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Biomedical subjects

A Lemonnier

Publications and source records attributed to A Lemonnier.

At least 73 records · Page 4Linked to original sources
PubMed1985

Cell density affects prolidase and prolinase activity and intracellular amino acid levels in cultured human cells.

Prolidase (EC 3.4.13.9) and prolinase (EC 3.4.13.8) and intracellular amino acid levels in cultured human cells increased when cell density rose. Firstly, two normal fibroblast strains were continuously cultured for 21 days and these parameters were measured on days 3, 7, 10, 14, 17 and 21 after plating. Prolidase, prolinase and amino acid levels varied considerably depending on the duration of culture and growth rate. Secondly, we studied the action of different cadmium and cobalt concentrations on prolidase activity. These two effectors altered this enzyme activity, but secondarily to modifying cell density. Thirdly, prolidase activity was investigated in 8 control amniotic cell strains, with a view to prenatal diagnosis of inherited prolidase deficiency, and we noted the same cell density interference. Due to the large variations related to cell density, we recommend specifying the number of cells per unit surface, and avoiding the term 'cells at confluency' which is unduly vague.

Amino Acids↗
PubMed1985

Determination of prolinase activity in plasma. Application to liver disease and its relation with prolidase activity.

We describe prolinase (EC 3.4.13.8) activity in human plasma for the first time. Optimum activity was obtained with prolylvaline as substrate and 0.02 mmol/L manganese concentration at pH 9.0. Moreover, preincubation with manganese was not required, contrary to prolidase (EC 3.4.13.9) activity. The mean value observed in 106 subjects without liver and renal disorders was 16 U/L +/- 14 (2 SD). We determined this plasma enzyme activity in patients with acute hepatitis and chronic liver disease. Plasma prolinase activity was strongly dependent upon cytolysis because of the high activity in liver and the low activity in plasma. Of 24 patients with chronic liver disease (4 chronic hepatitis and 20 cirrhosis) and without cytolysis, prolinase activity was slightly increased in only three patients, whereas prolidase activity was increased in 13. This could be due to a difference in the activation of these two enzymes in liver during the fibrotic process.

Clinical Enzyme Tests↗
PubMed1985

Some biochemical observations on gluconeogenesis from propionate in hepatocytes isolated from normal and biotin-deficients rats.

Propionate and pyruvate added to isolated normal and biotin-deficient adult rat hepatocytes increase the production of glucose. This production decreases about 30% on biotin deficiency. Malonate inhibits gluconeogenesis from propionate showing the metabolic transformation of propionyl-CoA via the Krebs cycle. Neither glucagon nor dibutyryl-cyclic AMP significantly stimulate gluconeogenesis.

Animals↗
PubMed1985

[Application of derivative spectrophotometry in biochemistry].

Derivative spectrophotometry is used to increase the specificity of spectroscopic measurements. The main advantages of this method are a better resolution of overlapping bands and a reduction in the effects of turbidity and coloured background. Its biochemical applications of this method are reviewed.

Amino Acids↗
PubMed1984

Evidence that cycloleucine affects the high-affinity systems of amino acid uptake in cultured human fibroblasts.

The influence of cycloleucine on kinetic parameters of uptake of L-alanine, L-proline and L-leucine into cultured human fibroblasts was examined under initial-rate conditions with substrate concentrations of 0.05-10 mM and 5 mM-cycloleucine. Kinetic data obtained by computer analysis showed that, in the absence of cycloleucine, cell uptake was heterogeneous for each amino acid. L-Alanine and L-leucine entered by two transport systems with different affinities; L-proline was taken up by one saturable transport system plus a diffusion-like process. This heterogeneity disappeared in the presence of cycloleucine, since the high-affinity systems were no longer detectable. The remaining process had the same kinetic constants as the low-affinity system for alanine and leucine and a KD similar to the diffusion constant for proline. The influence of cycloleucine on the amino acid uptake was not specific either to the amino acid concerned or to a particular transport system, since the three neutral amino acid-transport systems, A, ASC and L, were involved in these experiments. This influence was shown to be unaffected by the absence of Na+ (for leucine uptake). ATP content of the cells was identical in the presence or in the absence of cycloleucine.

Adenosine Triphosphate↗
PubMed1984

Methionine metabolism and ultrastructural changes with D-galactosamine in isolated rat hepatocytes.

The biochemical and morphological effects of 2, 10 and 100 mM of D-galactosamine (GalN) were studied in isolated rat hepatocytes during 2 h of incubation. Lactate dehydrogenase (LDH), alanine aminotransferase (ALAT) and cell viability did not change, whatever the concentration used. The variations observed, which were dose dependent, included a large drop in ATP levels and inhibition of RNA and protein synthesis. A very high concentration of GalN was necessary, however, to induce a significant decline in methionine adenosyltransferase activity compared to control cells. The use of L-[methyl-14C]methionine during cell incubation with GalN demonstrated a decrease of S-adenosyl-L-methionine (SAMe) and an accumulation of L-methionine content related to the GalN concentration. These results suggested that an hepatotoxic agent such as GalN was able to induce disturbances of methionine metabolism. Some of the ultrastructural changes observed were different from those previously found in vivo, in rats given GalN intraperitoneally, underlining the marked difference between in vivo and in vitro intoxication.

Alanine Transaminase↗
PubMed1984

Prolidase and prolidase deficiency.

Prolidase deficiency seems to be a rather rare metabolic disorder. However, many new cases can be detected because screening is easy to perform and enzymatic confirmation allows the differentiation from other iminodipeptidurias . Clinical symptoms are briefly reviewed, while biological considerations and prolidase properties are exhaustively described. Methods for investigating urinary iminodipeptides are given with results. Moreover, several collagen modifications observed in this disorder led us to formulate a hypothesis for their mechanism. Genetic considerations and treatment attempts are discussed.

Amino Acid Metabolism, Inborn Errors↗
PubMed1984

Analysis of dipeptides in urine by gas chromatography/mass spectrometry: implications for collagen breakdown in iminodipeptiduria following a study of the dipeptides by electron impact and chemical ionization.

Dipeptides in the urine of a patient suffering from dermatological purpura, associated with iminodipeptiduria , were determined by gas chromatography/mass spectrometry. The dipeptides were identified as R-proline and R'-hydroxyproline where R is any one of the residues, glycyl, alanyl, valyl, leucyl, isoleucyl, seryl, aspartyl, glutamyl, prolyl, phenylalanyl and R' is alanyl, valyl, leucyl or isoleucyl, seryl, prolyl, glutamyl, phenylalanyl. The predominance of proline- and hydroxyproline-containing dipeptides and the percentage distributions of the other amino acid residues, R and R', strongly implicate an abnormality of collagen metabolism. Structural assignments are confidently based on (a) gas chromatographic retention times, (b) electron impact mass spectra and automatic comparison with reference to spectra stored in a specialized library, (c) chemical ionization mass spectra with isobutane and methanol as reactant gases and (d) the use of deuteriated acetic anhydride as a derivatizing agent.

Adult↗
PubMed1984

Comparative use of fructose and glucose in human liver and fibroblastic cell cultures.

The effect of fructose as a substitute for glucose in cell culture media was investigated in human skin fibroblast and liver cell cultures. Cells were grown for between 2 and 10 days in identical flasks in four different media, containing 5.5 mmol X 1-1 and 27.5 mmol X 1-1 glucose and fructose, respectively. In the presence of fructose, cell growth was stimulated, but less in liver cells than fibroblasts. At Day 6, increases were observed in [3H]thymidine incorporation, protein levels, and amino acid consumption, and a reduction was noted in ATP levels. In media containing 5.5 mmol X 1-1 glucose or fructose, consumption of fructose was four times lower than that of glucose at Day 3 and did not rise until Day 6. In fructose media, the lactate production was very low (four to five times less than that of glucose) and the pH values were always higher. Some findings were different for the fibroblasts and liver cells, owing to the specific characteristics of these two cell types in culture; this applied especially to the effects of glucose and fructose concentrations of 27.5 mmol X 1-1. Several possible explanations for the stimulation of cell growth in fructose medium were discussed.

Adenosine Triphosphate↗
PubMed1984

Relationship between cell density and prolidase activity in human skin fibroblasts: effects of ascorbate and fructose.

To evaluate the influence of cell density on the activity of fibroblast prolidase (EC 3.4.13.9), we determined this activity in sparse and dense cultures. We also investigated, the effects of different concentrations of beta-D (-) fructose and L (+) ascorbate, which both increased cell density at confluency. For a fructose concentration of 25 mM, we observed that in the absence of glucose, intracellular total proteins increased 1.5-fold and prolidase specific activity, 1.8-fold. For ascorbate, a broad optimum concentration was found (range 0.01 - 0.50 mM). Addition to cultures of 0.1 mM ascorbate increased total proteins 1.4-fold, and doubled prolidase activity. This investigation was prompted by our previous results [J. Metab. Dis. 1983, 6, 27-31], confirmed here, and suggesting that increased prolidase activity at confluency was due to a rise in cell density.

Ascorbic Acid↗
PubMed1984

Plasma vitamin E levels in children with cholestasis.

Plasma vitamin levels were assayed in 58 children presenting with chronic cholestasis. In the infants who developed cholestasis during the first weeks of life, vitamin E levels dropped below normal values after the age of 4 months. In the older children, vitamin E levels were not correlated with the etiology of cholestasis but with the degree of cholestasis, as expressed by serum bilirubin, serum bile acids, and fat absorption coefficient. We did not find any relationship between vitamin E levels and other biological parameters such as alkaline phosphatases, triglycerides, phospholipids, and cholesterol. These results further support the importance of vitamin E deficiency in chronic cholestasis of infants and children.

Absorption↗
PubMed1984

[Genetic heterogeneity and the diagnosis of hepatic glycogenoses].

Glycogen storage diseases constitute a highly heterogeneous group of disorders, because of the many complex enzyme systems involved in glycogen metabolism, and also because of the diversity of molecular defects connected with gene mutations. To illustrate these features, the authors studied four types of liver glycogen storage diseases, respectively caused by deficiencies of glucose-6-phosphatase, debranching enzyme, phosphorylase and phosphorylase kinase. In each case, the role and functional characteristics of the enzyme system are described, as well as the bioclinical aspects of the deficiency. The only reliable way of diagnosing glycogen storage disease is by assaying the activity of the enzyme concerned. Assay procedure must take account of various factors, especially the progress made in understanding the nature and mechanism of action of enzyme systems, the possible tissular heterogeneity of the deficiency and the functional characteristics of certain enzymes.

Genetic Variation↗
PubMed1984

Plasma prolidase activity: a possible index of collagen catabolism in chronic liver disease.

We describe here an easy method of determining prolidase (EC 3.4.13.9) in plasma after preincubation with Mn2+ for 24 h at 37 degrees C to maximize prolidase activity. The mean activity in 338 patients who were either in hospital or outpatients was 900 U/L +/- 520 (2 SD), unrelated to sex or age. In 25 of these 338 samples tested, prolidase activity was between 1500 and 2000 U/L. It exceeded 2000 U/L in eight, all of whom were patients with chronic liver disease. Plasma prolidase activity was normal in cytolytic syndromes such as liver or heart disease. Of the 27 patients with cirrhosis, only five exhibited prolidase activity greater than 2000 U/L. Plasma prolidase activity was uncorrelated with six biochemical indexes to liver function (the aminotransferases, alkaline phosphatase, glutamyltransferase, total bilirubin, and serum albumin) or with the degree of cirrhotic fibrosis. We believe that plasma prolidase activity may be high only in the early stage of fibrosis. This hypothesis would be consistent with the data on rat-liver collagenolytic activities during CCl4 administration. Monitoring of plasma prolidase activity might be useful in evaluating fibrotic processes in chronic liver disease in the human.

Adolescent↗
PubMed1983

In-vitro responses to ascorbate and manganese in fibroblasts from a patient with prolidase deficiency and iminodipeptiduria: cell growth, prolidase activity and collagen metabolism.

After successful ascorbate and manganese treatment of a female patient with prolidase deficiency and iminodipeptiduria, we attempted to explain the mechanism of action of these drugs in vitro, using them preferentially on skin fibroblasts. Since in vivo, ascorbate and manganese seemed to be responsible for both biochemical and clinical improvement, they were also expected to activate prolidase activity in vitro. Cell growth and prolidase activity were accordingly observed in fibroblast cultures supplemented with these compounds. It seemed that only ascorbate accounted for the successful in vivo response. To understand the mechanism involved, we studied collagen metabolism and found a decreased proline pool, a massive increase of rapidly degraded collagen and moderate enhancement of type III collagen and type I trimer in the patient's fibroblasts. We believe that ascorbate allowed the prolidase-deficient cells to maintain a normal collagen pool by increasing collagen synthesis. Both the massive increase in cell growth in response to ascorbate and the bad response as regards the quality of the collagen produced confirm the secondary nature of this mechanism. However, the relationship between accelerated collagen catabolism and prolidase deficiency remains unclear.

Adult↗
PubMed1983

Transport of L-histidine by human diploid fibroblasts in culture.

The transport of L-histidine has been characterized in skin derived diploid human fibroblasts, cultured under strictly controlled conditions. The transport measurements were made on cells grown to subconfluency after 60 to 90 min timed preincubation. The data, at substrate concentrations ranging from 0.050 to 10 mmol/l, were analyzed by a computer program. A saturable transport system (Km = 0.25 mmol/l, Vmax = 17 nmol/mg protein per min) and a nonsaturable component of influx (Kd = 1.6 +/- 0.4 nmol/mg protein/min per mmol) were found. L-Histidine displayed no Na+ requirement at either low or high concentrations. Inhibition analysis demonstrated that L-histidine uptake at low concentration was poorly inhibited by amino acids known to be effective inhibitors of system A. The largest fraction of L-histidine uptake was inhibited by 2-amino-bicyclo (2,2,1)-heptane-2-carboxylic acid (BCH), leucine, and tryptophan. These results indicated that L-histidine is transported in human fibroblasts mainly by the Na+ independent system L. The differences between this cell type and others studied previously are discussed.

Amino Acids↗