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Biomedical subjects

G Fabre

Publications and source records attributed to G Fabre.

At least 73 records · Page 4Linked to original sources
PubMed1983

A new spin label method for the measurement of erythrocyte internal microviscosity.

A new spin-label method for the measurement of the internal microviscosity of erythrocyte is presented. The spin label used is 2,2',5,5'-tetramethyl-3-maleimidopyrrolidinyl-N-oxyl (MAL-5) which penetrates inside the red blood cell and binds covalently on cytoplasmic glutathione. After washing off the external label, 98% of the electron paramagnetic signal is due to the labelled glutathione. This signal allows one to measure the rotational correlation time of the label. A calibration curve established with spin-labelled glutathione in sucrose solutions of increasing viscosity is used to convert the measured rotation times into viscosity units. This method avoids the use of unphysiological salts like potassium ferricyanide, and permits the study of red blood cells in various suspension media. In normal human subjects, the mean value of microviscosity is 4.45 +/- 0.16 mPa . s at 20 degrees C in isotonic saline (25 subjects) and 6 +/- 0.25 mPa . s in plasma. The variations of microviscosity as a function of the osmolarity of the medium are explained according to a theoretical model taking into account the variations of the red blood cell volume and the viscometric properties of haemoglobin.

Adolescent↗
PubMed1983

High-performance liquid chromatographic analysis of MTX, 7-OH-MTX and MTX derivatives: application to intracellular metabolism in tumor cells (HT 29).

In order to study the intracellular metabolism of Methotrexate (MTX) and the cytotoxicity of the antifolates, a specific paired-ion HPLC method has been developed which permits the simultaneous determination of DAMPA, MTX, 7-OH-MTX, MTX-G1, MTX-G2 and MTX-G3. Cells were incubated with 3H-MTX. The MTX metabolites were extracted, purified on SEP-PAK cartridges and further analysed by high-performance liquid chromatography (HPLC). The stationary phase was constituted by a C18 muBondapak and the mobile phase by 5 mM phosphate buffer (pH 7.4) containing 2.5 mM tetrabutylammonium nitrate. The elution was performed with a linear methanol gradient (20--30%). HPLC fractions were collected and radioactivity evaluated by beta counting (retention times: DAMPA = 12.93 min; MTX = 18.29 min; 7-OH-MTX = 21.13 min; MTX-G1 = 22.69 min; MTX-G2 = 26.81 min; MTX-G3 = 30.61 min). This analytical procedure was applied to separate and characterize multiple forms of MTX polyglutamate derivatives in HT 29, a human adenocarcinoma cell line varying the incubation time (4--18 h) and MTX concentration (0.6--10.6 micrometer). The incorporation process seems to be characteristic of a cell line resistant to MTX. The incorporation were very low and after a 4-h exposure time only 5% of the MTX was converted to polyglutamates. Between 1.6 and 10.6 microM MTX, no difference was observed in the polyglutamization. The defect in the incorporation of the drug and in the metabolization process in vitro could partially explain the failure of the MTX treatment in colorectal cancer.

Adenocarcinoma↗
PubMed1983

[Changes in the rheologic properties of various samples of blood preserved at +4 degrees C].

Apparent stationary viscosity, viscoelasticity and thixotropy measured with a Couette viscosimeter have been determined on blood samples stored in eight anticoagulant and/or preservative solutions. All the results as well as those obtained in a previous morphological study show clearly the advantages of the SAG or PAGGS storage processes. It is observed that the rheological parameters of RBC stored in protein poor media are, between the 30 th and 35 th days, identical on CPD at the first week and with that measured in CPD concentrates between second and third week (for a same hematocrit value). We want to emphasize that the erythrocytes stored in these new media retain for a long time their ability to circulate in the capillary system.

Adenine↗
PubMed1983

In vitro formation of polyglutamyl derivatives of methotrexate and 7-hydroxymethotrexate in human lymphoblastic leukemia cells.

The intracellular synthesis of polyglutamyl derivatives of both methotrexate (4-amino-N-10-methylpteroylglutamic acid) and 7-hydroxymethotrexate, the primary plasma metabolite of methotrexate in humans, was evaluated in a methotrexate-sensitive, acute lymphoblastic leukemia cell line, MOLT 4. These studies were performed using a highly specific ion-pairing high-pressure liquid chromatography method which permits the simultaneous determination of methotrexate, 7-hydroxymethotrexate, and their corresponding polyglutamyl derivatives. When MOLT 4 cells were exposed to 1 microM methotrexate, the monoglutamate attained a steady state after 30 min, and polyglutamyl derivatives having from one to 4 additional glutamyl residues were observed over 4 hr. Four additional metabolites were also detected upon incubation with 1 microM 7-hydroxymethotrexate. On the basis of the retention times for these compounds relative to methotrexate polyglutamyl standards and since these metabolites reverted to 7-hydroxymethotrexate upon treatment with a preparation of hog kidney conjugase, they were identified as polyglutamyl derivatives of 7-hydroxymethotrexate. The identification of 7-hydroxymethotrexate polyglutamyl derivatives in vitro raises the possibility of an important new dimension in the pharmacological action of methotrexate. We investigated the effect of extracellular 7-hydroxymethotrexate on net methotrexate uptake and metabolism when cells were exposed simultaneously to 1 microM [3H]-methotrexate and unlabeled 7-hydroxymethotrexate. A decrease in the levels of both intracellular methotrexate and the corresponding polyglutamyl derivatives was noted for cells treated with 1 or 10 microM 7-hydroxymethotrexate. However, no appreciable effect of 7-hydroxymethotrexate on the amount of polyglutamyl derivatives formed relative to the total intracellular antifolate was noted. These studies show that in MOLT 4 cells (a) both methotrexate and 7-hydroxymethotrexate are rapidly converted to polyglutamyl derivatives, and (b) 7-hydroxymethotrexate interferes with net methotrexate accumulation and metabolism when present simultaneously in the extracellular medium. These results, moreover, suggest a potential role for 7-hydroxymethotrexate in modulating the biochemical effects of methotrexate in vivo.

Cell Line↗
PubMed1983

In vitro approach to 7-hydroxymethotrexate interaction on methotrexate metabolism as tool of pharmacokinetic study.

Methotrexate (MTX), a widely used antineoplastic agent, is metabolized to a non-active derivative, 7-OH-MTX, and to some active poly-gamma-glutamyl derivatives (MTX-Gn) which are retained within cells. Pharmacokinetic studies in humans indicate (i) a higher concentration of 7-OH-MTX than of MTX in plasma after a 24-h infusion and (ii) a time-dependent relationship for MTX and 7-OH-MTX kinetics in plasma and urine which might be explained by the variation of MTX metabolism. The intracellular metabolism of MTX and 7-OH-MTX has been investigated using a specific ion-paired method of high-performance liquid chromatography (HPLC) which permits the simultaneous determination of DAMPA, MTX, 7-OH-MTX and their respective polyglutamate derivatives. The formation of 7-OH-MTX polyglutamates and the possible effect of 7-OH-MTX on the transport and/or metabolism of the unchanged MTX in a human acute lymphoblastic leukaemia cell line (Molt 4) have been studied. After incubation of the cells to 1 microM (3H) 7-OH-MTX, four radiolabelled peaks, representing 75% of the intracellular 3H, were converted to 7-OH-MTX upon exposure to hog kidney conjugase indicating the formation of 7-OH-MTX polyglutamyl derivatives. The effects of 7-OH-MTX on MTX-PG formation were analysed after simultaneous incubation of the cells to 1 microM (3H) MTX and 10 microM 7-OH-MTX. The formation of the higher glutamyl derivatives, MTX-G3 and MTX-G4 was completely inhibited and the total intracellular accumulation of the MTX-PG's decreased by 35% compared to control. These data suggest that the 7-OH-MTX and the 7-OH-MTX-PG might modify the chemotherapeutic activity of this agent in vivo.

Cell Line↗
PubMed1982

[Comparative morphological study of erythrocytes in various types of whole blood and packed cell preparations stored in liquid state].

Morphological comparative study observed by scanning electron microscopy have been made on three blood stored in PVC plastic bags at 4 degrees C: 1. Whole blood in CPD, CPD-A,D (CPD added with 0,4 mM adenine and enriched with glucose 1,5 time the normal concentration) and ACD-Ad,Gua (ACD with 0,5 mM adenine and 0,5 mM guanosine); 2. Red blood cells obtained by centrifugation from blood collected in CPD, CPD-A,D and ACD-Ad, Gua; 3. Red blood cells from blood collected in CPD or ACD and resuspended in balanced, nutritious and enriched media: SAG (Saline, Adenine, Glucose) or PAGGS (Phosphate, Adenine, Guanosine, Glucose, Saline) (CPD/SAG, ACD and CPD/PAGGS). The results of statistical analysis show that the addition of adenine and guanosine in the preservative solutions improves the morphological quality of erythrocytes. On this view the resuspended media are the least traumatic. For example, in the concentrates CPD/PAGGS the level of normal or practically shape cells (discocytes and echinocytes I) nears 70 p. cent at the 6th week of storage. Thus the PAGGS system appears superior to SAG system.

Adenine↗
PubMed1981

[Transient rheological study of blood stored in a liquid state].

The rheological properties of blood stored in different conditions (in liquid state or as globular concentrates in the presence of the classical anticoagulant ACD or CPD solutions, or according to the SAG procedure) have beenn studied with a Couette viscosimeter. Apparent viscosity was measured at very low shear stress (0.05 s-1) as a function of the storage time. Furthemore the dynamic rheological parameters (thixotropy, viscoelasticity and rheofluidification) were determined. Blood viscosity increases strongly as a function of storage time, particularly on whole blood. Its variation is weaker on globular concentrates, particularly with SAG. Thixotropy, viscoelasticity and rheofluidification, on the contrary, are not strongly perturbated. These rheological modifications are closely related with the changaes occuring in the morphology and in the aggregation state of the stored red blood cells.

Adult↗
PubMed1977

[Technics for the study of a total leukocyte dialysate and its fractions].

The crude dialysate extracted from human white blood cells from a thousand unselected donors were studied by leukocyte migration inhibition test with measles antigen. This dialysate and its fractions eluted from Biogel P4 and P6 chromatography were compared towards rosette restoration test and nucleotide composition.

Cell Migration Inhibition↗
PubMedDate not supplied

Human hepatocytes as a key in vitro model to improve preclinical drug development.

Over past decades, numerous in vitro and/or ex vivo models have been developed to investigate drug metabolism. In the order of complexity we found the isolated perfused liver, hepatocytes in co-culture with epithelial cells, hepatocytes in suspension and in primary culture and subcellular hepatic microsomal fractions. Because they can be easily prepared from both animals (pharmacological and toxicological species) and humans (whole livers as well as biopsies obtained during surgery) hepatocytes in primary culture provide the most powerful model to better elucidate drug behavior at an early stage of preclinical development such as: the characterization of main biotransformation reactions, the identification of phase I and phase II isozymes involved in such reactions, the evaluation of inter-species differences allowing the selection of a second toxicological animal species more closely related to man on the basis of metabolic profiles, the detection of the inducing and/or inhibitory effects of a drug on metabolic enzymes, the prediction of drug interactions, the estimation of inter-individual variability in biotransformation reactions. The use of hepatocytes, and in particular those obtained from humans, at an early stage of drug development allows the obtention of more predictive preclinical data and a better knowledge of drug behavior in humans before the first administration of the drug in healthy volunteers.

Animals↗