BIOCHEMISTRY

Author: MOLINA JIMENEZ M. FRANCISCA.
Year: 2003.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE FARMACIA.
Place of preparation: FACULTAD DE FARMACIA.

Summary: In recent years, neurodegenerative diseases, including Parkinson's disease, have assumed great importance due to increased life expectancy in developed countries. The causes and mechanisms resulting in neuronal degeneration of the black substance is suffering from Parkinson's even unknown, although in this study highlights the involvement of oxidative stress and the mechanisms of apoptosis. The objectives of this study were to characterize an in vitro model to establish the relationship between Parkinson's disease, oxidative stress and neuronal death by apoptosis. Rotenone was used, complex I inhibitor of mitochondrial transport chain mail, to induce toxicity and found the ability neuroprotectora three antioxidant substances: fraxetrol, miricetina and N-acetylcysteine. All these alterations of cellular redox state leading to the neuronal death by apoptosis in dopaminergic cells from human SH-SY5Y that neuroblastoma was found by morphological changes characteristic, activation of caspase 3, proteolysis of poli-ADPribosa polymerase (PARP) and fragmentation internucleosomal DNA . The fraxetol fuez able to reverse this situation toxicity slowing oxidative stress through their ability recruiter of ERO, preventing apoptosis by inhibiting the proteolysis of PARP and DNA fragmentation, and increase cellular defenses by inducing gene expression of HSP70. In addition inhibit COX-2, a mediator in the inflammation process involved in neuronal death of GSH and inhibiting the activity of GPx also fragementación DNA, while the miricetina, despite its antioxidant effect, was not able to prevent death neuronal. It concludes that the in vitro model of Parkinson's disease developed in this study may be useful in evaluating potential therapies antioxidants tested in this disease.

Author: ADAME NAVARRETE M. YOLANDA.
Year: 2003.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of preparation: FACULTAD DE CC. QUҍMICAS.

Summary: In light intestinal stem substantial quantities of oxidants. The high reactivity of these molÃÂ © letters threatening the integrity of the mucosa, which may generate alterations at the level of the normal functions of the bowel. That is why this ÃÂ ³ rgano must have a powerful mechanism enzimÃÂ ¡tico of protecciÃÂ ³ n front of highly damaging agents. AsÃÂ, stated objectives for the study are intended to assess the impact exercises angente oxidanate fisiolÃÂ ³ gico, H2O2 on the funciÃÂ charges of secreciÃÂ charges and funciÃÂ ³ n barrier after coming into contact with the colon to travÃÂ © s of the luminal surface. AdemÃÂ ¡s, it is proposed to assess the participaciÃÂ charges of enzymes antioxidants endÃÂ ³ genas and especÃÂficas for H2O2, such as catalase, and glutathione peroxidase (GPx). The conclusions are derived from the study, conducted in Sprague-Dawley rat distal colon, are: 1, luminal-oxidants produced an increase in secreciÃÂ charges of Cl - probably in response to the activaciÃÂ charges nervous system entÃÂ © rich and disrupt the intestinal permeability. 2 - The distal colon of rats presents a mechanism enzimÃÂ ¡tico of protecciÃÂ charges against the H2O2 luminal where catalase desempeÃÂ ± a role crÃÂtico.

Author: BARDERAS MANCHADO RODRIGO.
Year: 2003.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD CIENCIAS QUҍMICAS.
Place of preparation: FAC. CC. QUҍMICAS DPTO. BIOQUҍMICA Y BIOLOGҍA MOLECULAR I..

Summary: The work of investigaciÃÂ charges that shapes the Doctoral Thesis submitted by Rodrigo Barderas, estÃÂ ¡No focused towards identificaciÃÂ charges and isolation of allergens relevant pÃÂ ³ lenes responsible for inducing allergies, your producciÃÂ ³ n recombinant and its possible utilizaciÃÂ charges in the diagnosis of this disorder and prophylaxis inmunolÃÂ ³ gico that increasingly has a higher incidence in paÃÂses developed. The investigaciÃÂ charges has focused on the study of four allergens from two sources polÃÂnicas little studied so far, the ash and white ash. L first because until ÃÂ eighth past aÃÂ ± os has not been a significant pollen to take into account that raise awareness among potential patients, and the second because ÃÂ eighth Only abounds in regions of Northern and Central Europe but estÃÂ ¡related structural and inmunolÃÂ ³ gicamente species plant important EspaÃÂ ± as to the olive tree. AsÃÂ, have been characterized, cloned and sequenced to know their complete primary structures, and there have been as proteÃÂnas recombinant allergens Che to 1, to 2 and Che Che-3 of ash and white Fra and 1 ash, determinÃÂ ¡ndose prevalence and implicaciÃÂ charges in cross-reactivity with other sources alergÃÂ © nicas. Also, as evidence essential to their utilizaciÃÂ charges have been validated by establishing equivalence structural and inmunolÃÂ ³ gica between allergens isolated from their natural sources and those expressed in the two systems producciÃÂ ³ n recombinant employees, Escherichia coli (Che Che-2 and to 3) and Pichia pastoris (Che-1 and Fra and 1). The expresiÃÂ ³ n recombinant allergens such permits have proteÃÂnas in large quantities and correctly folded that conducirÃÂ ¡na short term to a diagnosis and treatment mÃÂ ¡s effective disease alÃÂ © rgica in patients who are sensitive to these allergens.

Author: MATUS DE LA PARRA VALENZUELA ANA MARÍA.
Year: 2003.
University: VIGO [More theses of this university] [www.uvigo.es].
Place of defense: FACULTAD DE CIENCIAS.
Place of preparation: FACULTAD DE CIENCIAS.

Summary: France is the first producer of this kind in Europe, with an annual output of 135000 tons; that production depends almost in a natural part of the attraction of seeds, but the iregularidad in recruitment coupled with the high demand for youth the last few years has spurred the need for a more active role of the hatcheries supplying this industry. Between initiatives inthe study of reproductive success of this species is related to the quality of the gametes, this one the key points of seed production in criaderos.El present study seeks to learn more about the accumulation and utilization of energy reserves of various tissues during a reproductive cycle of Crassostrea gigas, examining these process on the basis of sex and the different stages marking its development gametogénico. Oysters from the bank ostrícola of Ronce in Bay Marenens-Olerón (La Tremblade, France) were collected every fortnight between March 1997 and February 1998.Se took sample of each of the principal organs that make up the soft parts of the animal ( gonad, múlculo adductor, papol labiales, mantle and digestive gland). seasonal variation and stages of maturity of the biochemical composition and classes lipídas was conducted in three of these organs: digestive gland, palpos labiales gonad and, in this latest addition tissue histological analysis was performed and identification of the various classes lipid and fatty acid composition of the fosfatidilcolina.Además there were two types of índicees status: one general and another of the various somatic tissues to determine the physiological status of individuals in the study. The growth of Crassostrea gigas is closely related to the availability of nutrients and the temperatura.En spring, temperatures and outcrops summer detemiann the development of gónada.En autumn, the drop in temperature and the high availability of microalgae is ralacionan with somatic growth in stature and peso.Las accumulated reserves in somatic tissues are used in winter, when the availability of food drops its minimum. The cycle gametogénico of Crasostrea gigas begins at the end of summer, with a period of restoration tissue reserves and a total absence of germ cells (stage 0) then begin training, proliferation and spread of gonias and training of gonadales follicles, which are observed indiferenciadas cells, interspersed with ovogonias and spermatogonial (stage I). In the vast majority of these individuals, sexually undifferentiated, developing oocytes previtelogénicos is faster and evidente.Durante phase or stage of winter hibernation the oyster, the cells differentiated reach suffer from a constant atresia.En spring, when the water temperature rises, the gametogenesis reactivates and differentiation occurs males, with a quick extentión and ramification of follicles ( stage II). While females also experience a rapid increase and development of oocytes previtelogénicos, they continue to show a high degree of atresia.En April, the males reaching a stadium IIIA1 which is characterized by an increase considerabgle volume follicular to occupy case the entire espacio.De April to June. Moment in males reach a stadium IIIA2 presents small and sucesicas emissions gametes (pulse-up), while the females remain a evedente atresia trained in gametes, which only slow start after the pulses of the males. Beginning in this time the hembreas reach a stadium IIIA1, which is evident oocytes vitelogénicos anchored in the pard of folículo.Posteriormente, females mature quickly, reaching Stadium IIIA2, like No males, are able to deliver some mature oocytes. As of this moment, the development of both sexes continues synchronously, ocn period of rapid restoration gon 8 adal (s 130th tadio IIIC), which begins after the spawning period of either sex during the month of July (IIIB). In mid-August, and after the placing, individuals suffer hemocitaria of residual germ cells, as well as genetic follicles (stadium IIID) and begins anew to form the fabric of giving reservation starts a new cycle gametogénico (stage 0 ). intersexual The nature of the gonad of Crassostrea gigas is corroborated by the appearance of a small proproción "hemafroditas functional" at the time of the resumption of gametogenesis, after the period of hibernation. Time which considerameo svr sexual differentiation. The index of general condition proposed C. Gigas is directly related to somatic growth and inversely with the development of reverse evolution gonadal.La condition index dela digestive gland respect alos rates condition the mantle, and palpos labiales adductor muscle, in times of outcrop chlorophyll ay sestón total indicate the intermediary role of the digestive gland in the transfer of food ingested and their accumulation in reserve inthe other somatic tissues. The main reserve energy Crassostrea gigas representing the glycogen, which is experiencing a marked seasonal variation in all tissues studied: increases in the digestive gland during outcrops fitoplanctónicos late verano-principios autumn and is rapidly transferred yñ accumulated in palpos and labial gonad (stage IA) and consumed completely case in early spring, with the revival of gametogenesis, coinciding with a significant increase in lipid and stage II of males and IIIA1 females. This suggests the use of reserves glucídicas in the synthesis of lipids at the moment, and a high demand for energy in the stages of germ cell division and vitellogenesis active in males and females, respectively. The digestive gland, in addition to an intermediary role in the transfer to other organs of lipids, ingested, appears to be an important organ of reserve component of this, in winter to mobilize palpos and gonad during somatic growth and the beginning of the cycle gonadal during the spring and development gametogénico. The slower consumption reservations glucídicas of palpos labiales, along with the increase of its total lipid content in early spring and its subsequent reduction, parallel to its increase in gonad during the last stage of maturation gonadal us to consider palpos labiales as body reserves to underpin the more advanced stages of gonadal maturation, mainly those of vitellogenesis and restoration, and suggests the importance of lipids of this body in key moments in the cycle of gonadal Crassostrea. Esters of colesteros of gonad and palpos labiales could entervenir in regulating the cycle of Crassotrea how gonadal hormone precursors possible, given its timely and significant increases, coinciding ocn the beginning of the cycle gametogénico (stage 0 and AI), with the revival of Spring gametogenesis and sexual differentiation (stage II) and the decline in atresia and the beginning of vitellogenesis in embras (stadium IIIA1). The high proportion of the phospholipid components of the gonad of fosfatidilinositol, fosfaditilsrina and fosfatidiletanolamina at the stadium IIID, fosfatidilinositol in stage II of males and fosfatidiletanolamina, fosfatidilcolinay plasmalógnos in eltadio IIIA1 females, could also suponeer molecular signals for the completion of a cycle gametogénico and the beginning of another, for the revival of gametogenesis and sex determination and for the disappearance of the artresia, respectively. The commposición fatty acid fraction of phosphatidylcholine, the phospholipid most abundant in the gonad of Crassostres gigas, reveals an increase in saturated fatty acids in times of abundance of nutrients, a posibler restoration of lipid membrane with the drop in temperature in winter, marked by a decline in the saturated fatty acids and the increase in monounsaturated, and an increase in polyunsaturated fatty acids durant ela maturation of gametes, parallel decline in monounsaturated fatty acids.

Author: CAMPANERO RHODES M. ASUNCIÓN.
Year: 2003.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE FARMACIA.
Place of preparation: INSTITUTO DE QUÍMICA FISICA "ROCASOLANO".

Summary: The epsermadhesinas are a family of proteins present in the seminal plasma of various mammalian species that cover the surface of sperm in ejaculation. Proteins are multivalent, capable of uniting various types of ligands not yet know their role, are believed to be implicads in more than one stage of the process of fertilization. PSP-Iy PSP-II are espermadhesinas pig most abundant in the seminal plasma, are forming a heterodímero not convalente whose structural features have been studied extensively. However, very little is known about the subunits PSP-Iy PSP-II isolated, and there are also dispute as to their capabilities binding ligands. In this thesis Doctroal has conducted a study of the structural organization, stability and state of aggregation and PSP-Iy PSP-II and have limited their capabilities binding ligands and the interdependence of the different interactions. Likewise, it has been explored in both protein a possible binding site to protease inhibitors and the area of binding to heparin in PSP-II and CDP-109, the majority of seminal plasma protein bovine compatible with Inter-relación observed between the various capacities ligands binding to these proteins. Finally, it has studied the effect of Zn2 +, a cation very abundant in seminal plasma on the structural organization and recognition of ligands of heterodímero and two subunits, revealing a regulatory role of the cation on the state of aggregation and binding capacity to heparin PSP-I. In addition, although there have not been any ligands for heterodímero PSP-I/PSP-II has shown that under certain physiological conditions (low protein concentration and pH or acid presence of Zn2 +) its dissociation occurs, enabling the functionality capabilities union of the individual subunits.

Author: RUIZ MACÍAS SERGIO.
Year: 2003.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE CIENCIAS BIOLÓGICAS.
Place of preparation: FACULTAD DE CIENCIAS BIOLÓGICAS.

Summary: In the work done in this Doctoral Thesis has tried to determine the role that family members of retinoblastoma are in the process of proliferation, differentiation and morphogenesis of carcinogenesis in mouse skin. To do so, it has done a thorough analysis of the skin of mice deficient in several lines of conventional kind or conditional, in one or several of the different family members of retinoblastoma. Animals deficient in p107 or p130 lacking epidermal phenotype. However, animals p107- /-; p130- /-have alterations in the terminal differentiation as well as follicular development. Changes in the hair follicle appear to be due to a decrease in the expression of morfógeno BMP4. Furthermore, these changes are reversible by transplanting skin deficient in p107 and p130 to a normal dermis animals NOD / SCID, demonstrating the importance of the mesenchymal component in the development of phenotypic animals p107- /-; p130- / -. Moreover, it was shown that the loss joint p107 and p130 led to the accumulation of beta-catenin nuclear basal keratinocytes in the epidermis. This was due to the overexpression of Frat one of the blocking protein kinase GSK3beta, which today is responsible for the degradation of beta-catenin. This phenotype also was restored using transplants above. The loss of pRb in the epidermis through the system Cre / LoxP produces severe alterations in the epidermal proliferation and differentiation. These alterations are compounded so dosis-dependietne by successive loss of alleles of p107 demonstrating the compensatory role of this protein in the absence of pRb. Animals deficient in pRb and p107 also have a dramatic phenotype in the hair follicle which involves the loss of hair. Furthermore, it has been shown that pRb has a dual role in the differentiation and, on the one hand, regulates the output cycle prior to the process of differentiation while on the other, engaged in the maintenance of the rule postmitótico of keratinocyte differential. By útlimo, analyzed the implication of pRb in the process of carcinogenesis chemical DMBA / TPA mouse skin. It was noted that in the absence of pRb, appeared fewer and smaller tumors than in the control animals. Despite this, tumors deficient in pRb were more malignant and had higher apoptotic rate, depending on the stabilization of p53 as compared with control tumors.

Author: MASIP ORDÓÑEZ MANUEL.
Year: 2003.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE CIENCIAS BIOLÓGICAS.
Place of preparation: FAC. DE CC. QUÍMICAS.

Summary: The alfa-sarcina is ribotoxina best feature cytotoxic proteins secreted by filamentous fungi of the genus Aspergillus. The alfa-sarcina which is produced by the fungus Aspergillus giganteus MDH 18894, was found during a research program conducted by the Department of Health of Michigan, in search of antibiotics and anti-tumor agents that could kill the cancer. This protein was very effective in inhibiting the growth of various tumor animals as sarcoma 180 and carcinoma 755. Just his name (alfa-sarcina) is due to his activity anti-sarcoma (Olson and Goerner, 1965; Olson et al., 1965). In this Doctoral Thesis has continued with the characterization of alfa-sarcina. Essentially, have been designed, prepared and purified a series of mutants in which various locations of the ribotoxina have been altered or delecionadas. Specifically, the mutant proteins that have been studied in this Doctoral Thesis, are the result of changes in the Arg121 (mutants R121K and R131Q), Leu145 (mutant L145F), the Asn54 (mutants N54D and N54Q), the region 122-129 (mutant alfaS-M), part of the N-terminal region mutant (7-18), l asustitución loop 2 of the alfa-sarcina the equivalent of RNasaU2 (mutants alfaS loopU2, alfaS (53-93 ) and alfaS loopU2/C132S / 'C76S'). This work has helped deepen the relations estructura-función this cytotoxin. Have been evaluated in detail how the mutations affect raised the folding and stability of the protein. It has studied the catalytic mechanism of alfa-sarcina through the characterization of the enzymatic activity of these mutants on different substrates, analyzing how they affected the ability ribonucleolítica dela enzyme in each case. In addition, it has valued the importance of anomalous pKa of His137, as it has studied the influence of pH on the catalytic mechanisms of alfa-sarcina. Finally, it has also been evaluated in detail the influence of these changes in the interaction of alfa-sarcina on vesicles fosfolipídicas, establecièndose the importance of Arg-121 in such interaction.

Author: ZULUAGA RODRÍGUEZ SUSANA.
Year: 2003.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE FARMACIA.
Place of preparation: FACULTAD DE FARMACIA.

Summary: The p38alfa MAPK is a serine / threonine kinase of the superfamily of MAPKs. The p38 MAPKs classically have been associated with processes of response to stress. However, more recent data indicate that can be activated in response to many other signals regulating various cellular functions including apoptosis and cell survival. Since there are four isoforms of p38 MAPKs, different isoforms may play different roles. Our work has focused on the study of the role of alpha isoform in the balance apoptosis / survival, using cells deficient in this kinase. The results presented in this report show that p38alfa sensitizes the cardiomioctos and mouse embryonic fibroblasts versus apoptosis activated by various stimuli through different mechanisms. On the one hand, p38alfa induces an increase in the levels of proteins pro-apoptóticas as Bax and Fas, probably through the reduction of the active form of the transcription factor STAT3 (fosfo-Ser727-Stat3). On the other hand, p38alfa MAPK negatively regulates the path of survival of ERK MAPKs and the PI3K/Akt. Specifically, the p38alfa MAPK mediaría the desfosforilación and inactivation of Akt through the activation of the serine / threonine phosphatase PP21A. This stimulation of activity in the PP2A occurs in regions caveolares membrane, where the catalytic subunit of PP2A (PP2Ac) remains united to caveolina-1 and Rac-1 forming a complex. The formation of this complex is dependent cell adhesion, so it might depend on the activation of integrins and / or cadherinas. In turn, the interaction between caveolina-1, PP2Ac and Rac-1 depends on the activation of tyrosine kinases.

Author: SÁNCHEZ BARRENA M. JOSÉ.
Year: 2004.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE CIENCIAS QUÍMICAS.
Place of preparation: INSTITUTO DE QUÍMICA-FÍSICA ROCASOLANO (CSIC).

Summary: Too much salt in the soil inhibits plant growth and cause huge losses in agricultural production worldwide. The route Salt Overly Sensitivie (SOS) plays a crucial role in salt tolerance in plants and the maintenance of low concentrations of sodium ion (Na +) in the cytosol, hence the establishment of the structural foundations of this process is of great importance to understand the molecular basis of the system and to be able in the future to give it a biotechnological application. The sensor Ca2 + SIS3 is a key protein in the path SOS, as it is responsible for initiating the response saline tolerance. SOS3 is able to feel the cytosolic signal of Ca2 +-induced stress saline, and then activate the kinase SOS2. In order to understand how SIS3 joins the Ca2 + and regulates the activity of SIS2, and ultimately understand how it works specifically upon the signal from the stress saline rather than other types of stress which may also induce changes in the concentration cytosolic Ca2 + , has dealt with the structural study of this protein. It has been determined the three-dimensional structure by X-ray diffraction of the complex SOS3 Ca2 + and SOS3 N2 + Ca2 + has been made esperimentos of analytical ultracentrifugation to characterize the state association of the protein in terms of various cations physiologically relevant, there have been experiments heat denaturation recorded dicrósmo circular to characterize the union of these metals have been carried out experiments hydrophobic interaction chromatography to study the hydrophobic nature of SOS3 and finally has compardo structurally SOS3 with their protein homologous to reveal structural determinants associated with their property. The integration of all results have led to propose a mechanism for response to Ca2 + and interaction with SIS2 whereby the binding of Ca2 + to SOS3 causes a change conformacional inducing dimerización of protein and an increase in the hydrophobic nature of the same. This allows interaction SOS3 with hydrophobic reason FISL of SOS2 and activating the role of the kinase.

Author: ZEINI MORENO MIRIAM.
Year: 2004.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE FARMACIA.
Place of preparation: INSTITUTO DE BIOQUÍMICA. CENTRO MIXTO CSIC-UCM. FACULTAD DE FARMACIA.

Summary: After surgical resection of 70% of the liver mass by hepatectomía partial (PH), liver triggers a cascade of signals, which initiated the recovery of a liver mass appropriate to the body's metabolic demands. Among the signs involved in the regeneration process are the expression of inducible proteins, NOS - 2 and COX-2. Nitric oxide (NO) and prostaglandins (PGs) released by these enzymes are enzymes that can deal with the generation process. In this paper, we analyzed the role of NO and PGs and their interrelationships in regulating the regenerative process. To that end, we explored the role of the release of PGs in an animal model without NOS - 2 (NOS - 2 KO): * deficiency simultaneous NO and PGs during the process of regenerating liver after PH carries a massive apoptosis hepatocyte proliferative and disability, which determine a high mortality rate for animals after the operation. This process is dependent on the activation of caspases, as the treatment inhibitor z-VAD protected animals from liver damage after PH. Although NO and PGs are not essential for recovery from liver tissue, modulate the regenerative process and cooperates with NO metabolites derived from COX-2 to complete the recovery process after PH. Since NO has been shown to be hepatoprotector and protects hepatocytes of apoptosis by different harm and injury mediated by cytokines after PH, we explored the effect of the release of NO in a pre-PH using a transgenic animal model that expressing NOS - 2 in liver and a model expression of NOS - 2 in liver injection hydrodynamics: * The release of NO in the liver prior to the PH cause a delay in the process of regeneration by the weak activation of NF-kB and Stat-3 and a lower release of TNF-alpha and IL-6. In addition, there was a delay in DNA synthesis and proliferation of hepatocytes. The regenerated liver mass is lower in animals that express NOS - 2 in the liver compared with their controls. * However, the release of NO in liver hepatocytes protects against the damage apoptótico mediated by an antibody anti-Fas due to inhibition of the activation of caspases. Both deficiency after the release of NO delaying the process of regeneration after PH. NO metabolites derived cooperates with COX-2 for the recovery of a liver mass adequate and has an impact hepatoprotector compared to the damage apoptótico mediated by Fas in the liver.

Author: BARRAL CATOIRA PATRICIA.
Year: 2004.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE QUÍMICAS.
Place of preparation: FACULTAD DE QUÍMICAS / DPTO. DE BIOQUÍMICA Y BIOLOGÍA MOLECULAR I.

Summary: The olive pollen is a major cause of alergía in countries around the Mediterranean, including Spain, Italy, Greece and Israel. The identification, isolation and characterization of allergen is one of the main objectives in the area of allergy research to be highly beneficial both for dignosis as therapy. In this Doctoral Thesis has been carried out isolation, immunological and molecular characterization, and expression of recombinant a new pollen allergen of Olive, Ole and 10. From a biochemist, Ole and 10 represents the first member of a new family of proteins from plants, which interacts with 1,3-beta-glucanos and defines a new set of modules union carbohydrate. From a clinical point of view, Ole and 10 is a major pollen allergen, which showed cross-reactivity with both protein pollen olive and other biological sources. Lastly had carried out the design and optimization of a protocol for the recombinant expression of Ole and 10 using two systems expression ecuariota: yeast P.pastori and insect cell cultures infected with baculovirus.

Author: LAINEZ MAS BEGOÑA.
Year: 2004.
University: BARCELONA [More theses of this university] [www.ub.es].
Place of defense: FACULTAD DE BIOLOGÍA.

Summary: The tumor necrosis factor, TNF is a cytokine pleiotrópica involved in a wide spectrum of inflammatory responses and the immune system. The biological effect of TNF receptors are mediated by two membrane receptor 1 TNF, TNFR1 And the receiver 2 TNF, TNFR2. These two receptors may present as soluble receptors formed from prosecution proteolítico extracellular region of the receptor membrane. It has been described high concentrations of soluble receptor of TNFR2 in the serum of multiple inflammatory diseases. In this paper, we describe for the first time an ísoforma of RNAm of TNFR2 human, produced by alternative splicing, which lacks the exons 7 and 8 encoding most of the cytoplasmic region and part of domininio intracitoplasmático of protein. The alternative splicing of the two exons change in the pattern of reading, which led to the emergence of a new stop codon after 6 new amino acids. It got this way, a cONA that codified protein TNFR2 formed by the same extracellular region and the same 5 first amino acid transmembrane domain, the TNFR2 native, followed by 6 new amino acids as the sole end carboxyl terminal. The study of the expression of the protein in cells tranfectadas with cONA this isoform, showed the nature of this soluble protein. The protein had a molecular weight of approximately 42 KOa. In an essay cytotoxicity induced by TNF, showed that this isoform could block apoptosis induced by TNF, suggesting its role as regulator of the action of TNF antagonist as its biological function. There was a ELlSA that quantifies the soluble receptor generated by alternative splicing. And concentrations were determined soluble receptor produced by splicing altemativo in the serum of healthy subjects and in patients of different inflammatory diseases. Our data show that the soluble receptor produced by alternative splicing is present in the serum of healthy individuals at low concentrations and high concentrations in subjects with sepsis and rheumatoid arthritis. It was determined that there was a negative association between soluble receptor produced by alternative splicing and various components of metabolic syndrome and also with insulin resistance. Moreover, it was observed that patients with arthritis reumatoíde with very high values of isoform, were probably a better response to therapy with antibodies anti-TNF-a.

Author: GASOL ESCUER EMMA.
Year: 2004.
University: BARCELONA [More theses of this university] [www.ub.es].
Place of defense: FACULTAD DE BIOLOGÍA.
Place of preparation: FACULTAD DE BIOLOGÍA DE LA UNIV.DE BARCELONA.

Summary: The family of amino acid transporters heteroméricos to pose the unique feature of being composed of two subunits: a heavy subunit and one subunit slight united by a disulfide bridge. Mutations in any of its members are responsible for aminoacidurias hereditary, as lisinuria intolerant protein (LPI) or cistinuria. At the beginning of this thesis, we sought to identify any new light subunit of this human family. By sequence homology search was donated the cDNA corresponding to a novel protein highly homologous to a mouse protein associated with the transport system xc-. The functional characterization confirmed that xCT human, in combination with 4F2hc, induces an activity of glutamate transport and cystine, sodio-independencia (system xc-). The profiles catchment substrates as a function of pH of the means of transport indicate that both the L-glutamato as L-cistina are transported on an anion, each with a net negative charge. By experiments Out substrate was determined the nature of this conveyor exchanger with a estequiometría of 1:1. Pattern of expression of xCT human by Northern blot shows a slight band only present in the brain. RT-PCR experiments were also positive for the pancreatic islets and cell lines for lines and epithelial tumor, while kidney and liver showed no sign. The physiological role of xCT appears to be implicated in the uptake of cystine for the synthesis of glutathione and its maintenance, giving an important role in oxidative stress situations. He then addressed the determination of the topology of xCT as model subunit lightly. Experiments inmunodetección demonstrate the intracellular localization of the two ends of the protein. Using the strategy of introducing cisteínas individual in the various loops and their accessibility to test reagents tiol-específicos, topology xCT is compatible with a model of 12 transmembrane domains. The results obtained in the area IL2-3 with two foreign waste accessibility (110Y 112) flanked by internal waste accessibility (102, 109 Y116) in the range of 15 amino acids make us think of a structure reentrant loop, with the residue HIlO as apex. The fact that these structures tend to be areas associated with the passage route of the substrate transporter, led us to examine the implication of the residue H 110. In summary, the results show that i) the biotinilación residue HII0C is blocked by the substrate and inhibitor not transportable 4SCPG ii) inactivation caused by MTSES in transporting hllOc is also protected by substrates with a ICso similar to the Km for each substrate iii) that protection was independent of temperature, and therefore does not involve major changes conforrnacionales; iv) although mutants Hll0C and Hll0D not alter the Km transporter or its substrate specificity, the replacement by a lysine inactive totally function xCT. Therefore, we have evidence that HIlO is close to the substrate binding site or passage route of the same. These results can be found in the following publications: J Biol Chem (2004) vol. 279,31228-36, J Biol Chem (2004) vol. 279, 11214-21, and Pflugers Arch. (2001) Vol. 442 (2) 286-296.

Author: FIGUERAS POLO M. TERESA.
Year: 2004.
University: BARCELONA [More theses of this university] [www.ub.es].
Place of defense: FACULTAD DE BIOLOGÍA.
Place of preparation: FACULTAD DE BIOLOGÍA.

Summary: During certain situations catabólicas such as cancer or sepsis, including happens is a common fact that the loss of muscle mass. Our research group has studied in depth the muscular associated with the wasting syndrome. The system has been identified proteolítico dependent ATP ubiquitina And as a mechanism involved in the activation of the muscle proteolysis. In addition, we have identified certain cytokines, particularly TNF-cx as elements contributing to development of the syndrome of cancer cachexia. The objective of this thesis was to deepen the knowledge of the mechanisms responsible for the loss of muscle mass in various pathological states. In particular, the role of TNF-cxen the cancer cachexia, sepsis i COPD (chronic obstructive pulmonary disease) i the possible therapeutic potential of cytokine anabólica IL-15. The approximations experimental atizadas has been: for the study of cancer cachexia use the model of tumor hematoma ascítico Yoshida AH-130, rat i muscle biopsies from patients with pancreatic cancer. Moreover, utilitzamos muscle biopsies from patients COPD. In addition we used a rat model of sepsis. The findings of this study are as follows: 1. In the wasting syndrome there is a loss of muscle protein as well as a valued asset apoptótico process as DNA fragmentation. 2. The TNF-cxestá involved in the induction of apoptosis in skeletal muscle since treatment with rolipram reverts partially DNA fragmentation observed in septic rats. 3. The increase in the expression of muscle receptors TNF-cxasí as the rise in the Bax/Bcl-2 (detected in experimental cancer) would be mechanisms involved in the induction of aopotosis and showing greater sensitivity of skeletal muscle to shares catabólicas of TNF. 4. In the experimental model of cancer cachexia noted an increase in the carbonization of muscle protein, as well as an increase in the content of iNOS and the UCPs. These facts show that skeletal muscle is subjected to oxidative stress during cachexia. 5. COPD patients show some muscle levels of reduced glutathione (GSH) lower than healthy individuals, which is most pronounced with decreasing body mass index. These patients can be seen an increase in muscle expression of yGCS that could act as a compensatory mechanism for the low levels of GSH. Oxidative stress muscle detected in COPD patients could be responsible for the increase in gene expression TNF-cxobservada in skeletal muscle. 6. In healthy individuals, the training program improved their redox state while in COPD patients is not improved else that oxidative stress muscle is much sharper. 7. The IL-15 acts as a cytokine anabólica clearly at the level of skeletal muscle exercise the following: reverses the protein degradation and blocks apoptosis. 8. Mechanisms associated with the therapeutic action of IL-15 could explain at normalizing the expression of the receptors TNF-cxy to inhibition of the activation of the system proteolítico dependent ATP And ubquitina in skeletal muscle at the same time the iNOS protein that decreases and increases the expression of UCP2 and UCP3 reversing oxidative stress in this tissue.

Author: GARCIA ALVAREZ GISELA.
Year: 2004.
University: BARCELONA [More theses of this university] [www.ub.es].
Place of defense: FACULTAD DE FARMÁCIA.
Place of preparation: BIOQUIMICA I BIOLOGÍA MOLECULAR -FARMACIA UB..

Summary: The decision to start a cell of the cell cycle, apoptosis, or go to survive is the result of the integration of different signals extrace1u1ares. Growth factors, contacts between cells, as well as various inducing apoptosis regulating a complex system of signal transduction pathways that induce activation of a large number of genes involved in the cellular response to the above processes anterionnente. One of the key proteins in the regulation of cell cycle and / or the entry into apoptosis is the transcription factor E2Fl. It has been suggested that activity levels E2F 1 might act as sensors entry or cell cycle arrest and apoptosis. These decisions not only the transcriptional activity is important but also the timing of E2F 1 with specific transduction pathways. This is the context in which it has been suggested that the activation of the path of fosfatidi1inosito1 3-quinasa (IP 3-quinasa) inhibits the effect apoptótico caused by the overexpression of E2Fl. Because the glycogen synthase quinasa-3-betá (GSK3B) is one of the most important physiological substrates of this route, experiments were conducted with the aim of analyzing the existence of an interaction between these two proteins. The results obtained in this thesis show that GSK3B fosfori1a factor detranscripción E2F1 human in vitro positions serine 403 and threonine 433. These wastes have already been described anterionnente as substrates for phosphorylation by the complex kinase TFIIH, specifically by one of its members: cdk7. Despite the fact that we can not detect fosfori1ación in vivo experiments immunoprecipitación confinnan the existence of a union between in vivo protein GSK3B and E2Fl. Using transient transfections, 'RNA interference (RNAi)', and the use of specific inhibitors of PI 3-quinasa and GSK3B, demonstrated that GSK3B regulates activity E2Fl through interaction with their dominiotransactivador and that the kinase activity of GSK3B not this is required for Therefore, our results would be integrated into a model in which the trans10cación of GSK3 the nucleus modularía activity E2FI and, therefore, the decision of a cell to enter the cell cycle or apoptosis directed towards. Historically, E2FI has been associated exclusively with his role as a transcription factor essential for the transition Sc / S. Results of recent years have changed that vision, involving E2F 1 on other cellular processes such as cell cycle arrest and apoptosis. It has been suggested that levels of this protein could be the sensor to determine the biological process to take place. It has also seen the need for other effectors signal transduction to define cellular processes. The activation of the path of IP 3-quinasa has proved to be decisive in decisions cell in which E2FI involved. The activation of this channel apoptótico inhibits the effect of overexpression of E2Fl. GSK3 ~ is one of the substrates fisiológicosde this path and has been involved in diferentesprocesos apoptotic. Whereas 10 above, our working assumption is that GSK3B is the signal, or the signal, which regulates the activity of E2FI and, consequently, cell division. Therefore, the objectives of this thesis are: -1. Analysis of phosphorylation in vitro and in vivo E2Fl by GSK3 (3. -2. Study of the interaction in vitro and in vivo between GSK3 (3 and E2Fl. -3. Study of the regulation of transcription factor E2Fl by part of GSK3 (3.

Author: ALONSO CEREZO M. CONCEPCIÓN.
Year: 2004.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE MEDICINA.
Place of preparation: UNIVERSIDAD COMPLUTENSE DE MADRID.

Summary: SUMMARY: Introduction: In recent years have documented the existence of variability in clinical practice. The variability is a phenomenon that can deteriorate the quality of care as a cost when due to the improper use of processes or resources. Objective: This raises the general objective of the study to know the causes that determine variability in the use of biochemical tests in patients dislipémicos in Primary Care. Materials and methods: comparing those guides international character, which are the bibliographic reference of the guides Spanish, and GPC. This is a cross-sectional study descriptive of the views of MAPy of coordinators in the use of laboratory tests in dislipemias Area 2 Madrid. There will always be a longitudinal study of type retrospective cohort on the use of evidence in lipid AP Area 2 Madrid by analyzing patient records. Results: The recommendations in the evaluation of cardiovascular risk performed by practice guidelines cynical analyzed are not homogeneous. The views of primary care physicians on the use of laboratory tests in screening, diagnosis and follow-up dislipemias are not uniform, and when there are more disparate dispute between the guides. There is underutilized or overuse of laboratory tests in dislipemias in the area 2 of Madrid in Primary Care. Conclusions The causes of variability in the use of laboratory tests in dyslipidaemia may be due to: firstly, the absence of a unified guide, adapted and updated in the primary care teams in the area 2 of Madrid. Second, the views of primary care physicians in the use of laboratory tests in dislipemias is not homogeneous, nor has a single criterion. Thirdly, the patient may add unsolicited evidence on the steering wheel of analytical application or not to go to health care. Fourth, the application of laboratory determinations using profiles produces analytical testing inappropriate. Fifth, address health area 2 Madrid has not provided during these years carrying out a program of clinical care in preventing cardiovascular risk.

Author: FERNÁNDEZ LÓPEZ LUCENDO M. ILUMINADA.
Year: 2004.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE CIENCIAS QUÍMICAS.
Place of preparation: CENTRO DE INVESTIGACIÓN (C.S.I.C.).

Summary: The galectinas proteins are a family of highly conserved through evolution, which are able to recognize specific carbohydrate compositions with the canonical structure (GalBl-GlcNAc) na through a carbohydrate recognition domain (CRD) from the highly conserved lower invertebrates to mammals. In recent years have been characterized structurally different galectinas. The galectina-l human (hGal-l) and galectinas lower organisms such as isoforms Cgl-ly Cgl-2 of the fungus Coprinus cinereus remain unresolved characterize this level. This work has been aimed primarily at determining the three-dimensional structures of this galectinas together to identify differences in the folding and evolutionary adaptation to the various elements of the secondary structure that determine the structure Quaternary. The folding general hGal-ly CG-14, eleven threads B adopting the cause of topological type "Jelly-roll" as a result of a strong partnership face to face between two sheets B with a provision of S threads (Fl-FS ) and 6 threads (Sl-S6) conformation antiparalela. It has been shown that h-Gall exists as dímero in solution by gel filtration analysis and mass spectrometry. The three-dimensional structure shows that the stabilization of dímero in hGal-l occurs mainly through a network of hydrogen bridges between the threads IF (N terminal) and Fl (C-terminal) of the two monomers. This network of bridges hydrogen is reinforced by the participation of the side chains of several hydrophobic residues of the SI and Fl threads that extend into the interior of the molecule creating a hydrophobic core. In chicken have identified two galectinas like "proto" (CG-14 and CG-16), whose expression varies depending on the fabric and the stage of development. These proteins differ significantly in terms of the binding specificity of oligosaccharides and tissue specific regulation. An important feature of the galectina CG-14 is that, unlike the h-Gall and CG-16, is a monomer in solution, despite the generally galectinas form dimers and oligomers. These statements are required for association occurring phenomena hemoaglutinación and the intersecting of ligands given that the majority of galectinas contains a single site reconnaissance carbohydrate (CRD) subunit. The intersecting oligosaccharides with different triggers various cellular signaling pathways. A detailed comparison of the amino acid sequence of CG-14 with galectinas typically diméricas, shows the existence of two cysteine residues at positions 2 and 7, which are not conserved in hGal-lo in CG-16. In CG14, the interaction between the two subunits is reinforced by the formation of an additional intermolecular disulfide bridge between residues Cys7 the thread Sl. Despite the fact that the crystallographic structure in the N-terminal region is not defined for the first two waste (Serl-Cys2), we have carried out the design of this region. Based on these data, we could propose a regulatory mechanism that allows the shifting of balance monomer I dimero, depending on the physiological conditions of the medium. Based on their molecular architecture, galectinas rungicas of Coprinus cinereus fall within the family of galectinas type "proto" with a peculiar organization Quaternary. Both share a folding in Jelly Role! But with subtle differences in the N-and C-terminal regions, which will regulate the spatial arrangement of the different oligomers. The insertion of 6 residues at the extreme amino leading to a new thread B (FO), increasing the number of strands of the film to six (FO-FS) available at the interface of dimero. As a result, prevents the dimerizaci 8 is being pr 5dd oduzca as h-Gall, CG-14. However, the dimerización in Cgl-l (-2) occurs via the C-terminal end (His14l-AlalSO) as a coupling between two arms, in conformation with a random coi!, Which are wrapped maintaining contacts nature basically hydrophobic. The low sequence homology in galectinas observed between animals and rungicas (MAYOR12%) is not transferred to the waste of carbohydrate binding domain, where it reflects a strict conservation. As a result of this conservation so selective, the overall architecture domain CRD in hGal-l, CG-14, Cgl-ly Cgl-2 is very similar. The conserved residues are found in all of them ply B S4, SS AND S6, on one side of the concave pair of blades B.

Author: BARRETINA GINESTA JORDI.
Year: 2004.
University: AUTÓNOMA DE BARCELONA [More theses of this university] [www.uab.es].
Place of defense: HOSPITAL UNIVERSTIARI GERMANS TRIAS I PUJOL.
Place of preparation: ESCUELA DE POSTGRADO.

Author: XIFRÓ COLLSAMATA FRANCESC XAVIER.
Year: 2004.
University: AUTÓNOMA DE BARCELONA [More theses of this university] [www.uab.es].
Place of defense: FACULTAD DE MEDICINA.
Place of preparation: ESCUELA DE POSTGRADO.

Summary: The granule neurons in the cerebellum (CGCs) needs the continued contribution of stimulations excitadoras to survive during the postnatal migration from the external granular layer to the internal granular layer. If CGCs not receive these stimulations, die by apoptosis. In cultivation, the CGCs die by apoptosis in the presence of physiological concentrations of potassium (5mM; K5), but survive in the presence of concentrations desporalizantes potassium (25mM; K25) or agonist glutamatérgico N-metil-D-aspártico (NMDA). Against this background, we note that the addition of the NMDA or K25 for only 24 hours in the CGCs immature (to 2 days in vitro; DIV) promotes the survival of CGCs over the DIVs (until 8 DIV). This neuroprotective effect would occur, at least in part through inhibition of caspase-3 (a protease key in triggering apoptosis). Using pharmacological inhibitors, we observed that inhibition of caspase-3 by the NMDA is dependent on the path PI3K/Akt and the tirosinas kinase, whereas the inhibition by K25 serious dependent on the PI3K/Akt and the CamKII. Part of the inhibition of caspase-3 by NMDA or K25 occur by inhibition of apoptosis via inbuilt, also called mitochondrial route. NMDA or K25 were able to inhibit different elements proapoptóticos related mitochondrial way, such as: decrease in the mitochondrial transmembrane potential, activation of protein propapotóticas the family Bcl-2 and release of proteins proapoptóticas mitochondria. In addition, they are able to activate proteins antiapoptóticas the family Bcl-2. All of these processes would be involved in the inhibition of caspase-3 that we see in the presence of NMDA or K25. Note also that, K25, via mitochondrial serious regulated, at least in part, by the way JNK. NMDA or K25 were able to inhibit this route temporarily in a pre-kinetic action on the various elements of the mitochondrial route, indicating that its effect on the route could be built through the inhibition of JNK way. Finally, we studied the possible role of track PI3K/Akt in the neuroprotective effects of NMDA or K25. This route has emerged as a key in multiple processes antipoptóticos. NMDA or K25 are not only able to activate this route, but their kinetics of activation is consistent with inhibition of proapoptóticos certain parameters, such as inhibition of caspasa-3. All these results suggest that only the presence of NMDA or K25 for only 24 hours is necessary for survival, long time, the CGCs. This protective effect would be through inhibition of caspasa-3. This inhibition occur on the one hand through the inhibition of apoptosis via inbuilt, and secondly, through the activation of the path of survival PI3K/Akt.

Author: ALÍA MORAL MARIO.
Year: 2004.
University: COMPLUTENSE DE MADRID [More theses of this university] [www.ucm.es].
Place of defense: FACULTAD DE CC. BIOLÓGICAS.
Place of preparation: FACULTAD DE CC. BIOLÓGICAS.

Summary: The antioxidants in the diet is considered beneficial to health because of their potential protective effect against oxidative stress, which has been implicated in the pathogenesis of many diseases such as cancer, heart disease and aterosclerosis.Varios epidemiological studies have shown that diets in the dominated fruits, vegetables or vegetables with high content in natural antioxidants, help reduce mortality in cardiovascular and cerebrovascular diseases, though its protective effects against the risk of cancer are less concluyentes.Las fruits are rich in natural antioxidants like For compounds polifenólicos.El potential effect of certain antioxidant polyphenols fruit extracts or concentrates them has been investigated in cultured cells ex vivo and in vivo in experimental animals and in humans. The overall objective of this work is the characterization ex vivo and in vivo response of the generator by antioxidant compounds from the diet polifenólicos assessed by measuring various biomarkers of redox state in basal conditions and after the induction of oxidative stress.