Members of the casein kinase 1 (CK1) family are important regulators of multiple signaling pathways. CK1α is a well-known negative regulator of the Wnt/β-catenin pathway, which promotes the degradation of β-catenin via its phosphorylation of Ser45. In contrast, the closest paralog of CK1α, CK1α-like, is a poorly characterized kinase of unknown function. In this study, we show that the deletion of CK1α, but not CK1α-like, resulted in a strong activation of the Wnt/β-catenin pathway. Wnt-3a treatment further enhanced the activation, which suggests there are at least two modes, a CK1α-dependent and Wnt-dependent, of β-catenin regulation. Rescue experiments showed that only two out of ten naturally occurring splice CK1α/α-like variants were able to rescue the augmented Wnt/β-catenin signaling caused by CK1α deficiency in cells. Importantly, the ability to phosphorylate β-catenin on Ser45 in the in vitro kinase assay was required but not sufficient for such rescue. Our compound CK1α and GSK3α/β KO models suggest that the additional nonredundant function of CK1α in the Wnt pathway beyond Ser45-β-catenin phosphorylation includes Axin phosphorylation. Finally, we established NanoBRET assays for the three most common CK1α splice variants as well as CK1α-like. Target engagement data revealed comparable potency of known CK1α inhibitors for all CK1α variants but not for CK1α-like. In summary, our work brings important novel insights into the biology of CK1α, including evidence for the lack of redundancy with other CK1 kinases in the negative regulation of the Wnt/β-catenin pathway at the level of β-catenin and Axin.

• MeSH
• alternativní sestřih MeSH
• beta-katenin * metabolismus   genetika   MeSH
• fosforylace MeSH
• GSK3B metabolismus   genetika   MeSH
• HEK293 buňky MeSH
• kasein kinasa Ialfa * metabolismus   genetika   MeSH
• kinasa 3 glykogensynthasy metabolismus   genetika   MeSH
• lidé MeSH
• protein Wnt3A metabolismus   genetika   MeSH
• signální dráha Wnt * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH

Thermogenesis in brown adipose tissue (BAT) uses intracellular triglycerides, circulating free fatty acids and glucose as the main substrates. The objective of the current study was to analyse the role of CD36 fatty acid translocase in regulation of glucose and fatty acid utilisation in BAT. BAT isolated from spontaneously hypertensive rat (SHR) with mutant Cd36 gene and SHR-Cd36 transgenic rats with wild type variant was incubated in media containing labeled glucose and palmitate to measure substrate incorporation and oxidation. SHR-Cd36 versus SHR rats showed significantly increased glucose incorporation into intracellular lipids associated with reduced glycogen synthase kinase 3β (GSK-3β) protein expression and phosphorylation and increased oxidation of exogenous palmitate. It can be concluded that CD36 enhances glucose transport for lipogenesis in BAT by suppressing GSK-3β and promotes direct palmitate oxidation.

• MeSH
• antigeny CD36 genetika   metabolismus   MeSH
• glukosa * metabolismus   MeSH
• GSK3B metabolismus   MeSH
• hnědá tuková tkáň * metabolismus   MeSH
• krysa rodu rattus MeSH
• mastné kyseliny metabolismus   MeSH
• palmitany metabolismus   MeSH
• potkani inbrední SHR MeSH
• potkani transgenní MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus 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

The infarct size-limiting effect elicited by cold acclimation (CA) is accompanied by increased mitochondrial resistance and unaltered β1-adrenergic receptor (AR) signaling persisting for 2 wk at room temperature. As the mechanism of CA-elicited cardioprotection is not fully understood, we examined the role of the salvage β2-AR/Gi/Akt pathway. Male Wistar rats were exposed to CA (8°C, 5 wk), whereas the recovery group (CAR) was kept at 24°C for additional 2 wk. We show that the total number of myocardial β-ARs in the left ventricular myocardium did not change after CA but decreased after CAR. We confirmed the infarct size-limiting effect in both CA and CAR groups. Acute administration of β2-AR inhibitor ICI-118551 abolished the protective effect in the CAR group but had no effect in the control and CA groups. The inhibitory Giα1/2 and Giα3 proteins increased in the membrane fraction of the CAR group, and the phospho-Akt (Ser473)-to-Akt ratio also increased. Expression, phosphorylation, and mitochondrial location of the Akt target glycogen synthase kinase (GSK-3β) were affected neither by CA nor by CAR. However, GSK-3β translocated from the Z-disk to the H-zone after CA, and acquired its original location after CAR. Our data indicate that the cardioprotection observed after CAR is mediated by the β2-AR/Gi pathway and Akt activation. Further studies are needed to unravel downstream targets of the central regulators of the CA process and the downstream targets of the Akt protein after CAR.NEW & NOTEWORTHY Cardioprotective effect of cold acclimation and that persisting for 2 wk after recovery engage in different mechanisms. The β2-adrenoceptor/Gi pathway and Akt are involved only in the mechanism of infarct size-limiting effect occurring during the recovery phase. GSK-3β translocated from the Z-line to the H-zone of sarcomeres by cold acclimation returns back to the original position after the recovery phase. The results provide new insights potentially useful for the development of cardiac therapies.

• MeSH
• aklimatizace MeSH
• beta-2-adrenergní receptory MeSH
• fosforylace MeSH
• GSK3B MeSH
• krysa rodu rattus MeSH
• potkani Wistar MeSH
• protoonkogenní proteiny c-akt metabolismus   MeSH
• reperfuzní poškození myokardu * MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: Alzheimer´s disease (AD) is characterized by a progressive neuronal degeneration caused by two pathological hallmarks, hyperphosphorylated tau protein aggregated into tau filaments and amyloid precursor protein derived beta amyloid peptides aggregated into extracellular amyloid plaques. All attempts so far to find effective drugs failed in clinical trials. AD is a multifactorial disease, so that selective drugs to target one AD-relevant structure alone may not be sufficient. OBJECTIVE: We built novel furopyridines with various substitution patterns to evaluate them as protein kinases inhibitors of enzymes related to tau pathology. METHODS: Furopyridine derivatives were synthesized and purified using column chromatography. The protein kinase inhibitory properties were determined in ATP-competition assays with determined affinity constants for the most active compounds. RESULTS: The compounds were prepared in simple two-component reactions of substituted 1,4- dihydropyridines and respective quinones to obtain various substitutions of the molecular furopyridine scaffold. The substituent effects on the determined kinase inhibitory properties of cdk1, cdk2, Fyn, JNK3 and gsk-3β are discussed. CONCLUSION: Various 3-substitutions were found most sensitive for the protein kinase inhibition depending on the length, nature and a substituent positioning within. We identified compounds as inhibitors of several kinases as a tool to potentially combat the disease progress in a multitargeting approach.

• MeSH
• Alzheimerova nemoc farmakoterapie   MeSH
• fosforylace účinky léků   MeSH
• GSK3B metabolismus   MeSH
• inhibitory proteinkinas chemie   farmakologie   terapeutické užití   MeSH
• lidé MeSH
• proteinkinasy metabolismus   MeSH
• proteiny tau metabolismus   MeSH
• pyridiny chemie   farmakologie   terapeutické užití   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Aging attenuates cardiac tolerance to ischemia/reperfusion (I/R) associated with defects in protective cell signaling, however, the onset of this phenotype has not been completely investigated. This study aimed to compare changes in response to I/R and the effects of remote ischemic preconditioning (RIPC) in the hearts of younger adult (3 months) and mature adult (6 months) male Wistar rats, with changes in selected proteins of protective signaling. Langendorff-perfused hearts were exposed to 30 min I/120 min R without or with prior three cycles of RIPC (pressure cuff inflation/deflation on the hind limb). Infarct size (IS), incidence of ventricular arrhythmias and recovery of contractile function (LVDP) served as the end points. In both age groups, left ventricular tissue samples were collected prior to ischemia (baseline) and after I/R, in non-RIPC controls and in RIPC groups to detect selected pro-survival proteins (Western blot). Maturation did not affect post-ischemic recovery of heart function (Left Ventricular Developed Pressure, LVDP), however, it increased IS and arrhythmogenesis accompanied by decreased levels and activity of several pro-survival proteins and by higher levels of pro-apoptotic proteins in the hearts of elder animals. RIPC reduced the occurrence of reperfusion-induced ventricular arrhythmias, IS and contractile dysfunction in younger animals, and this was preserved in the mature adults. RIPC did not increase phosphorylated protein kinase B (p-Akt)/total Akt ratio, endothelial nitric oxide synthase (eNOS) and protein kinase Cε (PKCε) prior to ischemia but only after I/R, while phosphorylated glycogen synthase kinase-3β (GSK3β) was increased (inactivated) before and after ischemia in both age groups coupled with decreased levels of pro-apoptotic markers. We assume that resistance of rat heart to I/R injury starts to already decline during maturation, and that RIPC may represent a clinically relevant cardioprotective intervention in the elder population.

• MeSH
• fosforylace MeSH
• GSK3B genetika   metabolismus   MeSH
• hemodynamika MeSH
• ischemické přivykání * MeSH
• krysa rodu rattus MeSH
• myokard metabolismus   MeSH
• potkani Wistar MeSH
• proteinkinasa C-epsilon genetika   metabolismus   MeSH
• protoonkogenní proteiny c-akt genetika   metabolismus   MeSH
• reperfuzní poškození myokardu metabolismus   patologie   MeSH
• stárnutí MeSH
• synthasa oxidu dusnatého, typ III genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Multitarget-directed ligands (MTDLs) are considered a promising therapeutic strategy to address the multifactorial nature of Alzheimer's disease (AD). Novel MTDLs have been designed as inhibitors of human acetylcholinesterases/butyrylcholinesterases, monoamine oxidase A/B, and glycogen synthase kinase 3β and as calcium channel antagonists via the Biginelli multicomponent reaction. Among these MTDLs, (±)-BIGI-3h was identified as a promising new hit compound showing in vitro balanced activities toward the aforementioned recognized AD targets. Additional in vitro studies demonstrated antioxidant effects and brain penetration, along with the ability to inhibit the aggregation of both τ protein and β-amyloid peptide. The in vivo studies have shown that (±)-BIGI-3h (10 mg/kg intraperitoneally) significantly reduces scopolamine-induced cognitive deficits.

• MeSH
• Alzheimerova nemoc * farmakoterapie   MeSH
• blokátory kalciových kanálů farmakologie   terapeutické užití   MeSH
• cholinesterasové inhibitory farmakologie   terapeutické užití   MeSH
• GSK3B MeSH
• lidé MeSH
• ligandy MeSH
• monoaminoxidasa metabolismus   MeSH
• vápníkové kanály 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

In analogy to antiviral acyclic nucleoside phosphonates, a series of 5-amino-3-oxo-1,2,4-thiadiazol-3(2H)-ones bearing a 2-phosphonomethoxyethyl (PME) or 3-hydroxy-2-(phosphonomethoxy)propyl (HPMP) group at the position 2 of the heterocyclic moiety has been synthesized. Diisopropyl esters of PME- and HPMP-amines have been converted to the N-substituted ureas and then reacted with benzoyl, ethoxycarbonyl, and Fmoc isothiocyanates to give the corresponding thiobiurets, which were oxidatively cyclized to diisopropyl esters of 5-amino-3-oxo-2-PME- or 2-HPMP- 1,2,4-thiadiazol-3(2H)-ones. The phosphonate ester groups were cleaved with bromotrimethylsilane, yielding N5-protected phosphonic acids. The subsequent attempts to remove the protecting group from N5 under alkaline conditions resulted in the cleavage of the 1,2,4-thiadiazole ring. Similarly, compounds with a previously unprotected 5-amino-1,2,4-thiadiazolone base moiety were stable only in the form of phosphonate esters. The series of twenty-one newly prepared 1,2,4-thiadiazol-3(2H)-ones were explored as potential inhibitors of cysteine-dependent enzymes - human cathepsin K (CatK) and glycogen synthase kinase 3β (GSK-3β). Several compounds exhibited an inhibitory activity toward both enzymes in the low micromolar range. The inhibitory potency of some of them toward GSK-3β was similar to that of the thiadiazole GSK-3β inhibitor tideglusib, whereas others exhibited more favorable toxicity profile while retaining good inhibitory activity.

• MeSH
• antitumorózní látky chemická syntéza   chemie   farmakologie   MeSH
• buněčné linie MeSH
• GSK3B antagonisté a inhibitory   metabolismus   MeSH
• inhibitory enzymů chemická syntéza   chemie   farmakologie   MeSH
• kathepsin K antagonisté a inhibitory   metabolismus   MeSH
• léky antitumorózní - screeningové testy MeSH
• lidé MeSH
• molekulární struktura MeSH
• nukleosidy chemická syntéza   chemie   farmakologie   MeSH
• organofosfonáty chemická syntéza   chemie   farmakologie   MeSH
• proliferace buněk účinky léků   MeSH
• thiadiazoly chemická syntéza   chemie   farmakologie   MeSH
• viabilita buněk účinky léků   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé 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