Androgen receptors in Klinefelter's syndrome.
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Adolescent↗
Biomedical subjects
U Keller
Publications and source records attributed to U Keller.
The initiation of peptide formation in the biosynthesis of actinomycin.
Actinomycin Synthetase II (ACMS II), which activates threonine and valine by a thioltemplate mechanism during the synthesis of the actinomycin half-molecule 4-methyl-3-hydroxyanthranilic acid (4-MHA) pentapeptide lactone, was purified to near homogeneity from Streptomyces chrysomallus. It is a single polypeptide chain of M(r) 280,000 and contains 4'-phosphopantetheine as a covalently bound prosthetic group. ACMS II charges itself with threonine but not with the 4-MHA analogue p-toluic acid via a specific sulfhydryl group at the expense of ATP. Charging of ACMS II with p-toluic acid in thioester linkage took place, however, only when actinomycin synthetase I (ACMS I), a 4-MHA-AMP ligase, was present. In the additional presence of L-threonine, enzyme-bound p-toluyl-L-threonine was formed on ACMS II. The latter compound was also formed when chemically synthesized p-toluic acid adenylate was added instead of ACMS I and p-toluic acid. This indicates that p-toluic acid adenylate is a free intermediate in the reaction and that charging of the enzyme and acylation of threonine are both catalyzed by ACMS II rather than by ACMS I. Chemically synthesized thioesters of p-toluic acid and coenzyme A, pantetheine, or beta-alanyl-cysteamine reacted with ACMS II, threonine, and ATP with formation of enzyme-bound p-toluyl-threonine. In contrast, p-toluyl-cysteamine thioester was inactive, which indicates structural constraints in the reactivity of free thioesters of p-toluic acid with ACMS II. Such constraints obviously require structural similarity of the artificial substrate to a p-toluic acid thioester formed on the enzyme's surface in the course of the reaction. Since free coenzyme A was not involved in the charging of p-toluic acid or in p-toluyl-threonine formation, the sulfhydryl group of the 4'-phosphopantetheine cofactor is most likely the primary acceptor of p-toluic acid (or 4-MHA) in the initiation of peptide lactone formation.
Amino Acid Sequence↗
[Intra-individual variations in triglyceride and total cholesterol levels in serum in mixed hyperlipidemia].
Swings in serum concentrations of total cholesterol and triglycerides were assessed retrospectively in 12 of 60 patients with mixed hyperlipidaemia (ten men, two women; mean age 44 [38-55] years). A second disease as a cause of the hyperlipidaemia had been excluded. Despite clinical stability and unchanged drug therapy the fasting values of triglycerides, measured enzymatically, in a minimum of four different samples (over several months) differed by more than 5 mmol/l, a mean of 16.6 (5-41.9) mmol/l. The lowest values were 2.7 (1.3-7.5) mmol/l, the highest 17.9 (6.4-44.6) mmol/l. Total cholesterol concentrations varied around 5.5 (1.6-31.7) mmol/l, minimal values 5.7 (4.2-8.9) mmol/l, maximal ones 12.0 (6.9-37.4) mmol/l. Six of the twelve patients consumed more than 60 g alcohol daily. The cause of the marked variations between individual samples is uncertain. Marked swings in triglyceride and total cholesterol concentrations are likely in mixed hyperlipidaemia. It is, therefore, essential to measure these concentrations repeatedly to assess correctly the diagnosis and treatment.
Adult↗
Noninvasive observation of hepatic glycogen formation in man by 13C MRS after oral and intravenous glucose administration.
The formation of glycogen in the liver of normal volunteers was followed noninvasively with 13C magnetic resonance spectroscopy (MRS) under two different conditions: a) intravenous infusion of [1-13C]glucose under hyperglycemic and hyperinsulinemic clamp conditions, and b) oral intake of glucose in the form of a bolus. For the intravenous infusion, [1-13C]glucose with an enrichment level of 99% was employed. The C1 signals of alpha- and beta-glucose could be detected in the human liver already after an infusion period of 8 min. However, an increase in the glycogen signal was observed only after a prolonged infusion of about 60 min. Changes in the glycogen signal correlated well with the time course of insulin and glucagon during the measurement. Experiments showed also that liver glycogen formation in man can be followed noninvasively by 13C-MRS using nonlabeled glucose or [1-13C]glucose with a low level of enrichment (6.6%). The use of nonlabeled glucose may therefore simplify the quantitation of net liver glycogen synthesis since it can be based directly on changes in the natural abundance 13C MRS glycogen signal, avoiding label dilution through the various metabolic pathways of glucose. The glucose uptake, estimated from the increase in the glycogen signal, was consistent with findings from more complex and invasive studies of glucose uptake in the liver. The average liver glycogen concentration in 12 h overnight fasted volunteers (n = 18) without any special dietary preparation was assessed to be 229 +/- 34 mM (minimum = 160 mM; maximum = 274 mM).
Administration, Oral↗
Pathophysiology of cancer cachexia.
Patients with advanced cancer and cachexia typically demonstrate modestly increased rates of energy expenditure in the presence of diminished food intake due to anorexia and to gastrointestinal disturbances. Rates of glucose production by the liver, gluconeogenesis and glycolysis to lactate (Cori cycle) are increased, fat mobilisation and oxidation are accelerated. There is a redistribution of body proteins away from muscle towards visceral proteins, resulting in marked muscle protein loss. Cancer cachexia differs from simple starvation and demonstrates metabolic similarities to sepsis or polytrauma. The metabolic response in the patient with cancer is largely due to mediators released by the tumour or by the host; recently the role of cytokines such as tumour necrosis factor alpha (TNF alpha), interleukin-1 (IL-1) and -6 (IL-6) and interferon gamma (INF gamma) has been emphasized. Catabolic hormones such as glucocorticoids and adrenaline have also been implicated. Cytokines have the potential to reproduce experimentally the clinical syndrome of cancer cachexia. There is evidence of increased production of several of them in certain types of cancer. There are overlapping activities of the cytokines TNF alpha, IL-1, IFN gamma and IL-6. The contribution of each of them to cancer cachexia remains unclear. Inhibition of cytokine activity using specific antibodies in cancer-bearing experimental animals demonstrated partial prevention of cachexia. A positive feedback between macrophage-derived IL-1 and tumour-derived IL-6 has been demonstrated recently in experimental cancer cachexia. Cytokines may support tumour growth by acting as growth factors.
Cachexia↗
Effects of recombinant human insulin-like growth factor I and insulin on counterregulation during acute plasma glucose decrements in normal and type 2 (non-insulin-dependent) diabetic subjects.
Insulin-like growth factor I (65 micrograms/kg) or insulin (0.1 IU/kg) were injected i.v. on two separate occasions in random order in normal and in Type 2 (non-insulin-dependent) diabetic subjects. Insulin-like growth factor I and insulin injection resulted in identical decrements of plasma glucose concentrations after 30 min but in delayed recovery after insulin-like growth factor I as compared to insulin in both groups (p < 0.05 insulin-like growth factor I vs insulin). Counterregulatory increases in plasma glucagon, adrenaline, cortisol and growth hormone concentrations after hypoglycaemia (1.9 +/- 0.2 mmol/l) in normal subjects were blunted after insulin-like growth factor I administration compared to insulin (p < 0.05). Plasma glucose in Type 2 diabetic subjects did not reach hypoglycaemic levels but the acute glucose decrease to 4.5 +/- 0.8 mmol/l was associated with significantly lower responses of plasma glucagon and adrenaline but higher cortisol levels after insulin-like growth factor I compared to insulin (p < 0.003). Plasma concentrations of non-esterified fatty acids and leucine decreased similarly after insulin-like growth factor I and insulin in both groups. The present results demonstrate that insulin-like growth factor I is capable of mimicking the acute effects of insulin on metabolic substrates (plasma glucose, non-esterified fatty acids, leucine). The decreases of plasma glucose were similar after both peptides in normal and in diabetic subjects who were presumably insulin resistant. Counterregulatory hormone responses to plasma glucose decrements differed, however, between insulin-like growth factor I and insulin and in the diabetic and the control subjects.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗
Comparison of the effects of recombinant human insulin-like growth factor-I and insulin on glucose and leucine kinetics in humans.
To compare the metabolic effects of elevated plasma concentrations of IGF-I and insulin, overnight-fasted normal subjects were studied twice, once receiving IGF-I and once insulin at doses that resulted in identical increases in glucose uptake during 8-h euglycemic clamping. Recombinant human IGF-I or insulin were infused in one group at high doses (30 micrograms/kg per h IGF-I or 0.23 nmol/kg per h insulin) and in another group at low doses (5 micrograms/kg per h IGF-I or 0.04 nmol/kg per h insulin). Glucose rate of disappearance (measured by [6,6-D2]-glucose infusions) increased from baseline by 239 +/- 16% during high dose IGF-I vs 197 +/- 18% during insulin (P = 0.021 vs IGF-I). Hepatic glucose production decreased by 37 +/- 6% during high dose IGF-I vs 89 +/- 13% during insulin (P = 0.0028 vs IGF-I). IGF-I suppressed whole body leucine flux ([1-13C]-leucine infusion technique) more than insulin (42 +/- 4 vs 32 +/- 3% during high doses, P = 0.0082). Leucine oxidation rate decreased during high dose IGF-I more than during insulin (55 +/- 4 vs 32 +/- 6%, P = 0.0001). The decreases of plasma concentrations of free fatty acids, acetoacetate, and beta-hydroxybutyrate after 8 h of IGF-I and insulin administration were similar. Plasma C-peptide levels decreased by 57 +/- 4% during high doses of IGF-I vs 36 +/- 6% during insulin (P = 0.005 vs IGF-I). The present data demonstrate that, compared to insulin, an acute increase in plasma IGF-I levels results in preferential enhancement of peripheral glucose utilization, diminished suppression of hepatic glucose production, augmented decrease of whole body protein breakdown (leucine flux), and of irreversible leucine catabolism but in similar antilipolytic effects. The data suggest that insulin-like effects of IGF-I in humans are mediated in part via IGF-I receptors and in part via insulin receptors.
3-Hydroxybutyric Acid↗
Effects of endotoxin on leucine and glucose kinetics in man: contribution of prostaglandin E2 assessed by a cyclooxygenase inhibitor.
The effects of endotoxin (E) administration on whole body protein and glucose metabolism were studied in normal volunteers. Injection of 4 ng/kg Escherichia coli E iv resulted in a relative increase in leucine flux (1-13C-leucine infusion technique) compared to controls [+0.12 +/- 0.10 vs. -0.45 +/- 0.23 mumol/kg.min after 360 min, P = 0.028, analysis of variance (ANOVA)], indicating increased proteolysis. Nonoxidative leucine flux was higher after E than after saline administration (0.08 +/- 0.11 vs. -0.47 +/- 0.18 mumol/kg.min, P = 0.007, ANOVA), suggesting increased amino acid incorporation into proteins. E caused a transient decrease of plasma glucose concentration (by 0.5 +/- 0.1 mmol/L after 150 min; P < 0.004 vs. saline controls) due to a relative increase in disappearance compared to appearance of glucose (6,6 D2-glucose infusion technique). These alterations were associated with increases in plasma concentrations of ACTH, beta-lipoprotein (beta-LPH), GH, cortisol, epinephrine, free fatty acid, beta-hydroxybutyrate, and decreases of plasma insulin. Pretreatment with ibuprofen, a cyclooxygenase inhibitor, blunted the effects of E on whole body leucine flux (P < 0.05 vs. E) and on nonoxidative leucine flux (P < 0.05 vs. E) but enhanced the E-induced decrease of plasma glucose concentration (P < 0.004 vs. E), due to a relative increase in glucose disappearance compared to appearance (P = 0.02). The increases in counterregulatory hormones (ACTH, beta-LPH, GH, cortisol, epinephrine) were also attenuated by ibuprofen. Thus, acute endotoxinemia results in a redistribution of whole body proteins due to an increase in both protein breakdown and amino acid incorporation into proteins and in decreased plasma glucose concentrations. The ibuprofen data suggested that these effects of E on leucine kinetics, but not on glucose metabolism, were prostaglandin E2-mediated.
Adult↗
Immunological identification of a 50 kDa Mr FK506-binding immunophilin as a component of the non-DNA binding, hsp90 and hsp70 containing, heterooligomeric form of the chick oviduct progesterone receptor.
p59/HBI, a Heat shock protein (hsp90)-Binding-Immunophilin of approximately 59 kDa, was originally detected in the heterooligomeric, non-DNA binding form of numerous mammalian steroid hormone receptors. P59/HBI belongs to the FKBP family since it binds the immunosuppressants FK506 and Rapamycin. Using immobilized FK506, we purified [3H]Org 2058-progesterone receptor (PR)-hsp90-complexes from chick oviduct cytosol in a species migrating at 9S in density gradient. This led to suppose that a protein present in these complexes is a FK506 binding protein. Following incubation of this FK506-affinity purified 9S-PR with BF4, a specific anti-chick hsp90 monoclonal antibody, a shift of the [3H]Org 2058-PR complexes from 9S to 11S has been observed, indicating the presence of hsp90, hsp70 also is included in the 9S-PR complexes as demonstrated by Western blotting and density gradient experiments. Two lines of evidence support the notion that an immunophilin of 50 kDa also associates to heterooligomeric 9S-PR complexes. Firstly, a shift at 11S of the FK506 eluted 9S-PR occurs in sucrose gradient after incubation with 419, a new polyclonal antibody raised against a peptide corresponding to the Hinge I region [Callebaut I., Renoir J. M., Lebeau M. C., Massol N., Burny A., Baulieu E. E., Mornon J. P. (1992) Proc. Natl. Acad. Sci. USA 89, 6270-6274] of the rabbit p59/HBI (amino acids Phe135 to Gly149). Secondly, in Western blotting experiments, both 419 and another anti-FKBP antibody, raised against purified Streptomyces Crysomallus FKBP12 [Pahl A. and Keller U. (1992) J. Bacteriol., 3, 325-333], allow detection of the same 50 kDa protein in the affinity purified PR.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Polymorphic haplotypes and recombination rates at the LDL receptor gene locus in subjects with and without familial hypercholesterolemia who are from different populations.
RFLPs at the low-density lipoprotein (LDL) receptor locus for TaqI, StuI, HincII, AvaII, ApaLI (5' and 3'), PvuII, and NcoI were studied in Swiss and German families with familial hypercholesterolemia (FH). A total of 1,104 LDL receptor alleles were analyzed using Southern blotting and new PCR-based techniques for detection of the TaqI, StuI, HincII, AvaII, NcoI RFLPs. Two hundred fifty-six independent haplotypes from 368 individuals of 61 unrelated Swiss families, as well as 114 independent haplotypes from 184 subjects of 25 unrelated German families, were constructed. In 76 families, clinical diagnosis of FH was confirmed by cosegregation analysis. Of the 43 unique haplotypes consisting of seven RFLPs detected in the Swiss and Germans, only 9 were common in both population samples. Analysis of linkage disequilibrium revealed nonrandom associations between several of the investigated RFLPs. ApaLI (5'), NcoI, PvuII, TaqI, and AvaII or HincII were particularly informative (cumulative informativeness .85). Relative frequencies, heterozygosity indexes, and PICs of the RFLPs from the Swiss and Germans were compared with values calculated from reported haplotype data for Italians, Icelanders, North American Caucasians, South African Caucasians, and Japanese. Pairwise comparisons of population samples by common RFLPs demonstrated unexpected differences even between geographically adjacent populations (e.g., the Swiss and Germans). Furthermore, genetic distances from the Germans to the other Caucasians were larger than to the Japanese. An unexpected lack of correlation between linkage disequilibria and physical distances was detected for the German and Japanese data, possibly because of nonuniform recombination with excessively high rates between exon 13 and intron 15. Hence, the present study revealed a striking variety of polymorphic haplotypes and heterogeneity of RFLP frequencies and recombination rates among the seven population samples.
Base Sequence↗
[Hypoglycemia and multiple myeloma].
We report the case of a 78-year-old patient with recurrent attacks of severe fasting and late postprandial hypoglycemia, whose plasma showed highly elevated concentrations of immunoreactive insulin evidenced by high titers of spontaneous insulin and proinsulin-binding antibodies. Insulin autoimmune syndrome was diagnosed. Further investigations revealed a multiple myeloma of the kappa-light chain type. The monoclonal insulin-binding antibodies were characterized as IgG2-subclass and were identical with the paraprotein, thereby confirming that the insulin-binding antibodies were in fact produced by the myeloma. Together with initial symptomatic treatment, plasmapheresis was performed repeatedly to reduce the antibody pool. Subsequently octreotide therapy proved successful. The underlying myeloma was treated by chemotherapy.
Aged↗
Insulin autoimmune syndrome and HLA-DR4.
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Autoimmune Diseases↗
Purification and characterization of actinomycin synthetase I, a 4-methyl-3-hydroxyanthranilic acid-AMP ligase from Streptomyces chrysomallus.
Actinomycin synthetase I was purified to homogeneiety from actinomycin-producing Streptomyces chrysomallus. The purified enzyme is a single polypeptide chain of M(r) 45,000. It catalyzes the formation of the adenylate of 4-methyl-3-hydroxyanthranilic acid (4-MHA) from the free acid and ATP in an equilibrium reaction. 4-MHA is the precursor of the chromophoric part of actinomycin. By using the 4-MHA analogue, 4-methyl-3-hydroxybenzoic acid, as a model substrate it could be established that the equilibrium constant Keq is independent on enzyme concentration, which suggests that no stoichiometric acyladenylate-enzyme complex is formed in contrast to observations made with aminoacyl adenylates formed by aminoacyl-tRNA synthetases or multifunctional peptide synthetases. Actinomycin synthetase I does not charge itself with substrate carboxylic acid via a covalent thioester bond as is usual for amino acid activation in non-ribosomal peptide synthesis. In addition, the enzyme does not act as an acyl-coenzyme A ligase as revealed by its inability to release AMP in the presence of 4-MHA or other structurally related aromatic carboxylic acids, coenzyme A and ATP. Additional analysis of the activation reaction showed that it is exothermic, whereas the free enthalpy change delta G0 is positive due to a negative entropy change indicating a strong influence of restriction of random motion on the course of the reaction. Determinations of Km and kcat of various substrate carboxylic acids revealed the highest kcat/Km ratio for the natural substrate 4-MHA. From these properties, actinomycin synthetase I represents the prototype of novel chromophore activating enzymes involved in non-ribosomal synthesis of chromopeptide lactones in streptomycetes.
Adenosine Triphosphate↗
Relationship between the degree of unsaturation of dietary fatty acids and adipose tissue fatty acids assessed by natural-abundance 13C magnetic resonance spectroscopy in man.
Natural-abundance 13C magnetic resonance spectroscopy was used for determining noninvasively the relative concentration of mono- and polyunsaturated fatty acids of adipose tissue in two groups of volunteers. The first consisted of subjects who had followed a fat-reduced diet for at least half a year before the 13C measurements. The second were control subjects who were on a usual high-fat diet. The ratio of unsaturated to total fatty acids in adipose tissue determined by 13C MRS correlated significantly with the same ratio in fat of the diet composition estimated by a dietician according to food records. The results indicate that 13C MRS is capable of assessing the degree of unsaturation of dietary fatty acids consumed during the preceding months.
Adipose Tissue↗
Treatment of diabetic ketoacidosis and non-ketotic hyperosmolar diabetic coma.
Although mortality of diabetic ketoacidosis (KA) has decreased during the past 20 yr to 1-2%, hyperosmolar non-ketotic coma (HNC) is still lethal in 20-30% of cases due to severe underlying conditions or to complications. The most frequent causes of death are infections and thromboembolic disorders. The strategies of initial treatment of KA and HNC are similar; in KA, insulin, fluid and electrolyte replacement have first priority. In HNC, rehydration and electrolyte administration are of primary importance. It is now generally recognized that insulin therapy is best performed using low doses (4-8 units/h); after institution of insulin treatment and rehydration there are rapid changes of fluid and electrolytes from the extra- into the intravascular space. In this situation it is a major therapeutic challenge to avoid complications due to hypokalaemia, hypophosphataemia, hypomagnesaemia and hypovolaemia. These complications should be avoided by adequate replacement, and particularly by regular clinical and laboratory monitoring. When blood glucose concentrations decrease below 14 mmols/l, blood glucose concentrations should initially be maintained at this level because rapid lowering below this level may increase the risk of brain oedema. Too-vigorous fluid replacement with crystalline solutions also increases the risk that brain oedema or complications like the adult respiratory distress syndrome will develop. If hypovolaemia persists in spite of adequate crystalloid solutions, colloid-containing fluids such as albumin should be administered. It is not established whether replacements of phosphate and magnesium have clinical benefits. Nevertheless, it is probably justified to administer phosphate and magnesium when their serum concentrations are below the normal range, particularly if the clinical situation is critical. Mortality from diabetic coma in industrialized countries may only be decreased by prophylaxis, i.e. by education of all diabetic patients and physicians to detect metabolic decompensation early.
Diabetic Coma↗
Streptomycetes possess peptidyl-prolyl cis-trans isomerases that strongly resemble cyclophilins from eukaryotic organisms.
A functionally active 17.5 kDa peptidyl-prolyl cis-trans isomerase was purified to homogeneity from Streptomyces chrysomallus, a Gram-positive filamentous bacterium. Characterization of the enzyme revealed inhibition and binding characteristics, against the immunsuppressive drug cyclosporin A, which were similar to cyclophilins from eukaryotes such as mammals, plants, fungi and yeasts, but different from those of cyclophilins from enterobacteria such as Escherichia coli. The amino acid sequence of the S. chrysomallus cyclophilin, as deduced from the gene sequence, revealed a striking degree of amino acid sequence identity with the corresponding 17 kDa proteins of humans (66%), Neurospora (70%) and yeast (69%). Comparison with cyclophilin sequences from the Gram-negative enterobacteria revealed much less homology (25% identity with E. coli b, 23% identity with E. coli a). Cyclophilin was detected in each of the four other Streptomyces species tested. The cyclophilins from the various streptomycetes differed in size, varying between 17 and 20.5 kDa. The cyclophilins were abundant in the Streptomyces cells, and present throughout growth.
Amino Acid Isomerases↗
Effect of acute acidosis and alkalosis on leucine kinetics in man.
The effects of acute pH changes on whole body leucine kinetics (1-13C-leucine infusion technique) were determined in normal subjects. Plasma insulin, glucagon, and growth hormone concentrations were kept constant by somatostatin and replacement infusions of the three hormones. When acidosis was produced by ingestion of NH4Cl (4 mmol kg-1 p.os; n = 8) arterialized pH decreased within 3 h from 7.39 +/- 0.01 to 7.31 +/- 0.01 (P less than 0.001) and leucine plasma appearance increased by 0.13 +/- 0.04 mumol kg-1 min-1 (P less than 0.02); in contrast, when alkalosis was produced by intravenous infusion of 4 mmol kg-1 NaHCO3 (n = 7, pH 7.47 +/- 0.01), leucine plasma appearance decreased by -0.09 +/- 0.04 mumol kg-1 min-1 (P less than 0.01 vs. acidosis). Whole body leucine flux also increased during acidosis compared to alkalosis (P less than 0.05), suggesting an increase in whole body protein breakdown during acidosis. Apparent leucine oxidation increased during acidosis compared to alkalosis (P = 0.05). Net forearm leucine exchange remained unaffected by acute pH changes. Plasma FFA concentrations decreased during acidosis by -107 +/- 67 mumol l-1 (P less than 0.05) and plasma glucose increased by 1.90 +/- 0.25 mmol l-1 (P less than 0.02); in contrast, alkalosis resulted in an increase in plasma FFA by 83 +/- 40 mumol l-1 (P less than 0.02; P less than 0.01 vs. acidosis), suggesting an increase in lipolysis; plasma glucose decreased compared to acidosis (P less than 0.01). The data demonstrate that acute metabolic acidosis and alkalosis, as they occur in clinical conditions, influence protein breakdown, and in the opposite direction, lipolysis.
Acidosis↗