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C Picard

Publications and source records attributed to C Picard.

At least 19 recordsLinked to original sources
PubMed1992

Characterization of the deamidated forms of recombinant hirudin.

Recombinant hirudin variant rHV2-Lys 47 (MW = 6906.5) was intentionally deamidated by incubation in pH 9 phosphate buffer at 37 degrees C. Anion-exchange HPLC analysis showed that 11 forms could be generated. These were isolated and purified by combined anion-exchange and reversed-phase HPLC. Acid-catalyzed carboxyl methylation was used to introduce a mass shift of +15 amu per deamidated residue present in the molecule before analysis by liquid secondary ion mass spectrometry (LSIMS). Methylation enhanced, in particular, the abundance of the sequence ions in the LSIMS spectra. This permitted the determination of both the number (three) and the localization of the deamidated residues: Asn 52, Asn 53, and a residue located in the N-terminal 1-39 domain. Complementary sequencing techniques proved that the latter residue was Asn 33. Altogether four mono-, three di-, and four tri-deamidated forms were identified. The heterogeneity of the forms having identical deamidation positions but being chromatographically separable is thought to arise from the generation of alpha- and beta-aspartyl iso forms during the nonenzymatic deamidation process.

Amino Acid Sequence↗
PubMed1992

High-performance liquid chromatographic determination of 3-hydroxypyridinium derivatives as new markers of bone resorption.

Hydroxylysylpyridinoline (HP) and lysylpyridinoline (LP) are intermolecular cross-linking amino acids of collagen and their urinary excretion reflects bone resorption. An isocratic high-performance liquid chromatographic assay using a reversed-phase column with a prefractionation step and fluorescence detection was developed. The accuracy and reproducibility were assessed by loading experiments and by double analysis of urinary samples. The recoveries after various loads were above 90% for HP and between 87 and 94% for LP, with an intra-assay relative standard deviation (R.S.D.) of 5% for HP and 8% for LP. The inter-assay R.S.D.s were 8% for HP and 12.4% for LP. The fasting and 24-h urinary excretions of HP and LP were measured in 40 healthy subjects (mean age 35 years) of both sexes. There was no difference between males and females. Mean adult normal values were 33.6 +/- 8.1 pmol/mumol creatinine for HP and 7.0 +/- 2.5 pmol/mumol creatinine for LP in morning fasting urine and 2-.9 +/- 7.0 pmol/mumol creatinine for HP and 5.8 +/- 1.9 pmol/mumol creatinine for LP after 24-h urinary collection. HP and LP excretions were significantly higher in morning fasting urine than in 24-h collections, in agreement with the physiological circadian rythm of bone resorption. This simplified and optimized procedure is a good method for the determination of pyridinolines and should be useful for the evaluation of bone resorption.

Adult↗
PubMed1992

Detection and enumeration of bacteria in soil by direct DNA extraction and polymerase chain reaction.

In order to develop a rapid and specific detection test for bacteria in soil, we improved a method based on the polymerase chain reaction (PCR). Each step of the protocol, including direct lysis of cells, DNA purification, and PCR amplification, was optimized. To increase the efficiency of lysis, a step particularly critical for some microorganisms which resist classical techniques, we used small soil samples (100 mg) and various lytic treatments, including sonication, microwave heating, and thermal shocks. Purification of nucleic acids was achieved by passage through up to three Elutip d columns. Finally, PCR amplifications were optimized via biphasic protocols using booster conditions, lower denaturation temperatures, and addition of formamide. Two microorganisms were used as models: Agrobacterium tumefaciens, which is naturally absent from the soil used and was inoculated to calibrate the validity of the protocol, and Frankia spp., an actinomycete indigenous to the soil used. Specific primers were characterized either in the plasmid-borne vir genes for A. tumefaciens or in the variable regions of the 16S ribosomal gene for Frankia spp. Specific detection of the inoculated A. tumefaciens strain was routinely obtained when inocula ranged from 10(7) to 10(3) cells. Moreover, the strong correlation we observed between the size of the inocula and the results of the PCR reactions permitted assessment of the validity of the protocol in enumerating the number of microbial cells present in a soil sample. This allowed us to estimate the indigenous population of Frankia spp. at 0.2 x 10(5) genomes (i.e., amplifiable target sequences) per g of soil.

Agrobacterium tumefaciens↗
PubMed1992

[A comparative study of the cost of open-circuit as opposed to closed-circuit ventilation].

The authors compared two open randomized groups of patients undergoing surgery through general anaesthesia. Group 1 consisted of 54 patients ventilated by a Siemens 900 B ventilator in open circuit, and group 2, 56 patients ventilated by an ELSA de Gambro ventilator in a closed circuit. Comparative hour cost for nitrous oxide (N2O), oxygen (O2) and halogen gas, Enflurane, Isoflurane, was noted. All patients received the same regimen of anaesthesia and the two groups were identical in age, weight, surgery, respiratory volume and ventilation time. The evaluation of comparative hour cost included specific materials of close circuit ventilator: CO2 filter (Aridus), Lime. Were excluded maintenance and gas consumption expenditures before patient connected to the ventilator. The total hour cost (O2, N2O, specific materials for close circuit, without halogen gas) was 8.23 FF in closed circuit against 13.28 FF in open circuit, an economy of 38.27%. Hence, for oxygen, the hour cost was 0.70 FF in open circuit against 0.27 FF in closed circuit (gain of 65.3%). For nitrous oxide, the hour cost in open circuit was 12.50 FF against 2.44 FF in closed circuit (80.5%). For Isoflurane, the open circuit hour cost was 41.38 FF against 22.44 FF in closed circuit (47%). For Enflurane, the open circuit hour cost was 14.17 FF against 5.94 FF in closed circuit (58.1%). And, lastly for Enflurane, open circuit hour cost was 14.17 FF against 5.94 FF in close circuit, gain of 58.1%. These "modest" economy against those found in previous studies can be explained by the long-time duration of ventilation, saturating time in open circuit more or less long, depending on the physician, specific materials for closed circuit ventilation--lime, CO2 filter--in not taken into account, the hour cost of O2 + NO2 goes from 8.23 FF to 2.71 FF, and the gain against the close circuit becomes 79.6%: reducing hour cost by 5 times. In order to improve the effective cost of close circuit, the authors proposed: the use of closed circuit ventilation for more than 3 hours surgery, gas saturation in closed circuit after denitrogenation--which demands the use of halogen infjectors, and lime in containers cheaper than disposable cartridges. Respecting the above criteria, the total hour cost in close circuit fell to 4.90 FF, gain of 63% against open circuit. For O2 et N2O, the hour cost goes from 1.34 FF in close circuit to 13.28 FF in open circuit, 90% economy.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1991

Liquid secondary ion mass spectrometry applied to structural confirmation of enzymically prepared C-terminal-truncated derivatives of recombinant hirudin.

The thrombin-specific inhibitor, hirudin variant rHV2-Lys 47 (rHirudin), is a 65-amino acid polypeptide produced by recombinant DNA technology in yeast. Previous studies have shown that the acidic C-terminal segment of hirudin is susceptible to enzymic degradation. To address the question of C-terminal-truncated forms of the protein in terms of by-products or metabolites, well-defined reference compounds are needed. We prepared nine derivatives by carboxypeptidase Y digestion of rHirudin followed by a two-step chromatographic purification. Liquid secondary ion mass spectrometric measurements performed on peptides collected after reversed-phase high-performance liquid chromatography showed three pure forms (1-64, 1-63 and 1-56) and three mixtures of two forms each (1-62 + 1-61, 1-58 + 1-57 and 1-55 + 1-54), which were readily distinguished from one another by their mass spectra. Further purification of these co-eluted samples was achieved by ion-exchange chromatography and their structures were confirmed by liquid secondary ion mass spectrometry. Preliminary studies conducted on intact rHirudin indicated that this is an excellent analytical tool for mass measurements of hirudin-related proteins. Indeed, it allowed rapid (within 10-15 min), precise (0.50 a.m.u. relative to expected value), reproducible (mean MH+ = 6907.64 +/- 0.42 a.m.u.), sensitive (up to 500 ng, i.e. 72 pmol) and specific measurement of the quasi-molecular ion (MH+) of the protein, and was thus readily applicable to the analysis of several derivatives.

Amino Acid Sequence↗
PubMed1991

[Erysipelas: epidemiological, clinical and therapeutic data (111 cases)].

A retrospective study of 111 patients admitted to the Dermatology department of the Bichat hospital, Paris, between 1981 and 1988 for treatment of erysipelas revealed the following data: 1. Erysipelas was located on the lower limbs in 88.3 p. 100 of the cases and on the face in only 9.8 p. 100. 2. Facilitating and/or aggravating factors were: portal of entry in 75 p. 100 of the cases; impairment of venous and lymphatic circulations (41 p. 100); diabetes mellitus (13.5 p. 100); alcoholism and its socio-economic consequences (29 p. 100); unnecessary prescription of anti-inflammatory agents (11 p. 100). 3. Insufficient consideration was given to the clinical diagnosis: in 7.2 p. 100 of the patients erysipelas was diagnosed either after failure of heparin therapy or because phlebography was normal; some clinical features, notably bullae (30 p. 100) or purpura on the lower limbs (13 p. 100), confused the physicians. Delayed treatment was the main cause of local complications, such as abscess (4 cases) or focal cutaneous necrosis (4 cases). Erysipelas was recurrent in 23.5 p. 100 of the patients. 4. Bacteriological data in this series were insufficient to establish percentages of responsible organisms. However, penicillin G in mean doses of 12 million units per day administered intravenously for 5.5 days, then intramuscularly for 10 days was effective as first-line treatment in 80 p. 100 of the cases. Penicillin therapy may fail in patients with insulin-dependent diabetes or belated treatment with complications. No thromboembolic complication was observed (89 p. 100 of patients with lower limb erysipelas had received anticoagulants). There was only one death due to a severe underlying condition.

Administration, Oral↗
PubMed1990

Treatment of the acquired immune deficiency syndrome-related Kaposi's sarcoma with bleomycin as a single agent.

A nonrandomized trial was conducted to assess the efficiency and toxicity of bleomycin as a single agent in treatment of non-life-threatening AIDS-related Kaposi's sarcoma (KS). Sixty patients were enrolled in this study. They all had a disseminated and progressive non-life-threatening AIDS-related KS associated with systemic symptoms and/or CD4 lymphocyte count less than 400/mm3. Thirty patients were treated with intramuscular bleomycin (5 mg/d for 3 days every 2 or 3 weeks) and 30 others with a slow continuous intravenous infusion of bleomycin (6 mg/m3/d for 4 days every 4 weeks). The mean duration of therapy was 5 months (range, 2 to 24 months). A partial response was observed in 29 patients (48.3%) and the disease was stabilized in 18 additional patients (30%). Bleomycin failed in 21.6% of patients. Therapy had to be discontinued in two patients because of side effects. Thus bleomycin as a single agent is a good alternative therapy for AIDS-related KS.

Acquired Immunodeficiency Syndrome↗
PubMed1990

Site specific radioiodination of recombinant hirudin.

Recombinant hirudin variant rHV2-Lys47 was radioiodinated using the chloramine-T method. Depending on the reaction pH, the two tyrosine residues, Tyr3 and Tyr63, responded differently to iodination but without change in total iodination yield. Of the incorporated -125 iodine 80% was located on Tyr3 at pH 7.4, but 65% was found on Tyr63 at pH 4. These distinct iodination patterns suggest the existence of a pH-dependent multimerization and/or important conformational changes in the tertiary structure with pH. Each radiotracer was purified to high specific activity by simple low-pressure chromatography including gel filtration and reverse-phase separation, both on short cartridges. The method was validated by reverse-phase and anion-exchange HPLC with on-line radioactivity detection. The iodination sites were characterized following carboxypeptidase Y cleavage coupled with radio-HPLC.

Amino Acid Sequence↗
PubMed1990

[Congenital mitral and tricuspid double stenosis].

The authors report the case of a child followed up from the age of 4 months to the age of 15 for asymptomatic congenital heart disease. Initial clinical examination showed the presence of a systolic murmur and a diastolic rumble in the xiphoid area with signs of right atrial dilatation-hypertrophy. During follow-up, an apical diastolic rumble was detected with left atrial dilatation from the age of 3. Doppler echocardiography carried out recently showed double mitral and tricuspid valve stenosis with two-dimensional appearances of doming of both valves and moderate transvalvular pressure gradients with slight elevation of pulmonary artery pressures on the Doppler study. The very early detection of the malformation and the continuous follow-up of this patient indicate the congenital and not rheumatic origin of this double valve stenosis.

Adolescent↗