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Biomedical subjects

A Romero

Publications and source records attributed to A Romero.

At least 55 records · Page 3Linked to original sources
PubMed2001

A proinflammatory peptide from Helicobacter pylori activates monocytes to induce lymphocyte dysfunction and apoptosis.

Infection with Helicobacter pylori causes chronic gastritis, which is characterized by a dense mucosal infiltration by inflammatory cells such as monocytes/macrophages. H. pylori-induced inflammation is a risk factor for the development of gastric adenocarcinoma, but the mechanisms involved in H. pylori-associated carcinogenesis are poorly understood. A cecropin-like H. pylori peptide, Hp(2-20), was found to be a monocyte chemoattractant and activated the monocyte NADPH-oxidase to produce oxygen radicals. The receptors mediating monocyte activation were identified as FPRL1 and the monocyte-specific orphan receptor FPRL2. Hp(2-20)-activated monocytes inhibited lymphocytes with antitumor properties, such as CD56+ natural killer (NK) cells and CD3epsilon+ T cells. The changes observed in NK cells and T cells--a reduced antitumor cytotoxicity, downregulation of CD3zeta expression, and apoptosis--were mediated by Hp(2-20)-induced oxygen radicals. Histamine, a gastric mucosal constituent, rescued NK cells and T cells from inhibition and apoptosis by suppressing Hp(2-20)-induced oxygen radical formation. We conclude that H. pylori expression of this monocyte-activating peptide contributes to its ability to attract and activate monocytes and reduces the function and viability of antineoplastic lymphocytes. These novel mechanisms may be subject to local, histaminergic regulation in the gastric mucosa.

Adenocarcinoma↗
PubMed2001

[Growth kinetics and proteases production of Pseudomonas fluorescens in raw milk at refrigeration].

The general use of refrigeration of raw milk has contributed to maintenance of its quality, but has induced the selection of a psychotrophic bacteria which during its growth produces heat-resistant enzymes responsible, in part, for the deterioration of long-life products. Given the condition of the prolonged refrigeration of the milk before the process, it was necessary to determine the growth curves for bacteria at temperatures between 2 degrees C and 10 degrees C and the kinetics of production of proteases in raw fresh milk, inoculated with Pseudomonas fluorescens RV1O, using an automatic fermentation system, with minimal agitation under conditions of controlled temperature and pH. The results show a development over 10(5) ufc/mL in the cultures at 6 degrees, 8 degrees and 10 degrees C during the first 30 h and proteasic activities over 30 microM p-NA/2 h getting levels of de 80-180 microM p-NA/2 h over 50 h of cultivation. Only the cultures at 2 degrees C appeared stable with inferior cell counts and without inducing enzymatic activity. At 4 degrees C an intermediate situation occurs.

Animals↗
PubMed2000

Chemo-enzymatic total synthesis of 3-epiaustraline, australine, and 7-epialexine.

Sequential enzymatic aldol reaction and bis-reductive amination leads to the total syntheses of tetrahydroxylated pyrrolizidine alkaloids, 3-epiaustaline (14), australine (1), and 7-epialexine (11). This approach allows for their rapid construction without the need for protecting group manipulation of the hydroxyl functionality. In addition, an improved procedure for the asymmetric epoxidation of divinyl carbinol (3) was described, and the product was used in a concise synthesis of the required triol 7 and ent-7.

Epoxy Compounds↗
PubMed2000

Laminin-coated poly(L-lactide) filaments induce robust neurite growth while providing directional orientation.

Cellular channels during development and after peripheral nerve injury are thought to provide guidance cues to growing axons. In tissue culture where these cues are absent, neurites from dorsal root ganglion neurons grow with a radial distribution. To induce directional axonal growth and to enhance the rate of axonal growth after injury, we have designed microfilaments of poly(L-lactide). We demonstrate that dorsal root ganglia grown on these filaments in vitro extend longitudinally oriented neurites in a manner similar to native peripheral nerves. The extent of neurite growth was significantly higher on laminin-coated filaments compared with uncoated and poly-L-lysine-coated filaments. As high as 5.8 +/- 0.2 mm growth was observed on laminin-coated filaments compared with 2.0 +/- 0.2 mm on uncoated and 2.2 +/- 0.3 mm on poly-L-lysine-coated filaments within 8 days. Schwann cells were found to grow on all types of filaments. They were, however, absent in the leading edges of growth on laminin-coated filaments. Photolysis of Schwann cells caused a significant reduction in the neurite length on all types of filaments. Laminin-coated filaments, however, induced significantly longer neurites compared with uncoated and/or poly-L-lysine-coated filaments even in the absence of Schwann cells. Our results suggest that laminin-coated poly(L-lactide) filaments are suitable for inducing directional and enhanced axonal growth. Implants designed by arranging these microfilaments into bundles should aid regenerating axons by providing guidance cues and channels to organize matrix deposition, cell migration, axon growth, and improve functional recovery.

Animals↗
PubMed2000

Dissecting the energetics of the apoflavodoxin-FMN complex.

Many flavoproteins are non-covalent complexes between FMN and an apoprotein. To understand better the stability of flavoproteins, we have studied the energetics of the complex between FMN and the apoflavodoxin from Anabaena PCC 7119 by a combination of site-directed mutagenesis, titration calorimetry, equilibrium binding constant determinations, and x-ray crystallography. Comparison of the strength of the wild type and mutant apoflavodoxin-FMN complexes and that of the complexes between wild type apoflavodoxin and shortened FMN analogues (riboflavin and lumiflavin) allows the dissection of the binding energy into contributions associated with the different parts of the FMN molecule. The estimated contribution of the phosphate is greatest, at 7 kcal mol(-1); that of the isoalloxazine is of around 5-6 kcal mol(-1) (mainly due to interaction with Trp-57 and Tyr-94 in the apoprotein) and the ribityl contributes least: around 1 kcal mol(-1). The stabilization of the complex is both enthalpic and entropic although the enthalpy contribution is dominant. Both the phosphate and the isoalloxazine significantly contribute to the enthalpy of binding. The ionic strength does not have a large effect on the stability of the FMN complex because, although it weakens the phosphate interactions, it strengthens the isoalloxazine-protein hydrophobic interactions. Phosphate up to 100 mM does not affect the strength of the riboflavin complex, which suggests the isoalloxazine and phosphate binding sites may be independent in terms of binding energy. Interestingly, we find crystallographic evidence of flexibility in one of the loops (57-62) involved in isoalloxazine binding.

Apoproteins↗
PubMed2000

Disulphide-bond pattern and molecular modelling of the dimeric disintegrin EMF-10, a potent and selective integrin alpha5beta1 antagonist from Eristocophis macmahoni venom.

The disulphide-bond pattern of the heterodimeric disintegrin EMF-10, a potent and selective integrin alpha(5)beta(1) antagonist from Eristocophis macmahoni venom, was established by combination of amino-acid analysis, N-terminal sequencing and collision-induced dissociation by nanoelectrospray ionization quadrupole ion-trap MS of fragments isolated by reversed-phase HPLC after degradation of EMF-10 with oxalic acid. Each EMF-10 subunit contains four intrachain disulphide bonds. Two interchain cystine residues join the EMF-10 polypeptides. The intrachain linkages are conserved in monomeric disintegrins. A molecular model of EMF-10 was built using averaged NMR co-ordinates of flavoridin as a template. The active hairpin loops of the EMF-10 subunits occupy opposite locations at the ends of an elongated disulphide-bond ladder. In the EMF-10 model the N-terminal polypeptide of EMF-10B is close to the RGD-loop of the EMF-10A subunit, suggesting that the N-terminal region of the B-subunit could potentially influence the biological activity of the A-subunit.

Amino Acid Sequence↗
PubMed2000

Biochemical characterization, cDNA cloning and protein crystallization of aryl-alcohol oxidase from Pleurotus pulmonarius.

Aryl-alcohol oxidase (AAO) involved in lignin degradation by Pleurotus pulmonarius has been purified and characterized. The enzyme was produced in glucose-peptone medium and isolated in a sole chromatographic step using Sephacryl S-200. The purified enzyme is an extracellular glycoprotein with 14% N-carbohydrate content and an estimated molecular mass of 70.5 kDa and pI of 3.95. The kinetic studies showed the highest enzyme affinity against p-anisyl alcohol, with constants similar to those of Pleurotus eryngii and Bjerkandera adusta AAO but different from the intracellular AAO described in Phanerochaete chrysosporium, which present the highest activity on m-anisyl alcohol. Simultaneously, the cDNA of P. pulmonarius AAO has been cloned and sequenced. The translation of this sequence consisted of 593 amino acids including a signal peptide of 27 amino acids. The comparison with other alcohol oxidases, 35% amino acid identity with glucose oxidase, showed highly conserved amino acid sequences in N-terminal and C-terminal regions, in spite of differences in substrate specificity. Crystallization of AAO, carried out for the first time using the P. pulmonarius enzyme, will permit to obtain a molecular model for this oxidase and establish some characteristic of its catalytic site and general structure.

Alcohol Oxidoreductases↗
PubMed2000

Thymidilate synthase and p53 primary tumour expression as predictive factors for advanced colorectal cancer patients.

The purpose of this work was to analyse the ability of p53 and thymidilate synthase (TS) primary tumour expression to retrospectively predict clinical response to chemotherapy and long-term prognosis in patients with advanced colorectal cancers homogeneously treated by methotrexate (MTX)-modulated-5-fluorouracil (5-FU-FA). A total of 108 advanced colorectal cancer patients entered the present retrospective study. Immunohistochemical p53 (pAb 1801 mAb) and TS (TS106 mAb) expression on formalin-fixed paraffin-embedded primary tumour specimens was related to probability of clinical response to chemotherapy, time to progression and overall survival. p53 was expressed in 53/108 (49%) tumours, while 54/108 (50%) showed TS immunostaining. No relationship was demonstrated between p53 positivity and clinical response to chemotherapy (objective response (OR): 20% vs 23%, in p53+ and p53- cases respectively) or overall survival. Percent of OR was significantly higher in TS-negative with respect to TS-positive tumours (30% vs 15% respectively; P < 0.04); simultaneous analysis of TS and p53 indicated 7% OR for p53-positive/TS-positive tumours vs 46% for p53-positive/TS-negative tumours (P < 0.03). Logistic regression analysis confirmed a significant association between TS tumour status and clinical response to chemotherapy (hazard ratio (HR): 2.91; 95% confidence interval (CI) 8.34-1.01; two-sided P < 0.05). A multivariate analysis of overall survival showed that only a small number of metastatic sites was statistically relevant (HR 1.89; 95% CI 2.85-1.26; two-sided P < 0.03). Our study suggests that immunohistochemical expression of p53 and TS could assist the clinician in predicting response of colorectal cancer patients to modulated MTX-5-FU therapy.

Adult↗
PubMed2000

A chloroform extract obtained from a decoction of Ficus carica leaves improves the cholesterolaemic status of rats with streptozotocin-induced diabetes.

The antidiabetic effects of Ficus carica leaf extracts have been reported previously. From the aqueous decoction of fig leaves, after treatment with HCI, centrifuging, treatment with sodium hydroxide (NaOH) and extraction with chloroform (CHCl3), the administration of the organic phase rats with streptozotocin-induced diabetes led to a decline in the levels of total cholesterol and an decrease in the total cholesterol/HDL cholesterol ratio (with respect to the control group), together with a reduction of the hyperglycaemia.

Animals↗
PubMed2000

Medicinal chemistry based on the sugar code: fundamentals of lectinology and experimental strategies with lectins as targets.

Theoretical calculations reveal that oligosaccharides are second to no other class of biochemical oligomery in terms of coding capacity. As integral part of cellular glycoconjugates they can serve as recognitive units for receptors (lectins). Having first been detected in plants, lectins are present ubiquitously. Remarkably for this field, they serve as bacterial and viral adhesins. Following a description of these branches of lectinology to illustrate history, current status and potential for medicinal chemistry, we document that lectins are involved in a wide variety of biochemical processes including intra- and intercellular glycoconjugate trafficking, initiation of signal transduction affecting e. g. growth regulation and cell adhesion in animals. It is thus justified to compare crucial carbohydrate epitopes with the postal code ensuring correct mail routing and delivery. In view of the functional relevance of lectins the design of high-affinity reagents to occupy their carbohydrate recognition domains offers the perspective for an attractive source of new drugs. Their applications can be supposed to encompass the use as cell-type-selective determinant for targeted drug delivery and as blocking devices in anti-adhesion therapy during infections and inflammatory disease. To master the task of devising custom-made glycans/glycomimetics for this purpose, the individual enthalpic and entropic contributions in the molecular rendezvous between the sugar receptor under scrutiny and its ligand in the presence of solvent molecules undergoing positional rearrangements need to be understood and rationally exploited. As remunerative means to this end, cleverly orchestrated deployment of a panel of methods is essential. Concerning the carbohydrate ligand, its topological parameters and flexibility are assessed by the combination of computer-assisted molecular-mechanics and molecular-dynamics calculations and NMR-spectroscopic measurements. In the presence of the receptor, the latter technique will provide insights into conformational aspects of the bound ligand and into spatial vicinity of the ligand to distinct side chains of amino acids establishing the binding site in solution. Also in solution, the hydrogen-bonding pattern in the complex can be mapped with monodeoxy and monofluoro derivatives of the oligosaccharide. Together with X-ray crystallographic and microcalorimetric studies the limits of a feasible affinity enhancement can be systematically probed. With galactoside-binding lectins as instructive mo del, recent progress in this area of drug design will be documented, emphasizing the general applicability of the outlined interdisciplinary approach.

Animals↗