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Biomedical subjects

P Perez

Publications and source records attributed to P Perez.

At least 19 recordsLinked to original sources
PubMed2006

Use of the yeast Saccharomyces cerevisiae as a pre-screening approach for assessment of chemical-induced phototoxicity.

Photoreactive chemicals can induce dermatological reactions when present in the skin exposed to sunlight. Thus, new chemicals absorbing above 290 nm should have their potential phototoxicity tested. In order to screen a large number of molecules with various physico-chemical properties, a microbiological method is helpful. To this end, the yeast Saccharomyces cerevisiae was evaluated for its ability to detect phototoxic compounds. Twelve products known to be phototoxic in vivo and previously used as standards for validating the regulatory test 3T3 NRU were used in this work. Eleven of them could be detected in the yeast assay and, among them, 5-methoxypsoralen (5-MOP), 8-methoxypsoralen (8-MOP), angelicin and, to a lower extend, tiaprofenic acid induced genetic alterations. Interestingly, a pre-incubation with yeast cells in the dark before exposure decreased the phototoxicity of 5-MOP and 8-MOP but had no effect on this of chlorpromazine and ketoprofen. Saccharomyces cerevisiae and Salmonella typhimurium (strains TA100 and TA102) were compared for the evaluation of 5-MOP and 8-MOP photogenotoxicity; only the yeast assay allowed to perform experiments in exposure conditions close to those encountered in environmental situations. Finally, an application of this experimental approach to the detection of traces of furocoumarins in fragrance materials was developed.

5-Methoxypsoralen↗
PubMed2002

Improvement of drought tolerance in maize: towards the functional validation of the Zm-Asr1 gene and increase of water use efficiency by over-expressing C4-PEPC.

Water availability is one of the major limiting factors for plant growth. Maize is particularly sensitive to water stress at reproductive stages with a strong impairment of photosynthesis and grain filling. Here, we describe the use of genetic transformation first to assess the role of a candidate gene Asr1-a putative transcription factor-as an explanation for genetically linked drought tolerance Quantitative Trait Loci (QTLs), and second to modify CO(2) fixation rates in leaves through changes of C(4) phosphoenolpyruvate carboxylase (C(4)-PEPC) activity. Transgenic Asr1 over-expressing lines show an increase in foliar senescence under drought conditions. The highest C(4)-PEPC overexpressing line exhibited an increase (+30%) in intrinsic water use efficiency (WUE) accompanied by a dry weight increase (+20%) under moderate drought conditions. Opposite effects were observed for transgenic plants under-expressing the corresponding proteins. The data presented here indicate the feasibility to increase the level of endogenous biochemical activities related to water economy and/or drought tolerance, and opens a way to develop maize varieties more tolerant to dry growing conditions.

Acclimatization↗
PubMed2002

Multivariate analysis of determinants of bacterial contamination of whole-blood donations.

BACKGROUND AND OBJECTIVES: Introduction of bacteria into blood components at the collection stage seems to be a frequent occurrence. We therefore assessed determinants of bacterial contamination of whole-blood donations to gain insight into contamination mechanisms and direct prevention. MATERIALS AND METHODS: A cross-sectional study was carried out on donors accepted for whole-blood donation in four French blood banks. Each blood bank used its own two-stage procedure for phlebotomy site preparation. Contamination was identified by culturing two 15-ml samples (collected aseptically at the outset of donation) in a BacT/Alert 240 system. Determinants were assessed by logistic regression analysis. RESULTS: Bacterial contamination, mainly by skin flora, occurred in 76 (2.2%) out of 3385 donations. Significant determinants were as follows: the blood bank (odds ratio [OR] range = 3.0-5.6, P < 0.001); lack of repetition of scrub (OR = 2.7, P = 0.032); and donor age > 35 years (OR = 1.8, P = 0.036). CONCLUSION: Systematic scrub repetition should be implemented to reduce bacterial contamination by skin flora at the collection stage. Further research is required to clarify the role of different antiseptic agents and of donor age.

Adult↗
PubMed2002

Manipulating PEPC levels in plants.

This review examines the current understanding of the structural, functional and regulatory properties of C4 and C3 forms of higher plant phosphoenolpyruvate carboxylase. The emphasis is on the interactive metabolic and post-translational controls acting on the enzyme in the physiological context of C4 photosynthesis and the anaplerotic pathway. A brief overview is given concerning the recent developments of PEPC-based genetic engineering of C3 plants with the aim of improving photosynthetic performance in normal and limiting environmental conditions. So far, in spite of achieving a considerable increase in PEPC levels, more work needs to be done with respect to the correct dosage and location before that goal is reached. Some unpublished results on the transformation of maize with a sorghum C4 PEPC cDNA are also presented. They show that it is possible to increase photosynthetic PEPC levels in this C4 plant and that the modification in enzyme content has a pleiotropic physiological impact and, notably, an improved water use efficiency when water is limited.

Biomass↗
PubMed2001

Sensitivity of the ProC global assay for protein C pathway abnormalities. clinical experience in 899 unselected patients with venous thromboembolism.

The ProC Global is a clotting assay designed to globally evaluate the functionality of the protein C (PC) pathway, which is found defective in up to 30% of the Caucasian patients with thrombophilia. It is based on the ability of endogenous activated protein C (APC), generated by activation of PC by a snake venom extract, to prolong an activated partial thromboplastin time (APTT). This retrospective study was carried out to evaluate the ability of this assay to distinguish patients with or without any PC pathway abnormalities in a cohort of 899 unselected patients with a history of thrombosis. The result of the ProC Global assay, expressed in PC activation time-normalized ratio (PCAT-NR), was significantly lower in patients in whom was previously demonstrated an abnormality of the PC pathway compared to those without. The cut-off level of PCAT-NR=0.75 was found to provide the best sensitivity-specificity ratio, since all the patients with the factor V (FV) Leiden mutation (n=71), APC resistance (n=3), PC deficiency (n=22), or combined defects (n=19) had a PCAT-NR below that value. The sensitivity of the ProC Global assay for a low protein S (PS) level (n=56) was only 66%, and was even weaker in the case of hereditary PS deficiency (46.6%, n=15). The assay did not perform well in samples from patients on oral anticoagulant treatment (OAT, n=64) or with liver failure (n=4), as the PCAT-NR was reduced in most cases, even in the absence of any abnormality. These results suggest that the ProC Global assay could be validly used as a first-step screening test for the FV Leiden-related APC resistance and PC deficiency in patients not on OAT. Given the moderate sensitivity of the assay for PS deficiency, this coagulation inhibitor must be determined in every case. However, the overall benefit of such a screening strategy is limited since more than 38% of the 659 patients without abnormality had a decreased PCAT-NR.

Adolescent↗
PubMed2001

The cytoplasmic and transmembrane domains of the p75 and Trk A receptors regulate high affinity binding to nerve growth factor.

Ligand-induced receptor oligomerization is an established mechanism for receptor-tyrosine kinase activation. However, numerous receptor-tyrosine kinases are expressed in multicomponent complexes with other receptors that may signal independently or alter the binding characteristics of the receptor-tyrosine kinase. Nerve growth factor (NGF) interacts with two structurally unrelated receptors, the Trk A receptor-tyrosine kinase and p75, a tumor necrosis factor receptor family member. Each receptor binds independently to NGF with predominantly low affinity (K(d) = 10(-9) m), but they produce high affinity binding sites (K(d) = 10(-11) m) upon receptor co-expression. Here we provide evidence that the number of high affinity sites is regulated by the ratio of the two receptors and by specific domains of Trk A and p75. Co-expression of Trk A containing mutant transmembrane or cytoplasmic domains with p75 yielded reduced numbers of high affinity binding sites. Similarly, co-expression of mutant p75 containing altered transmembrane and cytoplasmic domains with Trk A also resulted in predominantly low affinity binding sites. Surprisingly, extracellular domain mutations of p75 that abolished NGF binding still generated high affinity binding with Trk A. These results indicate that the transmembrane and cytoplasmic domains of Trk A and p75 are responsible for high affinity site formation and suggest that p75 alters the conformation of Trk A to generate high affinity NGF binding.

Animals↗
PubMed2001

p75 Co-receptors regulate ligand-dependent and ligand-independent Trk receptor activation, in part by altering Trk docking subdomains.

Neurotrophins signal via Trk tyrosine kinase receptors and a common receptor called p75. Nerve growth factor is the cognate ligand for TrkA, brain-derived neurotrophic factor for TrkB, and neurotrophin-3 (NT-3) for TrkC. NT-3 also binds TrkA and TrkB as a heterologous ligand. All neurotrophins bind p75, which regulates ligand affinity and Trk signals. Trk extracellular domain has five subdomains: a leucine-rich motif, two cysteine-rich clusters, and immunoglobulin-like subdomains IgG-C1 and IgG-C2. The IgG-C1 subdomain is surface exposed in the tertiary structure and regulates ligand-independent activation. The IgG-C2 subdomain is less exposed but regulates cognate ligand binding and Trk activation. NT-3 as a heterologous ligand of TrkA and TrkB optimally requires the IgG-C2 but also binds other subdomains of these receptors. When p75 is co-expressed, major changes are observed; NGF-TrkA activation can occur also via the cysteine 1 subdomain, and brain-derived neurotrophic factor-TrkB activation requires the TrkB leucine-rich motif and cysteine 2 subdomains. We propose a two-site model of Trk binding and activation, regulated conformationally by the IgG-C1 subdomain. Moreover, p75 affects Trk subdomain utilization in ligand-dependent activation, possibly by conformational or allosteric control.

Brain-Derived Neurotrophic Factor↗
PubMed2001

Association of Trk neurotrophin receptors with components of the cytoplasmic dynein motor.

Nerve growth factor (NGF) initiates its trophic effects by long-range signaling through binding, internalization, and transport of a ligand-receptor complex from the axon terminal to the cell body. However, the mechanism by which retrograde transport of NGF takes place has not been elucidated. Here we describe an interaction between the Trk receptor tyrosine kinase and a 14 kDa light chain of cytoplasmic dynein. After transfection in human embryonic kidney 293 cells, this 14 kDa dynein light chain was found to bind to TrkA, TrkB, and TrkC receptors. Mapping experiments indicated that the 14 kDa dynein light chain binds to the distal region of the TrkA juxtamembrane domain. Coimmunoprecipitation experiments in vivo indicate that Trk receptors are in a complex with the 14 kDa light chain and 74 kDa intermediate chain of dynein. Confirming the physiological relevance of this association, a marked accumulation of Trk with the 14 kDa and the 74 kDa dynein components was observed after ligation of the sciatic nerve. The association of Trk receptors with components of cytoplasmic dynein suggests that transport of neurotrophins during vesicular trafficking may occur through a direct interaction of the Trk receptor with the dynein motor machinery.

Animals↗
PubMed2001

Fluoroquinolones as chemical tools to define a strategy for photogenotoxicity in vitro assessment.

Today's lifestyle is often associated with frequent exposure to sunlight, but some xenobiotics used in drugs, cosmetics or food chemicals can produce adverse biological effects when irradiated. In particular, they can increase the risk of photogenotoxicity already due to UV radiation itself. There is thus a need to design appropriate approaches in order to obtain relevant data at the molecular and cellular level in this field. For ethical and practical reasons, in vitro models can be very convenient at least for first evaluation tests. Here, we propose a strategy based on complementary experiments to study the photogenotoxic potential of a compound. The fluoroquinolones BAYy3118 and lomefloxacin were used as standards to demonstrate the performance of each test: photoinduced interaction with supercoiled circular DNA, photomutagenicity in the yeast Saccharomyces cerevisae, induction of DNA photodamage in cultured human skin cells as revealed by comet assay, and finally induction of specific phototoxic stress responses such as p53 activation or melanogenesis stimulation. Such a strategy should help to ensure the safety of products likely to undergo environmental sunlight exposure.

Anti-Infective Agents↗
PubMed2001

Efficacy of a new collection procedure for preventing bacterial contamination of whole-blood donations.

BACKGROUND: Transfusion-related bacterial contamination is a serious problem. The introduction of bacteria into donations at the collection stage seems frequent, despite well-conducted phlebotomy site preparation. Additional preventive measures are required. STUDY DESIGN AND METHODS: The aim of this study was to assess the potential efficacy of excluding the first 15 mL of blood to reduce the bacterial contamination of donations. A special device allowed the aseptic collection of two samples at the beginning of donation: S1 (first 15 mL) and S2 (next 15 mL). Bacteriologic cultures of S1 and S2 were performed by using an automated system. The procedure's efficacy was measured by the proportion of positive donations in S1 that were then negative in S2. RESULTS: S1 and/or S2 were positive in 76 (2.2%) of 3385 donations. In about three-fourths of the culture-positive donations, contamination was detected in the first 15-mL sample only. Gram-positive cocci accounted for 81 percent of species, gram-positive bacilli for 14 percent, and gram-negative bacilli for 5 percent. The new procedure would have prevented the introduction of bacteria in 55 donations, reducing to 0.6 percent the risk of contamination from the first 15 mL collected. CONCLUSION: Although the final effect on blood component bacterial contamination rates cannot be derived from the study, excluding the first 15 mL of blood may reduce the rate of bacterial contamination in donations.

Adult↗
PubMed2001

Determinants of transfusion-associated bacterial contamination: results of the French BACTHEM Case-Control Study.

BACKGROUND: Transfusion-associated bacterial contamination (TABC), probably the most frequent transfusion-transmitted infection, may induce serious adverse events. Systematic information and documentation on determinants are lacking. STUDY DESIGN AND METHODS: The BACTHEM Study is a French matched case-control study assessing TABC determinants. Included were cases of TABC reported in France in a 2-year period, as determined from uniform definitions. Information on recipient-, blood component-, and donor-related potential determinants was collected on site. ORs were estimated by conditional logistic regression. RESULTS: Of the 158 cases of suspected TABC reported, 41 that involved transfusion with 25 RBCs and 16 platelet concentrates were included. Gram-negative rods accounted for nearly half of the bacteria species involved and for all six deaths. In comparison with the risk of TABC for patients receiving RBCs for anemia, the risk was higher for patients receiving RBCs for pancytopenia (OR, 7.3; 95% CI, 1.3-41.0) and for those receiving platelets for thrombocytopenia (OR, 5.3; 95% CI, 1.2-24.1). Other potential determinants were platelet transfusion for pancytopenia (OR, 4.5; 95% CI, 0.5-40.0), immunosuppressive treatment (OR, 2.8; 95% CI, 0.7-10.6), shelf-life of more than 1 day for platelets or 8 days for RBCs (OR, 2.6; 95% CI, 0.7-9.6), and more than 20 previous donations by donors (OR, 1.9; 95% CI, 0.7-5.3). CONCLUSION: This first comparative study revealed TABC determinants that suggest approaches for prevention.

Adult↗
PubMed2001

Coordination between fission yeast glucan formation and growth requires a sphingolipase activity.

css1 mutants display a novel defect in Schizosaccharomyces pombe cell wall formation. The mutant cells are temperature-sensitive and accumulate large deposits of material that stain with calcofluor and aniline blue in their periplasmic space. Biochemical analyses of this material indicate that it consists of alpha- and beta-glucans in the same ratio as found in cell walls of wild-type S. pombe. Strikingly, the glucan deposits in css1 mutant cells do not affect their overall morphology. The cells remain rod shaped, and the thickness of their walls is unaltered. Css1p is an essential protein related to mammalian neutral sphingomyelinase and is responsible for the inositolphosphosphingolipid-phospholipase C activity observed in S. pombe membranes. Furthermore, expression of css1(+) can compensate for loss of ISC1, the enzyme responsible for this activity in Saccharomyces cerevisiae membranes. Css1p localizes to the entire plasma membrane and secretory pathway; a C-terminal fragment of Css1p, predicted to encode a single membrane-spanning segment, is sufficient to direct membrane localization of the heterologous protein, GFP. Our results predict the existence of an enzyme(s) or process(es) essential for the coordination of S. pombe cell wall formation and division that is, in turn, regulated by a sphingolipid metabolite.

Amino Acid Sequence↗
PubMed2000

Esr genes show different levels of expression in the same region of maize endosperm.

Esr genes share high homology among each other, code for small hydrophilic proteins, and are expressed in a restricted region of maize endosperm surrounding the embryo. We show here that not only Esr2 but also Esr1 and Esr3 are expressed in maize, and that the relative contribution of Esr1, Esr2 and Esr3 to total Esr mRNA is 17%, 55% and 28%, respectively. DNA sequence analysis of putative promoter fragments ranging from 0.53 kb to 3.54 kb revealed the presence of retrotransposons related to the Zeon and Cinful families in the distal parts of the promoters. The proximal parts show high homology that extended over 504bp between Esr2 and Esr3, and 265bp between Esr1 and the other two genes. The most conspicuous potential cis element is a fully conserved tandem repeat of the sequence CTACACCA close to the respective open reading frames (ORFs). By the analysis of transgenic maize plants carrying promoter-Gus fusions, it was shown that all three cloned upstream fragments contain functional promoters, that the spatial activity of all three Esr promoters is identical, and that the cis element(s) responsible for the expression in the embryo surrounding region reside in the 265 bp upstream of the respective ORFs.

Base Sequence↗
PubMed2000

Screening for risk factors for thrombosis using a new generation of assays developed to evaluate the functionality of the protein C anticoagulant pathway.

These results, obtained in a small series of patients, suggest that both the ProC Global assay and the PCP Test would be suitable, using well-defined cut-off levels, to identify all the carriers of the Factor V Leiden mutation and all the patients with a protein C deficiency or with combined defects of the protein C pathway. For both assays, however, the sensitivity for protein S deficiency was below 60%. These results in selected patients are congruent with those previously reported in the literature about the ProC Global assay, the PCP Test, and other assays evaluating the functionality of the protein C anticoagulant pathway. All demonstrated a weak sensitivity to protein S deficiency, suggesting that protein S plays only a minor role as an APC cofactor in such global assays. A major discrepancy between the two evaluated assays was obtained in the group of patients without abnormality of the protein C pathway. Actually, using the ProC Global assay, more than 40% of the patients had a decreased PCAT-NR while presenting with none of the three tested abnormalities, whereas none of the studied patients had a ratio below 1.80, and only 5 of 143 (3.5%) had a ratio below 2.00 when using the PCP Test. The observation that around 40% of the control patients had a decreased PCAT-NR could suggest the influence of currently unknown defects of the protein C/protein S pathway on the ProC Global assay. It could also be hypothesized that the higher factor VIII levels already reported in patients with a history of thrombosis than in controls had a significant role in the low responsiveness, but this parameter was not tested in the authors' series. In that connection, it is also well established that elevated factor VIII levels both shorten the APTT and reduce the anticoagulant effect of heparin when evaluated using APTT. Actually, some of the samples investigated in this study were obtained during the acute phase of thrombosis. It is not possible to draw out the hypothesis of an association of biologically undetectable minor changes in various factors involved in the protein C anticoagulant pathway; all the individual factors would remain within their normal ranges. Finally, because 40% of the patients without abnormalities of the protein C pathway had a decreased PCAT-NR, the question arises whether the ProC Global assay might in itself be a biologic marker of thrombophilia, independent of its sensitivity for abnormalities of the protein C anticoagulant pathway. In that connection, the correlation between the result of the ProC Global assay and the risk for thromboembolism was recently evaluated by two different groups. In both cases, the preliminary results suggested that a decreased response to the ProC Global assay might be an independent risk factor for venous thrombosis. The two global assays could therefore have distinctly different applications. If the global assays are used in the hemostasis laboratory to screen for abnormalities of the protein C pathway, and thus to rationalize the use of specific assays, the PCP Test should be chosen, because of its high specificity. Because only 3.5% of the control patients had a ratio below 2.00 (and none had a ratio below 1.80), the PCP Test could be accurately used as a first-step assay in the laboratory screening for these abnormalities of the protein C anticoagulant pathway. Using such a flow chart, the specific assays for APC resistance or the identification of the factor V Leiden mutation and protein C would be performed only in case of a ratio below a cut-off defined using receiver operating characteristic (ROC)-analysis in unselected patients. Because of the weak sensitivity of this assay to both constitutional and acquired protein S deficiencies (below 15% using 1.80 as the cut-off level, or 60% using 2.00), the measurement of this parameter had to be performed in all cases. If, on the other hand, the assay is used to screen for risk factors for thrombosis, the ProC Global assay could b

Adolescent↗