Muriel Feigelson

The differential effects of glucocorticoid on tissue and plasma amino acid levels.

Abstract The alteration in free amino acid patterns in various responsive tissues and in plasma has been investigated 4 h after the administration of a relatively low dose of cortisone acetate. In liver, in which cortisone evokes elevated tyrosine transaminase ( L -tyrosine: 2-oxoglutarate aminotransferase, EC 2.6.1.5) levels, a marked depression in the level of tyrosine and an elevation in concentrations of glutamate, aspartate, and alanine are evident. In the liver, in which protein synthesis is augmented under corticoid influences, the concentrations of numerous other amino acids were depressed following cortisone administration. In tissues in which cortisone produces a decrease in protein content, namely, thymus and muscle, the steroid elicited smaller elevations in the concentrations of many amino acids in addition to marked increments in the amounts of glutamate, aspartate, and alanine. Thus the alterations in amino acid patterns in tissues responsive to glucocorticoid seem to be consequent to the hormonally induced elevation of tyrosine α-ketoglutarate transaminase in the liver and to protein loss from lymphoid tissues and muscle.

Metabolic effects of glucocorticoids as related to enzyme induction

Abstract The concept that glucocorticoid induction of the hepatic amino acid metabolizing enzymes, tyrosine-α-ketoglutarate transaminase and tryptophan pyrrolase, assumes an important role in the processes by which this hormone manifests many of its regulatory functions has been investigated from several points of view. In a search for the mechanisms by which glucocorticosteroids effect induction of these enzymes, inquiry into the effects of cortisone on the biosynthetic pathway leading to RNA was undertaken. It was found that incorporation in vivo of isotopically labeled precursors into liver RNA was rapidly and markedly enhanced after administration of a single low dose of cortisone, with a time course parallel to that of glucocorticoid induction of increased enzyme protein and activity. However, the following evidence has been accumulated which suggests that little, if any, of the observed stimulation in isotopic incorporation into RNA after cortisone administration is attributable to enhanced elaboration of RNA molecules from their nucleotide precursors: (1) the magnitude of cortisone induced stimulation of incorporation of various labeled precursors into liver RNA varies markedly, glycine- 14 C > inorganic 32 P > rotate- 3 H; (2) cortisone administration rapidly elevates incorporation in vivo of glycine- 14 C into liver acid soluble adenine nucleotides to a greater extent and with similar kinetics as incorporation of this precursor into RNA; (3) the stimulation of isotope incorporation into RNA of all liver subcellular compartments are of similar orders of magnitude and time course; (4) accumulation of increased quantities of hepatic RNA is observed only many hours after large doses of cortisone, at a time when enzyme induction has already subsided; (5) in vitro studies provide no evidence of glucocorticosteroid intervention in the synthesis of RNA on the DNA template, since neither binding of glucocorticoids with purified rat liver DNA nor alteration of the secondary structure of native DNA by these steroids could be detected. The only isotopic evidence suggesting possible enhancement of RNA elaboration following glucocorticoid administration is the observation of a slight stimulation in incorporation of orotate- 3 H into hepatic RNA which exceeds a yet smaller stimulation in incorporation of this isotope into a pyrimidine nucleotide precursor pool. That stimulation of hepatic purine biosynthesis does occur as a marked and early response to glucocorticoids has been established. Investigation of the mechanism of hormonal regulation of purine metabolism has revealed that intraperitoneal administration to adrenalectomized rats of l -glutamate, l -alanine, l -leucine, their d -counterparts, inorganic ammonium salts and l -glutamine, all mimic cortisone in the stimulation of glycine- 14 C incorporation into acid-soluble adenine. It has been accordingly proposed that glucocorticoid induction of transaminase activity may result in sequential expansion in the routes of amino nitrogen transport, leading to the observed enhancement in purine nucleotide biosynthesis—“purinoneogenesis”—as well as gluconeogenesis and increased urea output, all of which are characteristic of the cortisonized animal. Changes in liver amino acid patterns following cortisone administration are consistent with this working hypothesis: liver tyrosine levels are depressed; concentrations of glutamate and aspartate, amino acids pivotal in transamination, are elevated. Extension of these studies to extra hepatic target tissues, has revealed that simultaneous with stimulated glycine-2- 14 C incorporation into protein and acid-soluble purine in the liver, cortisone evokes a marked depression in these metabolic pathways in the lymphoid tissues, thymus and spleen.

Kinetics of liver nucleic acid turnovers during enzyme induction in the rat

Abstract The parental administration of tryptophan in the rat produces a transient elevation in liver protein synthesis and in liver tryptophan peroxidase activity which results in an increased incorporation of inorganic 32 P and [ 14 C]glycine into RNA. The relative kinetics of these phenomena indicate that there is a burst of RNA synthetic activity subsequent to the period of active enzyme protein synthesis and that this increased RNA synthesis persists for 8 h and then returns to control rates. A working hypothesis is proposed in which it is suggested that increased enzyme protein synthesis utilizes and consumes pre-existent RNA and that this RNA is sub subsequently replenished. This hypothesis implies that RNA serves as a reactant which is consumed, rather than as a perfect catalyst of protein synthesis.

Studies on the mechanisms of histidase development in rat skin and liver: I. Basis for tissue specific developmental changes in catalytic activity

Abstract Skin and liver histidases of the rat follow markedly differing developmental courses. In skin, the enzyme activity emerges in late fetal life, rises to a maximum during the first postnatal week and then declines; in liver, histidase activity initially appears in the neonatal period and progressively rises thereafter. The enzymes in both tissues, however, are immunologically identical, as demonstrated by Ouchterlony double diffusion, immunoelectrophoresis, and immunotitration analyses of histidase in adult animals. Furthermore, although crude preparations of skin and liver histidase differ slightly in electrophoretic mobility on acrylamide gels, the isoelectric points and Michaelis constants of the enzyme from both skin and liver are the same. During development neither dissociable activators nor inhibitors of the enzyme are evident, which might account for the divergent developmental patterns, nor are enzyme inhibitors found in tissues devoid of histidase activity. No immunologically cross-reacting materials are detectable in skin and liver at developmental stages prior to the emergence of enzyme activity, nor in tissues not expressing histidase catalytic activity. Furthermore, there is no evidence of elaboration of immunological variants of histidase in skin and liver during development. These data are compatible with the view that all developmental changes in catalytic activity are due to altered amounts of the same enzyme antigen.

Hypophyseal regulation of hepatic histidase during postnatal development and adulthood: II. Pituitary-estrogen interrelationship

Abstract Estrogen, which augments rat liver histidase activity in pre- and postpubertal rats, is ineffective in hypophysectomized animals, in which enzyme activities are already elevated. This failure of estrogen action has been demonstrated in hypophysectomized weanling, as well as adult females, the former of which manifest enzyme activities considerably below maximum possible values. The following agents, which are known to suppress histadase activity under certain conditions, were ineffective in permitting estrogen stimulation of histadase in hypophysectomized rats: glucocorticoid, testosterone, adrenocorticotrophin acting extra-adrenally and via the adrenals, growth, and preparations of whole pituitaries of bovine or female rat origin. The following materials, which do not themselves alter enzymic activity, were likewise incapable of permitting estrogenic enhancement of histadase activity in these animals: follicle-stimulating hormone and luteinizing hormone acting extra-gonadally, thyrotrophic hormone, and bovine posterior pituitary preparations. It is thus concluded that estrogen does not act directly on the liver to induced histadase activity, but requires participation of the hypophysis. However, estrogen does not seem to function by reversing any known pituitary derived suppressors of this enzyme, nor do pituitary factors seem to facilitate estrogen augmentation of the enzyme. These findings, however, would be compatible with the hydrolysis that estrogen may enhance liver histadase activity by inhibiting the secretion of some, as yet, unidentified enzyme-suppressing pituitary factor. Since the uterotrophic action of estrogen does not manifest hydrophyseal dependence, estrogenic effects on the uterus and on hepatic histadase are mediated by distinct mechanisms.

Effects of maternal diabetes on the levels, synthetic rates and activities of synthetic enzymes of surface-active phospholipids in perinatal rat lung.

Streptozotocin-induced maternal diabetes in the rat has been found to reduce selectively the content and synthetic rates of disaturated phosphatidylcholine and lysophosphatidylcholine in the lungs of term fetuses. Furthermore, the elevations in these parameters which occur during normal pulmonary maturation between the final gestational day and the first neonatal day are abolished or markedly curtailed, resulting in significantly reduced levels and synthetic rates of surfactant and its immediate precursor in the neonatal lung. In addition, complete or partial inhibition of the perinatal developmental rise in the activities of key enzymes catalyzing de novo phosphatidylcholine synthesis in the lung, viz., cholinephosphate cytidylyltransferase and cholinephosphotransferase, and of enzymes catalyzing reacylation of unsaturated to disaturated phospholipid, viz., lysophosphatidic acid and lysophosphatidylcholine acyltransferases, has been observed, resulting in reduced activities of these enzymes in the neonate. The observed reductions in surface-active phospholipid synthesis in the lungs of offspring of diabetic mothers may be related to these lowered enzyme activities, as well as to deficiencies in carbohydrate precursors available for phospholipid synthesis, as reported in previous studies. It is suggested that the hyperinsulinemia manifested in fetuses of diabetic mothers opposes the tendency of corticosteroids to enhance surface-active phospholipid synthesis, resulting in pulmonary surfactant deficiency and thus the propensity for neonatal respiratory distress.

Effects of maternal diabetes on the development of carbohydrate metabolizing enzymes in fetal rat liver

Abstract Effects of streptozotocin-induced maternal diabetes on fetal hepatic carbohydrate-metabolizing enzyme development and hormonal status has been explored in the rat. Hepatic glycogen synthase a activity of the normal fetus rose to a maximum at 20 days of gestation, then fell prior to parturition. In fetuses of diabetic mothers, this prepartum decline was curtailed, resulting in enhanced synthase a activity and increased glycogen content in fetal livers at term. Elevation in hepatic synthase a in fetuses of diabetic mothers was due, not to altered interconversion between existing synthase a and b, but to equivalent increases in both forms of the enzyme. Both hepatic and free plasma corticosterone levels were elevated in fetuses of diabetic mothers and may be responsible for the enhanced development of total glycogen synthase observed in these fetuses. In normal fetuses hepatic phosphofructokinase and pyruvate kinase activities also rose to maxima at 20 days, then declined prior to term. In fetuses of diabetic mothers pyruvate kinase activity attained higher than normal maximal levels and phosphofructokinase activity fell more gradually, thus resulting in elevations in both enzyme activities at term. Augmentations in these glycolytic enzymes are compatible with hyperinsulinemia observed in fetuses of diabetic mothers. The following conclusions may be drawn from these findings. During late fetal life developmental patterns of rate-limiting hepatic glycogen-synthesizing and glycolytic enzymes are adapted to glucose utilization. In the normal fetus these patterns reverse at term, thereby promoting glucose mobilization, which prepares the fetus for abrupt deprivation of maternal glucose at birth. Maternal diabetes results in retardation of these reversal processes, presumably due to elevations in fetal glucocorticoid and insulin levels. Glycogenolytic and glucogenic capacities are thereby impaired in these fetuses.

Hypophyseal regulation of hepatic histidase during postnatal development and adulthood. I. Pituitary suppression of histidase activity.

Abstract Hypophysectomy at 3 weeks of age effects precocious development augmentation of hepatic histidase activity in both the maturing male and female rat; pituitary excision likewise augments this enzymic activity in adult males and in ovariectomized but not in gonad-bearing, adult females. This enhanced histidase activity following hypophysectomy is entirely blocked when liver protein synthesis is inhibited by administration of l -ethionine. Attempts to ascertain the identity of the histidase repressive factor(s)_have implicated the anterior pituitary as their site of origin. Since hypophysectomy of castrated-adrenalectomized males results in elevation in histidase activity, it is thought that the hypophysis secretes principle(s) suppressive to hepatic histidase which are supplementary to and distinct from trophic influences on the testis and adrenal, which themselves also contribute to suppression of histidase under certain conditions. In hypophysectomized animals lacking target glands corresponding to particular pituitary trophic hormones, follicle-stimulating hormone and luteinizing hormone, alone and in combination, and thyrotrophic hormone are without effect, whereas growth hormone and adrenocorticotrophic hormone both suppress hepatic histidase activity. These, and possibly other as yet unidentified, hypophyseal repressive influences contribute to the establishment of the characteristic postnatal developmental pattern and to the maintenance of adult levels of hepatic histidase in the rat.

Alterations in nucleic acid turnovers in subcellular components during tryptophan peroxidase induction

Abstract Parental tryptophan administration to the rat leads to tryptophan peroxidase induction and to acceleration in the turnover rates of the RNA of the various liver subcellular fractions. The kinetics of these processes indicate that the stimulation in RNA turnover does not precede or accompany but, rather, is subsequent to the period of active enzyme protein synthesis. The maximum percentage increases in RNA turnover rates are: mitochondria and microsomal RNA, 260%; soluble RNA, 154%; nuclear RNA, 60%. However, due to the markedly different basal turnover rates of RNA in the various subcellular organelles, the absolute increase in total amount of RNA synthesized as a result of enzyme induction are in the order: nuclear > soluble = microsomal > mitochondrial. The kinetic data are compatible with the hypothesis that enzyme induction results in extra nuclear RNA utilization with subsequent resynthesis largely in the nucleus.

Effects of Maternal Diabetes on the Development of Carbohydrate-Metabolizing Enzymes, Glycogen Deposition and Surface Active Phospholipid Levels in Fetal Rat Lung

Streptozotocin-induced maternal diabetes has been shown to alter developmental patterns of carbohydrate-metabolizing enzyme activities, glycogen deposition and surfactant levels in late fetal rat lung in a tissue-specific manner, as follows: (a) marked reduction in glycogen synthase a activity, due to aberrant interconversion between active and inactive forms of the enzyme; less glycogen was thus accumulated; (b) lowered activities of hexokinase, phosphofructokinase and pyruvate kinase at term; (c) reduced disaturated phosphatidylcholine (surfactant) concentrations. The diminished synthesis and accumulation of glycogen and glycolytic capacity in the lungs of fetuses of diabetic mothers has been related to reduction in surfactant level, which underlies respiratory distress syndrome frequently encountered in neonates of diabetic pregnancies.