A. Lemonnier

Diagnostic value of serum gamma-glutamyl-transpeptidase activity in liver diseases in children

The clinical usefulness of serum gamma-glutamyl transpeptidase (gamma GT) assay for the diagnosis of liver disease in children was assessed retrospectively in 398 children investigated from 1981 to 1986, in whom diagnosis was ascertained according to currently accepted criteria including liver histology in each case. Serum gamma GT activity was within normal limits in 10 controls, in 19 children with portal vein obstruction, and in 10 of 12 children with congenital hepatic fibrosis. Serum gamma GT was raised in all children with biliary atresia, sclerosing cholangitis, paucity of interlobular bile ducts, and alpha 1-antitrypsin deficiency with jaundice. Serum gamma GT was normal in spite of patent clinical signs of cholestasis in 3 patients with benign recurrent intrahepatic cholestasis, 1 infant with post-hemolytic neonatal cholestasis, and in 22 of 28 patients with progressive idiopathic cholestasis akin to Byler disease. In the latter group, children with raised serum gamma GT displayed extensive portal fibrosis and bile duct proliferation on liver histology, while this was not a prominent feature in children with normal serum gamma GT. These results indicate (a) the value and limits of the assay for serum gamma GT activity in children with liver disease, (b) that raised serum gamma GT may be considered a fairly reliable index of bile duct damage, and (c) that serum gamma GT may prove a useful tool in separating two forms of progressive idiopathic cholestasis, with or without bile duct involvement.

High hepatic γ-glutamyltransferase (γ-GT) activity with normal serum γ-GT in children with progressive idiopathic cholestasis

Abstract γ-Glutamyl transferase (γ-GT) was assayed in the serum and liver hiopsies of children affected with either progressive idiopathic cholestasis (PIC, Bylers disease), or other types of cholestatic (biliary atresia, cholestasis of various origins) and non-cholestatic diseases. The mean liver γ-GT activity was increased significantly only in PIC and biliary atresia. In contrast, the serum γ-GT activity, raised in children with evident damage to the main bile ducts or to the interlobular bile ducts, was normal in children with PIC. Although the mechanism for such a discrepancy between high liver and normal serum γ-GT activties in PIC is still speculative, this peculiarity could prove to be of use in leading to a better understanding of the disease.

COMPARATIVE USE OF FRUCTOSE AND GLUCOSE IN HUMAN LIVER AND FIBROBLASTIC CELL CULTURES

SummaryThe 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·1−1 and 27.5 mmol·I−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·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·1−1. Several possible explanation for the stimulation of cell growth in fructose medium were discussed.

Comparative study of extracellular amino acids in culture of human liver and fibroblastic cells

SummaryAmino acid concentrations are studied in the extracellular media of ten series of human fibroblast and liver cell monolayer cultures. These two cell types consume and produce ostensively the same amino acids. Among the nonessential amino acids, the most significant variations involve serine and aspartate which are decreased; α-alanine, glutamate, ornithine and proline are, on the contrary, increased. Among the essential amino acids, leucine, isoleucine and glutamine are preferentially decreased. The variations of some amino acids are correlated with the cell density. The interrelations which may exist between the variations of these different amino acids are discussed. Furthermore, the glycolytic activity of the cells studied is very high: 85% of glucose consumed is found in the form of lactate.

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.

Comparative metabolic effects of fructose and glucose in human fibroblast cultures.

SummaryThe comparative metabolic effects of fructose and glucose were determined in human fibroblast cultures. Cells were grown in four different media containing 5.5 and 27.5 mM of glucose and fructose, respectively. For these two hexoses, we compared their uptake, consumption, and conversion into14CO2 and14C-lipids.D-Fructose was taken up in fibroblasts by an unsaturable process and its consumption was much smaller than that ofD-glucose. Whatever the experimental procedure, the glycogen content of cells grown in fructose media was significantly lower than of those grown in glucose media. Labeling of fructose and glucose with14C showed that more carbon from fructose than from glucose was incorporated into CO2 and glycerolipids. The relative distribution of14C in the different lipid fractions was similar for both hexoses.These results indicated that the pathways of intermediary metabolism in fibroblast cultures were influenced by the nature of the carbohydrate present in the culture medium and that fructose was a better lipogenic substrate than glucose in human fibroblast cultures.

Influence of two cell harvesting methods on intracellular ATP and amino acid concentrations in human fibroblast cultures

The intracellular ATP and amino acid concentrations were determined in human fibroblast cultures reaching confluence. The values obtained were very different, depending on the cell harvesting method: trypsinization or scraping. Trypsinization appeared to be the better method for measuring the ATP concentrations (21.25 +/- 0.96 nmol per mg cell protein), this level being much lower with scraping. On the contrary, scraping was the most appropriate method for amino acid measurement. This work underlines the importance of harvesting methods for metabolic studies in human cell cultures.

Changes in methionine metabolism induced by D-galactosamine in isolated rat hepatocytes

We studied several steps of methionine metabolism in isolated rat hepatocytes both with and without the presence of a hepatotoxic agent (D-galactosamine). By use of selective labelling either on methyl or on carboxyl groups, we showed that intracellular methionine is used preferentially for the methylation of phospholipids (42%) and nucleic acids (31%) via S-adenosylmethionine. In the presence of D-galactosamine, the incorporation of L-(14CH3) methionine into macromolecules is significantly inhibited (greater than 50%). This inhibition is associated with a decrease of S-adenosylmethionine and an increase of methionine in the injured cells. These results suggest that hepatotoxicity of galactosamine may be due in part to an inhibition of the methylation of nucleic acids and phospholipids. Consequently, we hypothesize that hypermethioninemia associated with human liver disease could be due, at least partly, to a defect in synthesis and/or utilization of S-adenosylmethionine by hepatocytes.

Antioxidant Properties of Vitamin E and Membrane Permeability in Human Fibroblast Cultures

α-tocopherol, the most active form of vitamin E (Vit. E) is considered as one of the chief cell components which maintains the structural and functional membrane integrity. It acts as a free radical scavenger preven ting the peroxidation of membrane fatty acids. The role of Vit. E in cell permeability is widely studied using artificial membranes, but only a few investigations were devoted to this effect in cell cultures 1,2.

Comparative use of glucose and fructose in cultured fibroblasts from patients with hereditary fructose intolerance.

The utilization of fructose and glucose by fibroblast cultures obtained from patients with hereditary fructose intolerance (HFI) was studied in comparison with fibroblast controls. The cell growth, the time course ofd-glucose ord-fructose uptake and the consumption of fructose were similar for both HFI and control cells. Some results showed significant differences between these two cell types: HFI cells consumed less glucose, produced less lactate and contained less glycogen than control cells. Furthermore, significantly less [U-14C]d-glucose and [U-14C]d-fructose was incorporated into lipids in HFI cells than in control cells. The mechanisms responsible for these differences observed between the two cell types are not known.