-TK domain, revealed the presence of (G2607A) polymorphism at exon 20. Statistical analysis showed significant difference in the frequencies of G2607A polymorphism between cancer cases and healthy controls (p=0.0001) and the frequencies of the GG and GA/AA genotypes among the cancer cases were 28% and 72%, respectively. Moreover, allelic frequencies of G2607A polymorphism showed significant difference between cancer cases and healthy controls (p=0.0025). Data analysis showed no significant association between G2607A polymorphism and patients' age, clinical stage and tumor grade (p > 0.05). However, a significant difference was found between this polymorphism and patients' sex that could be a sampling bias due to the very limited number of women with bladder cancer. Our findings highlight that, mutations in EGFR kinase domain is a rare event in bladder cancer, suggesting, that treatment of bladder cancer patients with TKI may not be effective. However, the EGFR G2607A polymorphism in exon 20 is frequent in bladder cancer cases and must be further explored for its relevance in the treatment of this disease.

Asunto(s)

Receptores ErbB/genética , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Mutación/genética , Neoplasias de la Vejiga Urinaria/genética , Adulto , Anciano , Secuencia de Bases , Estudios de Casos y Controles , Análisis Mutacional de ADN , Exones/genética , Femenino , Frecuencia de los Genes/genética , Humanos , Masculino , Persona de Mediana Edad , Marruecos , Polimorfismo de Nucleótido Simple/genética , Factores de Riesgo

RESUMEN

Proliferation and differentiation of retinal progenitor cells (RPCs) are tightly controlled by extrinsic cues and distinct combinations of transcription factors leading to the generation of retinal cell type diversity. In this context, we investigated the role of the protein tyrosine phosphatase interacting protein 51 (PTPIP51) in the differentiation of RPCs. The expression pattern of PTPIP51 was analyzed by immunostaining during post-natal retinal development in the rat. Ex vivo electroporation has been used to silence or misexpress PTPIP51 in post-natal retinal explants, and the retinal phenotype was investigated after 3-7days in vitro (div). PTPIP51 expression in the retina started postnatally and was maintained throughout adulthood, especially in retinal ganglion cells and in the inner segment of photoreceptor cells. Silencing of Ptpip51 expression in postnatal retina failed to modify the commitment of late RPCs in the different lineages but severely impaired the final differentiation of photoreceptors, observed by a decrease in the fraction of Rhodopsin-positive cells after 7div. By contrast, misexpression of PTPIP51 in early or late RPCs failed to modify the differentiation of the RPCs. Our data demonstrate that PTPIP51 is implicated in the differentiation process of immature photoreceptors. Because PTPIP51 is specifically localized in the inner segment, PTPIP51 may contribute to the complex stage of maturation of the apical segment of these cells.

Asunto(s)

Células-Madre Neurales/fisiología , Neurogénesis/fisiología , Células Fotorreceptoras de Vertebrados/fisiología , Proteínas Tirosina Fosfatasas/metabolismo , Retina/crecimiento & desarrollo , Retina/fisiología , Animales , Western Blotting , Electroporación , Técnica del Anticuerpo Fluorescente , Silenciador del Gen , Hibridación in Situ , Proteínas Tirosina Fosfatasas/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa , Técnicas de Cultivo de Tejidos

RESUMEN

Coagulase-negative staphylococci (CoNS) are the most frequent cause of late-onset sepsis (LOS) in neonatal intensive care units (NICUs). Staphylococcus epidermidis is usually considered the most prevalent CoNS in this setting. However, recent reports have identified Staphylococcus capitis, another CoNS, as an emerging cause of bacteremia in NICU wards. S. capitis is the main cause of LOS in several NICUs in France, whereas this species is rarely found in adult patients from the same hospitals. S. capitis isolates from NICU infants share several striking features: they all belong to the same pulsed-field gel electrophoresis type, designated as NRCS-A, which indicates their clonal relatedness; their resistance profile reflects adaptation to antimicrobial agents specifically used in NICUs, including resistance to beta-lactams and aminoglycosides but not to fluoroquinolones, and reduced susceptibility to vancomycin; and they are associated with more severe LOS than those caused by other CoNS. The molecular characterization of NICU S. capitis isolates from several countries has shown that S. capitis NRCS-A strains have disseminated in both Western Europe (France, the United Kingdom, and Belgium) and Australia. The dissemination of such multiresistant strains imposes difficult therapeutic choices on pediatricians. As a consequence of the recent strengthening of the French and European guidelines that regulate the interpretation of clinical vancomycin susceptibility in staphylococci, a non-negligible proportion of NICU CoNS isolates (including S. capitis as well as other CoNS species) that were usually reported as vancomycin-susceptible are now categorized as vancomycin-resistant. In such cases, practitioners are faced with uncomfortable alternatives: the continued use of vancomycin in spite of the pathogen being unambiguously reported as resistant to this molecule and the use of antimicrobial agents such as linezolid or daptomycin that retain an in vitro efficacy against CoNS but whose use in neonates has not received approval by the healthcare authorities. To cope with this emerging challenge, clinical investigations of the relative tolerance and efficacy of vancomycin, linezolid, and daptomycin in NICU infants infected with these newly reported vancomycin-resistant CoNS are urgently needed.

Asunto(s)

Sepsis/tratamiento farmacológico , Sepsis/microbiología , Infecciones Estafilocócicas/tratamiento farmacológico , Acetamidas/uso terapéutico , Antiinfecciosos/uso terapéutico , Daptomicina/uso terapéutico , Humanos , Recién Nacido , Unidades de Cuidado Intensivo Neonatal , Linezolid , Oxazolidinonas/uso terapéutico , Infecciones Estafilocócicas/microbiología , Resistencia a la Vancomicina

RESUMEN

The clinical presentation of adrenal hemorrhage varies, depending on the extent of hemorrhage as well as the amount of adrenal cortex involved by the hemorrhage. We report here a case of neonatal adrenal hemorrhage revealed by late onset of neonatal jaundice. This adrenal hemorrhage most probably resulted from shoulder dystocia. The aim of this work was to focus on the fact that jaundice can be caused by adrenal hemorrhage and to emphasize the crucial importance of abdominal ultrasound in cases of persistent jaundice.

Asunto(s)

Enfermedades de las Glándulas Suprarrenales/complicaciones , Hemorragia/complicaciones , Ictericia Neonatal/etiología , Enfermedades de las Glándulas Suprarrenales/sangre , Enfermedades de las Glándulas Suprarrenales/diagnóstico , Enfermedades de las Glándulas Suprarrenales/terapia , Insuficiencia Suprarrenal/sangre , Insuficiencia Suprarrenal/diagnóstico , Insuficiencia Suprarrenal/terapia , Hormona Adrenocorticotrópica/sangre , Distocia/diagnóstico , Femenino , Estudios de Seguimiento , Hemorragia/diagnóstico , Humanos , Hidrocortisona/uso terapéutico , Recién Nacido , Ictericia Neonatal/sangre , Ictericia Neonatal/diagnóstico , Ictericia Neonatal/terapia , Masculino , Embarazo , Ultrasonografía

RESUMEN

Optimising antifungal treatment requires the fast and species-specific identification of yeast isolates. We evaluated a modified protocol for the rapid identification of clinical yeast isolates using matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) technology. First, we evaluated a simplified extraction procedure using 54 clinical yeast isolates. Second, we validated a new protocol with this simplified extraction procedure and lower identification threshold by analysing 167 isolates with either MALDI-TOF or conventional identification techniques. MALDI-TOF analysis with both the standard and short extraction procedure yielded identical identification results, although the log-scores were lower with the latter. With the modified protocol, 163/167 (97.6%) isolates showed a correct identification as compared to conventional identification techniques. A total of 135 out of the 163 (82.8%) correct identifications showed log-scores above 1.7, which we considered as the minimum log-score for secure species identification. The rapid identification of clinical yeast isolates is crucial in patient management. The MALDI-TOF technique using a short extraction procedure can be an alternative for the labourious standard procedure, although the log-scores will be lower. The identification of clinical yeast isolates with the modified protocol is a practical and accurate alternative for conventional identification techniques. If the isolate shows a log-score below 1.7, the standard extraction procedure should be used.

Asunto(s)

Técnicas de Laboratorio Clínico/métodos , Micología/métodos , Micosis/diagnóstico , Micosis/microbiología , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Levaduras/química , Levaduras/aislamiento & purificación , Algoritmos , Humanos

RESUMEN

Desmoids tumors are rare. They often develop from the fascia and muscles of the abdominal wall. They are considered as benign, but endowed with local aggressiveness. Treatment is primarily surgical. Complete resection with large safety margins and sometimes complex reconstruction is necessary to reduce the risk of local reccurrence. WE report three cases of histology proven desmoids tumors of the abdominal wall treated between 2000 and 2007. Etiologic factors, diagnosis, surgical management and adjuvant therapy in case of incomplete resection or reccurrence are discussed.

Asunto(s)

Pared Abdominal/cirugía , Fibromatosis Agresiva/patología , Fibromatosis Agresiva/cirugía , Adulto , Femenino , Humanos

RESUMEN

AIMS: To study the effectiveness of a combination of cell-adsorbed bacteriocin (CAB; a suspension of producer cells on which maximum bacteriocin has been immobilized by pH adjustments) of a Lactobacillus curvatus strain with oregano or savory essential oil to control Listeria monocytogenes in pork meat at 4 degrees C. METHODS AND RESULTS: The antimicrobial activity of the CAB and six different essential oils was tested by the well diffusion assay against L. monocytogenes M, Escherichia coli 10536 and Salmonella serotype Typhi CWBI-H1. The anti-Listeria activity of the CAB and oregano or savory essential oils was also investigated in pork meat. The results of the well diffusion assay showed that CAB was only inhibitory to L. monocytogenes while savory and oregano essential oils were the most active against the three indicator bacteria. In pork meat, Listeria counts have declined from c. 10(2) CFU g(-1) to below the detectable limit during the first week of storage in samples treated with CAB or oregano essential oil and in those treated with CAB combined with oregano or savory essential oil. However, the counts of L. monocytogenes have increased after the third week of storage in all samples with the exception of those treated with the combination of CAB and oregano essential oil. The combination of CAB with savory essential oil resulted in a 2-week delay of the growth rebound compared with samples treated with CAB alone. CONCLUSIONS: Addition of oregano or savory essential oil exhibited a synergistic effect with CAB to control L. monocytogenes in pork meat during storage at 4 degrees C. SIGNIFICANCE AND IMPACT OF THE STUDY: The combination of CAB with oregano or savory essential oil may be effectively used in meat industry to enhance the safety and stability of meat products.

Asunto(s)

Antibacterianos/farmacología , Bacteriocinas/farmacología , Conservación de Alimentos , Lactobacillus/metabolismo , Listeria monocytogenes/efectos de los fármacos , Carne/microbiología , Aceites Volátiles/farmacología , Animales , Bacteriocinas/biosíntesis , Aceites de Plantas/farmacología , Porcinos

Asunto(s)

Hemocromatosis/genética , Antígenos de Histocompatibilidad Clase I/genética , Longevidad/genética , Proteínas de la Membrana/genética , Mutación/genética , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Femenino , Francia , Proteína de la Hemocromatosis , Heterocigoto , Homocigoto , Humanos , Masculino

RESUMEN

This study investigates the effects of SR141716, a selective CB(1) receptor antagonist that reduces food intake and body weight of rodents, on Acrp30 mRNA expression in adipose tissue. Acrp30, a plasma protein exclusively expressed and secreted by adipose tissue, has been shown to induce free fatty acid oxidation, hyperglycemia and hyperinsulinemia decrease, and body weight reduction. We report that N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboximide hydrochloride (SR141716) treatment once daily (10 mg/kg/d, i.p.) from 2 to 14 days reduced body weight and stimulated Acrp30 mRNA expression in adipose tissue of obese Zucker (fa/fa) rats. In parallel, the hyperinsulinemia associated with this animal model was reduced by SR141716 treatment. In cultured mouse adipocytes (3T3 F442A), SR141716 (25 to 100 nM) also induced an overexpression of Acrp30 mRNA and protein. In addition, in adipose tissue of CB(1)-receptor knockout mice, SR141716 had no effect on Acrp30 mRNA expression, demonstrating a CB(1) receptor mediating effect. Furthermore, RT-PCR analysis revealed that rat adipose tissue and 3T3 F442A adipocytes expressed CB(1) receptor mRNA. Relative quantification of this expression revealed an up-regulation (3- to 4-fold) of CB(1) receptor mRNA expression in adipose tissue of obese (fa/fa) rats and in differentiated 3T3 F442A adipocytes compared with lean rats and undifferentiated adipocytes, respectively. Western blot analysis revealed the presence of CB(1) receptors in 3T3 F442A adipocytes, and their expression was up-regulated in differentiated cells. These results show that SR141716 stimulated Acrp30 mRNA expression in adipose tissue by an effect on adipocytes, and reduced hyperinsulinemia in obese (fa/fa) rats. These hormonal regulations may participate in the body weight reduction induced by SR141716 and suggest a role of metabolic regulation in the antiobesity effect of SR141716.

Asunto(s)

Adipocitos/efectos de los fármacos , Tejido Adiposo/efectos de los fármacos , Péptidos y Proteínas de Señalización Intercelular , Obesidad/patología , Piperidinas/farmacología , Biosíntesis de Proteínas , Pirazoles/farmacología , Receptores de Droga/antagonistas & inhibidores , Células 3T3 , Adipocitos/metabolismo , Adiponectina , Tejido Adiposo/fisiopatología , Animales , Peso Corporal/efectos de los fármacos , Cannabinoides/antagonistas & inhibidores , Células Cultivadas , Modelos Animales de Enfermedad , Expresión Génica/efectos de los fármacos , Hiperinsulinismo/tratamiento farmacológico , Masculino , Ratones , Piperidinas/uso terapéutico , Proteínas/genética , Pirazoles/uso terapéutico , ARN Mensajero/biosíntesis , ARN Mensajero/efectos de los fármacos , Ratas , Ratas Zucker , Receptores de Cannabinoides , Rimonabant

RESUMEN

BACKGROUND: Although much progress has been made recently in characterising the proteins involved in duodenal iron trafficking, regulation of intestinal iron transport remains poorly understood. It is not known whether the level of mRNA expression of these recently described molecules is genetically regulated. This is of particular interest however as genetic factors are likely to determine differences in iron status among mouse strains and probably also contribute to the phenotypic variability seen with disruption of the haemochromatosis gene. AIMS: To investigate this issue, we examined concomitant variations in duodenal cytochrome b (Dcytb), divalent metal transporter 1 (DMT1), ferroportin 1 (FPN1), hephaestin, stimulator of Fe transport (SFT), HFE, and transferrin receptor 1 (TfR1) transcripts in response to different dietary iron contents in the four mouse strains C57BL/6, DBA/2, CBA, and 129/Sv. SUBJECTS: Six mice of each strain were fed normal levels of dietary iron, six were subjected to the same diet supplemented with 2% carbonyl iron, and six were fed an iron deficient diet. METHODS: Quantification of mRNAs isolated from the duodenum was performed using real time reverse transcription-polymerase chain reaction. RESULTS: There was a significant increase in mRNA expression of Dcytb, DMT1, FPN1, and TfR1 when mice were fed an iron deficient diet, and a significant decrease in mRNA expression of these molecules when mice were fed an iron supplemented diet. Strain to strain differences were observed not only in serum transferrin saturations, with C57BL/6 mice having the lowest values, but also in hepatic iron stores and in duodenal mRNA expression of Dcytb, DMT1, FPN1, hephaestin, HFE, and TfR1. CONCLUSIONS: The results favour some degree of genetic control of mRNA levels of these molecules.

Asunto(s)

Proteínas Portadoras/genética , Duodeno/metabolismo , Mucosa Intestinal/metabolismo , Hierro de la Dieta/administración & dosificación , ARN Mensajero/análisis , Enzimas Ubiquitina-Conjugadoras , Animales , Proteínas de Transporte de Catión/genética , Grupo Citocromo b/genética , Proteína de la Hemocromatosis , Antígenos de Histocompatibilidad Clase I/genética , Deficiencias de Hierro , Proteínas de Unión a Hierro/genética , Hígado/metabolismo , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos , Receptores de Transferrina/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Especificidad de la Especie , Transferrina/análisis

RESUMEN

Tachykinin NK2 receptors have been suggested to play an important role in the central nervous system. This study, using reverse transcription-polymerase chain reaction revealed a detectable expression of NK2 receptor mRNA in various human brain regions, including the caudate nucleus, the putamen, the hippocampus, the substantia nigra and the cerebral cortex. The distribution of NK2 receptor expression in the cortex revealed a major expression in frontal and temporal cortex compared to occipital and parietal areas. These results provide a molecular basis for considering a role of NK2 receptors in human pathophysiology.

Asunto(s)

Encéfalo/metabolismo , ARN Mensajero/metabolismo , Receptores de Neuroquinina-2/metabolismo , Anciano , Anciano de 80 o más Años , Humanos , Persona de Mediana Edad

RESUMEN

The aim of this article is to show how dysphonic voices can be characterized by means of a multivariate statistical analysis of flat vowel spectra. The spectral contour was obtained by means of a wavelet transform of the logarithmic magnitude spectrum, which was subsequently flattened to remove interspeaker variability related to the excitation and vocal tract filter functions. The results of the statistical analysis of flat spectra were the following. Firstly, principal components analysis produced markers that separated noisy from clean spectra. Secondly, the heuristic search for harmonic peaks or interharmonic dips could be omitted. Thirdly, conventional spectral markers of noise appeared as special instances of the markers that were derived statistically. Fourthly, the levels of visually assigned hoarseness and the first two principal components were significantly correlated. The assignment of different levels of (visual) hoarseness to different vowel timbres could be explained by the variability associated with the spectral contour.

Asunto(s)

Trastornos de la Voz/diagnóstico , Algoritmos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Análisis Multivariante , Fonética , Espectrografía del Sonido , Medición de la Producción del Habla/estadística & datos numéricos

RESUMEN

We examined the expression and presence of NK2 receptors in the septal area of rat brain, and investigated their functional role in the regulation of the septohippocampal cholinergic system. Using reverse transcription-polymerase chain reaction (RT-PCR) analysis, we showed the presence of NK2 receptor mRNA expression in the septal area, and detected septal NK2 binding sites by using a fluorescent-tagged neurokinin A (NKA) derivative. In vivo microdialysis was employed to explore the functional role of NK2 receptors in the release of hippocampal acetylcholine evoked by tactile stimulation in freely moving rats. Two sessions of stroking of the neck and back of the rat for 30 min, at 90 min intervals, produced a marked and reproducible increase in hippocampal acetylcholine release. This effect was dose-dependently prevented by intraperitoneal administration of the two selective non-peptide tachykinin NK2 receptor antagonists SR144190 (0.03-0.3 mg/kg, i.p.) and SR48968 (0.3 and 1 mg/kg, i.p.), but not by the inactive enantiomer of SR48968 (SR48965, 1 mg/kg) nor by the two non-peptide NK1 receptor antagonists SR140333 (3 mg/kg, i.p.) and GR205171 (1 mg/kg, i.p.). Furthermore, the intraseptal application of SR144190 (10(-8) M) reduced the sensory response. Finally, intraseptal perfusion of neurokinin A (0.01-10 microM) in anaesthetized rats produced a concentration-dependent increase in hippocampal acetylcholine release. The response to neurokinin A (0.1 microM) was prevented by SR144190 (0.03-0.3 mg/kg, i.p.) and SR48968 (0.3-1 mg/kg, i.p.). In conclusion, this study provides direct evidence for the role of endogenous NKA/substance P, through the activation of NK2 receptors, in regulating the septohippocampal cholinergic function.

Asunto(s)

Acetilcolina/metabolismo , Hipocampo/metabolismo , Receptores de Neuroquinina-2/fisiología , Sensación/fisiología , Tabique Pelúcido/metabolismo , Animales , Benzamidas/farmacología , Lóbulo Frontal/citología , Lóbulo Frontal/metabolismo , Masculino , Compuestos de Metilurea/farmacología , Microdiálisis , Morfolinas/farmacología , Neuroquinina A/metabolismo , Piperidinas/farmacología , Ratas , Ratas Sprague-Dawley , Receptores de Neuroquinina-2/antagonistas & inhibidores , Tabique Pelúcido/citología , Tacto/fisiología

RESUMEN

SR 31747A, defined as a sigma ligand, is a novel immunosuppressive agent that blocks proliferation of human and mouse lymphocytes. Using a radiolabeled chemical probe, we here purified a target of SR 31747A and called it SR 31747A-binding protein (SR-BP). Purified SR-BP retained its binding properties and migrated on SDS-polyacrylamide gel as a Mr 28,000 protein. Cloning of the cDNA encoding human SR-BP shows an open reading frame for a 223-amino acid protein, which is homologous to the recently cloned sigma 1 receptor. Interestingly, the deduced amino acid sequence was found to be related to fungal C8-C7 sterol isomerase, encoded by the ERG2 gene. The ERG2 gene product has been identified recently as the molecular target of SR 31747A that mediates antiproliferative effects of the drug in yeast. Northern blot analysis of SR-BP gene expression revealed a single transcript of 2 kilobases which was widely expressed among organs, with the highest abundance in liver and the lowest abundance in brain. Subcellular localization analysis in various cells, using a specific monoclonal antibody raised against SR-BP, demonstrated that this protein was associated with the nuclear envelope. When studying the binding of SR 31747A on membranes from yeast expressing SR-BP, we found a pharmacological profile of sigma 1 receptors; binding was displaced by (+)-pentazocine, haloperidol, and (+)-SKF 10,047, with (+)-SKF 10, 047 being a more potent competitor than (-)-SKF 10,047. Scatchard plot analysis revealed Kd values of 7.1 nM and 0.15 nM for (+)-pentazocine and SR 31747A, respectively, indicating an affinity of SR-BP 50-fold higher for SR 31747A than for pentazocine. Additionally, we showed that pentazocine, a competitive inhibitor of SR 31747A binding, also prevents the immunosuppressive effect of SR 31747A. Taken together, these findings strongly suggest that SR-BP represents the molecular target for SR 31747A in mammalian tissues, which could be critical for T cell proliferation.

Asunto(s)

Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Ciclohexanos/metabolismo , Proteínas de Unión al ADN/química , Receptores Opioides , Saccharomyces cerevisiae/metabolismo , Esteroide Isomerasas/química , Transactivadores/química , Secuencia de Aminoácidos , Animales , Antiinflamatorios no Esteroideos/metabolismo , Secuencia de Bases , Unión Competitiva , Proteínas Portadoras/aislamiento & purificación , Membrana Celular/metabolismo , Cromatografía de Afinidad , Cromatografía en Gel , Cromatografía por Intercambio Iónico , Clonación Molecular , Proteínas de Unión al ADN/metabolismo , Canal de Potasio ERG1 , Canales de Potasio Éter-A-Go-Go , Humanos , Cinética , Ratones , Datos de Secuencia Molecular , Receptores sigma/química , Receptores sigma/metabolismo , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Esteroide Isomerasas/metabolismo , Linfocitos T , Transactivadores/metabolismo , Regulador Transcripcional ERG , Receptor Sigma-1

RESUMEN

BACKGROUND: Townes-Brocks syndrome (TBS) is a rare autosomal dominant entity mainly characterized by ano-rectal, ear and extremities abnormalities with variable clinical expression. CASE REPORTS: The first case had ear and extremities, but not anorectal, abnormalities; a Pierre-Robin sequence with esophageal atresia was also observed. The second case had the classical triad of abnormalities also associated with tetralogy of Fallot which has been only once reported in the literature. CONCLUSIONS: Both cases are other examples of the frequent clinical variability observed in this syndrome explaining diagnostic difficulties in the absence of a specific marker.

Asunto(s)

Anomalías Múltiples/diagnóstico , Canal Anal/anomalías , Oído Externo/anomalías , Deformidades Congénitas de las Extremidades , Recto/anomalías , Anomalías Múltiples/genética , Atresia Esofágica/complicaciones , Atresia Esofágica/genética , Humanos , Recién Nacido , Masculino , Síndrome de Pierre Robin/complicaciones , Síndrome de Pierre Robin/genética , Síndrome , Tetralogía de Fallot/complicaciones , Tetralogía de Fallot/genética

RESUMEN

We report the molecular cloning of a beta 3-adrenergic receptor [beta 3-AR] cDNA from human brown adipose tissue. The cDNA-encoded protein is identical to the previously cloned beta 3-AR but with 6 additional amino acids at the C-terminus. The C-terminus is shared by the beta 3 receptors expressed in human neuroblastoma cells [SK-N-MC] [Mol. Pharmacol. 42 (1992) 964-970]. Furthermore, using a polymerase chain reaction strategy we have cloned and sequenced the beta 3-AR introns. Sequence analysis demonstrates that the human beta 3-AR gene comprises at least 3 exons and 2 introns and that the most abundant beta 3-AR transcripts encode a protein with an exon 3-derived C-terminus. Interestingly, although a similar organization has been found in rodent genes, the rat beta 3-AR transcripts encode a receptor with an exon 2-derived C-terminus.

Asunto(s)

Receptores Adrenérgicos beta/genética , Simpatomiméticos/metabolismo , Tejido Adiposo Pardo , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Exones/genética , Humanos , Intrones/genética , Datos de Secuencia Molecular , ARN Mensajero/genética , Receptores Adrenérgicos beta/biosíntesis , Proteínas Recombinantes/biosíntesis

RESUMEN

We report that the rat beta 3-adrenergic receptor (beta 3-AR) gene has an intron. The intron starts with an in-frame stop codon with the result that unspliced transcripts will encode a C-terminal truncated protein. The reported protein sequences of mouse and human beta 3-AR were both deduced from genomic DNA sequences. Given the heterogeneity at the C-termini of the otherwise highly similar rat, mouse and human sequences, we discuss the intriguing possibility that the beta 3-AR gene of the latter two species also contain an intron near the extremity of the open reading frame. A beta-adrenergic receptor (beta-AR) cDNA we have cloned from rat colonic tissue which has a sequence essentially identical to that previously reported for the rat adipose beta 3-AR cDNA [(1991) J. Chem. 266, 24053], encodes the spliced version of the beta 3-AR.

Asunto(s)

Intrones , Receptores Adrenérgicos beta/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Southern Blotting , Clonación Molecular , Codón , ADN/química , ADN/genética , Hibridación in Situ , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Ratas

RESUMEN

We have previously shown that basic fibroblast growth factor (bFGF) inhibits the FSH-induced differentiation of cultured rat granulosa cells, as manifested by prominent reduction of the LH receptor expression. We now investigate the possible sites and mechanism of action of bFGF. Whereas bFGF decreased the cAMP formation induced by FSH, it enhanced the cAMP production caused by cholera toxin and forskolin, suggesting that bFGF exerted its inhibitory action on cell differentiation at a step to cAMP production. Photoaffinity labeling with 8-azido-[32P]cAMP revealed that bFGF markedly reduced the FSH-induced increase in the level of regulatory subunit RII beta of the cAMP-dependent protein kinase (PKA) type II. In contrast to its striking effect on RII beta expression (70-80% inhibition), bFGF decreased PKA enzymatic activity by only 30%. On the other hand, transforming growth factor-beta (TGF beta) slightly amplified the stimulatory action of FSH and antagonized the bFGF inhibitory effect on both LH receptor expression and RII beta synthesis. We report that the protein kinase C (PKC) activator 12-O-tetradecanoylphorbol-13-acetate (TPA), which impaired granulosa cell differentiation, also abolished the RII beta synthesis induced by FSH. The activation of PKC by bFGF in granulosa cells was supported by the following findings: (i) bFGF markedly enhanced the production of diacylglycerol (2.3-fold stimulation at 5 min), the intracellular activator of PKC; (ii) bFGF promoted tight association of PKC to cellular membranes, a process that is believed to correlate with the enzyme activation; (iii) bFGF induced the phosphorylation of an endogenous M(r) 78,000/pI 4.7 protein that appears as a specific PKC substrate; (iv) bFGF mimicked the TPA-induced transmodulation of the epidermal growth factor (EGF) receptor, reducing by 36% the 125I-EGF binding on granulosa cells. We conclude that bFGF may exert its repressive action on RII beta synthesis, PKA activity, and granulosa cell differentiation by primarily targeting PKC activation.

Asunto(s)

Factor 2 de Crecimiento de Fibroblastos/farmacología , Células de la Granulosa/citología , Proteína Quinasa C/fisiología , Proteínas Quinasas/fisiología , Animales , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Cromatografía DEAE-Celulosa , AMP Cíclico/metabolismo , AMP Cíclico/fisiología , Diglicéridos/metabolismo , Activación Enzimática/efectos de los fármacos , Activación Enzimática/fisiología , Receptores ErbB/efectos de los fármacos , Femenino , Hormona Folículo Estimulante/farmacología , Células de la Granulosa/efectos de los fármacos , Células de la Granulosa/ultraestructura , Proteína Quinasa C/metabolismo , Ratas , Receptores de HL/análisis , Receptores de HL/efectos de los fármacos , Receptores de HL/fisiología , Acetato de Tetradecanoilforbol/farmacología

RESUMEN

AR4-2J, a rat pancreatic acinar-tumor cell line, was used to investigate long-term effects of basic fibroblast growth factor (bFGF) and somatostatin on pancreatic cancer cells. We observed that bFGF stimulated cell proliferation when cells were cultured in serum-free medium. The effect was dose-dependent with half-maximal and maximal effects at 25 pM and I nM bFGF, respectively. The somatostatin analog SMS 201-995 (SMS) decreased the growth-promoting effect of bFGF. The maximal effect was observed at I nM SMS and the half-maximal effect at 20 pM SMS. Characterization of bFGF receptor-binding properties with [125I]bFGF revealed that AR4-2J cells exhibited 2 classes of bFGF binding site with respective KD values of 47 pM and 3 nM and binding capacities of 14 fmol and 0.9 pmol/10(6) cells. High-affinity receptors correlated with bFGF stimulation of AR4-2J cell growth, suggesting that the effects of bFGF are receptor-mediated.

Asunto(s)

Factor 2 de Crecimiento de Fibroblastos/farmacología , Neoplasias Pancreáticas/patología , Somatostatina/farmacología , Animales , División Celular/efectos de los fármacos , Factor 2 de Crecimiento de Fibroblastos/antagonistas & inhibidores , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Ratas , Receptores de Superficie Celular/análisis , Receptores de Factores de Crecimiento de Fibroblastos , Células Tumorales Cultivadas

RESUMEN

Basic fibroblast growth factor (bFGF) is a potent mitogen for various cell types. We report here the first study of the effects of bFGF on a digestive tract-derived cell line. The effect of bFGF on the proliferation of AR4-2J cells, tumor cells of acinar pancreatic origin, was investigated together with modulation of ornithine decarboxylase (ODC) activity, an intracellular event involved in cell proliferation. bFGF caused a concentration-dependent stimulation of AR4-2J cell growth, with a half maximal effect (EC50) at 22 +/- 2 pM. ODC activity, assayed by the CO2-trapping method, was also increased by bFGF in a dose-dependent manner, reaching half-maximal stimulation at 20 pM. We conclude that bFGF is a very potent growth promoting factor for cells of pancreatic origin, already effective at picomolar concentrations. The parallelism between the growth assay and the ODC activity assay implicates the involvement of ODC activity in the pathway of the mitogenic effect of bFGF. The stimulation of ODC activity therefore seems to be a reliable early marker for cell proliferation in this model.

Asunto(s)

Factores de Crecimiento de Fibroblastos/fisiología , Neoplasias Pancreáticas/patología , Animales , División Celular , Línea Celular , Ornitina Descarboxilasa/metabolismo , Ratas , Células Tumorales Cultivadas/citología