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Biomedical subjects

V Stocchi

Publications and source records attributed to V Stocchi.

At least 127 records · Page 7Linked to original sources
PubMed1983

Red blood cell galactokinase activity and presenile cataracts.

Red blood cell galactokinase activity was measured in 70 patients with cataracts to assess a possible correlation between galactokinase activity levels and risk of cataract development. Among all, 15 patients developed cataracts during the first year of life, 25 patients under the age of 50 and 30 later in life. No cases of total or partial galactokinase deficiency were found. These results, taken together with the absence of cataracts in 9 patients with partial galactokinase deficiency render less certain the cause and effect relationship between partial galactokinase deficiency and the appearance of cataracts.

Adolescent↗
PubMed1983

Rabbit red blood cell hexokinase. Mechanism of decay during cell life-span.

Rabbit red blood cells contain hexokinase type I whereas in the reticulocyte two distinct molecular forms (HK Ia and Ib) are present. One (HK Ia) corresponds to hexokinase type I from other tissues, while the other differs from any previously reported isozyme. Rabbit bone marrow cells contain hexokinase type I and II. However, when the erythroid precursor cells become predominant over the non-erythroid cells (during phenylhydrazine anemia) a great increase of HK Ia can be observed concomitant with the appearance of HK Ib. Fractionation of the bone marrow cells on density gradients provides evidence that basophil erythroblasts and proerythroblasts contain only HK Ia while HK Ib appears at the reticulocyte stage. Maturation and ageing of circulating reticulocytes are associated with the decrease of hexokinase activity. Since the decay rate of HK Ib is about three times higher than the decay rate of HK Ia, the mature erythrocytes do not contain appreciable amounts of HK Ib. Furthermore, in vitro, HK Ia and Ib possess similar stabilities so that a cellular mechanism must be responsible of their in vivo different decay rates. This mechanism, as reported in this paper, is ATP-dependent, could be found in the soluble fraction, and is active only at the reticulocyte stage. These properties are similar to those of the ATP-dependent proteolytic system. Pure ubiquitin, an essential polypeptide of the ATP-dependent proteolytic system, is also able to catalyze the decay of hexokinase activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Red blood cell glucose metabolism in trisomy 10p: possible role of hexokinase in the erythrocyte.

Red blood cell glucose metabolism was investigated in a male patient with de novo trisomy 10p. According to previous evidence, when assigning hexokinase gene locus in the 10p11 leads to pter region, a triplex dosage effect of hexokinase activity (HK) was found, while all the other erythrocyte glycolytic enzymes were in the normal values range. Red blood cell glucose utilization was 2.87 mumole/hr/ml RBC as compared to 1.43 in normal controls; the rate of glucose metabolized through the hexose monophosphate shunt (HMPS) was unchanged. Glucose-6-phosphate, fructose-6-phosphate, fructose-1,6-diphosphate, and dihydroxyacetone phosphate increased with respect to normal controls, while normal levels of 3-phosphoglycerate, 2-phosphoglycerate, phosphoenolpyruvate, and ATP were found. The HK activity increased in all the red blood cell fractions obtained by density gradient ultracentrifugation. However, a small difference in the distribution of cells through the gradient was evident. The experiments reported in this article show that in the red blood cells of patients with trisomy 10p, an increased level of HK leads to higher concentrations of glucose-6-phosphate and to a faster glucose utilization in the Embden-Meyerhof pathway, while the HMPS rate is unchanged.

Blood Glucose↗
PubMed1982

Molecular forms of red blood cell hexokinase.

Mammalian red blood cell hexokinase has been shown to exist in two or more distinct molecular forms, which are separable by ion-exchange chromatography. Of these forms just one corresponds to hexokinase type I from other tissues, while the others differ from any previously reported hexokinase isozyme. Analysis of several molecular properties of the three major forms (Ia, Ib and Ic in the order of their elution from DE-52 columns) of hexokinase prepared from human red cells and of the two forms purified from rabbit reticulocytes, shows significant differences in the isoelectric point. The kinetic and regulatory characteristics, the molecular weight, the temperature and pH-dependence of the various isozymes were similar. The hexokinase isozymic pattern is largely dependent upon red blood cell age. Among all, hexokinase Ib is the predominant form in rabbit reticulocytes and becomes the minor component in the older cells; a similar situation has also been found in the human erythrocyte. At present the molecular basis of hexokinase heterogeneity remains unknown, however preliminary experimental findings indicate a post-translational modification as a possible mechanism.

Aging↗
PubMed1982

Multiple forms of human red blood cell hexokinase. Preparation, characterization, and age dependence.

Human red blood cell hexokinase (EC 2.7.1.1) has been shown to exist in multiple molecular forms which are separable by ion exchange chromatography. Of the major forms, designated hexokinase Ia, Ib, and Ic, only hexokinase Ia corresponds to hexokinase type I from human liver, while the others differ from every other previously reported hexokinase isozyme. Hexokinase Ib is the predominant form in the fetal erythrocytes, while it is present at lower levels in the red blood cells of adults. Analysis of the hexokinase isozymic pattern in red cells of different mean age shows that the level of hexokinase Ib is also dependent on the age of the cell. The three major forms of hexokinase have the same molecular weight of 100,000, by sedimentation velocity on sucrose density gradients, the same Michaelis constants, substrate and coenzyme specificity, pH-dependent activity, and the same thermal stability. The only significant differences were found in the isoelectric points which were 5.7 pH units for hexokinase Ia, 5.5 pH units for hexokinase Ib, and 5.35 pH units for hexokinase Ic. These data, together with that previously reported for rabbit erythrocytes (Stocchi, V., Magnani, M., Canestrari, F., Dachà, M., and Fornaini, G. (1981) J. Biol. Chem. 256, 7856-7861) suggest that the presence of multiple forms of hexokinase is a common phenomenon in mammalian red blood cells.

Adult↗
PubMed1982

Adult and fetal galactokinases in human red blood cells.

This paper reports the biochemical properties of galactokinase from fetal and adult human red blood cells. The specific activity of galactokinase is three times higher in the fetal red cells than in adult cells, shows a significant difference in the Michaelis constant toward galactose, and is more thermostable. On the other hand, no differences were found in molecular weight, electric charge, temperature and pH dependence between the two enzymes partly purified from fetal and adult erythrocytes. The possibility that these differences could be due to the shorter lifespan of the fetal erythrocytes (which could result in a higher proportion of young cells in the blood samples utilized) was investigated. Fetal and adult red blood cells were separated into fractions of different mean age by ultracentrifugation through density gradients. The kinetic properties and thermostability of galactokinase from fetal erythrocytes do not show any similarity with the same properties of the enzyme from young red blood cells. These results indicate that galactokinase from fetal erythrocytes show some biochemical properties that are typical signs distinguishing a fetal enzyme.

Adult↗
PubMed1981

Rabbit red blood cell hexokinase. Evidence for two distinct forms, and their purification and characterization from reticulocytes.

Rabbit hexokinase (EC 2.7.1.1) has been shown to exist in the soluble fraction of reticulocytes as two distinct molecular forms, designated hexokinase Ia and hexokinase Ib, which are separable by ion exchange chromatography and polyacrylamide gel electrophoresis. Hexokinase Ia was found to be similar to the brain enzyme, while hexokinase Ib differs from every other previously reported hexokinase isozyme. Reticulocyte hexokinase Ia and Ib have been purified 55,000-and 50,000-fold, respectively, by a combination of ion exchange chromatography, affinity chromatography, and preparative polyacrylamide gel electrophoresis, as proteins homogeneous by sodium dodecyl sulfate-gel electrophoresis. The native proteins have the same molecular weight of 105,000 by gel filtration and sedimentation velocity on sucrose density gradients. Sodium dodecyl sulfate-polyacrylamide gels have a molecular weight of 104,000, indicating that the two forms are monomers. Hexokinase Ia had a pI of 6.2 to 6.3 pH units while hexokinase Ib had a pI of 5.7 to 5.8 pH units by isoelectric focusing. The two enzymes were specific for Mg.ATP and Mg.ITP as the nucleotide substrates. Several hexoses could be phosphorylated by hexokinase Ia and Ib with different affinities.

Animals↗
PubMed1980

Action of oxidized and reduced glutathione on rabbit red blood cell hexokinase.

Reduced glutathione at 1 mM concentration is able to mantain rabbit red blood cell hexokinase (EC 2.7.1.1) in the reduced state with fully catalytic activity. At higher concentrations a marked inhibition is observed. In contrast, oxidized glutathione is a strong inhibitor of reduced erythrocyte hexokinase at all the concentration studied. Inactivation experiments show that some sulfhydryl groups reacting with oxidized glutathione are responsible for the enzyme inactivations. These findings suggest a cellular inter-relationship between redox and energetic metabolism coupled through glutathione at the hexokinase level.

Adenosine Triphosphate↗
PubMed1980

Rabbit red blood cell hexokinase. Purification and properties.

Rabbit red blood cell hexokinase (EC 2.7.1.1.) has been purified 300,000-fold by a combination of ion exchange chromatography, affinity chromatography, and preparative polyacrylamide gel electrophoresis. The hexokinase activity has been isolated in 35% yield as a protein that is homogeneous by polyacrylamide and sodium dodecyl sulfate gel electrophoresis. The highest specific activity obtained was 145 units/mg of proteins. The native protein has a molecular weight of 110,000 by gel filtration on Ultrogel AcA 44 and 112,000 by sedimentation velocity on sucrose density gradients. Sodium dodecyl sulfate-polyacrylamide gels gave a molecular weight of 110,000 indicating that hexokinase is a monomer. The enzyme had a pI of 6.20 to 6.30 pH units by isoelectric focusing. The enzyme was specific for Mg . ATP and Mg . ITP as the nucleotide substrates. Several hexokinase with different affinities.

Adenosine Triphosphate↗
PubMed1980

Erythrocytes of different ages: a new method of "in vivo" preparation.

This paper describes a new method for obtaining "in vivo" populations of homogeneous erythrocytes of a specific age. This method, proposed for experimental animals, is based on the induction of a large reticulocytosis (about 80%) by the administration of phenylhydrazine and the subsequent synchronized ageing of young red cells present, which is obtained by blocking the erythropoiesis with daily injections of Actinomycin D. The advantage of such a method over those published so far is that it produces substantial quantities of red blood cells of a specific age. A disadvantage however is that it is only possible to obtain cells of up to 30 days old.

Animals↗
PubMed1979

Decay pattern of rabbit erythrocyte hexokinase in cell aging.

As previously reported, during rabbit red blood cell aging glucose phosphorylating activities show several modifications. In the first period of the red cell life span the predominant form is similar to hexokinase II, while in the mature erythrocyte the predominant glucose phosphorylating activity resembles hexokinase I. In the oldest cells glucose phosphorylating activity has a low affinity (high Km) for glucose. In this paper the modifications of hexokinase in cell aging have been studied in vivo in a young erythrocyte population synchronized by actinomycin D, and in vitro in red cells separated in fractions according to different ages. Since protein synthesis is lacking in the mature red cell, we are inclined to explain the presence of low-affinity hexokinase activity in the oldest erythrocytes as an age-dependent transformation of a primary hexokinase.

Animals↗
PubMed1979

Comparative studies of galactokinase activity on mammal's red blood cells.

1. ATP: D-galactose-1-phosphotransferase activity was measured in human, pig, cow, rabbit, mouse and rat red blood cells. Mean values of galactokinase activity was markedly lower in the human and pig erythrocyte as compared to those of the other species. 2. The permeability to galactose of the red cells studied was always higher than galactose phosphorylation. 3. The affinity constants of galactokinase for galactose ranged from 119 to 291 microM and from 178 to 406 microM for ATPMg2-. 4. The thermostability values of the galactokinase of the species studied were similar. The pH-optimum is pH 7.5 for the human, mouse and rabbit enzyme and pH 8.0 for cow, pig and rat galactokinase.

Animals↗