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Biomedical subjects

E Brun

Publications and source records attributed to E Brun.

At least 37 records · Page 2Linked to original sources
PubMed1998

Calcium binding by the N-terminal cellulose-binding domain from Cellulomonas fimi beta-1,4-glucanase CenC.

The interaction of the N-terminal cellulose-binding domain, CBDN1, from Cellulomonas fimi beta-1,4-glucanase CenC with calcium was investigated using NMR spectroscopy and calorimetry. CBDN1 binds a single calcium ion with an equilibrium association constant of approximately 10(5) M-1 at 35 degreesC and pH 6.0. Binding is exothermic (-42 +/- 2 kJ mol-1) under these conditions and is accompanied by a small negative change in heat capacity (DeltaCp = -0.41 +/- 0.16 kJ mol-1 K-1). From an NMR line shape analysis, the rate constants for calcium association and dissociation were found to be (5 +/- 2) x 10(7) s-1 M-1 and (4.5 +/- 0.6) x 10(2) s-1, respectively. The rapid association kinetics indicate that the calcium-binding site on CBDN1 is accessible and, to the first approximation, preformed. Based on patterns of chemical shift perturbations, and structural comparisons with the Bacillus sp. 1, 3-1,4-beta-glucanases, the backbone carbonyl oxygens of Thr8, Gly30, and Asp142 and a side chain carboxyl oxygen of Asp142 are postulated to form the calcium-binding site of CBDN1. Consistent with the calcium-independent affinity of CBDN1 for cellopentaose, this exposed site is located on the face of CBDN1 opposite to that forming the oligosaccharide-binding cleft. The midpoint denaturation temperature of CBDN1 is increased by approximately 8 degreesC at pH 6.0 in the presence of saturating amounts of calcium, confirming that metal ion binding is thermodynamically linked to native-state stability.

Bacterial Proteins↗
PubMed1998

Determination of anti-Toxoplasma gondii immunoglobulin G avidity: adaptation to the Vidas system (bioMérieux).

The determination of specific anti-Toxoplasma gondii IgG avidity has been proposed to improve the determination of the date of toxoplasmic seroconversion in pregnant women. In this study, we adapted this serological technique to the Vidas system (bioMérieux) using 6 M urea as the dissociating agent. We studied 356 sera, including 42 sequential sera from sero-conversions in pregnant women. Our results show that the test is easy to use, and that an avidity index higher than 0.300 allows the exclusion of a recent infection acquired less than 4 months before serum sampling.

Animals↗
PubMed1998

A placental-derived tissue matrix as a bowel wall substitute in rats: preliminary study.

BACKGROUND: Occasionally, conventional closure of an intestinal wall defect (fistula) is not possible. The development of a biologic material that can be used in this setting would solve a significant clinical problem. We hypothesized that a collagen patch (connective tissue matrix [CTM]) designed to allow tissue regeneration was such a material. METHODS: To test this hypothesis, we performed a laparotomy on 75 male Sprague-Dawley rats (experiment A) and removed 25% of the anterior cecal wall (1 cm in diameter) to compare intestinal wound healing in four situations: (1) control (no repair), (2) fibrin glue repair, (3) primary repair, and (4) repair with CTM (human placental bilayer, types I, III, and IV). Animals were killed at 1, 2, 4, and 6 weeks. Healing was graded by bursting pressure expressed in millimeters of mercury, histologic score (0 to 4), and mortality rates. After this, we used the same protocol to remove 80% of the cecal wall (1.5 x 2.0 cm) in 19 animals (experiment B) to compare (5) fibrin glue repair alone with (6) CTM repair in a situation in which the defect was too large for primary repair. RESULTS: CTM repair of a lethal cecal wall defect (experiment A) is equivalent to either primary repair or fibrin glue repair. When the defect is too large for primary repair (experiment B), repair with fibrin glue also does not work. However, in this same setting, CTM repair is successful (p < 0.00001). CONCLUSIONS: Intestinal wall defects not suitable for conventional closure can be repaired successfully with a collagen patch, with histologic characteristics similar to those seen with primary repair.

Animals↗
PubMed1997

Solution structure of the cellulose-binding domain of the endoglucanase Z secreted by Erwinia chrysanthemi.

Two-dimensional proton nuclear magnetic resonance spectroscopy has been used to determine the three-dimensional structure of the 62 amino acid C-terminal cellulose-binding domain (CBD) of the endoglucanase Z (CBDEGZ), secreted by Erwinia chrysanthemi. An experimental data set comprising 958 interproton nOe-derived restraints was used to calculate 23 structures. The calculated structures have an average root-mean-square deviation between Cys4 and Cys61 of 0.91 +/- 0.11 A for backbone atoms and 1.18 +/- 0.12 A for the heavy atoms. The CBDEGZ exhibits a skiboot shape based mainly on a triple antiparallel beta-sheet perpendicular to a less-ordered summital loop. Three aromatic rings (Trp18, Trp43, and Tyr44) are localized on one face of the protein and are exposed to the solvent in a conformation compatible with a cellulose-binding site. Based on its original folding, we have been able to relate the CBD sequence to those of several domains of unknown function occurring in several bacterial chitinases as well as other proteins. This study also provides a structural basis for analyzing the secretion-related information specific to the CBDEGZ.

Amino Acid Sequence↗
PubMed1997

Early prediction of treatment outcome in head and neck cancer with 2-18FDG PET.

The development of alternative treatment regimens in clinical oncology has increased the need for early prediction of cancer therapy outcome. The aim of this study was, early in the treatment phase, to identify patients with advanced head and neck cancer, responding or not responding to initiated therapy. The tumour metabolic rate of glucose (MRgl) examined by 2-18FDG-PET was determined in 17 patients before and after the first weeks of either radiotherapy (16-35 Gy) or one course of combination chemotherapy. Metabolic values uptake values normalized to plasma activity integrals--were correlated to loco-regional outcome, as evaluated 5-6 weeks after completion of treatment. Initial low tumour MRgl (<20 micromol/min/100 g tissue), in primary lesions or regional metastases, predicted a local complete response. When a high initial tumour MRgl was found, the magnitude of the reduction of MRgl in the second PET examination might be an adjunct in predicting local tumour response.

Adult↗
PubMed1996

Insulin infusion amplifies 17 alpha-hydroxycorticosteroid intermediates response to adrenocorticotropin in hyperandrogenic women: apparent relative impairment of 17,20-lyase activity.

Recent data suggest that insulin is a modulator of ovarian and adrenal steroidogenesis and that, in the ovary of hyperandrogenic women, hyperinsulinemia might cause dysregulation of cytochrome P450c17 alpha activity. To further assess in vivo the effects of insulin on adrenal steroidogenesis, ACTH stimulation was carried out in 21 hyperandrogenic women during a 3-h hyperinsulinemic (80 mU/m2-min) euglycemic clamp. In all of these women the procedure was repeated during saline infusion as n control. In nonamenorrheic patients, the tests were performed in the early follicular phase of two different menstrual cycles. Serum cortisol, progesterone, 17-hydroxypregnenolone (17-OHJPREG). 17-hydroxyprogesterone (17-OHP), dehydroepiandrosterone (DHEA), and androstenedione (A) were measured after 2 h of insulin or saline infusion (zero time) and, subsequently, 30 and 60 min after an iv bolus of 0.25 mg ACTH-(1-24). At zero time, no difference was found in the serum steroid concentrations between the two protocols. ACTH-stimulated serum 17-OHPREG and, to a lesser extent, 17-OHP were significantly higher during insulin than during saline infusion (peaks, 60.6 +/- 9.0 vs. 40.7 +/- 7.9 and 7.7 +/- 7.7 vs. 6.6 +/- 0.6 nmol/L; P < 0.005 and P < 0.01, respectively). Serum DHEA was also slightly higher during hyperinsulinemia, although only after 30 min (54.5 +/- 3.0 vs. 48.2 +/- 4.2 nmol/L; P < 0.05). No statistically significant difference in the cortisol, progesterone, or androstenedione response to ACTH was found between the two protocols. ACTH-stimulated 17-OHPREG/DHEA and 17-OHP/A molar ratios, indexes of apparent 17,20-lyase activity, were significantly higher during the clamp studies than during saline infusion (by ANOVA, F = 12.8; P < 0.001 and F = 6.7; P < 0.005, respectively), suggesting an impaired enzyme activity. These in vivo data support the hypothesis that insulin potentiates ACTH-stimulated steroidogenesis. This effect of insulin seems to be associated with a relative impairment of 17,20-lyase activity.

17-Hydroxycorticosteroids↗
PubMed1996

The insulin resistance in women with hyperandrogenism is partially reversed by antiandrogen treatment: evidence that androgens impair insulin action in women.

To assess whether androgen excess per se might impair insulin action, insulin sensitivity was measured by a two-step (20 and 80 mU/m2.min) hyperinsulinemic euglycemic clamp combined with indirect calorimetry and tracer glucose infusion in 43 women (13 obese and 30 nonobese) with normal glucose tolerance and clinical evidence of increased androgen action (hirsutism and/or polycystic ovary syndrome) as well as 12 age- and body mass index-matched healthy controls. Hyperandrogenic women were studied basally and after 3-4 months of antiandrogen treatment with 3 different drugs: spironolactone (n = 23), flutamide (n = 10), or the GnRH agonist buserelin (n = 10). Six women given spironolactone were also reexamined after 1 yr of therapy. At baseline, insulin-mediated glucose uptake was lower in hyperandrogenic women than in controls (by ANOVA, F = 14.3; P < 0.001). Insulin resistance was observed in both ovarian and nonovarian hyperandrogenism, as distinguished by acute GnRH agonist testing. After antiandrogen therapy, insulin action on glucose metabolism significantly increased for both the patients as a whole (F = 7.4; P < 0.01) and each treatment group separately. However, insulin action remained lower than in controls and showed no further improvement in patients reevaluated after I yr of treatment. Increases in both oxidative and nonoxidative glucose metabolism accounted for the improvement in substrate disposal induced by antiandrogen drugs. The increase in the effectiveness of insulin was greater in the lean subjects, whereas the change was small and not statistically significant in the obese women. Response to treatment was more pronounced in women with nonovarian hyperandrogenism, particularly at the low insulin infusion rate. Endogenous glucose production in hyperandrogenic patients was similar to that in healthy women and was unaffected by therapy. In conclusion, antiandrogen treatment partially reversed the peripheral insulin resistance associated with hyperandrogenism regardless of which antiandrogen was used. These data strongly suggest that in women, androgen excess per se contributes to impairment of insulin action.

Adult↗
PubMed1995

Overproduction, purification and characterization of the cellulose-binding domain of the Erwinia chrysanthemi secreted endoglucanase EGZ.

EGZ is the major endoglucanase secreted by Erwinia chrysanthemi. Functional characterization indicates that it is made of a catalytic N-terminal domain linked to a C-terminal cellulose-binding domain (CBD) by a Ser/Thr-rich linker. A chimeric plasmid, in which the CBD-encoding region was fused downstream of the ompA signal sequence, was constructed and introduced into Escherichia coli. This allowed for the production of processed and disulfide-bonded CBD, mostly recovered from the culture supernatant of E. coli. One-dimensional NMR analysis of the purified CBD reveals that it folds into a well-structured domain. Moreover, comparison with the one-dimensional NMR analysis of full-length EGZ strongly suggests that the CBD folds autonomously, providing experimental support for the existence of domains of EGZ.

Binding Sites↗
PubMed1995

A phase I/II evaluation of metoclopramide as a radiosensitiser in patients with inoperable squamous cell carcinoma of the lung.

The feasibility of administering metoclopramide (MCA) as a radiosensitizer has been evaluated in 23 patients with a pathological or cytological diagnosis of a squamous cell carcinoma of the lung, clinically evaluated as inoperable. All patients received 40-60 Gy radiotherapy fractionated into 1.8 Gy fractions 5 times per week (Monday-Friday). Two MCA treatment regimens were used: (i) MCA at 2 mg/kg administered by intravenous-infusion 1-2 h prior to radiotherapy 3 times per week (Monday, Wednesday, Friday); and (ii) MCA at 1 mg/kg administered by intravenous infusion 1-2 h prior to radiotherapy 5 times per week (Monday-Friday). 11 of the 23 patients treated with radiotherapy and MCA had none to mild pneumonitis or fibrosis and another 8 of the 23 had moderate levels. No patient had their therapy interrupted due to radiation-related side-effects. The MCA-related side-effects were as expected, i.e. 78% of the patients experienced sedation/tiredness and 48% expressed restlessness/anxiety symptoms. Both the total dose and serum levels of MCA were significantly associated to the MCA side-effect profile. Tumour response, duration of tumour response and survival were significantly positively correlated to the total and weekly doses of MCA administered to the patients during their radiotherapy treatment. These favourable phase II data have justified the initiation of a phase II/III randomised multicentred trial being carried out in Europe to evaluate MCA as a radiosensitiser.

Aged↗
PubMed1995

A large pedigree with early-onset Alzheimer's disease: clinical, neuropathologic, and genetic characterization.

We present clinical, neuropsychological, and neuropathologic data on a large pedigree including 34 subjects with early-onset progressive dementia. The mean (+/- SD) age at onset was 46 +/- 3.5 years and the mean age at death 52.6 +/- 5.7 years. Twelve patients were clinically diagnosed as having probable Alzheimer's disease (AD) according to the NINCDS-ADRDA criteria. Neuropsychological evaluation, performed at a moderate stage of the disease, was available in six subjects and showed a classic pattern of cognitive deficit. Myoclonus and extrapyramidal signs were common, and seizures were present in all affected subjects. There were neuropathologic changes typical of AD in two brains. A significant lod score of 5.48 was observed at a recombination fraction of theta = 0.0 with the genetic marker D14S43, thereby establishing that the responsible gene was located on chromosome 14q24.3. These results suggest that epilepsy could represent a particular feature in AD families linked to chromosome 14q.

Age of Onset↗
PubMed1994

Periplasmic disulphide bond formation is essential for cellulase secretion by the plant pathogen Erwinia chrysanthemi.

Secretion to the cell exterior of cellulase EGZ and of at least six pectinases enables the Gram-negative Erwinia chrysanthemi to cause severe plant disease. The C-terminal cellulose-binding domain (CBD) of EGZ was found to contain a disulphide bond which forms, in the periplasm, between residues Cys-325 and Cys-382. Dithiothreitol (DTT)-treatment of native EGZ showed that the disulphide bond was dispensable, both for catalysis and cellulose binding. Adding DTT to E. chrysanthemi cultures led to immediate arrest of secretion of EGZ which accumulated in the periplasm where the CBD was eventually proteolysed. Site-directed mutagenesis that affected Cys residues involved in disulphide bond formation resulted in molecules that were catalytically active and able to bind to cellulose but were no longer secreted. Instead they accumulated in the periplasm. Interestingly, the region around EGZ Cys-325 is conserved in two pectinases secreted by the same pathway as EGZ. We conclude that the conserved Cys, and possibly adjacent residues, bear essential information for EGZ to be secreted and that periplasmic disulphide bond formation is an obligatory step which provides a pre-folded functional form of EGZ with secretion competence.

Amino Acid Sequence↗
PubMed1993

Endocarditis caused by Rochalimaea henselae.

A case of community-acquired, culture-negative, infective endocarditis was diagnosed in a 57-year-old construction worker. Small, pleomorphic gram-negative rods were seen in Brown-Hopps tissue gram stains and Warthin-Starry silver stains. The organism was identified as Rochalimaea henselae by polymerase chain reaction amplification and sequencing of the 16S rDNA gene sequence. This is the first report of infective endocarditis caused by R henselae.

Community-Acquired Infections↗