Effect of aluminum exposure on abnormal phosphorylationof tau protein in PC12 cells of rats / 预防医学

Objective @#To investigate the effect of aluminum exposure on expression of miR-497-5p, wingless murine breast cancer virus integration site family member 3a (Wnt3a), β-catenin protein, glycogen synthase kinase-3β (GSK-3β) protein and tau protein in rat adrenal pheochromocytoma PC12 cells, so as to provide insight into unraveling the mechanisms underlying aluminum exposure-induced abnormal phosphorylation of tau protein.@* Methods@# PC12 cells were exposed to Al(mal)3 at concentrations of 0, 100, 200, 400 μmol/L for 24 h. The viability of PC12 cells was measured using cell counting kit-8 (CCK-8) assay. The relative expression of miR-497-5p and Wnt3a was detected using a real-time fluorescent quantitative PCR (RT-qPCR) assay, and the expression of Wnt3a, β-catenin, GSK-3β, P-GSK-3β (Ser9), tau and p-tau (Ser396) proteins were determined using Western blotting. @*Results @#The viability of PC12 cells appeared a tendency towards a decline with the increase of aluminum dose (Ftrend=323.473, P=0.001). RT-qPCR assay detected that the relative miR-497-5p expression appeared a tendency towards a rise with the increase of aluminum dose (Ftrend=14.888, P=0.031), and the relative Wnt3a expression appeared a tendency towards a decline with the increase of aluminum dose (Ftrend=165.934, P<0.001). The miR-497-5p expression negatively correlated with the relative Wnt3a expression (r=-0.693, P=0.012). The expression of Wnt3a (Ftrend=357.656, P=0.001), β-catenin (Ftrend=208.750, P=0.001) and p-GSK-3β (Ser9) proteins (Ftrend=512.583, P<0.001) appeared a tendency towards a decline with the increase of aluminum dose, and the expression of GSK-3β (Ftrend=39.965, P<0.001), tau (Ftrend=277.929, P=0.006) and p-tau (Ser396) proteins (Ftrend=96.247, P=0.002) appeared a tendency towards a rise with the increase of aluminum dose. @*Conclusion@# Up-regulation of miR-497-5p and GSK-3β expression and down-regulation of Wnt3a and β-catenin expression may be a mechanism underlying aluminum exposure-induced abnormal phosphorylation of tau protein.

Medicinal plants with antimalarial activities mediated via glycogen synthase kinase-3 beta (GSK3β) inhibition

@#Many of the therapeutic effects of plant extracts and bioactive compounds appear related to their immunomodulatory effects and impact on the host immune system. The immune response is desirable to mitigate established infections and, in the case of severe malaria, is a feasible approach to dealing with the overwhelming cytokine response. Glycogen synthase kinase-3 (GSK3), a Ser/Thr kinase that is a central regulator of the cytokine response, is a promising antimalarial drug target. In this review, we discussed our ongoing research projects, which include assessing the antimalarial activities of medicinal plants and their bioactive compounds, immunomodulatory activities mediated by GSK3, and the potential inflammatory pathway involved in malarial infection.

Relationship between phosphorylation of GSK-3β and high glucose-caused abolition of cardioprotection induced by sevoflurane postconditioning / 中华麻醉学杂志

Objective:To evaluate the relationship between phosphorylation of glycogen synthase kinase-3β (GSK-3β) and high glucose-caused abolition of cardioprotection induced by sevoflurane postconditioning.Methods:H9c2 cells were incubated in normal glucose (5.56 mmol/L) DMEM culture medium or high glucose (33 mmol/L) DMEM culture medium.The cells were divided into 8 groups ( n=24 each) using a random number table method: normal control group (group NC), normal glucose-cultured hypoxia/reoxygenation (H/R) group (group NH/R), normal glucose-cultured sevoflurane postconditioning group (group NS), normal glucose-cultured GSK-3β inhibitor SB216763 group (group NSB), high glucose-cultured group (group HC), high glucose-cultured H/R group (group HH/R), high glucose-cultured sevoflurane postconditioning group (group HS) and high glucose-cultured GSK-3β inhibitor SB216763 group (group HSB). The model of cardiomyocyte H/R was established by subjecting cardiomyocytes to 3 h of hypoxia followed by reoxygenation.Immediately after onset of reoxygenation, cardiomyocytes were exposed to 2.4% sevoflurane for 30 min in Ns and HS groups.Before the beginning of reoxygenation, GSK-3β inhibitor SB216763 was added to the culture medium with the final concentration of 10 μmol/L in NSB and HSB groups.At 3 h of reoxygenation, the apoptosis rate was determined by Anexin V-PI flow cytometry, the expression of GSK-3β and phosphorylated GSK-3β (p-GSK-3β) was detected by Western blot, superoxide dismutase (SOD) activity was measured using xanthineoxidase method, and lactic dehydrogenase (LDH) activity and malondialdehyde (MDA) content were determined by colorimetric assay. Results:Compared with group NC, apoptosis rate, LDH activity and MDA content were significantly increased, and SOD activity was decreased in group NH/R and group HC, expression of GSK-3β was up-regulated, and expression of p-GSK-3β was down-regulated in group NH/R, expression of p-GSK-3β was up-regulated in group NS, and expression of p-GSK-3β was down-regulated in group HC ( P<0.05). Compared with group NH/R, apoptosis rate, LDH activity and MDA content were significantly decreased, and SOD activity was increased in group NS and NSB groups, and expression of GSK-3β was down-regulated, and expression of p-GSK-3β was up-regulated in group NS ( P<0.05). Compared with group HC, apoptosis rate, LDH activity and MDA content were significantly increased, SOD activity was decreased, expression of GSK-3β was up-regulated, and expression of p-GSK-3β was down-regulated in group HH/R ( P<0.05). Compared with group HH/R, apoptosis rate, LDH activity and MDA content were significantly decreased, and SOD activity was increased in group HSB ( P<0.05). Conclusion:The mechanism by which high glucose abolishes cardioprotection induced by sevoflurane postconditioning is related to inhibiting phosphorylation of GSK-3β.

The neuroprotection of cerebrolysin after spontaneous intracerebral hemorrhage through regulates necroptosis via Akt/ GSK3B signaling pathway

ABSTRACT Purpose: Spontaneous intracerebral hemorrhage (ICH) is a major cause of death and disability with a huge economic burden worldwide. Cerebrolysin (CBL) has been previously used as a nootropic drug. Necroptosis is a programmed cell death mechanism that plays a vital role in neuronal cell death after ICH. However, the precise role of necroptosis in CBL neuroprotection following ICH has not been confirmed. Methods: In the present study, we aimed to investigate the neuroprotective effects and potential molecular mechanisms of CBL in ICH-induced early brain injury (EBI) by regulating neural necroptosis in the C57BL/6 mice model. Mortality, neurological score, brain water content, and neuronal death were evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, Evans blue extravasation, Western blotting, and quantitative real-time polymerase chain reaction (PCR). Results: The results show that CBL treatment markedly increased the survival rate, neurological score, and neuron survival, and downregulated the protein expression of RIP1 and RIP3, which indicated that CBL-mediated inhibition of necroptosis, and ameliorated neuronal death after ICH. The neuroprotective capacity of CBL is partly dependent on the Akt/GSK3β signaling pathway. Conclusions: CBL improves neurological outcomes in mice and reduces neuronal death by protecting against neural necroptosis.

Glycogen synthase kinase-3 beta inhibitors protectagainst the acute lung injuries resulting from acute necrotizing pancreatitis

Abstract Purpose The research is intended for clarification of the efficacy as well as the underlying mechanism of GSK-3β inhibitors on the advancement of acute lung injuries in acute necrotizing pancreatitis (ANP) in rats. Methods Seventy-two rats were randomly divided into 6 groups: (1)ANP-vehicle; (2)ANP-TDZD-8;(3)ANP-SB216763;(4)Sham-vehicle;(5)Sham-TDZD-8;(6)Sham-SB216763; Blood biochemical test, histopathological examination and immunohistochemical analysis of rats pancreas and lung tissues were performed. The protein expression of GSK-3β, phospho-GSK-3β (Ser9), iNOS, ICAM-1, TNF-α, and IL-10 were detected in lung tissues by Western-blot. Results The outcomes revealed that the intervention of GSK-3β inhibitors alleviated the pathological damage of pancreas and lung (P<0.01), reduced serum amylase, lipase, hydrothorax and lung Wet-to-Dry Ratio, attenuated serum concentrations of IL-1β and IL-6 (P<0.01), inhibited the activation of NF-κB, and abated expression of iNOS, ICAM-1 and TNF-α protein, but up-regulated IL-10 expression in lung of ANP rats (P<0.01). The inflammatory response and various indicators in ANP-TDZD-8 groups were lower than those in ANP-SB216763 groups. Conclusions Inhibition of GSK-3β weakens acute lung injury related to ANP via the inhibitory function of NF-κB signaling pathway. Different kinds of GSK-3β inhibitors have different effects to ANP acute lung injury.

The Role of Histone Deacetylase Inhibitors in Regulation of Akt/GSK-3ß Signaling Pathway in Mice Following Transient Focal Cerebral Ischemia

Abstract Purpose: To investigate whether the neuroprotective effect of TSA on cerebral ischemia reperfusion injury is mediated by the activation of Akt/GSK-3β signaling pathway. Methods: Mice were randomly divided into four groups (n=15): sham group (S); ischemia reperfusion group (IR); ischemia reperfusion and pretreated with TSA group (IR+T); ischemia reperfusion and pretreated with TSA and LY294002 group (IR+T+L). The model of cerebral ischemia reperfusion was established by 1h of MCAO following 24h of reperfusion. TSA (5mg/kg) was intraperitoneally given for 3 days before MCAO, Akt inhibitor, LY294002 (15 nmol/kg) was injected by tail vein 30 min before the MCAO. Results: TSA significantly increased the expression of p-Akt, p-GSK-3β proteins and the levels of SOD, Bcl-2, reduced the infarct volume and the levels of MDA, ROS, TNF-α, IL-1β, Bax, Caspase-3, TUNEL and attenuated neurological deficit in mice with transient MCAO, LY294002 weakened such effect of TSA dramatically. Conclusions: TSA could significantly decrease the neurological deficit and reduce the cerebral infarct volume, oxidative stress, inflammation, as well as apoptosis during cerebral ischemia reperfusion injury, which was achieved by activation of the Akt/GSK-3β signaling pathway.

Baicalin attenuates high fat diet-induced insulin resistance and ectopic fat storage in skeletal muscle, through modulating the protein kinase B/Glycogen synthase kinase 3 beta pathway / 中国天然药物

Insulin resistance is the pathophysiological basis of many diseases. Overcoming early insulin resistance highly significant in prevention diabetes, non-alcoholic fatty liver, and atherosclerosis. The present study aimed at evaluating the therapeutic effects of baicalin on insulin resistance and skeletal muscle ectopic fat storage in high fat diet-induced mice, and exploring the potential molecular mechanisms. Insulin resistance in mice was induced with a high fat diet for 16 weeks. Animals were then treated with three different doses of baicalin (100, 200, and 400 mg·kg(-1)·d(-1)) for 14 weeks. Fasting blood glucose, fasting serum insulin, glucose tolerance test (GTT), insulin tolerance test (ITT), and skeletal muscle lipid deposition were measured. Additionally, the AMP-activated protein kinase/acetyl-CoA carboxylase and protein kinase B/Glycogen synthase kinase 3 beta pathways in skeletal muscle were further evaluated. Baicalin significantly reduced the levels of fasting blood glucose and fasting serum insulin and attenuated high fat diet induced glucose tolerance and insulin tolerance. Moreover, insulin resistance was significantly reversed. Pathological analysis revealed baicalin dose-dependently decreased the degree of the ectopic fat storage in skeletal muscle. The properties of baicalin were mediated, at least in part, by inhibition of the AMPK/ACC pathway, a key regulator of de novo lipogenesis and activation of the Akt/GSK-3β pathway, a key regulator of Glycogen synthesis. These data suggest that baicalin, at dose up to 400 mg·kg(-1)·d(-1), is safe and able to attenuate insulin resistance and skeletal muscle ectopic fat storage, through modulating the skeletal muscle AMPK/ACC pathway and Akt/GSK-3β pathway.

Icariin protects SH-SY5Y cells from formaldehyde-induced injury through suppression of Tau phosphorylation / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the neuroprotective effects of icariin on formaldehyde (FA)-treated human neuroblastoma SH-SY5Y cells and the possible mechanisms involved.</p><p><b>METHODS</b>SH-SY5Y cells were divided into FA treatment group, FA treatment group with icariin, and the control group. Cell viability, apoptosis, and morphological changes were determined by cell counting kit-8 (CCK 8), flow cytometry, and confocal microscopy, respectively. The phosphorylation of Tau protein was examined by western blotting.</p><p><b>RESULTS</b>FA showed a half lethal dose (LD50) of 0.3 mmol/L in SH-SY5Y cells under the experimental conditions. Icariin (1-10 µmol/L) prevented FA-induced cell death in SH-SY5Y cells in a dose-dependent manner, with the optimal effect observed at 5 µmol/L. After FA treatment, the absorbance in FA group was 1.31±0.05, while in the group of icariin (5 µmol/L) was 1.63±0.05. Examination of cell morphology by confocal microscopy demonstrated that 5 µmol/L icariin significantly attenuated FA-induced cell injury (P <0.05). Additionally, Icariin inhibited FA-induced cell apoptosis in SH-SY5Y cells. Results from western blotting showed that icariin suppressed FA-induced phosphorylation at Thr 181 and Ser 396 of Tau protein, while having no effect on the expression of the total Tau protein level. Furthermore, FA activated Tau kinase glycogen synthase kinase 3 beta (GSK-3β) by enhancement of Y216 phosphorylation, but icariin reduced Y216 phosphorylation and increased Ser 9 phosphorylation.</p><p><b>CONCLUSION</b>Icariin protects SH-SY5Y cells from FA-induced injury poßsibly through the inhibition of GSK-3β-mediated Tau phosphorylation.</p>

Glycogen synthase kinase 3β and cyclin D1 expression in cervical carcinogenesis

OBJECTIVE: Glycogen synthase kinase 3β (GSK3β) is a pluripotent protein kinase involved in the development of cancers through regulation of numerous oncogenic molecules. Cyclin D1, an important regulator of G1 to S phase transition in various cells, is one of target proteins that GSK3β regulate. Our objective was to assess the expression of GSK3β and cyclin D1 in cervical neoplasm of different histologic grades and to identify their correlation in cervical carcinogenesis. METHODS: Immunohistochemical analysis of GSK3β and cyclin D1 was performed in a total of 137 patients with 12 normal, 62 cervical intraepithelial neoplasia (CIN) (31 CIN1 and 31 CIN3) and 63 invasive cancers including 56 squamous cell carcinomas and 7 adenocarcinomas. RESULTS: The expression of GSK3β increased in parallel with the lesion grade, while that of cyclin D1 decreased with severity of the lesion (P<0.001). There was a significant inverse correlation between GSK3β and cyclin D1 expression in overall cervical neoplasia (Φ=-0.413, P<0.001). GSK3β expression was higher in squamous cell carcinoma than in adenocarcinoma (P=0.049). CONCLUSION: These results suggest that the expressional increase in GSK3β plays a role in cervical carcinogenesis and has inverse correlation with cyclin D1 expression in this process. In addition, GSK3β expression appears to be associated with the histologic type of cervical cancer, especially squamous cell carcinoma.

Effect of Xiaoyao Powder on Alzheimer's disease in hippocampal CA3 region of rats PP-2A, GSK-3βexpression / 中国免疫学杂志

Objective:To study the effect of Xiaoyao Powder on Alzheimer's disease in hippocampal CA 3βregion of rats PP-2A,GSK-3βexpression.Methods:Rats were randomly divided into four groups , including model group, western medicine group,Chi-nese medicine group were treated with intraperitoneal injection of D-gal and A-β1-42 peptide bilateral hippocampal injection mold-ing, physiological group only with equal volume of sterile saline intraperitoneal and hippocampal injection molding .The model was completed, normal group, model group were perfused with saline , western medicine group and Chinese medicine group were treated with oxiracetam solution and Xiaoyao decoction , four groups of intragastric volume was 0.5 ml/100 g, 1 time a day, continuous 28 d. After intragastric administration of isolated rat brain immediately , packet marking in 4℃4%glutaraldehyde solution of glass container , stored at 4℃.Repair piece, cut from the hippocampus, embedded in paraffin, sliced.Dilution of PP-2A for 1∶200, GSK-3βdilution of 1∶150.Images were analyzed by using Image-pro Plus 5 image analysis system, data were conducted by SPSS11.5 statistical analysis software, the comparison between 4 groups by single factor analysis of variance , between the two two groups was compared by LSD-t test, the test level of α=0.05.Results:The expression of PP-2A positive cell number and the positive area , average optical density, integral optical density index , in Xiaoyao Powder orally intervention compared with the model group increased significantly (P<0.01);GSK-3βabove indices were significantly decreased than that in model group after Xiaoyao Powder after intragastric ad -ministration (P<0.01).Conclusion:Xiaoyao Powder can up regulate the expression of PP-2A and down regulate expression of GSK-3β, may be condensed A-β1-42 peptide induced hyperphosphorylation of Tau has certain inhibition .

Amyloid-beta oligomers regulate the properties of human neural stem cells through GSK-3beta signaling

Alzheimer's disease (AD) is the most common cause of age-related dementia. The neuropathological hallmarks of AD include extracellular deposition of amyloid-beta peptides and neurofibrillary tangles that lead to intracellular hyperphosphorylated tau in the brain. Soluble amyloid-beta oligomers are the primary pathogenic factor leading to cognitive impairment in AD. Neural stem cells (NSCs) are able to self-renew and give rise to multiple neural cell lineages in both developing and adult central nervous systems. To explore the relationship between AD-related pathology and the behaviors of NSCs that enable neuroregeneration, a number of studies have used animal and in vitro models to investigate the role of amyloid-beta on NSCs derived from various brain regions at different developmental stages. However, the Abeta effects on NSCs remain poorly understood because of conflicting results. To investigate the effects of amyloid-beta oligomers on human NSCs, we established amyloid precursor protein Swedish mutant-expressing cells and identified cell-derived amyloid-beta oligomers in the culture media. Human NSCs were isolated from an aborted fetal telencephalon at 13 weeks of gestation and expanded in culture as neurospheres. Human NSCs exposure to cell-derived amyloid-beta oligomers decreased dividing potential resulting from senescence through telomere attrition, impaired neurogenesis and promoted gliogenesis, and attenuated mobility. These amyloid-beta oligomers modulated the proliferation, differentiation and migration patterns of human NSCs via a glycogen synthase kinase-3beta-mediated signaling pathway. These findings contribute to the development of human NSC-based therapy for AD by elucidating the effects of Abeta oligomers on human NSCs.

Suppression of Autophagy and Activation of Glycogen Synthase Kinase 3beta Facilitate the Aggregate Formation of Tau

Neurofibrillary tangle (NFT) is a characteristic hallmark of Alzheimer's disease. GSK3beta has been reported to play a major role in the NFT formation of tau. Dysfunction of autophagy might facilitate the aggregate formation of tau. The present study examined the role of GSK3beta-mediated phosphorylation of tau species on their autophagic degradation. We transfected wild type tau (T4), caspase-3-cleaved tau at Asp421 (T4C3), or pseudophosphorylated tau at Ser396/Ser404 (T4-2EC) in the presence of active or enzyme-inactive GSK3beta. Trehalose and 3-methyladenine (3-MA) were used to enhance or inhibit autophagic activity, respectively. All tau species showed increased accumulation with 3-MA treatment whereas reduced with trehalose, indicating that tau undergoes autophagic degradation. However, T4C3 and T4-2EC showed abundant formation of oligomers than T4. Active GSK3beta in the presence of 3-MA resulted in significantly increased formation of insoluble tau aggregates. These results indicate that GSK3beta-mediated phosphorylation and compromised autophagic activity significantly contribute to tau aggregation.

Clusterin protects H9c2 cardiomyocytes from oxidative stress-induced apoptosis via Akt/GSK-3beta signaling pathway

Clusterin is a secretory glycoprotein, which is highly up-regulated in a variety of normal and injury tissues undergoing apoptosis including infarct region of the myocardium. Here, we report that clusterin protects H9c2 cardiomyocytes from H2O2-induced apoptosis by triggering the activation of Akt and GSK-3beta. Treatment with H2O2 induces apoptosis of H9c2 cells by promoting caspase cleavage and cytochrome c release from mitochondria. However, co-treatment with clusterin reverses the induction of apoptotic signaling by H2O2, thereby recovers cell viability. The protective effect of clusterin on H2O2-induced apoptosis is impaired by PI3K inhibitor LY294002, which effectively suppresses clusterin-induced activation of Akt and GSK-3beta. In addition, the protective effect of clusterin is independednt on its receptor megalin, because inhibition of megalin has no effect on clusturin-mediated Akt/GSK-3beta phosphoylation and H9c2 cell viability. Collectively, these results suggest that clusterin has a role protecting cardiomyocytes from oxidative stress and the Akt/GSK-3beta signaling mediates anti-apoptotic effect of clusterin.

Melatonin Potentiates the Neuroprotective Properties of Resveratrol Against Beta-Amyloid-Induced Neurodegeneration by Modulating AMP-Activated Protein Kinase Pathways

BACKGROUND AND PURPOSE: Recent studies have demonstrated that resveratrol (RSV) reduces the incidence of age-related macular degeneration, Alzheimer's disease (AD), and stroke, while melatonin (MEL) supplementation reduces the progression of the cognitive impairment in AD patients. The purpose of this investigation was to assess whether the co-administration of MEL and RSV exerts synergistic effects on their neuroprotective properties against beta-amyloid (Abeta)-induced neuronal death. METHODS: The neuroprotective effects of co-treatment with MEL and RSV on Abeta1-42 -induced cell death, was measured by MTT reduction assay. Abeta1-42 caused an increase in intracellular levels of reactive oxygen species (ROS), as assessed by H2-DCF-DA dye, and a reduction of total glutathione (GSH) levels and mitochondrial membrane potential, as assessed using monochlorobimane and rhodamine 123 fluorescence, respectively. Western blotting was used to investigate the intracellular signaling mechanism involved in these synergic effects. RESULTS: We treated a murine HT22 hippocampal cell line with MEL or RSV alone or with both simultaneously. MEL and RSV alone significantly attenuated ROS production, mitochondrial membrane-potential disruption and the neurotoxicity induced by Abeta1-42. They also restored the Abeta1-42-induced depletion of GSH, back to within its normal range and prevented the Abeta1-42-induced activation of glycogen synthase kinase 3beta (GSK3beta). However, co-treatment with MEL and RSV did not exert any significant synergistic effects on either the recovery of the Abeta1-42-induced depletion of GSH or on the inhibition of Abeta1-42-induced GSK3beta activation. Abeta1-42 treatment increased AMP-activated protein kinase (AMPK) activity, which is associated with subsequent neuronal death. We demonstrated that MEL and RSV treatment inhibited the phosphorylation of AMPK. CONCLUSIONS: Together, our results suggest that co-administration of MEL and RSV acts as an ef-fective treatment for AD by attenuating Abeta1-42-induced oxidative stress and the AMPK-dependent pathway.

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

OBJECTIVE: Glycogen synthase kinase-3beta (GSK-3beta) 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-3beta activity may be increased in the brain of patients with major depressive disorders (MDD). A recent animal study reported increased GSK-3beta messenger ribonucleic acid (mRNA) level in the hippocampus of those with depression. However, few studies have investigated GSK-3beta activity in the brain of patients with MDD. METHODS: In order to test whether patients with MDD have an increase in GSK-3beta activity in the brain compared to normal controls, we explored GSK-3beta 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-3beta 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-3beta 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-3beta 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.

Interaction between Neuronal Depolarization and MK-801 in SH-SY5Y Cells and the Rat Cortex

OBJECTIVE: The interaction between MK-801, a model of psychosis and KCl-induced depolarization or electroconvulsive shock (ECS), a therapeutic model of electroconvulsive therapy (ECT), was investigated in SH-SY5Y cells and the rat frontal cortex. METHODS: SH-SY5Y cells were pretreated with 1 microM MK-801 for 15 min, followed by cotreatment with 100 mM KCl for 5 min. MK-801 was reintroduced after the KCl was washed out, and the samples were incubated before harvesting. For the experiments in rats, male Sprague-Dawley rats were treated with MK-801 followed by ECS. Immunoblot analyses of glycogen synthase kinase 3beta (GSK3beta) (Ser9), AKT (Ser473) and extracellular legulated kinase (ERK)1/2 in SH-SY5Y cells and the rat frontal cortex were performed. RESULTS: KCl-induced neuronal depolarization resulted in the transient dephosphorylation of AKT (Ser473) and GSK3beta (Ser9), followed by increased phosphorylation of the enzymes in SH-SY5Y cells. Cotreatment with MK-801 and KCl inhibited the initial dephosphorylation of AKT and GSK3beta produced by KCl-induced neuronal depolarization. Similarly, ECS resulted in the transient dephosphorylation of AKT (Ser473) and GSK3beta (Ser9), whereas cotreatment with MK-801 inhibited the initial dephosphorylation of AKT (Ser473) and GSK3beta (Ser9) produced by ECS in the rat frontal cortex. No significant interaction was observed between MK-801 and KCl in the dephosphorylation of ERK1/2. CONCLUSION: These results suggest that an antagonistic interplay between MK-801 and neuronal depolarization by KCl or ECS is involved the regulation of AKT (Ser473) and GSK3beta (Ser9) phosphorylation.

Effect of Lithium Chloride on Proliferation and Differentiation of Neural Stem Cells / 中国康复理论与实践

@#Objective To investigate the effect of lithium chloride(LiCl),an inhibitor of glycogen synthase kinase-3beta(GSK-3beta),on proliferation and differentiation of neural stem cells(NSCs).Methods The NSCs were isolated from cortex of rat fetus and expanded in culturing system.Their morphological changes and attachment process were observed under microscope.The cell cycle dynamics of NSCs was examined with flow cytometry.And the expression of GSK-3β and β-catenin was examined quantitatively with Western blot.Results The culturing NSCs treated with LiCl were usually floated and much dispersed in the media.Many of the neurospheres became small and the time of attachment after serum induction became longer.Using flow cytometry,it was detected that the proportion of G1 phase NSCs declined gradually accompanying the increased concentration of LiCl,while the percentage of S and G2/M phase cells showed an increasing trend.Western blotting results revealed β-catenin expression increased whereas Gsk-3βdecreased gradually under the treatment of LiCl and also showed a dose dependent manner.Conclusion These results suggest that LiCl may promote the proliferation of NSCs and prevent them from differentiating,which may partly involve the activation of wnt/β-catenin signaling pathway.

Expression of DNMT1,?-catenin and P-GSK-3? in colon carcinoma / 西安交通大学学报(医学版)

Objective To examine DNMT1,?-catenin and P-GSK-3? expressions in tissues of colon carcinoma compared with tumor clinicopathological parameters including differentiation and metastasis.Methods SYBR Green real-time PCR and immunoblotting method were used to detect these gene expressions in 62 tissues of colon carcinoma and 21 tissues of normal colon.Results Overexpressions of DNMT1,?-catenin,and P-GSK-3? were found in colon carcinoma tissues.The expression of DNMT1 was found to be higher in poorly differentiated tissues.The results also demonstrated statistical significance in the expression of ?-catenin involving differentiation and metastasis,but no statistical significance in the expression of P-GSK-3?.Conclusion These results show that DNMT1,?-catenin and P-GSK-3? expressions are regulated throughout colon cancer progression.