Glycogen Synthase Kinase-3beta Activity Research Articles

Microdose lithium improves behavioral deficits and modulates molecular mechanisms of memory formation in female SAMP-8, a mouse model of accelerated aging.

Alzheimer's disease (AD) is the most common neuronal disorder that leads to the development of dementia. Until nowadays, some therapies may alleviate the symptoms, but there is no pharmacological treatment. Microdosing lithium has been used to modify the pathological characteristics of the disease, with effects in both experimental and clinical conditions. The present work aimed to analyze the effects of this treatment on spatial memory, anxiety, and molecular mechanisms related to long-term memory formation during the aging process of a mouse model of accelerated aging (SAMP-8). Female SAMP-8 showed learning and memory impairments together with disruption of memory mechanisms, neuronal loss, and increased density of senile plaques compared to their natural control strain, the senescence-accelerated mouse resistant (SAMR-1). Chronic treatment with lithium promoted memory maintenance, reduction in anxiety, and maintenance of proteins related to memory formation and neuronal density. The density of senile plaques was also reduced. An increase in the density of gamma-aminobutyric acid A (GABAA) and α7 nicotinic cholinergic receptors was also observed and related to neuroprotection and anxiety reduction. In addition, this microdose of lithium inhibited the activation of glycogen synthase kinase-3beta (GSK-3β), the classical mechanism of lithium cell effects, which could contribute to the preservation of the memory mechanism and reduction in senile plaque formation. This work shows that lithium effects in neuroprotection along the aging process are not related to a unique cellular mechanism but produce multiple effects that slowly protect the brain along the aging process.

GSK-3β inhibition alleviates arthritis pain via reducing spinal mitochondrial reactive oxygen species level and inflammation.

Pain is the main symptom of osteoarthritis, which severely reduces the patients' quality of life. Stimulated neuroinflammation and elevated mitochondrial oxidative stress are associated arthritis pain. In the present study, arthritis model was established by intra-articular injection of complete Freund's adjuvant (CFA) on mice. Knee swelling, pain hypersensitivity and motor disability were observed in CFA-induced mice. In spinal cord, neuroinflammation was triggered and presented as severe infiltration of inflammatory cells and up-regulated expressions of glial fibrillary acidic protein (GFAP), nuclear factor-kappaB (NF-κB), PYD domains-containing protein 3 (NLRP3), cysteinyl aspartate specific proteinase (caspase-1) and interleukin-1 beta (IL-1β). Mitochondrial function was disrupted and characterized as elevated expressions of B-cell lymphoma 2 (Bcl-2)-associated X protein (Bax), dihydroorotate dehydrogenase (DHODH) and cytochrome C (Cyto C), and reduced expressions of Bcl-2 and Mn-superoxide dismutase (Mn-SOD) activity. Meanwhile, as a potential target for pain management, glycogen synthase kinase-3 beta (GSK-3β) activity was up-regulated in CFA induced mice. To explore potential therapeutic options for arthritis pain, GSK-3β inhibitor TDZD-8 was intraperitoneally injected for three days on CFA mice. Animal behavioral tests found that TDZD-8 treatment elevated mechanical pain sensitivity, suppressed spontaneous pain and recovered motor coordination. Morphological and protein expression analysis indicated that TDZD-8 treatment decreased spinal inflammation score and inflammatory related protein levels, recovered mitochondrial related protein levels, and increased Mn-SOD activity. In summary, TDZD-8 treatment inhibits GSK-3β activity, reduces mitochondrial mediated oxidative stress, suppresses spinal inflammasome response, and alleviates arthritis pain.

Different Effects and Mechanisms of Selenium Compounds in Improving Pathology in Alzheimer’s Disease

Owing to the strong antioxidant capacity of selenium (Se) in vivo, a variety of Se compounds have been shown to have great potential for improving the main pathologies and cognitive impairment in Alzheimer's disease (AD) models. However, the differences in the anti-AD effects and mechanisms of different Se compounds are still unclear. Theoretically, the absorption and metabolism of different forms of Se in the body vary, which directly determines the diversification of downstream regulatory pathways. In this study, low doses of Se-methylselenocysteine (SMC), selenomethionine (SeM), or sodium selenate (SeNa) were administered to triple transgenic AD (3× Tg-AD) mice for short time periods. AD pathology, activities of selenoenzymes, and metabolic profiles in the brain were studied to explore the similarities and differences in the anti-AD effects and mechanisms of the three Se compounds. We found that all of these Se compounds significantly increased Se levels and antioxidant capacity, regulated amino acid metabolism, and ameliorated synaptic deficits, thus improving the cognitive capacity of AD mice. Importantly, SMC preferentially increased the expression and activity of thioredoxin reductase and reduced tau phosphorylation by inhibiting glycogen synthase kinase-3 beta (GSK-3β) activity. Glutathione peroxidase 1 (GPx1), the selenoenzyme most affected by SeM, decreased amyloid beta production and improved mitochondrial function. SeNa improved methionine sulfoxide reductase B1 (MsrB1) expression, reflected in AD pathology as promoting the expression of synaptic proteins and restoring synaptic deficits. Herein, we reveal the differences and mechanisms by which different Se compounds improve multiple pathologies of AD and provide novel insights into the targeted administration of Se-containing drugs in the treatment of AD.

Translocating proteins compartment-specifically alter the fate of epithelial-mesenchymal transition in a compartmentalized Boolean network model

Regulation of translocating proteins is crucial in defining cellular behaviour. Epithelial-mesenchymal transition (EMT) is important in cellular processes, such as cancer progression. Several orchestrators of EMT, such as key transcription factors, are known to translocate. We show that translocating proteins become enriched in EMT-signalling. To simulate the compartment-specific functions of translocating proteins we created a compartmentalized Boolean network model. This model successfully reproduced known biological traits of EMT and as a novel feature it also captured organelle-specific functions of proteins. Our results predicted that glycogen synthase kinase-3 beta (GSK3B) compartment-specifically alters the fate of EMT, amongst others the activation of nuclear GSK3B halts transforming growth factor beta-1 (TGFB) induced EMT. Moreover, our results recapitulated that the nuclear activation of glioma associated oncogene transcription factors (GLI) is needed to achieve a complete EMT. Compartmentalized network models will be useful to uncover novel control mechanisms of biological processes. Our algorithmic procedures can be automatically rerun on the https://translocaboole.linkgroup.hu website, which provides a framework for similar future studies.

Regorafenib Regulates AD Pathology, Neuroinflammation, and Dendritic Spinogenesis in Cells and a Mouse Model of AD.

The oral multi-target kinase inhibitor regorafenib, which targets the oncogenic receptor tyrosine kinase (RTK), is an effective therapeutic for patients with advanced gastrointestinal stromal tumors or metastatic colorectal cancer. However, whether regorafenib treatment has beneficial effects on neuroinflammation and Alzheimer’s disease (AD) pathology has not been carefully addressed. Here, we report the regulatory function of regorafenib in neuroinflammatory responses and AD-related pathology in vitro and in vivo. Regorafenib affected AKT signaling to attenuate lipopolysaccharide (LPS)-mediated expression of proinflammatory cytokines in BV2 microglial cells and primary cultured microglia and astrocytes. In addition, regorafenib suppressed LPS-induced neuroinflammatory responses in LPS-injected wild-type mice. In 5x FAD mice (a mouse model of AD), regorafenib ameliorated AD pathology, as evidenced by increased dendritic spine density and decreased Aβ plaque levels, by modulating APP processing and APP processing-associated proteins. Furthermore, regorafenib-injected 5x FAD mice displayed significantly reduced tau phosphorylation at T212 and S214 (AT100) due to the downregulation of glycogen synthase kinase-3 beta (GSK3β) activity. Taken together, our results indicate that regorafenib has beneficial effects on neuroinflammation, AD pathology, and dendritic spine formation in vitro and in vivo.

GSK3β and Tau Protein in Alzheimer's Disease and Epilepsy.

Alzheimer’s disease (AD) is the most common form of dementia present in older adults; its etiology involves genetic and environmental factors. In recent years, epidemiological studies have shown a correlation between AD and chronic epilepsy since a considerable number of patients with AD may present seizures later on. Although the pathophysiology of seizures in AD is not completely understood, it could represent the result of several molecular mechanisms linked to amyloid beta-peptide (Aβ) accumulation and the hyperphosphorylation of tau protein, which may induce an imbalance in the release and recapture of excitatory and inhibitory neurotransmitters, structural alterations of the neuronal cytoskeleton, synaptic loss, and neuroinflammation. These changes could favor the recurrent development of hypersynchronous discharges and epileptogenesis, which, in a chronic state, favor the neurodegenerative process and influence the cognitive decline observed in AD. Supporting this correlation, histopathological studies in the brain tissue of temporal lobe epilepsy (TLE) patients have revealed the presence of Aβ deposits and the accumulation of tau protein in the neurofibrillary tangles (NFTs), accompanied by an increase of glycogen synthase kinase-3 beta (GSK3β) activity that may lead to an imminent alteration in posttranslational modifications of some microtubule-associated proteins (MAPs), mainly tau. The present review is focused on understanding the pathological aspects of GSK3β and tau in the development of TLE and AD.

Effect of phosphoglycogen synthase kinase-3 β induced by low dose of tunicamycin on mitochondrial permeability transition pore in ischemia-reperfusion myocardium

Objective To evaluate the effect of glycogen synthase kinase-3beta (GSK-3β) activity on mitochondrial permeability transition pore (mPTP) in the protective effects of preconditioning with endoplasmic reticulum stress (ERS) induced by low dose of tunicamycin (TM) against myocardial ischemia reperfusion (I/R) injury. Methods Forty adult male sprague-dawley rats were randomly divided into four groups: Sham, TM (Rats were treatmented by TM 0.6 mg/kg at 30 min before model established), I/R, and TM+ LY [Rats were treatmented by TM 0.6 mg/kg and protein kinase B (Akt) inhibitor LY294002 10 mg/kg]. The model of I/R were established by occlusion of left anterior descending coronary artery for 0.5 h followed by reperfusion for 2 h. Hemodynamic parameters were recorded after coronary artery perforation, 1 h and 2 h after reperfusion. After 2 hours of reperfusion, the expression of phosphorylated Akt (p-Akt), p-GSK-3β and the openness of mPTP were detected in the hearts. Results Compared with base line, left ventricular systolic pressure (LVSP), maximum rate of increase or decrease of left ventricular pressure (±dp/dt max) decreased, and left ventricular end-diastolic pressure (LVEDP)increased in I/R group, TM group and TM+ LY group, while the change was more significantly in I/R group. The expression of p-Akt and p-GSK-3 beta protein in TM and TM+ LY group was higher than that in I/R group, and the expression of p-Akt and p-GSK-3β protein in TM+ LY group was lower than that in TM group. At 2 hours after reperfusion, degree of variation of absorbance value (ΔA) of the mPTP of Sham, I/R, TM and TM+ LY group were (78.9±9.2)%, (36.1±4.8)%, (61.4±8.3)% and (46.3 ±5.2)%. The ΔA of TM group was larger than that of I/R group (t=3.280, P<0.01). Compared with TM group, the ΔA of TM+ LY group was relatively small (t=2.720, P<0.01). Conclusion The protective effect of preconditioning with ERS induced by low dose TM is related to the inhibition of mPTP opening by regulating GSK-3β activity. Key words: Myocardial ischemia reperfusion injury; Endoplasmic reticulum stress; Preconditioning; Glycogen synthase kinase-3β; Mitochondrial permeability transition pore

The Regulating Role of SB216763 in Pseudomonas aeruginosa Keratitis.

Glycogen synthase kinase 3 beta (GSK3b) is a multifunctional molecule, which plays a critical role in the regulation of various signaling pathways including cell proliferation, growth and development, and inflammation. However, whether GSK3b is involved in the pathological process of Pseudomonas aeruginosa keratitis remains unknown. First, western blots were performed to measure the phosphorylated level of GSK3β at Ser9 (inactive form) in an animal model of Pseudomonas aeruginosa keratitis. Second, the keratitis model received the GSK3β inhibitor SB216763, and the inflammation of cornea was evaluated by clinical scores and slit photos. The expressions of inflammatory cytokines were assessed by real-time PCR, and the corneal bacterial burden was determined by plate count. The phosphorylated level of GSK3β at Ser9 in the cornea markedly decreased after Pseudomonas aeruginosa infection. The inhibition of GSK3β by SB216763 significantly ameliorated the progress of corneal disease and alleviated corneal opacity. SB216763 suppressed the expression of inflammatory cytokines IL-6 and IL-1b, but exhibited no effects on TNF-a and IL-10 expression. SB216763 dramatically decreased cornea bacterial burden at 5 days after infection with Pseudomonas aeruginosa. The activity of GSK3b was enhanced in Pseudomonas aeruginosa keratitis. The inhibition of GSK3β by SB216763 promoted host resistance against Pseudomonas aeruginosa keratitis, via down regulating inflammatory cytokines and bacterial burden.

Nerve growth factor prevents arsenic-induced toxicity in PC12 cells through the AKT/GSK-3β/NFAT pathway.

The potential risk of arsenic-related neurodegeneration has been a growing concern. Arsenic exposure has been reported to disrupt neurite growth and neuron body integrity in vitro; however, its underlying mechanism remains unclear. Previously, we showed that arsenic sulfide (AS) exerted cytotoxicity in gastric and colon cancer cells through regulating nuclear factor of the activated T cells (NFAT) pathway. The NFAT pathway regulates axon path finding and neural development. Using neural crest cell line PC12 cells as a model, here we show that AS caused mitochondrial membrane potential collapse, reactive oxygen species production, and cytochrome c release, leading to mitochondria-mediated apoptosis via the AKT/GSK-3β/NFAT pathway. Increased glycogen synthase kinase-3 beta (GSK-3β) activation leads to the inactivation of NFAT and its antiapoptotic effects. Through inhibiting GSK-3β activity, both nerve growth factor (NGF) and Tideglusib, a GSK-3β inhibitor partially rescued the PC12 cells from the AS-induced cytotoxicity and restored the expression of NFATc3. In addition, overexpression of NFATc3 stimulated neurite outgrowth and potentiated the effect of NGF on promoting the neurite outgrowth. Collectively, our results show that NFATc3 serves as the downstream target of NGF and plays a key role in preventing AS-induced neurotoxicity through regulating the AKT/GSK-3β/NFAT pathway in PC12 cells.

Abstract 4618: TGF-B upregulates GSK3B and mediates NK dysfunction in cancer

Abstract Transforming growth factor-B (TGF-B) is an important regulator of tumorigenesis. Initially, it was discovered to have anti-cancer activities by promoting apoptosis and preventing hyperproliferation of cells. However, abundant presence of TGF-B in the tumor microenvironment is known to hinder cancer eradication by suppressing anti-tumor activities of immune cells. The molecular switches that regulate the role of TGF-B are not fully understood. One main target of TGF-B-mediated immune cell dysfunction are natural killer (NK) cells. NK cells are innate immune cells that lyse virally-infected and cancer cells. Recently, we have discovered that glycogen synthase kinase 3-beta (GSK3B) upregulation in NK cells leads to NK dysfunction. Another study has indicated that GSK3 phosphorylates SMAD3 to inhibit TGF-B-mediated growth dysfunction in AML12 cells. In addition, GSK3B inhibition has been linked to increased sensitivity of cancer cells to NK killing but the role of GSK3B in NK function is understudied. This study examines how the role that GSK3B has on TGF-B-mediated NK cell dysfunction. Methods: NK cells were isolated from the peripheral blood of healthy donors and expanded in vitro for 14 days. NK cells were either treated with or without 5ng/mL TGF-B for 72 hours, and GSK3B expression, and the expression of NK activating receptors was measured via flow cytometry. In addition, GSK3B expression is abrogated using a siRNA in NK cells. The NK cells were treated with or without TGF-B, and NK cell cytotoxicity is measured with a fluorometric cytotoxicity assay. Results: TGF-B pretreatment of NK cells led to 2-fold GSK3B expression in NK cells. TGF-B pretreatment led to decreased expression of NK activating receptors NKp46 and NKG2D. Also, TGF-B led to decreased expression of lymphocyte function-associated antigen-1 (LFA-1), an adhesion molecule necessary for NK cells to adhere to target cells prior to lysis of the target cells. TGF-B pretreatment of NK cells led to 33.3% reduced killing of both leukemic and solid tumor targets. Pharmacologic and genetic abrogation of GSK3 minimized TGF-B-mediated NK dysfunction as measured by calcein-AM based cytotoxicity assays. Importantly, inhibiting GSK3B activity with TGF-B treatment rescued NK cell cytotoxic function. Discussion: Our data suggests that TGF-B leads to upregulated GSK3B expression, and downregulation of NK activating receptors. In addition, TGF-B reduced NK killing of cancer cells. We believe that TGF-B-mediated NK dysfunction is facilitated by GSK3B upregulation. Further studies are underway to determine how TGF-B affects the expression of other NK receptors, and to elucidate the mechanism by which TGF-B directs GSK3B upregulation. Citation Format: Evelyn Ojo, Folashade Otegbeye, Stephen Moreton, David Wald. TGF-B upregulates GSK3B and mediates NK dysfunction in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4618. doi:10.1158/1538-7445.AM2017-4618

P7C3 inhibits GSK3\u03b2 activation to protect dopaminergic neurons against neurotoxin-induced cell death in vitro and in vivo

Parkinson’s disease (PD) is the second most prevalent neurodegenerative disease. Although its pathogenesis remains unclear, mitochondrial dysfunction plays a vital role in the pathology of PD. P7C3, an aminopropyl carbazole, possesses a significant neuroprotective ability in several neurodegenerative disorders, including PD. Here, we showed that P7C3 stabilized mitochondrial membrane potential, reduced reactive oxygen species production, and inhibited cytochrome c release in MES23.5 cells (a dopaminergic (DA) cell line) exposed to 1-methyl-4-phenylpyridinium (MPP+). In MES23.5 cells, P7C3 inhibited glycogen synthase kinase-3 beta (GSK3β) activation induced by MPP+. P7C3 also inhibited p53 activity and repressed Bax upregulation to protect cells from MPP+ toxicity. In addition, the activation of p53 was significantly attenuated with the inhibition of GSK3β activity by P7C3. Furthermore, P7C3 blocked GSK3β and p53 activation in the midbrain, and prevented DA neuronal loss in the substantia nigra in 1-methyl-4-phenyl-1,2,3,4-tetrahydropyridine mice. Thus, our study demonstrates that P7C3 protects DA neurons from neurotoxin-induced cell death by repressing the GSK3β-p53-Bax pathway both in vitro and in vivo, thus providing a theoretical basis for P7C3 in the potential clinical treatment of PD.

Erratum: Dihydromyricetin protects neurons in an MPTP-induced model of Parkinson's disease by suppressing glycogen synthase kinase-3 beta activity.

Erratum: Dihydromyricetin protects neurons in an MPTP-induced model of Parkinson's disease by suppressing glycogen synthase kinase-3 beta activity

Salidroside protects rat liver against ischemia/reperfusion injury by regulating the GSK-3β/Nrf2-dependent antioxidant response and mitochondrial permeability transition

Salidroside (Sal) is a natural antioxidant that elicits cardioprotective and neuroprotective effects in vivo and in vitro; however, its impact on hepatic ischemia/reperfusion (I/R) injury remains unclear. The purpose of this study was to investigate the hepatoprotective effects of salidroside against segmental (70%) warm hepatic I/R injury in rats. Animals were randomized into Sham, Sham+salidroside pretreatment (Sal), Sham+Sal+carboxyatractyloside (CATR), Sham+CATR, I/R, I/R+Sal, I/R+Sal+CATR and I/R+CATR groups. The hepatic artery, left portal vein and median liver lobes were occluded for 60min and then unclamped to allow reperfusion. Pretreatment with salidroside (20mg/kg/day for 7 days, intraperitoneally) significantly decreased serum alanine aminotransferase (sALT) and serum aspartate aminotransferase (sAST) levels after 6h and 24h of reperfusion and protected the liver against I/R-induced injury. However, this protective effect could be reversed by CATR, a mitochondrial permeability transition pore (MPTP) opener (5mg/kg 30min before I/R insult, intraperitoneally). Mechanistic studies have revealed that salidroside inhibits glycogen synthase kinase-3 beta (GSK-3β) activity and enhances the NF-E2-related factor (Nrf2)-dependent antioxidant response by activating the Akt signaling pathway, thereby reducing mitochondrial reactive oxygen species generation, increasing MPTP resistance and preventing apoptosis by suppressing cytochrome c release and caspase activation during reperfusion. Therefore, salidroside ameliorates hepatocyte death and apoptosis through activation of the GSK-3β/Nrf2-dependent antioxidant response and subsequent MPTP inhibition. These results provide experimental evidence supporting the clinical use of salidroside for hepatoprotection in surgical settings.

Early neurotrophic pharmacotherapy rescues developmental delay and Alzheimer\u2019s-like memory deficits in the Ts65Dn mouse model of Down syndrome

Down syndrome (DS), caused by trisomy 21, is the most common genetic cause of intellectual disability and is associated with a greatly increased risk of early-onset Alzheimer’s disease (AD). The Ts65Dn mouse model of DS exhibits several key features of the disease including developmental delay and AD-like cognitive impairment. Accumulating evidence suggests that impairments in early brain development caused by trisomy 21 contribute significantly to memory deficits in adult life in DS. Prenatal genetic testing to diagnose DS in utero, provides the novel opportunity to initiate early pharmacological treatment to target this critical period of brain development. Here, we report that prenatal to early postnatal treatment with a ciliary neurotrophic factor (CNTF) small-molecule peptide mimetic, Peptide 021 (P021), rescued developmental delay in pups and AD-like hippocampus-dependent memory impairments in adult life in Ts65Dn mice. Furthermore, this treatment prevented pre-synaptic protein deficit, decreased glycogen synthase kinase-3beta (GSK3β) activity, and increased levels of synaptic plasticity markers including brain derived neurotrophic factor (BNDF) and phosphorylated CREB, both in young (3-week-old) and adult (~ 7-month-old) Ts65Dn mice. These findings provide novel evidence that providing neurotrophic support during early brain development can prevent developmental delay and AD-like memory impairments in a DS mouse model.

Role and regulation of Glycogen synthase kinase-3 beta in bovine spermatozoa.

The serine/threonine kinase Glycogen synthase kinase 3 (GSK-3) is a master switch that regulates a multitude of cellular pathways, including the acrosome reaction in sperm. In epididymal sperm cells, for example, GSK-3 activity correlates with inhibition of motility-yet no direct pathways connecting GSK-3 activation with loss of motility have been described. Indeed, the details of how GSK-3 is regulated during sperm capacitation and the acrosome reaction remains obscure. To this end, we addressed the involvement of the GSK-3 beta isoform in several known pathways that contribute to motility and the acrosome reaction. We established that Protein kinase A (PKA) is the main regulator of GSK-3β in sperm, as pre-treatment of cells with a GSK-3 inhibitor prior to addition of H89, an inhibitor of PKA, attenuated the motility loss induced by blocking PKA activity. Both induced and spontaneous acrosome reactions also occurred less frequently in sperm treated with GSK-3 inhibitors. Finally, we observed a slow decline in phosphorylation of GSK-3β on Ser 9, which represents an inhibited state, during sperm capacitation; this phenotype is reversed during the induced acrosome reaction, in parallel to activation of Protein phosphatase 1. These results suggest that maintenance of sperm motility and acrosome reaction timing are mediated by PKA through the regulation of GSK-3 beta activity. Mol. Reprod. Dev. 84: 8-18, 2017. © 2016 Wiley Periodicals, Inc.

Effect of inhibition of GSK-3β activity on sevoflurane postconditioning-induced cardioprotection in diabetic rats

Objective To investigate the effect of inhibition of glycogen synthase kinase-3 beta(GSK-3β)activity on sevoflurane postconditioning-induced cardioprotection in diabetic rats. Methods Healthy adult male Sprague-Dawley rats, weighing 250-300 g, in which diabetes mellitus was induced by intraperitoneal 1% streptozotocin 60 mg/kg combined with high-fat and high-sucrose diet and confirmed by blood glucose level >16.7 mmol/L.Forty rats with diabetes mellitus were divided into 5 groups(n=8 each)using a random number table: sham operation group(S group), ischemia-reperfusion(I/R)group, sevoflurane postconditioning group(SP group), GSK-3β inhibitor SB216763 group(SB group), and sevoflurane postconditioning plus SB216763 group(SS group). Myocardial ischemia was induced by 30 min occlusion of the left anterior descending branch of the coronary artery followed by 120 min reperfusion.The rats inhaled sevoflurane with the end-tidal concentration of 2.5% for 5 min starting from 1 min before reperfusion in group SP.SB216763 0.2 mg/kg was injected via the caudal vein at 1 min before reperfusion in group SB.In group SS, the rats inhaled sevoflurane with the end-tidal concentration of 2.5% for 5 min starting from 1 min before reperfusion, and SB216763 0.2 mg/kg was injected via the caudal vein at 1 min before reperfusion.At 120 min of reperfusion, blood samples were collected from the carotid artery for determination of serum creatine kinase-MB(CK-MB)activity and cardiac troponin I(cTnI) concentrations.Myocardial specimens were collected at 120 min of reperfusion for microscopic examination of the pathological changes and for determination of myocardial infarct size(by 2, 3, 5-triphenyltetrazolium chloride staining)and phosphorylated GSK-3β(p-GSK-3β)expression(by Western blot). Results Compared with group S, the myocardial infarct size and serum CK-MB activity and cTnI concentration were significantly increased, and the expression of p-GSK-3β was significantly down-regulated in I/R, SP, SB and SS groups(P 0.05). Compared with group SB, the myocardial infarct size and serum CK-MB activity and cTnI concentration were significantly decreased, and the expression of p-GSK-3β was significantly up-regulated in group SS(P<0.05). The pathological changes of myocardium were significantly attenuated in SB and SS groups as compared with group I/R and group SP. Conclusion Inhibition of GSK-3β activity can improve sevoflurane postconditioning-induced cardioprotection in diabetic rats. Key words: Anesthetics, inhalation; Diabetes mellitus; Glycogen synthase kinase 3; Myocardial reperfusion injury

Dihydromyricetin protects neurons in an MPTP-induced model of Parkinson's disease by suppressing glycogen synthase kinase-3 beta activity.

It is general believed that mitochondrial dysfunction and oxidative stress play critical roles in the pathology of Parkinson's disease (PD). Dihydromyricetin (DHM), a natural flavonoid extracted from Ampelopsis grossedentata, has recently been found to elicit potent anti-oxidative effects. In the present study, we explored the role of DHM in protecting dopaminergic neurons. Male C57BL/6 mice were intraperitoneally injected with 1-methyl4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) for 7 d to induce PD. Additionally, mice were treated with either 5 or 10 mg/kg DHM for a total of 13 d (3 d before the start of MPTP, during MPTP administration (7 d) and 3 d after the end of MPTP). For the saline or DHM alone treatment groups, mice were injected with saline or DHM for 13 d. On d 14, behavioral tests (locomotor activity, the rotarod test and the pole test) were administered. After the behavioral tests, the mice were sacrificed, and brain tissue was collected for immunofluorescence staining and Western blotting. In addition, MES23.5 cells were treated with MPP+ and DHM, and evaluated using cell viability assays, reactive oxygen species (ROS) measurements, apoptosis analysis and Western blotting. DHM significantly attenuated MPTP-induced mouse behavioral impairments and dopaminergic neuron loss. In the MES23.5 cells, DHM attenuated MPP+-induced cell injury and ROS production in a dose-dependent manner. In addition, DHM increased glycogen synthase kinase-3 beta phosphorylation in a dose- and time-dependent manner, which may be associated with DHM-induced dopaminergic neuronal protection. The present study demonstrated that DHM is a potent neuroprotective agent for DA neurons by modulating the Akt/GSK-3β pathway, which suggests that DHM may be a promising therapeutic candidate for PD.

The Neuroprotective Effects of Justicidin A on Amyloid Beta25-35-Induced Neuronal Cell Death Through Inhibition of Tau Hyperphosphorylation and Induction of Autophagy in SH-SY5Y Cells.

Justicidin A is a structurally defined arylnaphthalide lignan, which has been shown anti-cancer activity; however, the neuroprotective effect of justicidin A is still untested. In this study, we investigated the action of justicidin A on amyloid beta (Aβ)25-35-induced neuronal cell death via inhibition of the hyperphosphorylation of tau and induction of autophagy in SH-SY5Y cells. Pretreatment with justicidin A significantly elevated cell viability in cells treated with Aβ25-35. Western blot data demonstrated that justicidin A inhibited the Aβ25-35-induced up-regulation the levels of hyperphosphorylation of tau in SH-SY5Y cells. In addition, treatment with justicidin A significantly induced autophagy as measured by the increasing LC3 II/I ratio, an important autophagy marker. These studies showed that justicidin A inhibited activity of glycogen synthase kinase-3beta (GSK-3β), which is an important kinase in up-stream signaling pathways; inhibited hyperphosphorylation of tau in AD; and enhanced activity of AMP-activated protein kinase (AMPK), which is the key molecule for both hyperphosphorylation of tau and induction of autophagy. These data provide the first evidence that justicidin A protects SH-SY5Y cells from Aβ25-35-induced neuronal cell death through inhibition of hyperphosphorylation of tau and induction of autophagy via regulation the activity of GSK-3β and AMPK, and they also provide some insights into the relationship between tau protein hyperphosphorylation and autophagy. Thus, we conclude that justicidin A may have a potential role for neuroprotection and, therefore, may be used as a therapeutic agent for AD.

Memantine Attenuates Alzheimer's Disease-Like Pathology and Cognitive Impairment.

Deficiency of protein phosphatase-2A is a key event in Alzheimer’s disease. An endogenous inhibitor of protein phosphatase-2A, inhibitor-1, I1 PP2A, which inhibits the phosphatase activity by interacting with its catalytic subunit protein phosphatase-2Ac, is known to be upregulated in Alzheimer’s disease brain. In the present study, we overexpressed I1 PP2A by intracerebroventricular injection with adeno-associated virus vector-1-I1 PP2A in Wistar rats. The I1 PP2A rats showed a decrease in brain protein phosphatase-2A activity, abnormal hyperphosphorylation of tau, neurodegeneration, an increase in the level of activated glycogen synthase kinase-3beta, enhanced expression of intraneuronal amyloid-beta and spatial reference memory deficit; littermates treated identically but with vector only, i.e., adeno-associated virus vector-1-enhanced GFP, served as a control. Treatment with memantine, a noncompetitive NMDA receptor antagonist which is an approved drug for treatment of Alzheimer’s disease, rescued protein phosphatase-2A activity by decreasing its demethylation at Leu309 selectively and attenuated Alzheimer’s disease-like pathology and cognitive impairment in adeno-associated virus vector-1-I1 PP2A rats. These findings provide new clues into the possible mechanism of the beneficial therapeutic effect of memantine in Alzheimer’s disease patients.

Hesperidin induces apoptosis and triggers autophagic markers through inhibition of Aurora-A mediated phosphoinositide-3-kinase/Akt/mammalian target of rapamycin and glycogen synthase kinase-3 beta signalling cascades in experimental colon carcinogenesis

Abnormalities in the homeostasis mechanisms involved in cell survival and apoptosis are contributing factors for colon carcinogenesis. Interventions of these mechanisms by pharmacologically safer agents gain predominance in colon cancer prevention. We previously reported the chemopreventive efficacy of hesperidin against colon carcinogenesis. In the present study, we aimed at investigating the potential of hesperidin over the abrogated Aurora-A coupled pro-survival phosphoinositide-3-kinase (PI3K)/Akt signalling cascades. Further, the role of hesperidin over apoptosis and mammalian target of rapamycin (mTOR) mediated autophagic responses were studied. Azoxymethane (AOM) induced mouse model of colon carcinogenesis was involved in this study. Hesperidin treatment was provided either in initiation/post-initiation mode respectively. Hesperidin significantly altered AOM mediated anti-apoptotic scenario by modulating Bax/Bcl-2 ratio together with enhanced cytochrome-c release and caspase-3, 9 activations. In addition, hesperidin enhanced p53–p21 axis with concomitant decrease in cell cycle regulator. Hesperidin treatment caused significant up-regulation of tumour suppressor phosphatase and tensin homologue (PTEN) with a reduction in the expression of AOM mediated p-PI3K and p-Akt. Additionally, hesperidin administration exhibited inhibition against p-mTOR expression which in turn led to stimulation of autophagic markers Beclin-1 and LC3-II. Aurora-A an upstream regulator of PI3K/Akt pathway was significantly inhibited by hesperidin. Furthermore, hesperidin administration restored glycogen synthase kinase-3 beta (GSK-3β) activity which in turn prevented the accumulation of oncoproteins β-catenin, c-jun and c-myc. Taken together, hesperidin supplementation initiated apoptosis via targeted inhibition of constitutively activated Aurora-A mediated PI3K/Akt/GSK-3β and mTOR pathways coupled with autophagic stimulation against AOM induced colon carcinogenesis.