Steady-state expression of self-regulated genes.

MOTIVATION: Regulatory gene networks contain generic modules such as feedback loops that are essential for the regulation of many biological functions. The study of the stochastic mechanisms of gene regulation is instrumental for the understanding of how cells maintain their expression at levels commensurate with their biological role, as well as to engineer gene expression switches of appropriate behavior. The lack of precise knowledge on the steady-state distribution of gene expression requires the use of Gillespie algorithms and Monte-Carlo approximations. METHODOLOGY: In this study, we provide new exact formulas and efficient numerical algorithms for computing/modeling the steady-state of a class of self-regulated genes, and we use it to model/compute the stochastic expression of a gene of interest in an engineered network introduced in mammalian cells. The behavior of the genetic network is then analyzed experimentally in living cells. RESULTS: Stochastic models often reveal counter-intuitive experimental behaviors, and we find that this genetic architecture displays a unimodal behavior in mammalian cells, which was unexpected given its known bimodal response in unicellular organisms. We provide a molecular rationale for this behavior, and we implement it in the mathematical picture to explain the experimental results obtained from this network.

Appropriate management of special situations in Crohn's disease (upper gastro-intestinal; extra-intestinal manifestations; drug safety during pregnancy and breastfeeding): results of a multidisciplinary international expert panel-EPACT II

Introduction: High-grade evidence is lacking for most therapeutic decisions in Crohn's disease. Appropriateness criteria were developed for upper gastro-intestinal, extra-intestinal manifestations and drug safety during conception, pregnancy and breastfeeding in patients with Crohn's disease, to assist the physician in clinical decision making. Methods: The European Panel on the Appropriateness of Crohn's Disease Therapy (EPACT II), a multidisciplinary international European expert panel, rated clinical scenarios based on evidence from the published literature and panelists' own clinical expertise. Median ratings (on a 9-point scale) were stratified into three categories: appropriate (7-9), uncertain (4-6 with or without disagreement) and inappropriate (1-3). Experts were also asked to rank appropriate medications by priority. Results: Proton pump inhibitors, steroids, azathioprine/6-mercaptopurine and infliximab are appropriate for upper gastro-duodenal Crohn's disease; for stenosis, endoscopic balloon dilation is the first-tine therapy, although surgery is also appropriate. Ursodeoxycholic acid is the only appropriate treatment for primary sclerosing cholangitis. Infliximab is appropriate for Pyoderma gangrenosum, ankylosing spondylitis and uveitis, steroids for Pyoderma gangrenosum and ankylosing spondylitis, adalimumab for Pyoderma gangrenosum and ankylosing spondylitis, cyclosporine-A/tacrolimus for Pyoderma gangrenosum. Mesalamine, sulfasalazine, prednisone, azathioprine/6-mercaptopurine, ciprofloxacin, and probiotics, may be administered safety during pregnancy or for patients wishing to conceive, with the exception that mate patients considering conception should avoid sulfasalazine. Metronidazol is considered safe in the 2nd and 3rd trimesters whereas infliximab is rated safe in the 1st trimester but uncertain in the 2nd and 3rd trimesters. Methotrexate is always contraindicated at conception, during pregnancy or during breastfeeding, due to its known teratogenicity. Mesalamine, prednisone, probiotics and infliximab are considered safe during breastfeeding. Conclusion: EPACT II recommendations are freely available online (www.epact.ch). The validity of these criteria should now be tested by prospective evaluation. (C) 2009 European Crohn's and Colitis Organisation. Published by Elsevier B.V. All rights reserved.

Homeostatic and circadian contribution to EEG and molecular state variables of sleep regulation.

STUDY OBJECTIVES: Besides their well-established role in circadian rhythms, our findings that the forebrain expression of the clock-genes Per2 and Dbp increases and decreases, respectively, in relation to time spent awake suggest they also play a role in the homeostatic aspect of sleep regulation. Here, we determined whether time of day modulates the effects of elevated sleep pressure on clock-gene expression. Time of day effects were assessed also for recognized electrophysiological (EEG delta power) and molecular (Homer1a) markers of sleep homeostasis. DESIGN: EEG and qPCR data were obtained for baseline and recovery from 6-h sleep deprivation starting at ZT0, -6, -12, or -18. SETTING: Mouse sleep laboratory. PARTICIPANTS: Male mice. INTERVENTIONS: Sleep deprivation. RESULTS: The sleep-deprivation induced changes in Per2 and Dbp expression importantly varied with time of day, such that Per2 could even decrease during sleep deprivations occurring at the decreasing phase in baseline. Dbp showed similar, albeit opposite dynamics. These unexpected results could be reliably predicted assuming that these transcripts behave according to a driven damped harmonic oscillator. As expected, the sleep-wake distribution accounted for a large degree of the changes in EEG delta power and Homer1a. Nevertheless, the sleep deprivation-induced increase in delta power varied also with time of day with higher than expected levels when recovery sleep started at dark onset. CONCLUSIONS: Per2 and delta power are widely used as exclusive state variables of the circadian and homeostatic process, respectively. Our findings demonstrate a considerable cross-talk between these two processes. As Per2 in the brain responds to both sleep loss and time of day, this molecule is well positioned to keep track of and to anticipate homeostatic sleep need. CITATION: Curie T; Mongrain V; Dorsaz S; Mang GM; Emmenegger Y; Franken P. Homeostatic and circadian contribution to EEG and molecular state variables of sleep regulation. SLEEP 2013;36(3):311-323.

Paroxetine increases steady-state concentrations of (R)-methadone in CYP2D6 extensive but not poor metabolizers

Steady-state blood concentrations of (R)- methadone (i.e., the active form), (S)-methadone, and (R,S)-methadone were measured before and after introduction of paroxetine 20 mg/day during a mean period of 12 days in 10 addict patients in methadone maintenance treatment. Eight patients were genotyped as CYP2D6 homozygous extensive metabolizers (EMs) and two patients as poor metabolizers (PMs). Paroxetine significantly increased concentrations of both enantiomers of methadone in the whole group (mean increase for (R)-methadone +/- SD, 26 +/- 32%; range, -14% to +83%, p = 0.032; for (S)-methadone, 49 +/- 51%; range, -29% to +137%, p = 0.028; for (R,S)-methadone, 35 +/- 41%; range, -20% to +112%, p = 0.032) and in the group of eight EMs (mean increase, 32%, p = 0.036; 53%, p = 0.028; and 42%, p = 0.036, for (R)-methadone, (S)-methadone, and (R,S)-methadone, respectively). On the other hand, in the two PMs, (S)-methadone but not (R)-methadone concentrations were increased by paroxetine (mean increases of 36% and 3%, respectively). Paroxetine is a strong CYP2D6 inhibitor, and these results confirm previous studies showing an involvement of CYP2D6 in methadone metabolism with a stereoselectivity toward the (R)-enantiomer. Because paroxetine is a mild inhibitor of CYP1A2, CYP2C9, CYP2C19, and CYP3A4, increase of (S)-methadone concentrations in both EMs and PMs could be mediated by inhibition of any of these isozymes.

Steady State Concentrations of the Enantiomers of Mianserin and Desmethylmianserin in Poor and in Homozygous and Heterozygous Extensive Metabolizers of Debrisoquine.

AB Summary: Steady state concentrations of (S)- and (R)-mianserin and desmethylmianserin were measured in 21 homozygous extensive metabolizers (as determined by genotyping for mutations 3 [or A] and 4 [or B]), in seven heterozygous extensive metabolizers and in one poor metabolizer of debrisoquine, as well as in one patient receiving very high doses of mianserin (360 mg/day) and fluoxetine (160 mg/day), a strong cytochrome P450IID6 inhibitor. The mean dose of mianserin was (mean +/- SD, range: 67 +/- 63, 10 to 360 mg/day). High dispersions of the (S)/(R)-mianserin and desmethylmianserin ratios were observed (mean +/- SD, range: 2.10+/- 1.01, 0.64 to 4.76, and 0.29 +/- 0.14, 0.08 to 0.57, respectively). The highest (S)/(R)-mianserin ratio was calculated for the poor metabolizer (4.76) agreeing with those results of a single-dose study with poor and extensive metabolizers of debrisoquine, in that the cytochrome P450IID6 is probably involved in the metabolism of mianserin with an enantioselectivity for the (S)-enantiomer. Nevertheless, the mean concentration-to-dose ratios for (S)- or (R)-mianserin or desmethylmianserin were not significantly different between homozygous and heterozygous extensive metabolizers, and no particular values were measured in the poor metabolizer nor in the patient receiving fluoxetine. Furthermore, the(S)/(R)-mianserin ratio measured in the PM was only slightly higher than the second highest ratio (3.85) of an homozygous extensive metabolizer, whereas no particular value (2.92) was calculated for the patient taking fluoxetine. Finally, no significant differences in (S)/(R)-mianserin or(S)/(R)-desmethylmianserin were calculated between homozygous and heterozygous extensive metabolizers. Although the number of patients included in this study is too low to allow definite conclusions, the results suggest that the debrisoquine genotype has only a moderate influence on the steady state concentrations of the enantiomers of mianserin and desmethylmianserin. (C) Lippincott-Raven Publishers