RESUMEN
Barite chimneys are known to form in hydrothermal systems where barium-enriched fluids generated by leaching of the oceanic basement are discharged and react with seawater sulfate. They also form at cold seeps along continental margins, where marine (or pelagic) barite in the sediments is remobilized because of subseafloor microbial sulfate reduction. We test the possibility of using multiple sulfur isotopes (δ34S, Δ33S, ∆36S) of barite to identify microbial sulfate reduction in a hydrothermal system. In addition to multiple sulfur isotopes, we present oxygen (δ18O) and strontium (87Sr/86Sr) isotopes for one of numerous barite chimneys in a low-temperature (~20 °C) venting area of the Loki's Castle black smoker field at the ultraslow-spreading Arctic Mid-Ocean Ridge (AMOR). The chemistry of the venting fluids in the barite field identifies a contribution of at least 10% of high-temperature black smoker fluid, which is corroborated by 87Sr/86 Sr ratios in the barite chimney that are less radiogenic than in seawater. In contrast, oxygen and multiple sulfur isotopes indicate that the fluid from which the barite precipitated contained residual sulfate that was affected by microbial sulfate reduction. A sulfate reduction zone at this site is further supported by the multiple sulfur isotopic composition of framboidal pyrite in the flow channel of the barite chimney and in the hydrothermal sediments in the barite field, as well as by low SO4 and elevated H2S concentrations in the venting fluids compared with conservative mixing values. We suggest that the mixing of ascending H2- and CH4-rich high-temperature fluids with percolating seawater fuels microbial sulfate reduction, which is subsequently recorded by barite formed at the seafloor in areas where the flow rate is sufficient. Thus, low-temperature precipitates in hydrothermal systems are promising sites to explore the interactions between the geosphere and biosphere in order to evaluate the microbial impact on these systems.
Asunto(s)
Sulfato de Bario/análisis , Sulfato de Bario/química , Respiraderos Hidrotermales , Isótopos de Azufre/análisis , Bacterias Reductoras del Azufre/aislamiento & purificación , Regiones Árticas , Hierro/química , Fenómenos Microbiológicos , Sulfuros/químicaRESUMEN
Lipid lowering therapy by statins and antiaggregation have become the basis of any anti-atherosclerotic prophylaxis either as primary or secondary prophylaxis. As several recent papers indicated immunosuppressive properties of statins we investigated changes in lymphocyte subpopulations, apoptosis markers, and cellular immune response towards mitogens after a short-term therapy with atorvastatin and clopidogrel. Nine healthy volunteers (four male, five female, age ranging from 26 to 43 years) were treated with 20 mg atorvastatin for 4 weeks and for 2 additional weeks with 20 mg atorvastatin and 75 mg clopidogrel after oral consent was given. Lymphocyte subpopulations were counted by flow cytometry. To assess cellular in vitro immune function, lymphocyte transformation tests with four mitogens (PHA, ConA, PWM, and OKT3) were performed. Absolute leucocyte counts remained unchanged as well as the granulocyte, monocyte, lymphocyte, and lymphocyte subpopulation counts. There were no detectable changes in markers of cell activation (HLA-DR, CD25, CD69, and CD86) or apoptosis (CD95, annexin). Cellular in vitro responses towards four mitogens did not show significant changes after atorvastatin nor after atorvastatin plus clopidogrel treatment.In conclusion, our data show that atorvastatin is not an immunosuppressive drug under therapeutical conditions.
Asunto(s)
Ácidos Heptanoicos/efectos adversos , Inhibidores de Hidroximetilglutaril-CoA Reductasas/efectos adversos , Inmunidad Celular/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/efectos adversos , Pirroles/efectos adversos , Ticlopidina/efectos adversos , Adulto , Atorvastatina , Clopidogrel , Femenino , Humanos , Inmunosupresores/efectos adversos , Técnicas In Vitro , Recuento de Leucocitos , Activación de Linfocitos/efectos de los fármacos , Subgrupos Linfocitarios/efectos de los fármacos , Subgrupos Linfocitarios/inmunología , Masculino , Persona de Mediana Edad , Inhibidor 1 de Activador Plasminogénico/sangre , Ticlopidina/análogos & derivadosRESUMEN
UNLABELLED: It is generally accepted that growth factors play an important role in the pathogenesis of proliferative diabetic retinopathy. Since platelet-derived growth factor AB (PDGF AB) is known to be involved in many angiogenetic and proliferative processes, it was the aim of our study to elucidate the role of PDGF AB in the angiogenetic process in proliferative diabetic retinopathy. We measured PDGF AB concentrations in the vitreous of 23 patients with proliferative diabetic retinopathy, 4 of them with additional rubeosis iridis as an indicator of very high vasoproliferative activity. Control measurements were done in 19 patients without diabetic or ischemic eye diseases and also in 4 non-diabetic patients with ischemic proliferative retinopathy with rubeosis iridis. To exclude PDGF remnants in the vitreous due to vitreous bleeding we additionally measured platelet factor 4 concentrations as a stable marker of activated thrombocytes in the vitreous. RESULTS: Significantly elevated concentrations of PDGF AB were found in the vitreous of patients with proliferative diabetic retinopathy, with higher levels in individuals with additional rubeosis iridis compared to controls. However, concentrations of PDGF AB were also elevated in ischemic non-diabetic retinopathy, supporting the concept that ischemia might be a strong stimulator of growth factor production in the retina. Platelet factor 4 was not detectable in any of the vitreous samples included in the study. In summary, our results indicate that the growth factor PDGF plays an important role in the pathogenesis of proliferative diabetic retinopathy, probably in synergistic action with other growth factors like IGF I, IGF II, VEGF and TNF alpha.
Asunto(s)
Retinopatía Diabética/metabolismo , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Cuerpo Vítreo/metabolismo , Líquidos Corporales/metabolismo , Humanos , Factor Plaquetario 4/análisis , Factor de Crecimiento Derivado de Plaquetas/análisis , Valores de Referencia , Enfermedades de la Retina/metabolismoRESUMEN
The luteinizing, follicle-stimulating, and thyroid-stimulating hormone receptors belong to the huge family of G-protein-coupled receptors. Identification of either activating or inactivating mutations of these receptors has led to a fundamental improvement in our understanding of glycoprotein hormone/receptor interaction. Furthermore, clinical phenotypes such as precocious puberty, follicle-stimulating hormone (FSH) insensitivity syndrome, and congenital hypthyroidism are now being explained by mutated glycoprotein hormone receptors. Since there is an ongoing worldwide search for certain clinical phenotypes that might be caused by mutations of these receptors, there is a demand for strategies and techniques that can be used to screen patients in a effective and reliable way. This article focuses, therefore, on patient selection and techniques for the detection of mutations of glycoprotein hormone receptors, and compiles useful laboratory protocols to conduct such studies.
Asunto(s)
Análisis Mutacional de ADN/métodos , Receptores de HFE/química , Receptores de HL/química , Receptores de Tirotropina/química , Alelos , Animales , Células COS , Cromosomas Humanos Par 14 , AMP Cíclico/biosíntesis , Relación Dosis-Respuesta a Droga , Mutación de Línea Germinal , Humanos , Mutación , Polimorfismo de Longitud del Fragmento de Restricción , Polimorfismo Conformacional Retorcido-Simple , Estructura Terciaria de Proteína , Receptores de HFE/genética , Receptores de HL/genética , Receptores de Tirotropina/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Transducción de SeñalRESUMEN
The heterotrimeric Gs protein-adenylyl cyclase (AC) cascade plays a pivotal role in controlling hormone secretion by endocrine glands. Consequently, deficiency of the alpha-subunit of Gs leads to endocrine hypofunction and hypoplasia in the affected cells whereas AC hyperactivity results from activating point mutations within the Gs-alpha gene. The latter, termed gsp oncogenes, are found primarily in a subset of growth hormone (GH)-secreting human pituitary tumours (somatotrophinomas) and are thus associated with excessive GH secretion. We present here evidence that another type of defect in human somatotrophinomas may be overexpression of the Gs-alpha subunit. Immunohistochemistry using an antibody against recombinant human Gs-alpha revealed high levels of expression in 25 of 39 somatotrophinomas but weak staining in normal human pituitary cells. These results were confirmed by Western blot analysis. Additionally, cholera toxin-mediated ADP-ribosylation in the presence of 32P-labelled NAD+ resulted in an autoradiographic signal intensity which correlated directly with magnitude of immunostaining and amount of antigen shown by Western blot analysis, providing evidence for overexpression of functionally active subunit. Finally, reconstitution assays were applied and directly demonstrated the increased activity of overexpressed Gs-alpha. In vivo, the effect of Gs-alpha on AC activity may be partially counterregulated by high levels of inhibitory G protein that also occurred in these tumours. In culture, GH-releasing hormone (GHRH) had markedly reduced effects on GH secretion by somatotrophinomas exhibiting Gs-alpha overexpression, whereas powerful stimulation occurred in weakly staining tumours. In contrast to these observations with Gs-alpha, immunostaining for the phospholipase C-coupled G11-alpha subunit was relatively weak in all somatotrophinomas studied and synthetic GH-releasing peptide, which acts via a specific G11-coupled receptor, led to powerful and consistent stimulation of GH secretion by different tumours. These results indicate that Gs-alpha overexpression is associated with dysfunction in hormone secretion by some somatotrophinomas.
Asunto(s)
Adenoma/metabolismo , Subunidades alfa de la Proteína de Unión al GTP Gs/metabolismo , Hormona de Crecimiento Humana/metabolismo , Neoplasias Hipofisarias/metabolismo , Adenoma/genética , Adenosina Difosfato Ribosa/metabolismo , Adenilil Ciclasas/metabolismo , Adolescente , Adulto , Anciano , Resistencia a Medicamentos , Femenino , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Hormona Liberadora de Hormona del Crecimiento/farmacología , Humanos , Inmunohistoquímica , Técnicas In Vitro , Masculino , Persona de Mediana Edad , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Oncogenes , Neoplasias Hipofisarias/genéticaRESUMEN
INTRODUCTION: Some studies showed that in multinodular goiters clonal and polyclonal nodules coexist. The clonality of nodules in recurrent goiters is, however, still unknown and may contribute to help explain the pathogenesis of this thyroid disease. METHODS AND RESULTS: The clonality of 14 nodules derived from recurrent goiters was assessed by means of an X-chromosome-inactivation method. Of 14 nodules, 10 showed a polyclonal pattern, 3 were clonal and, in 1 case, the result remained unclear. The mean age of the patients with recurrent goiter at the time of their first operation was significantly lower than the mean age of 50 patients who underwent thyroid surgery for the first time over the same period of time (34.6+/-10.9 years vs 53.7+/-13.5 years; P<0.05). The mean interval between first and second operation was 18 years. CONCLUSION: The finding that nodules in recurrent goiters are predominantly polyclonal suggests that these lesions have their origin in a de novo proliferation of different cohorts of thyrocytes due to unknown growth stimulating molecular events.
Asunto(s)
Bocio Nodular/genética , Bocio Nodular/cirugía , Adolescente , Adulto , Células Clonales , Compensación de Dosificación (Genética) , Femenino , Humanos , Persona de Mediana Edad , Recurrencia , ReoperaciónRESUMEN
Previous structural analyses of diphosphoinositol polyphosphates in biological systems have relied largely on NMR analysis. For example, in Dictyostelium discoideum, diphosphoinositol pentakisphosphate was determined by NMR to be 4- and/or 6-PPInsP5, and the bisdiphosphoinositol tetrakisphosphate was found to be 4, 5-bisPPInsP4 and/or 5,6-bisPPInsP4 [Laussmann, Eujen, Weisshuhn, Thiel and Vogel (1996) Biochem. J. 315, 715-720]. We now describe three recent technical developments to aid the analysis of these compounds, not just in Dictyostelium, but also in a wider range of biological systems: (i) improved resolution and sensitivity of detection of PPInsP5 isomers by microbore metal-dye-detection HPLC; (ii) the use of the enantiomerically specific properties of a rat hepatic diphosphatase; (iii) chemical synthesis of enantiomerically pure reference standards of all six possible PPInsP5 isomers. Thus we now demonstrate that the major PPInsP5 isomer in Dictyostelium is 6-PPInsP5. Similar findings obtained using the same synthetic standards have been published [Laussmann, Reddy, Reddy, Falck and Vogel (1997) Biochem. J. 322, 31-33]. In addition, we show that 10-25% of the Dictyostelium PPInsP5 pool is comprised of 5-PPInsP5. The biological significance of this new observation was reinforced by our demonstration that 5-PPInsP5 is the predominant PPInsP5 isomer in four different mammalian cell lines (FTC human thyroid cancer cells, Swiss 3T3 fibroblasts, Jurkat T-cells and Chinese hamster ovary cells). The fact that the cellular spectrum of diphosphoinositol polyphosphates varies across phylogenetic boundaries underscores the value of our technological developments for future determinations of the structures of this class of compounds in other systems.
Asunto(s)
Dictyostelium/química , Fosfatos de Fosfatidilinositol/química , Células 3T3 , Animales , Cromatografía Líquida de Alta Presión , Cricetinae , Humanos , Isomerismo , Células Jurkat , Cinética , Hígado/enzimología , Mamíferos , Ratones , Resonancia Magnética Nuclear Biomolecular/métodos , Fosfatos de Fosfatidilinositol/metabolismo , Pirofosfatasas/metabolismo , RatasRESUMEN
Whereas in normal human thyroid tissue total cell mass is maintained by a balance between cell proliferation and apoptosis, the programmed cell death, in thyroid tumors this equilibrium is disrupted. In tumor cells, an augmented proliferation rate is no longer counterbalanced by an equally enhanced apoptosis resulting in an increased netto growth rate. To investigate regulation of apoptosis in thyroid tumors, we analyzed the expression of apoptosis-related proteins of the bcl-2 family in human thyroid tissues and in the human thyroid carcinoma cell lines FTC 133, HTC, HTC-TSHr and HTh74. In comparison to normal tissue, we detected an increased expression of the anti-apoptotic protein bcl-2 in adenomas, whereas follicular carcinomas showed various expression of bcl-2 with decreased levels in 32% of cases. BclxL expression was comparable in all tissues examined. The pro-apoptotic protein bax was expressed at lower levels in carcinomas than in adenomas, whereas bak and bclx were expressed in the same order of magnitude in all tissues examined. In contrast, thyroid carcinoma cell lines exhibited a relatively strong expression of bclxL, but a weak expression of bcl-2. In all four cell lines, the amounts of the pro-apoptotic proteins bax, bak and bclx were higher than in most tumor tissues. Our data show that in thyroid tumors expression of members of the bcl-2 protein family is not uniform. Rather, the expression pattern of pro- and anti-apoptotic proteins in thyroid tumors is heterogeneous. This may, at least in part, reflect the futile attempt of tumor cells to counterbalance the action of growth-promoting factors in thyroid tumor-igenesis.