Regulation by Glycogen Synthase Kinase-3 of Inflammation and T Cells in CNS Diseases

Elevated markers of neuroinflammation have been found to be associated with many psychiatric and neurodegenerative diseases, such as mood disorders, Alzheimer’s disease, and multiple sclerosis (MS). Since neuroinflammation is thought to contribute to the pathophysiology of these diseases and to impair responses to therapeutic interventions and recovery, it is important to identify mechanisms that regulate neuroinflammation and potential targets for controlling neuroinflammation. Recent findings have demonstrated that glycogen synthase kinase-3 (GSK3) is an important regulator of both the innate and adaptive immune systems’ contributions to inflammation. Studies of the innate immune system have shown that inhibitors of GSK3 profoundly alter the repertoire of cytokines that are produced both by peripheral and central cells, reducing pro-inflammatory cytokines, and increasing anti-inflammatory cytokines. Furthermore, inhibitors of GSK3 promote tolerance to inflammatory stimuli, reducing inflammatory cytokine production upon repeated exposure. Studies of the adaptive immune system have shown that GSK3 regulates the production of cytokines by T cells and the differentiation of T cells to subtypes, particularly Th17 cells. Regulation of transcription factors by GSK3 appears to play a prominent role in its regulation of immune responses, including of NF-κB, cyclic AMP response element binding protein, and signal transducer and activator of transcription-3. In vivo studies have shown that GSK3 inhibitors ameliorate clinical symptoms of both peripheral and central inflammatory diseases, particularly experimental autoimmune encephalomyelitis, the animal model of MS. Therefore, the development and application of GSK3 inhibitors may provide a new therapeutic strategy to reduce neuroinflammation associated with many central nervous system diseases

Regulation of Cell Survival Mechanisms in Alzheimer's Disease by Glycogen Synthase Kinase-3

A pivotal role has emerged for glycogen synthase kinase-3 (GSK3) as an important contributor to Alzheimer's disease pathology. Evidence for the involvement of GSK3 in Alzheimer's disease pathology and neuronal loss comes from studies of GSK3 overexpression, GSK3 localization studies, multiple relationships between GSK3 and amyloid β-peptide (Aβ), interactions between GSK3 and the microtubule-associated tau protein, and GSK3-mediated apoptotic cell death. Apoptotic signaling proceeds by either an intrinsic pathway or an extrinsic pathway. GSK3 is well established to promote intrinsic apoptotic signaling induced by many insults, several of which may contribute to neuronal loss in Alzheimer's disease. Particularly important is evidence that GSK3 promotes intrinsic apoptotic signaling induced by Aβ. GSK3 appears to promote intrinsic apoptotic signaling by modulating proteins in the apoptosis signaling pathway and by modulating transcription factors that regulate the expression of proteins involved in apoptosis. Thus, GSK3 appears to contribute to several neuropathological mechanisms in Alzheimer's disease, including apoptosis-mediated neuronal loss

Pro- and Anti-inflammatory Biomarkers as Predictors of Response to Valproate in Patients with Comorbid Alcohol Use and Bipolar Disorder-Preliminary Findings

Objective/Hypothesis: Bipolar disorder (BD) has the highest association with alcohol and other substance use disorders compared to other major psychiatric disorders. This patient population is particularly challenging to treat. We have previously shown that some patients with co-occurring alcohol use and bipolar disorders respond to the GABAergic agonist valproate (VPA), which is known to modulates the dopaminergic system, and also as an epigenetic modifier. Predictors of therapeutic response to VPA in patients with AUD/BD are not known, and the subgroup which would benefit from VPA is still to be identified. Recent evidence suggests that AUD promotes a pro-inflammatory state while VPA increases levels of anti-inflammatory factors. We hypothesized that VPA has an anti-inflammatory effect and that patients with AUD/BD who respond to VPA have higher baseline inflammatory indices. Methods: Nine patients with DSM-IV-defined diagnoses of AUD and BD (AUD/BD) were enrolled in the study. Patients received a course of VPA for 3 months at an average dose of 1000 mg a day in addition to receiving either naltrexone of 50 mg daily or placebo. Blood was collected prior to the initiation of VPA and throughout the treatment study. Liver function tests and trough VPA serum concentrations were evaluated periodically. Alcohol use outcome was assessed using the Timeline Follow-Back for Recent Drinking. The use of other drugs was monitored through regular urine drug screen. The primary alcohol use outcome was changes in proportion of weekly heavy drinking days (defined as ³ 5 drinks per day for men and ³ 4 drinks per day for women). Plasma levels of cytokines were measured using Multiplex Immunoassay, in accordance to the manufacturers’ recommendations. Results: We found that about one half of enrolled patients responded to VPA. Screening of pro- and anti-inflammatory cytokines showed that responders had higher levels of the chemokine SDF-1a/CXCL12a and the pro-inflammatory marker C-reactive protein (CRP) and lower levels of anti-inflammatory factor matrix metalloproteinase-10 (MMP-10) (p \u3c 0.05). Screening of cytokines in samples before and after treatment with VPA showed that VPA increased levels of anti-inflammatory factors interleukin-10 (IL-10) and MMP-10 (p \u3c 0.05) and tended to decrease levels of pro-inflammatory CRP (p \u3e 0.05). Discussion: Pro- and anti-inflammatory biomarkers may serve as predictors of treatment response to VPA in patients with combined AUD/BD. Our preliminary results also suggest that therapeutic effect of VPA may be in part due to anti-inflammatory action of VPA. Larger studies may be indicated to validate these findings

Involvement of Innate and Adaptive Immune Systems Alterations in the Pathophysiology and Treatment of Depression

Major depressive disorder (MDD) is a prevalent and debilitating disorder, often fatal. Treatment options are few and often do not provide immediate relief to the patients. The increasing involvement of inflammation in the pathology of MDD has provided new potential therapeutic avenues. Cytokine levels are elevated in the blood and cerebrospinal fluid of MDD patients whereas immune cells often exhibit an immunosuppressed phenotype in MDD patients. Blocking cytokine actions in patients exhibiting MDD show some antidepressant efficacy. However, the role of cytokines, and the immune response in MDD patients remain to be determined. We reviewed here the roles of the innate and adaptive immune systems in MDD, as well as potential mechanisms whereby the immune response might be regulated in MDD

First flower inclusion and fossil evidence of Cryptocarya (Laurales, Lauraceae) from Miocene amber of Zhangpu (China)

Lauraceae have one of the oldest fossil records of angiosperms with the earliest known evidence from the mid-Cretaceous. However, most of these records are based on leaves, especially from the Cenozoic of Asia, which are often challenging to assign to extinct or extant genera or species. In contrast, fossils of reproductive organs are more informative, but remain scarce. We here described the first Cenozoic Lauraceae flower of Asia and confirmed the presence of Cryptocarya in the Miocene Zhangpu flora (Fujian Province, south-eastern China) based on an amber inclusion. We scanned the specimen using synchrotron radiation-based micro-computed tomography (SRμCT) and then compared the fossil with extant flowers of the genus. The present fossil flower is small, bisexual, and polysymmetric, with a whorled and trimerous perianth and androecium along with a hypanthium around the gynoecium. The perianth comprises six undifferentiated tepals, the androecium consists of nine stamens and three innermost staminodes, and the gynoecium of a single carpel with a superior, unilocular (and uniovulate) ovary. Our study also shows that the fossil shares an unusual position of the typical staminal glands and a short androecial tube on the rim of the hypanthium with at least one extant Australian species of Cryptocarya, which have not been reported before. Nowadays, Lauraceae are still present in tropical to subtropical regions, mostly in American and Asian rainforests. The discovery of many Lauraceae leaf fossils in Zhangpu, as well as the amber flower of this study, is consistent with the current reconstruction of the amber source environment as a megathermal seasonal rainforest during the Mid-Miocene

GSK3 Influences Social Preference and Anxiety-Related Behaviors during Social Interaction in a Mouse Model of Fragile X Syndrome and Autism

BACKGROUND: Nearly 1% of children in the United States exhibit autism spectrum disorders, but causes and treatments remain to be identified. Mice with deletion of the fragile X mental retardation 1 (Fmr1) gene are used to model autism because loss of Fmr1 gene function causes Fragile X Syndrome (FXS) and many people with FXS exhibit autistic-like behaviors. Glycogen synthase kinase-3 (GSK3) is hyperactive in brains of Fmr1 knockout mice, and inhibition of GSK3 by lithium administration ameliorates some behavioral impairment in these mice. We extended our studies of this association by testing whether GSK3 contributes to socialization behaviors. This used two mouse models with disrupted regulation of GSK3, Fmr1 knockout mice and GSK3 knockin mice, in which inhibitory serines of the two isoforms of GSK3, GSK3alpha and GSK3beta, are mutated to alanines, leaving GSK3 fully active. METHODOLOGY/PRINCIPAL FINDINGS: To assess sociability, test mice were introduced to a restrained stimulus mouse (S1) for 10 min, followed by introduction of a second restrained stimulus mouse (S2) for 10 min, which assesses social preference. Fmr1 knockout and GSK3 knockin mice displayed no deficit in sociability with the S1 mouse, but unlike wild-type mice neither demonstrated social preference for the novel S2 mouse. Fmr1 knockout mice displayed more anxiety-related behaviors during social interaction (grooming, rearing, and digging) than wild-type mice, which was ameliorated by inhibition of GSK3 with chronic lithium treatment. CONCLUSIONS/SIGNIFICANCE: These results indicate that impaired inhibitory regulation of GSK3 in Fmr1 knockout mice may contribute to some socialization deficits and that lithium treatment can ameliorate certain socialization impairments. As discussed in the present work, these results suggest a role for GSK3 in social behaviors and implicate inhibition of GSK3 as a potential therapeutic

HDAC6 Regulates LPS-Tolerance in Astrocytes

Inflammatory tolerance is a crucial mechanism that limits inflammatory responses in order to avoid prolonged inflammation that may damage the host. Evidence that chronic inflammation contributes to the neuropathology of prevalent neurodegenerative and psychiatric diseases suggests that inflammatory tolerance mechanisms are often inadequate to control detrimental inflammation in the central nervous system. Thus, identifying mechanisms that regulate neuroinflammatory tolerance may reveal opportunities for bolstering tolerance to reduce chronic inflammation in these diseases. Examination of tolerance after repeated lipopolysaccharide (LPS) treatment of mouse primary astrocytes demonstrated that histone deacetylase (HDAC) activity promoted tolerance, opposite to the action of glycogen synthase kinase-3 (GSK3), which counteracts tolerance. HDAC6 in particular was found to be critical for tolerance induction, as its deacetylation of acetyl-tubulin was increased during LPS tolerance, this was enhanced by inhibition of GSK3, and the HDAC6 inhibitor tubacin completely blocked tolerance and the promotion of tolerance by inhibition of GSK3. These results reveal opposing interactions between HDAC6 and GSK3 in regulating tolerance, and indicate that shifting the balance between these two opposing forces on inflammatory tolerance can obliterate or enhance tolerance to LPS in astrocytes

Glycogen synthase kinase-3 inhibition disrupts nuclear factor-kappaB activity in pancreatic cancer, but fails to sensitize to gemcitabine chemotherapy

<p>Abstract</p> <p>Background</p> <p>Aberrant activation NF-kappaB has been proposed as a mechanism of drug resistance in pancreatic cancer. Recently, inhibition of glycogen synthase kinase-3 has been shown to exert anti-tumor effects on pancreatic cancer cells by suppressing NF-kappaB. Consequently, we investigated whether inhibition of GSK-3 sensitizes pancreatic cancer cells to the chemotherapeutic agent gemcitabine.</p> <p>Methods</p> <p>GSK-3 inhibition was achieved using the pharmacological agent AR-A014418 or siRNA against GSK-3 alpha and beta isoforms. Cytotoxicity was measured using a Sulphorhodamine B assay and clonogenic survival following exposure of six different pancreatic cancer cell lines to a range of doses of either gemcitabine, AR-A014418 or both for 24, 48 and 72 h. We measured protein expression levels by immunoblotting. Basal and TNF-alpha induced activity of NF-kappaB was assessed using a luciferase reporter assay in the presence or absence of GSK-3 inhibition.</p> <p>Results</p> <p>GSK-3 inhibition reduced both basal and TNF-alpha induced NF-kappaB luciferase activity. Knockdown of GSK-3 beta reduced nuclear factor kappa B luciferase activity to a greater extent than GSK-3 alpha, and the greatest effect was seen with dual knockdown of both GSK-3 isoforms. GSK-3 inhibition also resulted in reduction of the NF-kappaB target proteins XIAP, Bcl-X_L, and cyclin D1, associated with growth inhibition and decreased clonogenic survival. In all cell lines, treatment with either AR-A014418, or gemcitabine led to growth inhibition in a dose- and time-dependent manner. However, with the exception of PANC-1 where drug synergy occurred with some dose schedules, the inhibitory effect of combined drug treatment was additive, sub-additive, or even antagonistic.</p> <p>Conclusion</p> <p>GSK-3 inhibition has anticancer effects against pancreatic cancer cells with a range of genetic backgrounds associated with disruption of NF-kappaB, but does not significantly sensitize these cells to the standard chemotherapy agent gemcitabine. This lack of synergy might be context or cell line dependent, but could also be explained on the basis that although NF-kappaB is an important mediator of pancreatic cancer cell survival, it plays a minor role in gemcitabine resistance. Further work is needed to understand the mechanisms of this effect, including the potential for rational combination of GSK3 inhibitors with other targeted agents for the treatment of pancreatic cancer.</p

Non-redundant activity of GSK-3α and GSK-3β in T cell-mediated tumor rejection

Glycogen synthase kinase-3 (GSK-3) is a positive regulator of PD-1 expression in CD8+ T cells and GSK-3 inhibition enhances T cell function and is effective in the control of tumor growth. GSK-3 has two co-expressed isoforms, GSK-3α and GSK-3β. Using conditional gene targeting, we demonstrate that both isoforms contribute to T cell function to different degrees. Gsk3b−/− mice suppressed tumor growth to the same degree as Gsk3a/b−/− mice, whereas Gsk3a−/− mice behaved similarly to wild-type, revealing an important role for GSK-3β in regulating T cell-mediated anti-tumor immunity. The individual GSK-3α and β isoforms have differential effects on PD-1, IFNγ, and granzyme B expression and operate in synergy to control PD-1 expression and the infiltration of tumors with CD4 and CD8 T cells. Our data reveal a complex interplay of the GSK-3 isoforms in the control of tumor immunity and highlight non-redundant activity of GSK-3 isoforms in T cells, with implications for immunotherapy

Increased Glycogen Synthase Kinase-3β mRNA Level in the Hippocampus of Patients with Major Depression: A Study Using the Stanley Neuropathology Consortium Integrative Database

Objective Glycogen synthase kinase-3 beta (GSK-3 beta) has become recognized as a broadly influential enzyme affecting diverse range of biological functions, including gene expression, cellular architecture, and apoptosis. The results of previous studies suggest that GSK-3 beta activity may be increased in the brain of patients with major depressive disorders (MDD). A recent animal study reported increased GSK-3 beta messenger ribonucleic acid (mRNA) level in the hippocampus of those with depression. However, few studies have investigated GSK-3 beta activity in the brain of patients with MDD. Methods In order to test whether patients with MDD have an increase in GSK-3 beta activity in the brain compared to normal controls, we explored GSK-3 beta expression level in all brain regions by using the Stanley Neuropathology Consortium Integrative Database (SNCID), which is a web-based method of integrating the Stanley Medical Research Institute data sets. Results The level of GSK-3 beta mRNA expression in the hippocampus was significantly increased in the MDD group (n=8) compared with the control group (n=12, p<0.05). Spearman's test also reveals that GSK-3 beta mRNA expression levels were significantly correlated with nitric oxide synthase 1 (NOS1)(rho=0.70, p<0.0001) and stathmin-like 3 (STMN3)(rho=0.70, p<0.0001) in the hippocampus. Conclusion Our results correspond with the results of previous animal studies that reported increased GSK-3 beta activity in the hippocampus of those with depression. Our findings also suggest that oxidative stress-induced neuronal cell death and abnormal synaptic plasticity in the hippocampus may play important roles in the pathophysiology of major depression. Psychiatry Investig 2010;7:202-207This research was supported by a grant of the Korea Health 21 R & D project, Ministry of Health and Welfare, Republic of Korea (A050047). We would like to thank the SMRI collaborators who allow SNCID to be freely available to all users, and also all investigators generating original data in the SMRIDB, in particular Dr. William Deakin and Carla Toro