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A three-year prospective study was conducted to assess the efficacy and safety of transjugular liver biopsy with a Trucut-type needle employing an automated device. Fifty-two consecutive patients (39 women and 13 men), ages 46.3+/-14.6 years, in whom percutaneous liver biopsy was contraindicated were included in the study. An 18-gauge transjugular hepatic needle with a 20-mm throw length, a cutting cannula at the distal end, and an automated trigger device on the proximal end was used. All procedures were performed under fluoroscopic control, and the following variables were assessed: (1) number of passes, (2) size and average number of the obtained fragments, (3) number of portal triads, and (4) adequacy of the specimen for histologic diagnosis. The procedure was successful in 49/52 patients, and all samples obtained were satisfactory for histologic analysis even when cirrhosis or bridging fibrosis were present. Mean biopsy specimen length was 1.7+/-0.88 cm; mean number of passes was 2.42+/-1.01, the mean number of biopsy fragments and portal triads per attempt were 2.5+/-1.01 and 6.24+/-3.18, respectively. No major complications were observed. Transjugular hepatic biopsy with this new cutting system is an effective and safe procedure in patients with contraindication for percutaneous liver biopsy.

The glucose-induced switch between glycogen phosphorylase and glycogen synthase in the liver is investigated by means of a theoretical approach based on a minimal, bicyclic cascade model involving the reversible phosphorylation of the two enzymes. The aim of the analysis is to evaluate the contribution of different factors to the sequential changes in the activity of glycogen phosphorylase and glycogen synthase observed following the addition of suprathreshold amounts of glucose.

Liver biopsies from 27 patients with malignant diseases and virus B carriers state were studied. The age of the patients ranged from 2 to 14 years. ELISA tests for virus B hepatitis were performed in all patients. Three of them resulted HBV positive at admission, whereas 24 became positive after anti-tumoral treatment onset. One patient was also virus C positive. Morphological disorders findings comprises: symptomless acute hepatitis in 7 patients (including the B-C carrier), minimal hepatocytes damage was found in 17 patients as the result of their carriers state and where also certain drug-induced hepatotoxic effect might be present. In this group there were also 7 biopsies with minimal portal region damage. Three biopsies were reported as normal liver tissue. This results confirmed the significance of the study of serological hepatitis viral test and hepatic biopsy in high risk group patients, where it is also necessary the vaccination against HBV before treatment onset.

Rat liver contains four hexokinase isoenzymes, one of which, despite often being called 'glucokinase', is no more specific for glucose than the others. However, it does differ from them in displaying a sigmoid kinetic response to glucose, requiring much higher glucose concentrations for activity, and being insensitive to physiological concentrations of glucose 6-phosphate.

Rat liver 'glucokinase' (hexokinase D) catalyses the phosphorylation of fructose with a maximal velocity about 2.5-fold higher than that for the phosphorylation of glucose. The saturation function is hyperbolic and the half-saturation concentration is about 300 mM. Fructose is a competitive inhibitor of the phosphorylation of glucose with a Ki of 107 mM. Fructose protects hexokinase D against inactivation by 5,5'-dithiobis-(2-nitrobenzoic acid), and the apparent dissociation constants are about 300 mM in the presence of different concentrations of the inhibitor. The co-operativity of the enzyme in the phosphorylation of glucose can be abolished by addition of fructose to the reaction medium. Fructose appears to be no better as a substrate for the other mammalian hexokinases than it is for hexokinase D. It is proposed that the name 'glucokinase' ought to be reserved for enzymes that are truly specific for glucose, such as those of micro-organisms and invertebrates, and that liver glucokinase must be called hexokinase D (or hexokinase IV) within the classification EC 2.7.1.1.

Hexokinase D, also called hexokinase IV or glucokinase, is the isoenzyme characteristic of liver. In spite of its common name of glucokinase it phosphorylates also other sugars besides glucose; in particular, it phosphorylates fructose with similar specificity to that shown by the other hexokinases. Although hexokinase D is a monomeric protein it displays positive cooperativity with glucose and mannose. In contrast, the kinetic behaviour with 2-deoxyglucose and fructose is Michaelian. Mannose, fructose, 2-deoxyglucose and N-acetylglucosamine are competitive inhibitors of glucose phosphorylation and suppress the cooperativity. The cooperative behaviour can also be suppressed by the presence of glycerol at the assay medium at concentrations over 20%, with a decrease in the K0.5. Neither glycerol nor the inhibitors affect the monomeric state of the enzyme. Hexokinase D exhibits an intrinsic fluorescence at about 326 nm due to tryptophan residues. The binding of glucose to the enzyme enhances the native fluorescence by about 15%. A dissociation constant for glucose of about 3.5 mM was estimated; this value decreased to about 0.5 mM glucose in the presence of glycerol. These and other results are discussed on the basis of steady-state models which assume that hexokinase D exists mainly in one conformation state (EI) in the absence of ligands, and that the binding of glucose or mannose induces a conformational transition to a new conformation EII with higher affinity for the sugar substrates. It is postulated that differences in the velocities of the conformational transitions induced by the different sugar substrates give account of the differences in kinetic behaviour with the different sugar substrates.

Glucokinases obtained from the liver of several species of mammals and amphibians exhibit sigmoidal saturation functions for glucose. Hill coefficients (nH) are about 1.5, and half-saturation values (K0.5) lie between 1.5 and 8.5 mmol/l. The nH and K0.5 values are constant throughout the purification steps of rat glucokinase. A dimeric form of rat glucokinase appearing in aged preparations exhibits michaelian kinetics. Sigmoidal kinetics is considered as an adaptive feature of glucokinases to increase the efficiency of the liver uptake of glucose at the changeable concentrations in the blood resulting from variations in the amount of dietary glucose.

The cellular distribution of hexokinase isoenzymes, N-acetylglucosamine Kinase and pyruvate kinases in rat liver was studied. Hepatocytes and non-parenchymal cells with high viability and almost no cross-contamination were obtained by perfusion in situ of the liver with collagenase, with the use of an enriched cell-culture medium in all steps of cell isolation. Separation of hexokinase isoenzymes was done by DEAE-cellulose chromatography, and enzyme activities were measured by a specific radioassay. Cytosol from isolated hepatocytes contained high-affinity hexokinases A, B and C, in addition to hexokinase D. The last-mentioned represented about 95% of total glucose-phosphorylating activity. Only hexokinase A was found associated t the particulate fraction. Isolated non-parenchymal cells contained only hexokinases A, B and C. N-Acetylglucosamine kinase was measured with a specific radioassay and was found as a single enzyme form in both hepatocytes and non-parenchymal cells, with higher activities in the former. Pyruvate kinase isoenzyme L was present only in the hepatocytes and isoenzyme K only in the non-parenchymal liver cells, confirming that they are good cellular markers.

The effect of dietary and hormonal variations on the specific activities of hexokinase isoenzymes, N-acetylglucosamine kinase and pyruvate kinase isoenzymes in parenchymal and non-parenchymal liver cells was studied. Hexokinase D was markedly decreased in hepatocytes from animals fasted or fed on the carbohydrate-free diet as well as from diabetic rats, attaining a constant low level of about 17% of normal values. Pyruvate kinase L was also diminished in hepatocytes under the same experimental conditions. In contrast, the three high-affinity hexokinase isoenzymes A, B and C remained without variation in total amount or in their relative proportions in hepatocytes and non-parenchymal liver cells isolated from animals under the various conditions studied. N-Acetylglucosamine kinase activities also did not change either in parenchymal or in non-parenchymal liver cells under all conditions. The results are discussed in relation to the significance of N-acetylglucosamine kinase and the various hexokinase isoenzymes for the phosphorylation of glucose after dietary and hormonal manipulations.