Biomedical subjects

F Morel

Publications and source records attributed to F Morel.

At least 109 records · Page 6Linked to original sources

PubMed1995

Changes in the respiratory chain complexes activities and in the mitochondrial DNA content during ageing in D. subobscura.

The time course (age 0-8 weeks) of the enzyme activities of respiratory chain complexes I, III and IV and of citrate synthase, and the cell mitochondrial/nuclear DNA content ratio were studied in Drosophila subobscura. The activities of the three respiratory complexes decreased with age, but with different kinetics. The activities of complexes I and III remained nearly stable between weeks 0 and 3 (falling by 6% and 15%, respectively), and then gradually decreased; after 8 weeks residual activities were about 50% of the initial value for complexes I and III. The activity of complex IV fell in the first week, decreasing continually to week 8, where residual activity was 30% of the initial value. No significant age-related change in citrate synthase activity was observed. Mitochondrial DNA (measured by mitDNA/nucDNA) increased linearly up to week 5 (2.6-fold) and then dropped by 40% in week 6 though it remained higher than initial values.

Aging↗

PubMed1995

Turnover of glutathione S-transferase alpha mRNAs is accelerated by 12-O-tetradecanoyl phorbol-13-acetate in human hepatoma and colon carcinoma cell lines.

The phorbol ester, 12-O-tetradecanoyl phorbol-13-acetate (TPA), known to induce murine glutathione S-transferase (GST) Ya, was examined for its effect on the expression of human GST alpha. Unexpectedly, 24-h treatment of the human hepatoma cell line HepG2 with 100 nmol/l TPA caused a decrease of the GST alpha mRNA level to below 5% of controls, i.e. opposite to the known response in the mouse. The level of mRNA for GST Mu was also decreased, but the mRNAs of c-jun and jun-B were elevated after 2 h. The decrease of GST alpha mRNAs was inhibited by staurosporine, suggesting an involvement of protein kinase C. Inhibition of transcription and translation by actinomycin D and cycloheximide also partially inhibited the effect of TPA on the expression of GST alpha. In the presence of actinomycin D, GST alpha mRNA halflife was 14.5 h, compared to 3.5 h in the presence of TPA. The calcium ionophore A23187 caused a loss of GST alpha mRNAs to levels almost as low as those obtained with TPA. The effects of TPA and the calcium ionophore were also observed in CaCo2 colon carcinoma cells. As a consequence of the decrease of mRNA levels, GST alpha protein levels and total GST enzyme activity were also diminished. Also, the morphology of the cells was changed after 3 h exposure to TPA. These data suggest that human GST alpha expression can be regulated at the level of mRNA stability by a pathway involving protein kinase C.

Animals↗

PubMed1995

A mutation located at the 5' splice junction sequence of intron 3 in the p67phox gene causes the lack of p67phox mRNA in a patient with chronic granulomatous disease.

Chronic granulomatous disease (CGD) is due to a functional defect of the O2(-)-generating NADPH oxidase of neutrophils. Mutations resulting in CGD have been shown to occur in only four genes, thus identifying the main components of the oxidase complex, namely the two subunits of a membrane-bound cytochrome b and two cytosolic factors of activation of 67 kD (p67phox) and 47 kD (p47phox). The present study deals with the biochemical and genetic analysis of the defect in a patient suffering from a p67phox-deficient form of CGD. The p67phox deficiency was ascertained by immunochemistry and the ability of recombinant p67phox to restore NADPH oxidase activity using a cell-free system of oxidase activation. The cellular extracts from the proband contained no p67phox protein and no p67phox mRNA when assayed by Western and Northern blot analysis. However, reverse transcription of mRNA and subsequent cDNA amplification by polymerase chain reaction using specific p67phox primers showed that trace amounts of a p67phox mRNA deleted for exon 3 were synthesized in the patient immortalized B lymphocytes. Sequence analysis of the genomic DNA showed a T-to-C transition at position +2 of intron 3. This point mutation in the consensus 5' splice site of the intron 3 was probably responsible for lack of accumulation of mRNA and also for the skipping of exon 3 detected in the few mRNA molecules that escaped cellular degradation.

Adult↗

PubMed1995

Sites of arginine synthesis and urea production along the nephron of a desert rodent species, Meriones shawi.

The distribution of arginine synthase and arginase activities along the successive nephron segments of Meriones kidney was measured in vitro with single tubule enzymatic microtechniques making use of either L-[ureido-14C] Citrulline (0.108 mM) or L-[guanidino-14C]arginine (0.2 mM) as the respective substrates. Arginase activity (fmol urea formed per min per mm of tubule) was very low (5-25 fmol.min-1.mm-1) in most nephron segments including the early portions of proximal convoluted tubules (early PCT). It increased progressively after 3 mm of the PCT to reach a value of 200 fmol.min-1.mm-1 in the cortical portion of the straight proximal tubule (CPST), with a further increase, along the pars recta, of up to 250 fmol.min-1.mm-1 in the outer medullary portion (OSPST). In addition, arginase activity in OSPST and the adjacent descending thin limb (DTL) was higher in juxtamedullary nephrons (JN) than in the corresponding portions of superficial nephrons (SN). Arginine synthase activity (fmol arginine formed per mm of tubule per min) was present in proximal tubules exclusively, with a gradient decreasing along the PCT (about 600 fmol.min-1.mm-1 in the 1st mm, 65 fmol.min-1.mm-1 in CPST and 30 fmol.min-1.mm-1 in OSPST). It has been checked that CPST and OSPST (where the two enzyme systems are present) are able to convert citrulline directly into urea with a yield of 65%.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

[Temporomandibular joint luxation and Ehlers-Danlos disease. Apropos of a case].

First described by Tschernogobow in 1981, Ehlers-Danlos syndrome is usually observed in white males. Symptoms results from defective collagen synthesis. Diagnosis is based on clinical presentation. There are 9 different clinical groups. Maxillofacial manifestations are usually seen in type VIII Ehlers-Danlos syndrome. The clinical case presented here illustrates the problems involving the temporomandibular joints and focuses on an assessment of proposed therapeutic options.

Adolescent↗

PubMed1994

Further characterization of the gelatinase-containing particles of human neutrophils.

In addition to azurophil and specific granules, a third storage compartment is known to exist in the neutrophils. This compartment which consists of morphologically heterogeneous particles is characterized by a high specific activity in gelatinase. A gelatinase enriched fraction was prepared by subcellular fractionation of neutrophil homogenates using rate zonal centrifugation. This fraction was enriched in diamine oxidase. Among the proteins released from the neutrophils upon stimulation by formyl peptides, those belonging to the gelatinase enriched fraction were determined after removal of the proteins from specific and azurophil granules by selective immunoadsorption. Gelatinase was recovered together with tetranectin, beta 2-microglobulin and diamine oxidase in the same fraction. Differences in the kinetics of release of gelatinase and diamine oxidase versus vitamin B-12-binding protein suggest that the proteins belong to distinct subcellular structures.

Amine Oxidase (Copper-Containing)↗

PubMed1994

Choice of fixation and denaturation for the triple labelling of intra-cytoplasmic antigen, bromodeoxyuridine and DNA. Application to bone marrow plasma cells.

A triple staining method of intra-cytoplasmic antigen, bromodeoxyuridine (BrdU), and DNA for fluorescence image analysis is described. Several kinds of fixation and DNA denaturation methods were tested to obtain a technique suitable for heterogeneous tissues. The model chosen was the analysis of plasma cells in bone marrow. The fluorochromes used were fluorescein isothiocyanate (FITC) for intra-cytoplasmic antigens (light chain immunoglobulins), aminomethylcoumarin acetic acid (AMCA) for BrdU, and propidium iodide (PI) for DNA. The quality of the staining was analysed according to: (1) cell morphology with a good preservation of the chromatin structure, (2) intensity of light chains and of BrdU labelling, and (3) the quality of DNA staining judged from a DNA histogram. For most of the analysed tissues, fixation with methanol followed by 0.5% paraformaldehyde and denaturation by an NaOH concentration adapted to the tissue gave good results. However, in our model fixation by methanol, followed by methanol/acetic acid and denaturation of DNA by 0.03 N NaOH was the solely satisfactory technique. A good correlation (P < 0.001) was found with the plasma cell BrdU labelling index obtained with our reference immuno-enzymatic technique. Quantification of DNA content showed a satisfactory G1 peak coefficient of variation (CV) in diploid cells and a 4C to 2C ratio equal to 2. With this technique, the nuclear and cytoplasmic structures of both myeloid cells and plasma cells were well preserved, while their sensitivity to DNA denaturation was quite different.

Antigens↗

PubMed1994

Urea production by kidney collecting ducts in vitro: effect of amino acid addition.

Urea production by cortical (CCD) and medullary (OMCD) collecting ducts of the rat kidney was measured in vitro by incubating single microdissected pieces of tubule in the presence of L-[guanido-14C]arginine (0.2 mM). The [14C]urea released from the cells was hydrolysed in presence of urease added to the incubation medium and the 14CO2 formed was trapped in KOH and counted. The effect of various amino acids (AA) on urea production was investigated by adding unlabelled AA (either in combination or singly) at concentrations close to those present in blood plasma. A mixture of 17 AA decreased urea production from [14C]arginine by 46% in CCD and by 58% in OMCD. When lysine and proline were omitted from the mixture, the inhibition was less marked (19% in CCD and 43% in OMCD, respectively). When AA were tested singly, lysine induced the larger inhibition (40% in CCD and 45% in OMCD), than ornithine and glutamine (about 15% each, in CCD and OMCD), whereas proline inhibition (7% in CCD, 10% in OMCD) was not statistically significant. Branched-chain amino acids (BCAA) in combination (leucine, isoleucine and valine) also markedly reduced urea production by CCD and OMCD. Their effect was dose dependent. Solubilization of CCD and OMCD cell membranes with Triton X-100 resulted in a twofold increase in urea production by control samples; the relative inhibition (per cent) induced by BCAA was enhanced, whereas that induced by lysine was decreased. The data suggest that, in living tubules, the inhibition obtained with lysine resulted, for a large part, from competition between lysine and arginine for cell uptake via a common membrane carrier, whereas the inhibition induced by BCAA corresponded to an effect on arginase activity itself.

Amino Acids↗

PubMed1994

Proliferative response of B chronic lymphocytic leukemia lymphocytes stimulated with IL2 and soluble CD23.

The in vitro proliferative response of purified B-chronic lymphocytic leukemia (B-CLL) lymphocytes cultured in the presence of soluble CD23 (sCD23) with or without IL2 was compared to the responses induced by phorbol 12-myristate 13-acetate (PMA), Staphylococcus aureus strain Cowan I (SAC), IL1, IL2, IL4, IL6 and the combination of IL2 and interferon (IFN) alpha or IFN gamma. As expected, B-CLL lymphocytes proliferated with PMA, SAC and IL2 with a clear enhancement of the IL2-induced response by IFN alpha or IFN gamma. They failed to proliferate in response to sCD23, IL1, IL4 or IL6 alone nor to the combinations of sCD23 and any of the 3 latter cytokines. However, sCD23 significantly increased the proliferation of B-CLL cells induced by IL2, suggesting a protective effect of sCD23 on apoptosis. Serum levels of sCD23 and CD23 membrane expression were high in every patient which is compatible with the hypothesis of an autocrine or paracrine activation loop. Detectable CD23 expression was lost in all cultures except for that stimulated by PMA. Only supernatants of PMA-stimulated cultures contained high sCD23 levels.

Adult↗

PubMed1994

Gene structure and regulation of expression of human glutathione S-transferases alpha.

Current knowledge on the structure of human alpha-class glutathione S-transferase (GST) genes and the regulation of their expression is reviewed. The alpha-class GST comprises several genes and pseudogenes localised in a tight cluster on chromosome 6. Although the human GSTA1 and GSTA2 genes have the same number of exons and introns as their rat and mouse counterparts, the sequences of the 5'-flanking regions of the human alpha-class genes are significantly different from the rodents, suggesting different mechanisms of regulation between human and rodents. The expression of GST alpha is altered in a variety of tumors and several lines of evidence implicate the alpha-class GSTs in chemotherapeutic drug resistance. Finally, the induction of human GSTs by drugs or nutritional constituents would justify an interest for developing chemointervention strategies in populations highly exposed to carcinogens like aflatoxin B1 and polycyclic aromatic hydrocarbons.

Animals↗

PubMed1994

Aspartate aminotransferase macroenzyme complex in serum identified and characterized.

Macromolecular aspartate aminotransferase was found in the serum of an apparently healthy patient. This complex was composed of aspartate aminotransferase (AST; EC 2.6.1.1) and immunoglobulin. Electrophoresis of the patient's serum showed an abnormal band migrating between mitochondrial (m) and cytosolic (s) AST. The macromolecular complex was purified by gel filtration on Sephacryl S300. The molecular mass of the complex was estimated to be 250 kDa, suggesting that the complex probably consists of one immunoglobulin molecule associated with one AST molecule. By immunoelectrophoresis, the immunoglobulin was found to be an IgG with kappa-lambda type light chain. When we used polyclonal antibodies against human mAST or sAST, the sAST antibodies strongly inhibited the AST activity of the macrocomplex, whereas the mAST antibodies had no effect. Thus the AST molecule of the macrocomplex is an sAST type.

Aged↗

PubMed1993

Use of human hepatocyte cultures for drug metabolism studies.

Among the in vitro models developed to investigate drug metabolism isolated hepatocytes have become the most powerful model. Human hepatocytes can be prepared from whole livers and surgical wedge biopsies. When placed in culture they retain their specific drug metabolizing activities including inducible cytochrome P450 enzymes for several days. Primary human hepatocyte cultures are now increasingly used for studying drug behavior during preclinical development, e.g. drug interactions, stereoselective drug metabolism and drug metabolic profiles. As a rule there is a good in vivo/in vitro correlation in drug biotransformation activity. The main metabolites found in vivo are recovered in vitro. However, quantitative differences are frequently observed.

Animals↗

PubMed1993

The O2- generating oxidase of B lymphocytes: Epstein-Barr virus-immortalized B lymphocytes as a tool for the identification of defective components of the oxidase in chronic granulomatous disease.

The O2- generating NADPH oxidase of human Epstein-Barr virus immortalized B lymphocytes (EBV-B lymphocytes) and the NADPH oxidase of human neutrophils were compared. The capacity of the oxidase of EBV-B lymphocytes to generate O2- is 100-fold less than that of neutrophils. Like the oxidase of neutrophils, the oxidase of EBV-B lymphocytes is decreased or abolished in chronic granulomatous disease (CGD). Activation of neutrophil oxidase in an heterologous cell-free system, using human neutrophil membranes and EBV-B lymphocyte cytosol from healthy and CGD patients, combined with immunoblotting investigations of the cytosolic activating factors p47 and p67 involved in O2- production, suggests that neutrophils and EBV-B lymphocytes possess similar complements of cytosolic factors p47 and p67. Cytochrome b -245, the major membrane redox component of the O2- generating oxidase, is only slightly expressed in the membrane of EBV-B lymphocytes. A sensitive and specific immunocytochemical method for detection of the two subunits of cytochrome b -245 is described; it shows that both subunits are virtually absent in EBV-B lymphocytes from CGD patients deficient in the large subunit.

Animals↗

PubMed1993

Human neutrophil gelatinase is a collagenase type IV.

Secreted gelatinase from human neutrophils was purified as a 94 kDa polypeptide. Gelatinolytic and type IV collagenolytic activities of the purified protein were measured and compared. Immunoglobulins purified from antisera raised against gelatinase inhibited both the gelatinase and type IV collagenase activities. There was no cross-reaction in the inhibition with type I collagenase while the three metalloproteases were similarly inhibited by recombinant tissue inhibitor of metalloproteases. Purified gelatinase degraded denatured type I and native type IV collagens; there was no proteolysis of native type I collagen.

Collagenases↗

PubMed1993

Mitochondrial genome expression in a mutant strain of D. subobscura, an animal model for large scale mtDNA deletion.

A mitochondrial mutant strain of D. subobscura has two mitochondrial genome populations (heteroplasmy): the first (20-30% of the population, 15.9 kb) is the same as could be found in the wild type; the second (70-80% of the population, 11 kb) has lost by deletion several genes coding for complex I and III subunits, and four tRNAs. In human pathology, this kind of mutation has been correlated with severe diseases such as the Kearns-Sayre syndrome, but the mutant strain, does not seem to be affected by the mutation (1). Studies reported here show that: a) Transcripts from genes not concerned by the mutation are present at the same level in both strains. b) In contrast, transcript concentrations from genes involved in the deletion are significantly decreased (30-50%) in the mutant. c) Deleted DNA was expressed as shown by the detection of the fusion transcript. d) The mtDNA/nuc.DNA ratio is 1.5 times higher in the mutant strain than in the wild type. The mutation leads to change in the transcript level equilibrium. The apparent innocuousness of the mutation may suggest some post-transcriptional compensation mechanisms. This drosophila strain is an interesting model to study the consequence of this type of mitochondrial genome deletion.

Animals↗

PubMed1993

Preferential increase of glutathione S-transferase class alpha transcripts in cultured human hepatocytes by phenobarbital, 3-methylcholanthrene, and dithiolethiones.

In rodents, a diversity of compounds are able to protect against acute and chronic toxicities of various xenobiotics including carcinogens, at least in part through induction of drug-metabolizing enzymes including glutathione S-transferase (GST) enzymes. We have posed the question as to whether or not these compounds also induce GSTs in human liver. Primary human hepatocyte cultures were exposed to phenobarbital, 3-methylcholanthrene, and two dithiolethiones [1,2-dithiole-3-thione and its 5-(2-pyrazinyl)-4-methyl derivative, oltipraz], and steady-state mRNA levels of GST classes alpha, mu, and pi were determined by Northern blot analysis. After 3 daily treatments, the two dithiolethiones were the most potent inducers; phenobarbital was also effective but to a lesser extent and 3-methylcholanthrene increased GST mRNA in only 2 of the 6 samples, although it stimulated cytochrome P-450 1A2 mRNA in all cell preparations. Whatever the compound only GSTA1 and/or A2 transcripts were induced. GST M1 mRNAs were not responsive or only slightly responsive, and GST P1 mRNAs, which were mostly undetectable in control cells, were not affected by treatment with any of the four chemicals. Large individual variations were observed in the level of induction of GST A1 and/or A2 mRNAs, and no sex difference could be demonstrated. These results clearly indicate that phenobarbital, 3-methylcholanthrene, and dithiolethiones are able to markedly increase mRNA levels of GST in human hepatocytes and that the GST alpha class is preferentially involved.

Cells, Cultured↗

PubMed1993

Effects of sCD23 on proliferation of leukemic cells from a patient with chronic myelogenous leukemia during blast crisis.

The present study has attempted to further delineate the growth factor requirements of peripheral blasts of a patient with CML in acute phase. Phenotypic analysis of leukemic blasts from this patient before culture has shown a homogenous population of CD34+ cells at the onset of blast crisis. In the second and third samples the percentage of CD34+ DR+ blast cells decreased slightly and up to 32% of cells in the third sample expressed the CD19 antigen. Optimal proliferation of cells derived from the first sample required the presence of exogenous sCD23 and to a lesser extent IL7. The stimulatory effects of sCD23 and IL7 were clearly reduced 4 months later and no longer detected after 6 months. This variability in growth factor response along with disease progression may be related to phenotypic differentiation. There was no evidence for lymphoid or myeloid maturation after 4 days of liquid culture. Our results in conjunction with previous studies are in agreement with sCD23-involvement in the complex control of proliferative processes at both normal and leukemic stages, demonstrating that cytokines are critical in determining CML cell proliferation.

Acute-Phase Reaction↗

PubMed1993

P-glycoprotein expression in human, mouse, hamster and rat hepatocytes in primary culture.

P-Glycoprotein (P-gp), the multidrug resistance (MDR) gene product, has previously been shown to be functional and overexpressed in adult rat hepatocytes during primary culture. In the present study, we examined P-gp expression in human, mouse and hamster hepatocytes cultured in conditions similar to those used for rat liver cells. Northern blotting and doxorubicin P-gp-mediated efflux analyses revealed that liver parenchymal cells from these three species exhibited only limited increased MDR mRNA levels with no intracellular decreased drug retention, thus suggesting that the marked functional P-gp induction observed in rat hepatocytes could reflect a species-specific response of these cells to an unfamiliar environment. An 8 h exposure to cycloheximide, which strongly induced MDR mRNAs in rat liver cells, also increased, although to a far lesser extent, MDR mRNA levels in human, mouse and hamster hepatocytes, indicating that P-gp expression in normal liver parenchymal cells could be at least partly negatively regulated by a labile protein factor.

ATP Binding Cassette Transporter, Subfamily B, Mem↗