Proteome analysis is a fundamental step in systematic functional genomics. Here we have resolved 8,767 proteins from the mouse brain proteome by large-gel two-dimensional electrophoresis. We detected 1,324 polymorphic proteins from the European collaborative interspecific backcross. Of these, we mapped 665 proteins genetically and identified 466 proteins by mass spectrometry. Qualitatively polymorphic proteins, to 96%, reflect changes in conformation and/or mass. Quantitatively polymorphic proteins show a high frequency (73%) of allele-specific transmission in codominant heterozygotes. Variations in protein isoforms and protein quantity often mapped to chromosomal positions different from that of the structural gene, indicating that single proteins may act as polygenic traits. Genetic analysis of proteomes may detect the types of polymorphism that are most relevant in disease-association studies.

A thorough, simultaneous biopsychosocial diagnostic assessment enables the early recognition of non-specific, functional, and somatoform bodily complaints. The appropriate treatment depends on the severity of the condition. Effective treatment requires the patient's active cooperation and the collaboration of all treating health professionals under the overall management of the patient's primary-care physician.

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Objective: Selective inhibitors of 11b-hydroxysteroid-dehydrogenase type I may be of therapeutical interest for two reasons: i) 9a-Fluorinated 11-dehydrosteroids like 11-dehydro-dexamethasone (DH-D) are rapidly activated by human kidney 11b-hydroxysteroid-dehydrogenase type II (11b-HSD-II) to dexamethasone (D). If the same reaction by hepatic 11b-HSD-I could be selectively inhibited, DH-D could be used for selective renal immunosuppressive therapy. ii) Reduction of cortisone to cortisol in the liver may increase insulin resistance in type 2 diabetes mellitus, and inhibition of the enzyme may lead to a decrease in gluconeogenesis.Therefore, we characterized the metabolism of DH-D by human hepatic 11b-HSD-I and tried to find a selective inhibitor of this isoenzyme. Methods: For kinetic analysis of 11b-HSD-I, we used microsomes prepared from unaffected parts of liver segments, resected because of hepatocarcinoma or metastatic disease. For inhibition experiments, we also tested 11b-HSD-II activity with human kidney cortex microsomes. The inhibitory potency of several compounds was evaluated for oxidation and reduction in concentrations from 10 ¹9 to 10 ¹5 mol/l. Results: Whereas D was not oxidized by human liver microsomes at all, cortisol was oxidized to cortisone with a maximum velocity (V max ) of 95 pmol/mg per min. The reduction of DH-D to D (V max = 742 pmol/ mg per min, Michaelis-Menten constant (K m ) = 1.6 mmol/l) was faster than that of cortisone to cortisol (V max =187 pmol/mg per min). All reactions tested in liver microsomes showed the characteristics of 11b-HSD-I: K m values in the micromolar range, preferred cosubstrate NADP(H), no product inhibition. Of the substances tested for inhibition of 11b-HSD-I and -II, chenodeoxycholic acid was the only one that selectively inhibited 11b-HSD-I (IC 50 for reduction: 2.8 × 10 ¹6 mol/l, IC 50 for oxidation: 4.4 × 10 ¹6 mol/l), whereas ketoconazole preferentially inhibited oxidation and reduction reactions catalyzed by 11b-HSD-II. Metyrapone, which is reduced to metyrapol by hepatic 11b-HSD-I, inhibited steroid reductase activity of 11b-HSD-I and -II and oxidative activity of 11b-HSD-II. These findings can be explained by substrate competition for reductase reactions and by product inhibition of the oxidation, which is a well-known characteristic of 11b-HSD-II. Conclusions: Our in vitro results may offer a new concept for renal glucocorticoid targeting. Oral administration of small amounts of DH-D (low substrate affinity for 11b-HSD-I) in combination with chenodeoxycholic acid (selective inhibition of 11b-HSD-I) may prevent hepatic first pass reduction of DH-D, thus allowing selective activation of DH-D to D by the high affinity 11b-HSD-II in the kidney. Moreover, selective inhibitors of the hepatic 11b-HSD-I, like chenodeoxycholic acid, may become useful in the therapy of patients with hepatic insulin resistance including diabetes mellitus type II, because cortisol enhances gluconeogenesis.

Metabolites and lipids are the final products of enzymatic processes, distinguishing the different cellular functions and activities of single cells or whole tissues. Understanding these cellular functions within a well-established model system requires a systemic collection of molecular and physiological information. In the current report, the green alga Chlamydomonas reinhardtii was selected to establish a comprehensive workflow for the detailed multi-omics analysis of a synchronously growing cell culture system. After implementation and benchmarking of the synchronous cell culture, a two-phase extraction method was adopted for the analysis of proteins, lipids, metabolites and starch from a single sample aliquot of as little as 10-15 million Chlamydomonas cells. In a proof of concept study, primary metabolites and lipids were sampled throughout the diurnal cell cycle. The results of these time-resolved measurements showed that single compounds were not only coordinated with each other in different pathways, but that these complex metabolic signatures have the potential to be used as biomarkers of various cellular processes. Taken together, the developed workflow, including the synchronized growth of the photoautotrophic cell culture, in combination with comprehensive extraction methods and detailed metabolic phenotyping has the potential for use in in-depth analysis of complex cellular processes, providing essential information for the understanding of complex biological systems.

The maternal adrenal cortex seems to be involved in the adaptation to pregnancy. To study in detail adrenocortical secretion during pregnancy, we measured plasma aldosterone, corticosterone, 11-deoxycorticosterone, progesterone, 17-hydroxyprogesterone, 11-deoxycortisol, cortisol, and cortisone simultaneously by RIA after extraction and automated Sephadex LH-20 chromatography of 10 normal pregnant women longitudinally throughout pregnancy at weeks 8-10, 14-17, 21-24, 28-32, and 38 as well as at the time of admission to the delivery room. The mean plasma progesterone and 17-hydroxy-progesterone concentrations increased from 37.2 +/- 6.5 (+/- SE) and 8.2 +/- 1.0 nmol/L, respectively, in early gestation to maximum levels of 138.0 +/- 25.7 and 22.8 +/- 2.2 nmol/L at week 38 (P less than 0.01). Plasma glucocorticoid levels rose 2- to 3-fold (P less than 0.01) from weeks 8-10 (corticosterone, 18.5 +/- 5.4; 11-deoxycortisol, 1.9 +/- 0.2; cortisone, 24.2 +/- 4.2; cortisol, 195.5 +/- 37.6 nmol/L) to week 38 (corticosterone, 42.9 +/- 11.2; 11-deoxycortisol, 4.6 +/- 0.5; cortisone, 71.5 +/- 13.6; cortisol, 420 +/- 63 nmol/L). Similarly, plasma mineralocorticoid levels increased 5- to 7-fold (P less than 0.01) from weeks 8-10 (11-deoxycorticosterone, 0.69 +/- 0.12; aldosterone, 0.41 +/- 0.08 nmol/L) to maximum levels at week 38 (5.3 +/- 0.9 and 2.1 +/- 0.3 nmol/L, respectively). At the time of admission to the delivery room, plasma 11-deoxycortisol, corticosterone, and cortisol concentrations were higher (P less than 0.02) than at 38 weeks, but plasma progestin and mineralocorticoid concentrations were not. We conclude that the source of the elevated maternal corticosteroid levels in pregnancy in addition to the estrogen-mediated rise in corticosteroid-binding globulin is the maternal adrenal cortex itself. The peak glucocorticoid levels at admission to the delivery room reflect increased maternal and fetal stress with the onset of labor.

The nutritional regimen of patients with phenylketonuria (PKU) comprises a diet of natural proteins and phenylalanine (Phe)-free amino acid (AA) mixture. The main daily protein requirement is covered by a Phe-free AA mixture. In an adult with PKU, the consumption of the daily AA requirement in one single dose at breakfast caused nausea and vomiting. Therefore, four studies were designed to investigate the adverse and metabolic effects resulting from large intakes of AA mixtures used in the treatment of PKU patients with respect to the following: (1) biochemical effects following consumption of one single dose of Phe-free AA mixture in healthy persons; (2) transient metabolic changes caused by different individual regimens of AA intake in healthy persons and in one PKU patient; (3) nitrogen excretion in PKU patients taking the AA mixture in two or three portions; and (4) catabolic metabolism of AA in a PKU patient. In healthy subjects following the ingestion of the AA mixture in one bolus there was an increase in the blood levels of the given AA and also an increase in blood insulin concentration and a decrease in blood glucose concentration. These changes were less marked when the AA mixture was divided into three portions per day. In contrast, in a PKU patient following the ingestion of AA there was an increase in blood glucose. The urinary nitrogen excretion was greater in PKU patients when one compared to three portions of AA mixture was taken. The consumption of the daily requirement of AA mixture in one single does produced increased catabolism in a PKU patient. In conclusion it is recommended that the total daily amount of AA mixture should be divided into a minimum of three portions.

CWP is defined by the criteria of the American College of Rheumatology (ACR--strong consensus). FMS can be diagnosed for clinical purposes by symptom-based criteria (without tender point examination) as well as by the ACR criteria (strong consensus).