Circadian systems provide a fitness advantage to organisms by allowing them to adapt to daily changes of environmental cues, such as light/dark cycles. The molecular mechanism underlying the circadian clock has been well characterized. However, how internal circadian clocks are entrained with regular daily light/dark cycles remains unclear. By collecting and analyzing indirect calorimetry (IC) data from more than 2000 wild-type mice available from the International Mouse Phenotyping Consortium (IMPC), we show that the onset time and peak phase of activity and food intake rhythms are reliable parameters for screening defects of circadian misalignment. We developed a machine learning algorithm to quantify these two parameters in our misalignment screen (SyncScreener) with existing datasets and used it to screen 750 mutant mouse lines from five IMPC phenotyping centres. Mutants of five genes (Slc7a11, Rhbdl1, Spop, Ctc1 and Oxtr) were found to be associated with altered patterns of activity or food intake. By further studying the Slc7a11tm1a/tm1a mice, we confirmed its advanced activity phase phenotype in response to a simulated jetlag and skeleton photoperiod stimuli. Disruption of Slc7a11 affected the intercellular communication in the suprachiasmatic nucleus, suggesting a defect in synchronization of clock neurons. Our study has established a systematic phenotype analysis approach that can be used to uncover the mechanism of circadian entrainment in mice.
- MeSH
- cirkadiánní rytmus genetika MeSH
- komplexy ubikvitinligas genetika MeSH
- mutace MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- proteiny vázající telomery genetika MeSH
- receptory oxytocinu genetika MeSH
- represorové proteiny genetika MeSH
- serinové endopeptidasy genetika MeSH
- strojové učení MeSH
- transportní systém aminokyselin y+ genetika MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Granulosa cells (GCs) have many functions in the endocrine system. Most notably, they produce progesterone following ovulation. However, it has recently been proven that GCs can change their properties when subjected to long‑term culture. In the present study, GCs were collected from hyper‑stimulated ovarian follicles during in vitro fertilization procedures. They were grown in vitro, in a long‑term manner. RNA was collected following 1, 7, 15 and 30 days of culture. Expression microarrays were used for analysis, which allowed to identify groups of genes characteristic for particular cellular processes. In addition, reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) was performed to validate the obtained results. Two ontological groups characteristic for processes associated with the development and morphogenesis of the heart were identified during the analyses: 'Heart development' and 'heart morphogenesis'. The results of the microarrays revealed that the highest change in expression was demonstrated by the lysyl Oxidase, oxytocin receptor, nexilin F‑actin binding protein, and cysteine‑rich protein 3 genes. The lowest change was exhibited by odd‑skipped related transcription factor 1, plakophilin 2, transcription growth factor‑β receptor 1, and kinesin family member 3A. The direction of changes was confirmed by RT‑qPCR results. In the present study, it was suggested that GCs may have the potential to differentiate towards other cell types under long‑term in vitro culture conditions. Thus, genes belonging to the presented ontological groups can be considered as novel markers of proliferation and differentiation of GCs towards the heart muscle cells.
- MeSH
- buněčná diferenciace genetika MeSH
- buněčné kultury * MeSH
- buněčný rodokmen genetika MeSH
- folikulární buňky cytologie metabolismus MeSH
- kultivované buňky MeSH
- lidé MeSH
- lysyloxidasa genetika MeSH
- morfogeneze genetika MeSH
- ovariální folikul cytologie metabolismus MeSH
- ovulace genetika MeSH
- progesteron genetika MeSH
- receptory oxytocinu genetika MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
The metabolic action of oxytocin has recently been intensively studied to assess the ability of the peptide to regulate energy homeostasis. Despite the obvious weight-reducing effect of oxytocin observed in experimental studies, plasma oxytocin levels were found to be unchanged or even elevated in human obesity. The aim of our study was to evaluate the changes in the oxytocin system in Zucker rats, an animal model closely mirroring morbid obesity in humans. Plasma oxytocin levels were measured in obese Zucker rats and lean controls by enzyme immunoassay after plasma extraction. The expression of oxytocin and oxytocin receptor (OXTR) was assessed at the mRNA and protein levels by quantitative real-time PCR and immunoblotting respectively. Plasma and tissue activity of oxytocinase, the main enzyme involved in oxytocin degradation, were measured by fluorometric assay using an arylamide derivate as the substrate. Obese Zucker rats displayed a marked reduction in plasma oxytocin levels. Elevated liver and adipose tissue oxytocinase activity was noticed in obese Zucker rats. Hypothalamic oxytocin gene expression was not altered by the obese phenotype. OXTR mRNA and protein levels were upregulated in the adipose tissue of obese animals in contrast to the reduced OXTR protein levels in skeletal muscle. Our results show that obesity is associated with reduced plasma oxytocin due to increased peptide degradation by liver and adipose tissue rather than changes in hormone synthesis. This study highlights the importance of the oxytocin system in the pathogenesis of obesity and suggests oxytocinase inhibition as a candidate approach in the therapy of obesity.
- MeSH
- cystinylaminopeptidasa krev MeSH
- játra metabolismus MeSH
- kosterní svaly metabolismus MeSH
- krysa rodu rattus MeSH
- lidé MeSH
- morbidní obezita genetika metabolismus MeSH
- oxytocin krev MeSH
- potkani Zucker MeSH
- proteolýza MeSH
- receptory oxytocinu genetika metabolismus MeSH
- tuková tkáň metabolismus MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
OBJECTIVES: Oxytocin (OT) is a neuropeptide acting both as a peripheral hormone and in the brain as neurotransmitter and neuromodulator. In addition to its well-known effects on milk-ejection and uterine contraction, OT was shown to exert neuroendocrine regulation of heart functions. The aim of this study was to investigate the expression of mRNA of OT receptors (OTR) in rat hearts by real-time quantitative PCR (qPCR). The study was performed in Sprague-Dawley (SD) and Lewis (LE) rat strains, the latter having lower activity of HPA axis. METHODS: We used adult male SD and LE rats. OTR mRNA expression was detected in all heart chambers by comparing their threshold cycle values (CT) to CT of reference gene β-actin. The relative expression ratios were calculated using the 2-ΔΔCT method. The specificity of reaction of primary antibody with OTRs was tested by Western Blot and localization of OTR in the heart compartments was performed by immunofluorescence with commercial OTR specific antibodies. RESULTS: We found expression of OTR mRNA in all heart compartments. The expression of OTR mRNA in both atria (LA, RA) was much higher than in the ventricles (RV, LV). By using two-way ANOVA we found no statistical differences between corresponding compartments of SD and LE rats. Immunohistochemical studies showed that OTR staining is not related to neuronal tissue and findings from left atrium indicate that prevalent localization of OTR is on cell membranes of cardiomyocytes. CONCLUSIONS: The finding of expression of OTR mRNA by real-time qPCR and proof of OTR staining by immunohistochemistry in all heart compartments indicate that OT and its receptors may have function as a cardiovascular hormone. The differences in the HPA axis activity, as is exemplified in Sprague-Dawley and Lewis rat strain, do not project in the expression of OTR mRNA under basal condition. The effect of activity of HPA on OTR expression should be studied under stimulated conditions as it was performed in the behavioral studies.
- MeSH
- krysa rodu rattus MeSH
- messenger RNA metabolismus MeSH
- myokard cytologie metabolismus MeSH
- oxytocin metabolismus MeSH
- potkani inbrední LEW MeSH
- potkani Sprague-Dawley MeSH
- receptory oxytocinu genetika metabolismus MeSH
- systém hypofýza - nadledviny fyziologie MeSH
- systém hypotalamus-hypofýza fyziologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Novel sites of oxytocin receptor expression have recently been detected in central nervous system, cardiomyocytes, endothelial cells, various carcinoma cells, etc. These and other discoveries have greatly expanded the classical biological roles of oxytocin, which are stimulation of uterine smooth muscle contraction at parturition and milk ejection during lactation. It is becoming clear that the great diversity of oxytocin actions in the brain and peripheral organs is paralleled by activation of a diversity of signalling pathways. On the other hand, until now only one single oxytocin receptor type has been detected. This receptor belongs to G protein-coupled receptors and in dependence on cell conditions it binds to different G proteins; this phenomenon is called receptor-G protein promiscuity. Thus, in the same cells oxytocin can activate multiple responses at the same time. Recently, the oxytocinergic system has also been implicated in the growth modulation of various neoplastic cells, where it may inhibit or stimulate cell proliferation in dependence on cell type and activated metabolic pathways. The discovery of novel oxytocin receptor-linked signalling cascades brings interesting knowledge opening new avenues for research in oncology and molecular pharmacology with perspectives of finding new therapeutic agents.
- MeSH
- financování organizované MeSH
- inositol-1,4,5-trisfosfát metabolismus MeSH
- lidé MeSH
- nádorové procesy MeSH
- nádory metabolismus patologie MeSH
- oxytocin genetika metabolismus MeSH
- podjednotky proteinů genetika metabolismus MeSH
- proliferace buněk MeSH
- protoonkogenní proteiny c-akt metabolismus MeSH
- receptory oxytocinu genetika metabolismus MeSH
- signální transdukce fyziologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- přehledy MeSH
