Filipe Silva

Role of the Human ST6GalNAc-I and ST6GalNAc-II in the Synthesis of the Cancer-Associated Sialyl-Tn Antigen

The Sialyl-Tn antigen (Neu5Acα2–6GalNAc-O-Ser/Thr) is highly expressed in several human carcinomas and is associated with carcinoma aggressiveness and poor prognosis. We characterized two human sialyltransferases, CMP-Neu5Ac:GalNAc-R α2,6-sialyltransferase (ST6GalNAc)-I and ST6GalNAc-II, that are candidate enzymes for Sialyl-Tn synthases. We expressed soluble recombinant hST6GalNAc-I and hST6GalNAc-II and characterized the substrate specificity of both enzymes toward a panel of glycopeptides, glycoproteins, and other synthetic glycoconjugates. The recombinant ST6GalNAc-I and ST6GalNAc-II showed similar substrate specificity toward glycoproteins and GalNAcα-O-Ser/Thr glycopeptides, such as glycopeptides derived from the MUC2 mucin and the HIVgp120. We also observed that the amino acid sequence of the acceptor glycopeptide contributes to the in vitro substrate specificity of both enzymes. We additionally established a gastric cell line, MKN45, stably transfected with the full length of either ST6GalNAc-I or ST6GalNAc-II and evaluated the carbohydrate antigens expression profile induced by each enzyme. MKN45 transfected with ST6GalNAc-I showed high expression of Sialyl-Tn, whereas MKN45 transfected with ST6GalNAc-II showed the biosynthesis of the Sialyl-6T structure [Galβ1–3 (Neu5Acα2–6)GalNAc-O-Ser/Thr]. In conclusion, although both enzymes show similar in vitro activities when Tn antigen alone is available, whenever both Tn and T antigens are present, ST6GalNAc-I acts preferentially on Tn antigen, whereas the ST6GalNAc-II acts preferentially on T antigen. Our results show that ST6GalNAc-I is the major Sialyl-Tn synthase and strongly support the hypothesis that the expression of the Sialyl-Tn antigen in cancer cells is due to ST6GalNAc-I activity.

Energy analysis during biped walking

Describes the dynamic analysis of biped locomotion systems. A planar biped is modelled and studied focusing on the problem of energy efficiency. First, the motion of the system is characterised in terms of a set of locomotion variables, namely: step length, hip height, hip ripple, hip offset, foot clearance and link lengths. Afterwards, three cost functions related to energy are proposed: absolute power, power dispersion and power lost. The aim is to understand the influence of these locomotion variables on the energy flow. The performance measures are discussed and the results compared with those observed in nature.

MUC1 gene polymorphism in the gastric carcinogenesis pathway

MUC1 like most mucin genes shows extensive length polymorphism in the central core region. In a previous study it was shown that individuals with small MUC1 alleles/genotypes have an increased risk for development of gastric carcinoma. Our aim was to see if MUC1 gene polymorphism was involved in susceptibility for the development of conditions that precede gastric carcinoma: chronic atrophic gastritis (CAG) and intestinal metaplasia (IM). We evaluated MUC1 polymorphism in a population of 174 individuals with chronic gastritis (CG) displaying (CAG) and/or intestinal metaplasia (IM). The population of patients with CG shows MUC1 allele frequencies significantly different from the gastric carcinoma patients and blood donors population. A significantly lower frequency of CAG and IM was observed in MUC1 VNTR heterozygotic patients. Within the group of patients with IM, MUC1 large VNTR homozygotes show a significantly higher frequency of complete IM while small VNTR homozygotes show a significantly higher frequency of incomplete IM. These findings show that MUC1 polymorphism may define different susceptibility backgrounds for the development of conditions that precede gastric carcinoma: chronic atrophic gastritis (CAG) and intestinal metaplasia (IM).

Role of site-specific promoter hypomethylation in aberrant MUC2 mucin expression in mucinous gastric carcinomas

In the present work we investigated the methylation levels of mucin gene (MUC)2 promoter region in normal gastric mucosa and mucinous gastric carcinoma in order to access if the observed de novo expression of the intestinal mucin MUC2 in mucinous gastric carcinoma is associated with changes in MUC2 promoter methylation status. The results obtained by methylation-sensitive single nucleotide primer extension suggest that MUC2 expression in gastric cells is regulated by promoter methylation and further indicate that two specific cytosine guaine dinucleotide (CpG) sites may play a particularly important regulatory role.

Goal-oriented biped walking based on force interaction control

Addresses the problem of modelling and control of a biped robot by combining Cartesian-based position and force control algorithms. The walking cycle is divided in two phases: single support, in which one leg is in contact with the ground and the other leg swings forward, and double support, in which the forward leg absorbs the impact and gradually accepts the robots weight. The contact of the foot with the constrained surface is modelled through linear and nonlinear spring-damper systems. The proposed control approach is based on simple motion goals taking into account the reaction forces between the feet and the ground. The control algorithm is tested through several experiments and its effectiveness and robustness is discussed.

Influence of flow rate variation on the development of Escherichia coli biofilms

This work investigates the effect of flow rate variation on mass transfer and on the development of Escherichia coli biofilms on a flow cell reactor under turbulent flow conditions. Computational fluid dynamics (CFD) was used to assess the applicability of this reactor for the simulation of industrial and biomedical biofilms and the numerical results were validated by streak photography. Two flow rates of 374 and 242xa0Lxa0h−1 (corresponding to Reynolds numbers of 6,720 and 4,350) were tested and wall shear stresses between 0.183 and 0.511xa0Pa were predicted in the flow cell reactor. External mass transfer coefficients of 1.38xa0×xa010−5 and 9.64xa0×xa010−6xa0mxa0s−1 were obtained for the higher and lower flow rates, respectively. Biofilm formation was favored at the lowest flow rate because shear stress effects were more important than mass transfer limitations. This flow cell reactor generates wall shear stresses that are similar to those found in some industrial and biomedical settings, thus it is likely that the results obtained on this work can be used in the development of biofilm control strategies in both scenarios.

Kinematic aspects of robotic biped locomotion systems

This paper presents the kinematic study of robotic biped locomotion systems. The main purpose is to determine the kinematic characteristics and the system performance during walking. For that objective, the prescribed motion of the biped is completely characterised in terms of five locomotion variables: step length, hip height, maximum hip ripple, maximum foot clearance and link lengths. In this work, we propose three methods to quantitatively measure the performance of the walking robot: locomobility measure, perturbation analysis and low-pass frequency response. These performance measures are discussed and compared in determining the robustness and effectiveness of the resulting locomotion.

Extended structural variation of a pentanucleotide repeat in the GSTP1 gene: Characterisation in a normal population and in thyroid and gastric tumours

The promoter region of the human GSTP1 gene contains a polymorphic short tandem repeat (STR) locus consisting of pentanucleotide repeat units (ATAAA). In this work we report the existence of a total of 26xa0alleles in a Caucasian population. While differences in size (ranging from one to five base pairs) were responsible for the major variation, in five size-defined classes, two alternative sequences were found. Automatic fragment sizing and sequencing analysis revealed that this polymorphism is of a highly complex nature in contrast with previous reports. A genetic population study was carried out on a random sample from Portugal showing no deviation from Hardy-Weinberg equilibrium. Somatic instability studies were also performed on gastric and thyroid tumours using this STR: no instability was detected in thyroid tumour tissues when compared with their normal counterpart but in gastric tumour tissues microsatellite instability (MSI) was detected in 9.6% of the cases and loss of heterozygosity (LOH) also in 9.6% of the cases studied. The results obtained with GSTP1 in gastric cancer were compared with previously reported data on MSI using BAT-26 and several dinucleotide repeat markers.

Segregation analysis of tetra‐ and pentanucleotide short tandem repeat polymorphisms: Deviation from Mendelian expectations

Short tandem repeat (STR) polymorphisms are powerful tools for linkage studies, chromosome mapping and population analysis. The instability of these microsatellite regions is a prevailing event in several tumors and human genetic diseases and, despite various reports associating instability‐related genes and meiosis control, the dynamics of these STR regions in normal cells/individuals has frequently been disregarded. Having previously assayed somatic instability in gastric cancer for some tetra‐ and pentanucleotide STRs and given the increased application of this type of marker for routine forensic expertise, we report the results of an extensive analysis of segregation in nuclear families of a normal population for the same loci. No mutations were detected in 2374 parent/offspring allelic transfers at TH01, TPO, VWA31/A, MBPB, and CD4 STR loci. Nonsignificant differences were found between gene frequencies of parental and offspring generations. However, the segregation analysis revealed significant deviation from Mendelian expectations for: VWA31/A locus — alleles 19, 17 and 14 and TH01 locus — allele 6. In particular, parental meiosis strongly favored specific allele transmission, depending upon the sex of the offspring. Specific mating types are apparently responsible for most of these abnormal segregations. These results suggest selective factors working either at the gametic or zygotic levels.

Polipeptídeos e proteínas com influência na qualidade da espuma da cerveja e métodos analíticos utilizados no seu estudo

A review of polypeptides and proteins that influence, direct or indirectly, beer foam quality, as well as the most relevant analytical methods used in their study, are presented. Protein Z, LTP1 and hordein/glutelin fragments originated from malt have a direct influence on beer foam quality. Other proteins, like malt hordeins and albumins and wheat puroindolines, are, to some degree, also important for beer foam quality. Protein hydrophobicity is pointed out as a key parameter to enhance foam quality. Electrophoretic, chromatographic and immunological analytical methods are currently used to study polypeptides and proteins present in barley, malt, wort, beer, and foam. Best results are obtained when combinations of these methods are applied.