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Brain‐derived neurotrophic factor (BDNF) content and signaling has been identified as one potential regulator of amyloid precursor protein (APP) processing. Recently published work has demonstrated that BDNF reduces BACE1 activity while also elevating the inhibition of GSK3β in the prefrontal cortex of male C57BL/6J mice. These results provide evidence that BDNF alters APP processing by reducing BACE1 activity, which may act through GSK3β inhibition. The purpose of this study was to further explore the role of GSK3β in BDNF‐induced regulation on BACE1 activity. We utilized a cell culture and an in vitro activity assay model to pharmacologically target BDNF and GSK3β signaling to confirm its involvement in the BDNF response. Treatment of differentiated SH‐SY5Y neuronal cells with 75 ng/mL BDNF resulted in elevated pTrkB content, pAkt content, pGSK3β content, and reduced BACE1 activity. An in vitro BACE1 activity assay utilizing mouse prefrontal cortex (n = 6/group) supplemented with BDNF, BDNF + ANA12 (Trkb antagonist), or BDNF + wortmannin (Akt inhibitor) demonstrated that BDNF reduced BACE1 activity; however, in the presence of TrkB or Akt inhibition, this effect was abolished. An in vitro ADAM10 activity assay utilizing mouse prefrontal cortex (n = 6/group) supplemented with BDNF, BDNF + ANA12 (Trkb antagonist), or BDNF + wortmannin (Akt inhibitor) demonstrated that BDNF did not alter ADAM10 activity. However, inhibiting BDNF signaling reduced ADAM10 activity. Collectively these studies suggest that GSK3β inhibition may be necessary for BDNF‐induced reductions in BACE1 activity. These findings will allow for the optimization of future therapeutic strategies by selectively targeting TrkB activation and GSK3β inhibition. [ABSTRACT FROM AUTHOR]

Glucose is the main brain fuel in fed conditions, while astrocytic glycogen is used as supplemental fuel when the brain is stimulated. Brain glycogen levels are decreased shortly after induced seizures in rodents, but little is known about how glycogen levels are affected interictally in chronic models of epilepsy. Reduced glutamine synthetase activity has been suggested to lead to increased brain glycogen levels in humans with chronic epilepsy. Here, we used the mouse pilocarpine model of epilepsy to investigate whether brain glycogen levels are altered, both acutely and in the chronic stage of the model. One day after pilocarpine‐induced convulsive status epilepticus (CSE), glycogen levels were higher in the hippocampal formation, cerebral cortex, and cerebellum. Opposite to expected, this was accompanied by elevated glutamine synthetase activity in the hippocampus but not the cortex. Increased interictal glycogen amounts were seen in the hippocampal formation and cerebral cortex in the chronic stage of the model (21 days post‐CSE), suggesting long‐lasting alterations in glycogen metabolism. Glycogen solubility in the cerebral cortex was unaltered in this epilepsy mouse model. Glycogen synthase kinase 3 beta (Gsk3b) mRNA levels were reduced in the hippocampal formations of mice in the chronic stage, which may underlie the elevated brain glycogen content in this model. This is the first report of elevated interictal glycogen levels in a chronic epilepsy model. Increased glycogen amounts in the brain may influence seizure susceptibility in this model, and this warrants further investigation. [ABSTRACT FROM AUTHOR]

Allergic rhinitis (AR) is a chronic inflammatory disease of the nasal mucosa. M2 macrophage polarization can reduce inflammation and repair tissue injury during AR development. Studies have substantiated the involvement of miRNAs in AR pathogenesis. Herein, the molecular mechanism of miR‐214‐3p in AR development was explored. To mimic the AR environment, ovalbumin (OVA) was used to treat macrophages. MiR‐214‐3p and glycogen synthase kinase 3 beta (GSK3B) expression in nasal mucus tissues and macrophages was assessed by RT‐qPCR. The M2 phenotypic signature of CD206 in macrophages was assessed by flow cytometry. The protein expression of GSK3B and M2 macrophage markers (ARG‐1 and IL‐10) was evaluated by western blotting. The correlation between miR‐214‐3p and GSK3B was validated by a luciferase reporter assay. We found that miR‐214‐3p was overexpressed in macrophages and nasal mucus tissues from AR patients. MiR‐214‐3p facilitated M2 polarization of macrophages upon OVA stimulation. Mechanistically, miR‐214‐3p targeted the GSK3B 3′ untranslated region in macrophages. In addition, GSK3B was downregulated in macrophages and nasal mucus tissues from AR patients. In rescue assays, GSK3B downregulation reversed the inhibitory effects of miR‐214‐3p silencing on M2 polarization of macrophages treated with OVA. Overall, miR‐214‐3p facilitates M2 macrophage polarization by targeting GSK3B. [ABSTRACT FROM AUTHOR]

microRNAs (miRNAs) have recently been recognized as playing an important role in bone‐associated diseases. This study investigated whether the reduced miR‐155‐5p in steroid‐associated osteonecrosis of the femoral head (ONFH) attenuated osteogenic differentiation and cell proliferation by targeting GSK3B. Bone marrow was collected from the proximal femurs of patients with steroid‐associated ONFH (n = 10) and patients with new femoral neck fracture (n = 10) and mesenchymal stem cells (MSCs) were isolated. The expression profile, the biological function of miR‐155‐5p, and the interaction between miR‐155‐5p and GSK3B were investigated by cell viability measurement, western blot, real‐time polymerase chain reaction, luciferase reporter assay, and Alizarin Red S (ARS) staining of MSCs. The MSCs that were obtained from the femoral neck fracture group and from the steroid‐associated ONFH group were transfected with or without miR‐155‐5p. We found that, in ONFH samples, the level of mature miR‐155‐5p was significantly lower than that of control samples. By inhibiting GSK3B, miR‐155‐5p promoted the nuclear translocation of β‐catenin, increased the expression of osteogenesis‐related genes, and facilitated the proliferation and differentiation of MSCs. Restoring the expression of GSK3B in MSCs partially reversed the effect of miR‐155‐5p. These findings suggest that reduced miR‐155‐5p in steroid‐associated ONFH attenuates osteogenic differentiation and cell proliferation by increased levels of GSK3B and inhibition of Wnt signaling. [ABSTRACT FROM AUTHOR]

Lithium ion, commonly used as the carbonate salt in the treatment of bipolar disorders, has been identified as an inhibitor of several kinases, including Glycogen Synthase Kinase‐3β, for almost 20 years. However, both the exact mechanism of enzymatic inhibition and its apparent specificity for certain metalloenzymes are still a matter of debate. A data‐driven hypothesis is presented that accounts for the specificity profile of kinase inhibition by lithium in terms of the presence of a unique protein environment in the magnesium‐binding site. This hypothesis has been validated by the discovery of two novel potential targets for lithium, namely NEK3 and MOK, which are related to neuronal function. [ABSTRACT FROM AUTHOR]

Objective: AKT1 and GSK3B take part in one of the intracellular cascades activated by the D2 dopamine receptor (DRD2). This receptor is antagonized by antipsychotics and plays a role in the pathogenesis of antipsychotic‐induced tardive dyskinesia (TD). The present study investigated association of several polymorphisms in the two candidate genes, AKT1 and GSK3B, with TD in antipsychotic‐treated patients with schizophrenia. Methods: DNA samples from 449 patients from several Siberian regions (Russia) were genotyped, and the results were analyzed using chi‐squared tests and analyses of variance. Results: Antipsychotic‐induced TD was not associated with either of the tested functional polymorphisms (rs334558, rs1130214, and rs3730358). Conclusions: Despite regulation of AKT1 and GSK3B by DRD2, we found no evidence that these two kinases play a major role in the pathogenesis of antipsychotic‐induced TD. These results agree with previously published data and necessitate further exploration of other pathogenic mechanisms, such as neurotoxicity due to excessive dopamine metabolism. [ABSTRACT FROM AUTHOR]

There is growing evidence that the impaired IGF-I system contributes to neurodegeneration. In this study, we examined the spinal cords of the EAE, the animal model of multiple sclerosis, to see if the expression of the IGF-I system is altered. To induce EAE, C57/BL6 mice were immunized with the Hooke lab MOG kit, sacrificed at the peak of the disease and their spinal cords were examined for the immunoreactivities (ir) of the IGF-I, IGF binding protein-1 (IGFBP-1) and glycogen synthase kinase 3b (GSK3b), as one major downstream molecule in the IGF-I signaling. Although neurons in the non EAE spinal cords did not show the IGF-I immunoreactivity, they were numerously positive for the IGFBP-1. In the inflamed EAE spinal cord however, the patterns of expressions were reversed, that is, a significant increased number of IGF-I expressing neurons versus a reduced number of IGFBP-1 positive neurons. Moreover, while nearly all IGF-I-ir neurons expressed GSK3b, some expressed it more intensely. Considering our previous finding where we showed a significant reduced number of the inactive (phosphorylated) but not that of the total GSK3b expressing neurons in the EAE spinal cord, it is conceivable that the intense total GSK3b expression in the IGF-I-ir neurons belongs to the active form of GSK3b known to exert neuroinflammatory effects. We therefore suggest that the altered expression of the IGF-I system including GSK3b in spinal cord neurons might involve in pathophysiological events during the EAE. [ABSTRACT FROM AUTHOR]

BACKGROUND: Cancer stem cells (CSCs) are involved in both tumourigenesis and in tumour recurrence after therapy. In head and neck squamous cell carcinoma (HNSCC), there are two biologically different CSC phenotypes both of which express high levels of CD44 but differ in their expression levels of epithelial-specific antigen (ESA). One phenotype is CD44high/ESAhigh and has epithelial features (Epi-CSCs), while the other is CD44high/ESAlow, has undergone epithelial to mesenchymal transition (EMT-CSCs), has mesenchymal features and is migratory (Biddle et al., 2011). CSCs are resistant to therapeutically induced apoptosis but the molecular mechanisms by which they develop apoptotic resistance remains unclear. However, glycogen synthase kinase 3β (GSK3β) contributes to regulation of both the selfrenewal and switching of these two CSC phenotypes (Shigeishi et al., 2013). METHODS: CD44high/ESAlow, CD44high/ESAhigh and CD44low cells were FACS sorted from the HNSCC cell line LUC4, and 5-FU-induced apoptosis was analysed by Annexin V staining followed by flow cytometry analysis. RESULTS: CD44high/ESAlow cells exhibited marked resistance to 5-FU-induced apoptosis and had high expression of dihydropyrimidine dehydrogenase (DPD). The DPD inhibitor, 5-chloro-2, 4-dihydroxypyridine (CDHP) significantly enhanced 5-FU-induced apoptosis of CD44high/ ESAlow cells. Inhibition of GSK3β induced CD44high/ ESAlow cells to undergo mesenchymal-to-epithelial transition (MET) to CD44high/ESAhigh cells and pre-existing CD44high/ESAhigh cells to differentiate. Apoptosis induced by 5-FU was thus facilitated. Combination of both CDHP and GSK3β inhibitors markedly enhanced 5-FU-induced apoptosis of CD44high/ESAlow cells. CONCLUSIONS: Our results suggest potentially new approaches for the elimination of the therapy resistant HNSCC CSC population. [ABSTRACT FROM AUTHOR]

Glycogen synthase kinase 3 beta (Gsk3b) acts as a negative modulator in endothelial cells through the Wnt/β-catenin/PI3K/AKT/Gsk3b axis in cancer- induced angiogenesis. However, the function of Gsk3b during embryonic angiogenesis remains unclear. Here, either gsk3b knockdown by morpholino or Gsk3b loss of activity by LiCl treatment had serious phenotypic consequences, such as defects in the positioning and patterning of intersegmental blood vessels and reduction of vegfaa121 and vegfaa165 transcripts. In embryos treated with the phosphatidylinositol 3-kinase inhibitor, angiogenesis was severely inhibited, along with reduced Wnt, phosphorylated AKT and phosphorylated Gsk3b, suggesting that the remaining Gsk3b in somites could still degrade b-catenin, resulting in decreased vascular endothelial growth factor Aa(VegfAa) expression. However, in gsk3b-mRNA-overexpressed embryos, intersegmental vessels ectopically sprouted by the increase in phosphorylated-Gsk3b which prevented the degradation of β-catenin and promoted the increase in phosphorylated AKT activity, thus increasing VegfAa expression in somites. Interestingly, the Gsk3b-dependent cross-talk between PI3K/AKT and Wnt/β-catenin suggests that Wnt/b-catenin and PI3K/AKT interaction controls embryonic angiogenesis by a positive feedback loop rather than a hierarchical framework such as that found in cancer-induced angiogenesis. Thus, both active and inactive forms of Gsk3b mediate the cooperative signaling between Wnt/β-catenin and PI3K/AKT to control VegfAa expression in somites during angiogenesis in zebrafish embryos. [ABSTRACT FROM AUTHOR]

Recent evidence suggests that variation in the SNCA, MAPT, and GSK3B genes interacts in affecting risk for Parkinson disease (PD). In the current study, we attempt to validate previously published findings, evaluating gene-gene interactions between SNCA, MAPT, and GSK3B in association with PD. Three Caucasian PD patient-control series from the United States, Ireland, and Norway (combined n = 1020 patients and 1095 controls) were genotyped for SNCA rs356219, MAPT H1/H2-discriminating SNP rs1052553, and GSK3B rs334558 and rs6438552. Our findings indicate that as previously reported, the SNCA rs356219-G allele and MAPT rs1052553 (H1 haplotype) were both associated with an increased risk of PD, whilst contrary to previous reports, GSK3B variants were not. No pair-wise interaction was observed between SNCA, MAPT, and GSK3B; the risk effects of SNCA rs356219-G and MAPT rs1052553-H1 were seen in a similar manner across genotypes of other variants, with no evidence suggesting synergistic, antagonistic, or deferential effects. In the Caucasian patient-control series examined, risk for PD was influenced by variation in SNCA and MAPT but not GSK3B. Additionally, those three genes did not interact in determining disease risk. [ABSTRACT FROM AUTHOR]

Expression of the c-Myc proto-oncoprotein is tightly regulated in normal cells. Phosphorylation at two conserved residues, threonine58 (T58) and serine62 (S62), regulates c-Myc protein stability. In cancer cells, c-Myc can become aberrantly stabilized associated with altered T58 and S62 phosphorylation. A complex signalling cascade involving GSK3β kinase, the Pin1 prolyl isomerase, and the PP2A-B56α phosphatase controls phosphorylation at these sites. We report here a novel role for the tumour suppressor scaffold protein Axin1 in facilitating the formation of a degradation complex for c-Myc containing GSK3β, Pin1, and PP2A-B56α. Although knockdown of Axin1 decreases the association of c-Myc with these proteins, reduces T58 and enhances S62 phosphorylation, and increases c-Myc stability, acute expression of Axin1 reduces c-Myc levels and suppresses c-Myc transcriptional activity. Moreover, the regulation of c-Myc by Axin1 is impaired in several tested cancer cell lines with known stabilization of c-Myc or loss of Axin1. This study provides critical insight into the regulation of c-Myc expression, how this can be disrupted in three cancer types, and adds to our knowledge of the tumour suppressor activity of Axin1. [ABSTRACT FROM AUTHOR]

Certain effects of melatonin on senescence were investigated. The experimental model used was 10-month-old senescence-accelerated mouse prone 8 (SAMP8). The mice in the experiment were administered melatonin (10 mg/kg) from the age of 1 month. Results showed that chronic administration of melatonin decreased cell loss in the cerebral cortex and reduced oxidative damage in protein and lipids. There are several studies suggesting that the activation of the cdk5/p35 pathway at its cleavage to cdk5/p25 may play a role in hyperphosphorylation of tau during aging and neurodegenerative diseases. Melatonin not only reduced the cerebral aging disturbances, but also prevented tau hyperphosphorylation present in the experimental model used in this study. Melatonin reduced cdk5 expression, as well as the cleavage of p35 to p25. The other tau kinase studied, GSK3 β, showed a reduction in this activity in comparison with SAMP8 nontreated SAMP8. These data indicate that melatonin possesses neuroprotective properties against cerebral damage gated to senescence. Moreover, these data suggest that the cdk5/GSK β signaling cascade has a potential role as a target for neurodegenerative diseases related to aging. [ABSTRACT FROM AUTHOR]

The glycogen synthase kinase 3B (GSK3B) is an important target protein of several antidepressants, such as lithium, a mood stabilizer. Recent studies associated structural variations of the GSK3B gene to bipolar disorder (BP), although replications were not conclusive. Here we present data on copy number variations (CNVs) of the GSK3B gene probing the 9th exon region in 846 individuals (414 controls, 172 patients with major depressive disorder (MDD) and 260 with BP). A significant accumulation (odds ratio: 5.5, P = 0.00051) of the amplified exon 9 region was found in patients (22 out of 432) compared to controls (4 of 414). Analyzing patient subgroups, GSK3B structural variants were found to be risk factors of BP particularly (P = 0.00001) with an odds ratio of 8.1 while no such effect was shown in the MDD group. The highest odds (19.7 ratio) for bipolar disorder was observed in females with the amplified exon 9 region. A more detailed analysis of the identified GSK3B CNV by a set of probes covering the GSK3B gene and the adjacent NR1I2 and C3orf15 genes showed that the amplified sequences contained 3' (downstream) segments of the GSK3B and NR1I2 genes but none of them involved the C3orf15 gene. Therefore, the copy number variation of the GSK3B gene could be described as a complex set of structural variants involving partial duplications and deletions, simultaneously. In summary, here we confirmed significant association of the GSK3B CNV and bipolar disorder pointing out that the copy number and extension of the CNV varies among individuals., (© 2014 Wiley Periodicals, Inc.)