BACKGROUND: Glycogen synthase kinase-3β (GSK3β), cAMP-response element-binding protein (CREB) and brain-derived neurotrophic factor (BDNF) play critical roles in neuronal survival, synaptic plasticity and memory and participate in the pathophysiology of both depressive disorder and Alzheimer's disease (AD). METHODS: This study was designed to determine the association of GSK3β activity, CREB activity and BDNF concentration in peripheral blood of patients with AD with or without depressive symptoms and in depressive patients without AD. GSK3β activity in platelets, CREB activity in lymphocytes and BDNF concentration in plasma, platelet-rich plasma or platelets were measured in 85 AD patients (36 of whom displayed co-morbid depressive symptoms), 65 non-AD patients with depressive disorder and 96 healthy controls. AD patients were clinically assessed for stage of dementia, cognitive impairment and severity of depressive symptoms. Depressive patients were clinically assessed for severity of depression. RESULTS: We observed increased CREB activity and GSK3β activity in AD with depressive symptoms or in AD at mild stage of dementia. Decreased BDNF concentration was found in platelet-rich plasma of AD patients at moderate to severe stages of dementia or in AD without depressive symptoms. An association was revealed of the severity of cognitive impairment with the increase of GSK3β in the platelets of AD patients with mild dementia. In depressive patients, a lower concentration of phosphorylated GSK3β was associated with a higher severity of depression. Association was confirmed between severity of depression, CREB activation, and BDNF concentration in drug-naïve depressive patients. CONCLUSION: Our data demonstrated that AD is accompanied by increased CREB activity in lymphocytes and a decreased concentration of BDNF in platelet-rich plasma. The decreased BDNF concentration appears to correlate with moderate to severe stages of dementia in AD. Observation of decreased phosphorylation of GSK3β in platelets of both AD patients with depressive symptoms and depressive patients after treatment confirms the role of increased GSK3β activity in the pathophysiology of both AD and depressive disorder. Associations were confirmed between AD and platelet GSK3β activity, lymphocyte CREB activity and plasma BDNF. CREB activity and platelet BDNF concentration seems to be related to depressive disorder.

• MeSH
• Alzheimerova nemoc krev   komplikace   psychologie   MeSH
• deprese krev   komplikace   psychologie   MeSH
• kinasa 3 glykogensynthasy krev   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mozkový neurotrofický faktor krev   MeSH
• neuropsychologické testy MeSH
• protein vázající CREB krev   MeSH
• psychiatrické posuzovací škály MeSH
• senioři MeSH
• studie případů a kontrol MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

B-Myb, a highly conserved member of the Myb transcription factor family, is expressed ubiquitously in proliferating cells and controls the cell cycle dependent transcription of G2/M-phase genes. Deregulation of B-Myb has been implicated in oncogenesis and loss of genomic stability. We have identified B-Myb as a novel interaction partner of the Mre11-Rad50-Nbs1 (MRN) complex, a key player in the repair of DNA double strand breaks. We show that B-Myb directly interacts with the Nbs1 subunit of the MRN complex and is recruited transiently to DNA-damage sites. In response to DNA-damage B-Myb is phosphorylated by protein kinase GSK3β and released from the MRN complex. A B-Myb mutant that cannot be phosphorylated by GSK3β disturbs the regulation of pro-mitotic B-Myb target genes and leads to inappropriate mitotic entry in response to DNA-damage. Overall, our work suggests a novel function of B-Myb in the cellular DNA-damage signalling.

• MeSH
• ATM protein metabolismus   MeSH
• biologické modely MeSH
• buněčné linie MeSH
• DNA vazebné proteiny chemie   metabolismus   MeSH
• dvouřetězcové zlomy DNA MeSH
• enzymy opravy DNA chemie   metabolismus   MeSH
• fosforylace MeSH
• GSK3B metabolismus   MeSH
• homologní protein MRE11 chemie   metabolismus   MeSH
• interakční proteinové domény a motivy MeSH
• jaderné proteiny chemie   metabolismus   MeSH
• lidé MeSH
• mitóza genetika   MeSH
• multiproteinové komplexy metabolismus   MeSH
• mutace MeSH
• oprava DNA MeSH
• poškození DNA * MeSH
• proteiny buněčného cyklu chemie   genetika   metabolismus   MeSH
• regulace genové exprese MeSH
• sekvence aminokyselin MeSH
• signální transdukce * MeSH
• trans-aktivátory chemie   genetika   metabolismus   MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Reperfusion therapies for ischaemic stroke can induce secondary injury accompanied by neuronal death. The Y-box binding protein 1 (YBX1), an oncoprotein, is critical for regulating tumour cell proliferation and apoptosis. Thus, we wanted to know whether YBX1 could regulate neuronal cell apoptosis caused by cerebral ischaemia/reperfusion (I/R). We established a model of cerebral I/R-induced injury in vitro by oxygen-glucose deprivation/reoxygenation (OGD/R) treatment and determined YBX1 expression using Western blot. Next, the effect of YBX1 on the apoptosis and viability of OGD/R-treated PC12 cells was evaluated by flow cytometry, MTT assay, and Western blot. Besides, the release of lactate dehydrogenase (LDH) and the activity of catalase (CAT) and superoxide dismutase (SOD) were detected to evaluate oxidative stress of PC12 cells induced by OGD/R. The regulatory roles of YBX1 in the AKT/GSK3β pathway were examined by Western blot. As a result, OGD/R treatment down-regulated YBX1 expression in PC12 cells. YBX1 over-expression attenuated the growth inhibition and apoptosis of PC12 cells induced by OGD/R. Besides, the increase of LDH release and the decrease of SOD and CAT activities caused by OGD/R were reversed by YBX1 over-expression. Moreover, YBX1 over-expression could activate the AKT/GSK3β pathway in OGD/ R-treated PC12 cells. Therefore, YBX1 could protect against OGD/R-induced injury in PC12 cells through activating the AKT/GSK3β signalling pathway, and thus YBX1 has the potential to become a therapeutic target for cerebral I/R-induced injury.

• MeSH
• apoptóza MeSH
• buňky PC12 MeSH
• cévní mozková příhoda * MeSH
• DNA vazebné proteiny MeSH
• glukosa MeSH
• GSK3B * MeSH
• ischemie mozku * MeSH
• krysa rodu rattus MeSH
• kyslík MeSH
• oxidační stres MeSH
• protoonkogenní proteiny c-akt * MeSH
• reperfuzní poškození * MeSH
• viabilita buněk MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

CD8+ T cells protect against tumors and intracellular pathogens. The inflammatory cytokines IL-2, IL-15, and IL-7 are necessary for their expansion. However, elevated serum levels of these cytokines are often associated with cancer, poorer prognosis of cancer patients, and exhaustion of antigen-expanded CD8+ T cells. The impact of acute conditioning of antigen-expanded CD8+ T cells with these cytokines is unknown. Here, we generated antigen-expanded CD8+ T cells using dendritic cells and PC-3 cells. The cells were acutely (18-24 h) conditioned with IL-2 and either the GSK3β inhibitor TWS119, the mTORC1 inhibitor rapamycin, or the mTORC1/2 inhibitor Torin1, then their immediate and post-re-expansion (distal) cytokine responses after antigen rechallenge were evaluated. We found that acute IL-2 conditioning upregulated the immediate antigen-induced cytokine response of the tested cells. Following their re-expansion, however, the cells showed a decreased cytokine response. These IL-2 conditioning-mediated impacts were counteracted with TWS119 or rapamycin but not with Torin1. Our data revealed that the acute conditioning of antigen-expanded CD8+ T cells with IL-2 modulates the GSK3β-mTORC signaling axis. This modulation differentially affected the immediate and distal cytokine responses of the cells. The acute targeting of this signaling axis could, therefore, represent a novel strategy for the modulation of antigen-expanded CD8+ T cells.

• Publikační typ
• časopisecké články MeSH

BACKGROUND AND PURPOSE: Opioids affect the circadian clock and may change the timing of many physiological processes. This study was undertaken to investigate the daily changes in sensitivity of the circadian pacemaker to an analgesic dose of morphine, and to uncover a possible interplay between circadian and opioid signalling. EXPERIMENTAL APPROACH: A time-dependent effect of morphine (1 mg·kg(-1) , i.p.) applied either during the day or during the early night was followed, and the levels of phosphorylated ERK1/2, GSK3β, c-Fos and Per genes were assessed by immunohistochemistry and in situ hybridization. The effect of morphine pretreatment on light-induced pERK and c-Fos was examined, and day/night difference in activity of opioid receptors was evaluated by [(35) S]-GTPγS binding assay. KEY RESULTS: Morphine stimulated a rise in pERK1/2 and pGSK3β levels in the suprachiasmatic nucleus (SCN) when applied during the day but significantly reduced both kinases when applied during the night. Morphine at night transiently induced Period1 but not Period2 in the SCN and did not attenuate the light-induced level of pERK1/2 and c-Fos in the SCN. The activity of all three principal opioid receptors was high during the day but decreased significantly at night, except for the δ receptor. Finally, we demonstrated daily profiles of pERK1/2 and pGSK3β levels in the rat ventrolateral and dorsomedial SCN. CONCLUSIONS AND IMPLICATIONS: Our data suggest that the phase-shifting effect of opioids may be mediated via post-translational modification of clock proteins by means of activated ERK1/2 and GSK3β.

• MeSH
• cirkadiánní hodiny účinky léků   MeSH
• cirkadiánní proteiny Period metabolismus   MeSH
• cirkadiánní rytmus účinky léků   MeSH
• fosforylace účinky léků   MeSH
• kinasa 3 glykogensynthasy metabolismus   MeSH
• krysa rodu rattus MeSH
• mitogenem aktivovaná proteinkinasa 1 metabolismus   MeSH
• mitogenem aktivovaná proteinkinasa 3 metabolismus   MeSH
• mitogenem aktivované proteinkinasy metabolismus   MeSH
• morfin farmakologie   MeSH
• nucleus suprachiasmaticus účinky léků   metabolismus   MeSH
• potkani Wistar MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

As with other drugs or pharmaceuticals, opioids differ in their rewarding or analgesic effects depending on when they are applied. In the previous study, we have demonstrated the day/night difference in the sensitivity of the major circadian clock in the suprachiasmatic nucleus to a low dose of morphine, and showed the bidirectional effect of morphine on pERK1/2 and pGSK3β levels in the suprachiasmatic nucleus depending on the time of administration. The main aim of this study was to identify other brain structures that respond differently to morphine depending on the time of its administration. Using immunohistochemistry, we identified 44 structures that show time-of-day specific changes in c-Fos level and activity of ERK1/2 and GSK3β kinases in response to a single dose of 1 mg/kg morphine. Furthermore, comparison among control groups revealed the differences in the spontaneous levels of all markers with a generally higher level during the night, that is, in the active phase of the day. We thus provide further evidence for diurnal variations in the activity of brain regions outside the suprachiasmatic nucleus indicated by the temporal changes in the molecular substrate. We suggest that these changes are responsible for generating diurnal variation in the reward behavior or analgesic effect of opioid administration.

• MeSH
• cirkadiánní rytmus fyziologie   MeSH
• GSK3B metabolismus   MeSH
• krysa rodu rattus MeSH
• MAP kinasový signální systém fyziologie   MeSH
• morfin farmakologie   MeSH
• mozek účinky léků   metabolismus   MeSH
• opioidní analgetika farmakologie   MeSH
• potkani Wistar MeSH
• protoonkogenní proteiny c-fos metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The intrinsic period of circadian clock in the suprachiasmatic nucleus is entrained to a 24-h cycle by external cues, mainly light. Previous studies have shown that light applied at night induces robust phosphorylation of extracellular-signal-regulated kinase that is necessary to process the light pulse into the phase shift of the clock phase. In this study, we show the persistent downregulation of phosphorylated extracellular-signal-regulated kinase and transient downregulation of phosphorylated glycogen synthase kinase-3beta in the ventrolateral part of the suprachiasmatic nucleus to photic stimuli starting at 2 h after the beginning of the light pulse. As both kinases are involved in regulation of circadian clockwork, we hypothesize that these changes may contribute to the phase-shifting effect of light at night.

• MeSH
• cirkadiánní hodiny * MeSH
• kinasa 3 glykogensynthasy metabolismus   MeSH
• krysa rodu rattus MeSH
• MAP kinasový signální systém MeSH
• mitogenem aktivovaná proteinkinasa 1 metabolismus   MeSH
• mitogenem aktivovaná proteinkinasa 3 metabolismus   MeSH
• nucleus suprachiasmaticus metabolismus   fyziologie   MeSH
• potkani Wistar MeSH
• reakční čas * MeSH
• světelná stimulace MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

AIMS: Circadian clocks in the hippocampus (HPC) align memory processing with appropriate time of day. Our study was aimed at ascertaining the specificity of glycogen synthase kinase 3-beta (GSK3β)- and glucocorticoid (GC)-dependent pathways in the entrainment of clocks in individual HPC regions, CA1-3, and dentate gyrus (DG). METHODS: The role of GCs was addressed in vivo by comparing the effects of adrenalectomy (ADX) and subsequent dexamethasone (DEX) supplementation on clock gene expression profiles (Per1, Per2, Nr1d1, and Bmal1). In vitro the effects of DEX and the GSK3β inhibitor, CHIR-99021, were assessed from recordings of bioluminescence rhythms in HPC organotypic explants of mPER2Luc mice. RESULTS: Circadian rhythms of clock gene expression in all HPC regions were abolished by ADX, and DEX injections to the rats rescued those rhythms in DG. The DEX treatment of the HPC explants significantly lengthened periods of the bioluminescence rhythms in all HPC regions with the most significant effect in DG. In contrast to DEX, CHIR-99021 significantly shortened the period of bioluminescence rhythm. Again, the effect was most significant in DG which lacks the endogenously inactivated (phosphorylated) form of GSK3β. Co-treatment of the explants with CHIR-99021 and DEX produced the CHIR-99021 response. Therefore, the GSK3β-mediated pathway had dominant effect on the clocks. CONCLUSION: GSK3β- and GC-dependent pathways entrain the clock in individual HPC regions by modulating their periods in an opposite manner. The results provide novel insights into the mechanisms connecting the arousal state-relevant signals with temporal control of HPC-dependent memory and cognitive functions.

• MeSH
• cirkadiánní hodiny * genetika   MeSH
• cirkadiánní proteiny Period genetika   metabolismus   MeSH
• cirkadiánní rytmus MeSH
• glukokortikoidy metabolismus   farmakologie   MeSH
• GSK3B metabolismus   MeSH
• gyrus dentatus metabolismus   MeSH
• hipokampus metabolismus   MeSH
• kinasa 3 glykogensynthasy metabolismus   farmakologie   MeSH
• krysa rodu rattus MeSH
• myši MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Cytochrome P450 (CYP) expression and activity are not homogeneous in the liver lobules. Indeed, CYPs are mainly expressed and induced in centrilobular hepatocytes. The wingless-type MMTV integration site family (WNT)/β-catenin pathway was identified as a major regulator of this zonal organization. We have recently demonstrated that in primary human hepatocytes (PHHs), the expression of CYP2E1, CYP1A2, and aryl hydrocarbon receptor (AhR), but not of CYP3A4, is regulated by the WNT/β-catenin pathway in response to WNT3a, its canonical activator. Here, we investigated whether glycogen synthase kinase 3β (GSK3β) inhibitors, which mimic the action of WNT molecules, could be used in PHHs to activate the β-catenin pathway to study CYP expression. We assessed the activity of 6BIO (6-bromoindirubin-3'-oxime), CHIR99021 (6-((2-((4-(2,4-dichlorophenyl)-5-(4methyl-1H-imidazol-2-yl)pyrimidin-2-yl)amino)ethyl)amino) nicotinonitrile), and GSK3iXV (Pyridocarbazolo-cyclopentadienyl Ruthenium complex GSK3 inhibitor XV) that belong to structurally different families of GSK3β inhibitors. Using small interfering RNAs, reporter gene assays, and molecular docking predictions, we demonstrated that GSK3β inhibitors can activate the WNT/β-catenin pathway in PHHs to regulate CYP2E1 expression. We also found that 6BIO and GSK3iXV are AhR full agonists that participate, through AhR signaling, to CYP1A2 induction. Conversely, CHIR99021 is an AhR partial agonist, and a pregnane X receptor ligand and partial agonist, thus regulating CYP1A2 and CYP3A4 gene expression in a β-catenin-independent manner. In conclusion, GSK3β inhibitors can activate the WNT/β-catenin pathway in PHHs. Nevertheless, their role in CYP regulation should be analyzed with caution as these molecules can interact with xenosensors.

• MeSH
• beta-katenin agonisté   antagonisté a inhibitory   genetika   metabolismus   MeSH
• enzymová indukce účinky léků   MeSH
• hepatocyty cytologie   účinky léků   metabolismus   MeSH
• indoly farmakologie   MeSH
• induktory cytochromu P450 chemie   metabolismus   farmakologie   MeSH
• inhibitory proteinkinas chemie   metabolismus   farmakologie   MeSH
• kinasa 3 glykogensynthasy antagonisté a inhibitory   metabolismus   MeSH
• kultivované buňky MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• organokovové sloučeniny farmakologie   MeSH
• oximy farmakologie   MeSH
• pyridiny farmakologie   MeSH
• pyrimidiny farmakologie   MeSH
• receptory aromatických uhlovodíků agonisté   chemie   genetika   metabolismus   MeSH
• rekombinantní fúzní proteiny chemie   metabolismus   MeSH
• reportérové geny účinky léků   MeSH
• RNA interference MeSH
• signální dráha Wnt účinky léků   MeSH
• simulace molekulového dockingu MeSH
• steroidní receptory agonisté   genetika   metabolismus   MeSH
• systém (enzymů) cytochromů P-450 chemie   genetika   metabolismus   MeSH
• transkripční faktory bHLH agonisté   chemie   genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH

Actin-associated proteins regulate multiple cellular processes, including proliferation and differentiation, but the molecular mechanisms underlying these processes are unclear. Here, we report that the actin-binding protein filamin A (FlnA) physically interacts with the actin-nucleating protein formin 2 (Fmn2). Loss of FlnA and Fmn2 impairs proliferation, thereby generating multiple embryonic phenotypes, including microcephaly. FlnA interacts with the Wnt co-receptor Lrp6. Loss of FlnA and Fmn2 impairs Lrp6 endocytosis, downstream Gsk3β activity, and β-catenin accumulation in the nucleus. The proliferative defect in Flna and Fmn2 null neural progenitors is rescued by inhibiting Gsk3β activity. Our findings thus reveal a novel mechanism whereby actin-associated proteins regulate proliferation by mediating the endocytosis and transportation of components in the canonical Wnt pathway. Moreover, the Fmn2-dependent signaling in this pathway parallels that seen in the non-canonical Wnt-dependent regulation of planar cell polarity through the Formin homology protein Daam. These studies provide evidence for integration of actin-associated processes in directing neuroepithelial proliferation.

• MeSH
• beta-katenin metabolismus   MeSH
• buněčná diferenciace MeSH
• buněčná membrána fyziologie   MeSH
• buněčné linie MeSH
• endocytóza fyziologie   MeSH
• filaminy genetika   metabolismus   MeSH
• GSK3B antagonisté a inhibitory   metabolismus   MeSH
• HEK293 buňky MeSH
• jaderné proteiny genetika   metabolismus   MeSH
• LDL receptor related protein 6 metabolismus   MeSH
• lidé MeSH
• mikrocefalie genetika   MeSH
• mikrofilamentové proteiny genetika   metabolismus   MeSH
• myši knockoutované MeSH
• myši MeSH
• proliferace buněk genetika   fyziologie   MeSH
• proteiny Wnt metabolismus   MeSH
• signální dráha Wnt fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH