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Biomedical subjects

A Burchell

Publications and source records attributed to A Burchell.

At least 73 records · Page 4Linked to original sources
PubMed1995

An altered T2 beta translocase of the glucose-6-phosphatase system in the membrane of the endoplasmic reticulum from livers of Ehrlich-ascites-tumour-bearing mice.

The inhibitory interactions of orthophosphate (P1) with the glucose-6-phosphatase system of intact microsomes derived from the livers of normal and Ehrlich-ascites-tumour-bearing mice reveal the appearance of a novel form of the T2 beta translocase component of the glucose-6-phosphatase system in tumour-stressed mice. Kinetic studies, with and without 20 mM P1, show a strictly classical competitive inhibition, with a K1,P1 of 4.2 mM, with disrupted microsomes from both control and tumour-bearing mouse liver. Inhibition was also observed with intact microsomes from livers of control mice, and contributions by both competitive and non-competitive components of inhibition were quantified by calculation of Kis,P1 and Kii,P1 values respectively. However, little inhibition was noted with intact microsomes from the livers of tumour-bearing mice. It is concluded that this novel form of T2 beta is less able to transport Pi, from the cytosol to the endoplasmic reticulum lumen, perhaps because of the tumour-related increased Km for Pi transport in this direction.

Animals↗
PubMed1995

The in vivo regulation of hepatic and renal glucose-6-phosphatase by thyroxine.

The hepatic and renal microsomal glucose-6-phosphatase enzymes are situated with their active site in the lumen of the endoplasmic reticulum and for normal enzyme activity in vivo transport systems are needed for the substrates and products of the enzyme. We have shown that thyroxine activates the kidney glucose-6-phosphatase enzyme and the liver glucose 6-phosphate transport systems. In contrast, in hypophysectomised and adrenalectomised animals, thyroxine activates the transport systems and the enzyme in both liver and kidney.

Adrenalectomy↗
PubMed1995

The ontogeny of the glucose-6-phosphatase enzyme in human embryonic and fetal red blood cells.

We have shown for the first time that the microsomal glucose-6-phosphatase enzyme protein is present in human embryonic and fetal red blood cells and the ontogeny of its expression has been determined. In the earliest embryos, red cells are predominantly of the primitive megaloblastic type. Circulating red cells in the primitive megaloblastic series are predominantly nucleated and glucose-6-phosphatase immunopositive. Non-nucleated, immunoreactive megaloblastic cells are in a minority. In fetuses > 12 weeks gestation, the erythrocytes are of the definitive normoblastic series and in the transitional period of switch-over in late embryonic-early fetal life, up to 30% of glucose-6-phosphatase immunopositive cells are definitive normoblastic in type, with a variable contribution from nucleated and non-nucleated cells. Thereafter, the number of immunopositive cells in the definitive normoblastic series decreases such that after 12 weeks gestation it is less than 5%. The fact that a predominantly hepatic protein in adults (glucose-6-phosphatase) is present in embryonic and fetal red blood cells, particularly nucleated red cells, raises the possibility of diagnosis of disorders of liver protein expression in nucleated fetal red cells isolated from the first trimester maternal circulation.

Embryo, Mammalian↗
PubMed1995

Effects of hypophysectomy and thyroxine on the expression of hepatic oestrogen, hydroxysteroid and phenol sulphotransferases.

Sulphation in rats, and other mammals, is carried out by a family of sulphotransferase isoenzymes, which can be further subdivided into oestrogen, hydroxysteroid and phenol sulphotransferases. We have examined the effects of hypophysectomy on the activity and expression of representative members of the three major sulphotransferase sub-families in male Wistar rat liver cytosols, and have found that the different sub-families are subject to differential regulation by pituitary hormones. Our data show that in male rat liver hydroxysteroid sulphotransferases activity was increased, oestrogen sulphotransferases activity was not altered and phenol sulphotransferases activity was reduced. Further, we have studied the effect on sulphotransferase expression of administration of thyroxine and dexamethasone to hypophysectomized rats. Treatment of hypophysectomized rats with thyroxine virtually abolished oestrogen sulphotransferase activity in male rat liver but had no effect on hydroxysteroid sulphotransferase or phenol sulphotransferase activity. Treatment of hypophysectomized rats with dexamethasone had no effect on sulphotransferase activities. Quantitative immunoblot analysis of liver cytosols showed that these changes in enzyme activity were related to changes in levels of the respective enzyme proteins.

Animals↗
PubMed1995

Fatty acyl-CoA esters inhibit glucose-6-phosphatase in rat liver microsomes.

In native rat liver microsomes glucose 6-phosphatase activity is dependent not only on the activity of the glucose-6-phosphatase enzyme (which is lumenal) but also on the transport of glucose-6-phosphate, phosphate and glucose through the respective translocases T1, T2 and T3. By using enzymic assay techniques, palmitoyl-CoA or CoA was found to inhibit glucose-6-phosphatase activity in intact microsomes. The effect of CoA required ATP and fatty acids to form fatty acyl esters. Increasing concentrations (2-50 microM) of CoA (plus ATP and 20 microM added palmitic acid) or of palmitoyl-CoA progressively decreased glucose-6-phosphatase activity to 50% of the control value. The inhibition lowered the Vmax without significantly changing the Km. A non-hydrolysable analogue of palmitoyl-CoA also inhibited, demonstrating that binding of palmitoyl-CoA rather than hydrolysis produces the inhibition. Light-scattering measurements of osmotically induced changes in the size of rat liver microsomal vesicles pre-equilibrated in a low-osmolality buffer demonstrated that palmitoyl-CoA alone or CoA plus ATP and palmitic acid altered the microsomal permeability to glucose 6-phosphate, but not to glucose or phosphate, indicating that T1 is the site of palmitoyl-CoA binding and inhibition of glucose-6-phosphatase activity in native microsomes. The type of inhibition found suggests that liver microsomes may comprise vesicles heterogeneous with respect to glucose-6-phosphate translocase(s), i.e. sensitive or insensitive to fatty acid ester inhibition.

Acyl Coenzyme A↗
PubMed1995

Identification of an ATP-dependent copper transport system in endoplasmic reticulum vesicles isolated from rat liver.

1. This paper identifies and characterizes an ATP-dependent copper transport system in endoplasmic reticulum vesicles isolated from male rat liver. 2. The transporter has a Km of 2.5 +/- 1.2 mumol 1(-1) copper glutathione (CuGSH) and a Vmax of 4.5 +/- 1.3 nmol (mg protein)-1 (5 min)-1 for copper. 3. At a copper concentration of 2 mumol l-1, ATP dependence reaches saturation, with a Km for ATP of 4.7 +/- 2.4 mmol l-1 and a Vmax of 2.8 +/- 0.6 nmol (mg protein)-1 (5 min)-1. 4. The uptake is dependent on ATP hydrolysis, since a low energy analogue of ATP, adenosine 5'-[beta-gamma-methylene] triphosphate tetralithium (AMP.PCP), has no effect on copper uptake. 5. The transporter is a P-type ATPase, since vanadate inhibits uptake with a high degree of specificity (100 mumol l-1 inhibits uptake by 50% at a copper concentration of 2 mumol l-1).

Adenosine Triphosphatases↗
PubMed1995

Multiple transport protein defects in a patient with glycogen storage disease type 1: GSD 1b/1c beta.

A male child presented at 5 months of age with vomiting, diarrhoea, hypoglycaemia and hepatomegaly. Histology on a frozen liver biopsy suggested glycogen storage disease (GSD), while biochemical analyses confirmed an elevated glycogen content and normal activities of the GSD enzymes with the proviso that a variant of GSD 1 should be considered. The patient presented at 9 months of age with severe lactic acidosis and hypoglycaemia. A glucagon tolerance test and galactose load test on the patient produced no glycaemic response. A second biopsy was obtained and appropriately handled for the investigation of variants of the glucose-6-phosphatase enzyme (G6Pase) complex. Results showed that the patient had a deficiency of two transport proteins of the G6Pase complex, namely glucose-6-phosphate translocase and pyrophosphate translocase, i.e. GSD 1b/1c beta. These results were confirmed by additional kinetic analyses which provided confirmation of the double translocase deficiency. Evidence for inhibitors to these translocases was not found. The patient's treatment has resulted in the hypoglycaemia now being well controlled; however, at 3 years of age, height and weight are markedly lagging and he is moderately developmentally delayed. Neutropenia has not been found and neutrophil function is normal. Double enzyme deficiencies are very rare and possible explanations which might lead to this phenotype are considered. This, to the authors' knowledge, is the first report of a double translocase deficiency causing GSD type 1.

Antiporters↗
PubMed1995

Induction of hepatic estrogen sulfotransferase expression by hypophysectomy in female rats.

We have examined the effects of hypophysectomy and treatment with thyroxine (T4) on enzyme activity and expression (as determined by immunoblot analysis) of members of the three principal sulfotransferase (ST) sub-families (phenol STs, PST; estrogen STs, EST; hydroxysteroid STs, HST) in cytosols prepared from female Wistar rat livers. The results demonstrate that in female rat liver cytosol, EST activity was decreased by treatment with T4, increased following hypophysectomy and that treatment of hypophysectomized animals with T4 also greatly reduced EST activity. T4 had no significant effect on PST or HST activity in normal animals, but it decreased HST activity in hypophysectomized rat liver cytosol. Immunoblot analysis of these cytosols with antibodies recognising HST and PST indicated that where changes in enzyme activity occurred they mirrored changes in enzyme protein expression. In normal adult female rat livers, EST protein is not expressed, and the small residual activity results predominantly from the action of HST. Hypophysectomy induced EST activity and the expression of EST enzyme protein in female rat liver cytosol, and T4 treatment of hypophysectomized animals reduced the activity to below normal levels without reducing the corresponding enzyme protein levels, indicating that T4 regulation of EST in females is via a post-translational mechanism.

Animals↗
PubMed1995

The human adrenal microsomal glucose-6-phosphatase system.

Microsomal glucose-6-phosphatase (EC 3.1.3.9) is a multicomponent enzyme system traditionally thought only to be present in gluconeogenic tissues. The enzyme is associated with transport systems, for its substrate glucose-6-phosphate, and its products phosphate and glucose. It has been shown, using immunohistochemical methods and monospecific antibodies, that the component proteins of the enzyme system are present in human embryonic and fetal adrenal gland and are predominantly located in the fetal zone with lesser reactivities in the definitive zone. In addition, specific glucose-6-phosphatase activity was shown, and the rates of entry of glucose-6-phosphate, phosphate, and glucose into microsomes isolated from human fetal adrenals were measured. Although the complete enzyme system is present, the ratio of the component activities and comparison with human fetal and adult liver indicate that the regulation of the adrenal and liver glucose-6-phosphatase systems is different. In the human postnatal adrenal, immunoreactivies to the protein components decrease dramatically and are confined predominantly to the zona reticularis, suggesting a specialized role for adrenal glucose-6-phosphatase in fetal life.

Adrenal Glands↗
PubMed1995

Differential expression of hepatic oestrogen, phenol and dehydroepiandrosterone sulphotransferases in genetically obese diabetic (ob/ob) male and female mice.

Sulphotransferases (STs) are a family of closely related enzymes playing a key role in regulation of the bioavailability and activity of important endogenous molecules such as steroid hormones. A relationship between the expression of steroid STs and the diabetic state has been demonstrated in various laboratory animal models, and steroid sulphates such as dehydroepiandrosterone sulphate are known to have anti-diabetic properties. In order to further our understanding of the molecular basis for the association of steroid hormone sulphation and diabetes, we have examined the expression of oestrogen, phenol and dehydroepiandrosterone (DHEA) STs in mice carrying the obesity mutation (ob), which in the homozygous state (ob/ob) produces mice which are obese and diabetic. Our data show that, in male mice, ST activities towards oestrone (E1), oestriol (E3), DHEA and the xenobiotic 1-naphthol are elevated in ob/ob mice, whereas in female mice, only the oestrogen ST activities were elevated, with the DHEA and 1-naphthol ST activities reduced. Using antibodies directed against oestrogen ST, it was demonstrated that the induction of E1 and E3 ST activity in ob/ob mice correlated with the expression of an ST isoenzyme not constitutively expressed in control mouse liver.

Animals↗
PubMed1995

The glucose-6-phosphatase system in human development.

The classical role of glucose-6-phosphatase in liver and kidney is the production of glucose for release into blood. In liver, glucose-6-phosphatase catalyses the terminal step of glycogenolysis and gluconeogenesis. Abnormally low hepatic glucose-6-phosphatase activity is found in human genetic deficiencies i.e. glycogen storage disease type I and in cases of developmental delay, found predominantly in preterm infants. In contrast, abnormally high liver glucose-6-phosphatase occurs in poorly controlled or untreated diabetes mellitus. Hepatic glucose-6-phosphatase is an integral endoplasmic reticulum (and nuclear membrane) protein and it is part of a multicomponent system. Its active site is situated inside the lumen of the endoplasmic reticulum and transport proteins are needed to allow its substrates glucose-6-phosphate (and pyrophosphate) and its products phosphate and glucose to cross the endoplasmic reticulum membrane. In addition, a calcium binding protein is also associated with the glucose-6-phosphatase enzyme. Immunohistochemical studies, in combination with image analysis, have shown that glucose-6-phosphatase is present in liver and kidney and also in specific cell types in a variety of human tissues, for example Leydig cells in the testis and some astrocytes in the brain. Where practicable, enzymatic analysis, direct transport assays and/or immunological detection of the endoplasmic reticulum glucose and phosphate transport proteins have been used to demonstrate the presence and activity of the whole glucose-6-phosphatase system. The distribution of the human glucose-6-phosphatase system changes dramatically during development with a different spatial and temporal pattern in each tissue. The most unexpected localization was in circulating, predominantly nucleated, embryonic and early fetal red blood cells.

Glucose-6-Phosphatase↗
PubMed1994

Immunohistochemical localisation of glucose-6-phosphatase in developing human kidney.

The objective of our study was to determine the cellular localisation of glucose-6-phosphatase in developing human kidney using monospecific antiserum and a standard immunohistochemical method (peroxidase-antiperoxidase, PAP) on formalin fixed and paraffin embedded tissue. In embryonic and early fetal development of the metanephric kidney, glucose-6-phosphatase is located primarily in derivatives of the ureteric bud such as the pelvis, calyces and collecting ducts. In mid-fetal life as nephrons evolve and develop they become increasingly immunoreactive to glucose-6-phosphatase, such that in mature metanephric kidney the proximal tubules are highly reactive for glucose-6-phosphatase with other elements of the nephron also immunopositive albeit at lower reactivities. In addition the parietal layer of Bowman's capsule and some cells of the visceral layer are immunopositive. Only with the development of nephrons does the early predominance of glucose-6-phosphatase immunoreactivity to ureteric bud derivatives change: in mature kidney the reactivity in the collecting ducts is a small proportion of the total. In proximal tubular cells the distribution of glucose-6-phosphatase immunoreactivity is relatively uniform throughout development in contrast to collecting ducts where in fetal life this reactivity is displaced to the apices and basal areas by intracellular glycogen deposits. The mesonephric kidney has a similar pattern of glucose-6-phosphatase immunoreactivity to that of metanephric kidney. The availability of monospecific antiserum to glucose-6-phosphatase and immunohistochemical methods now allows an alternative approach to cellular localisation. Many of the difficulties in the fixation of tissue and assay of glucose-6-phosphatase activity inherent in conventional histochemical methods are avoided by such methods.

Female↗
PubMed1994

Sulfation of endogenous compounds and xenobiotics--interactions and function in health and disease.

Sulfation is a major detoxication mechanism for endogenous compounds and xenobiotics performed by a family of sulfotransferase isoenzymes. Understanding the normal cellular functions of these different sulfotransferases and the way in which endogenous and exogenous factors are able to influence their activity and expression will provide us with the information necessary to develop novel therapeutic strategies for conditions where sulfation may be implicated. This concept is discussed and is illustrated by examples including adverse drug reactions, fetal development and cancer.

Amino Acid Sequence↗