Search

Cellular responses following cerebral ischemia/reperfusion injury are critical to recovery and survival after ischemic stroke. Understanding of these cellular responses can help the design of therapies to protect brain tissue and promote recovery after stroke. One of these cellular responses may be mediated by the AKT (protein kinase B) signal transduction pathway. This study was aimed to investigate the cerebral ischemia-induced alterations of AKT signaling and the upstream molecular pathways.We modeled cerebral ischemia by middle cerebral artery occlusion in 2-3-month-old male C57BL/6J mice and then analyze the brain samples by using quantitative Western blots and phosphorylation/activation-dependent kinase antibodies. Cerebral ischemia was confirmed by staining of brain slices with 1% 2,3,5-triphenyltetrazolium chloride (TTC) and Nissl, as well as neurological assessments of the mice 24 h after ischemia-reperfusion surgery.We found marked downregulation of AKT within 12 h of cerebral ischemia/reperfusion, which leads to overactivation of glycogen synthase kinase-3β (GSK-3β). Furthermore, we found that the downregulation of AKT was mediated by downregulation of mTORC2 (the complex 2 of the mechanistic target of rapamycin) instead of its common upstream kinases, phosphatidylinositol 3-kinase and phosphoinositide-dependent kinase-1.Our findings provide new insight into the cellular responses to ischemia/reperfusion brain injury and will help develop new treatments targeting the AKT signaling pathway for the treatment of ischemic stroke.

Neurofibrillary tangle (NFT) of hyperphosphorylated tau is a hallmark of Alzheimer's disease (AD) and related tauopathies. Tau lesion starts in the trans-entorhinal cortex, from where it spreads to limbic regions, followed by neocortical areas. The regional distribution of NFTs associates with the progression of AD. Accumulating evidence suggests that proteopathic tau can seed tau aggregation in a prion-like fashion in vitro and in vivo. Inhibition of tau seeding activity could provide a potential therapeutic opportunity to block the propagation of tau pathology in AD and related tauopathies.In the present study, we investigated the role of 77G7, a monoclonal tau antibody to the microtubule-binding repeats, in repressing the seeding activity of proteopathic tau.We found that 77G7 had a higher affinity toward aggregated pathological tau fractions than un-aggregated tau derived from AD brain. 77G7 inhibited the internalization of tau aggregates by cells, blocked AD O-tau to capture normal tau, and to seed tau aggregation in vitro and in cultured cells. Tau pathology induced by hippocampal injection of AD O-tau in 3xTg-AD mice was suppressed by mixing 77G7 with AD O-tau. Intravenous administration of 77G7 ameliorated site-specific hyperphosphorylation of tau induced by AD O-tau in the hippocampi of Tg/hTau mice.These findings indicate that 77G7 can effectively suppress the seeding activity of AD O-tau and thus could be developed as a potential immunotherapeutic drug to inhibit the propagation of tau pathology in AD and related tauopathies.

Background: Abnormal hyperphosphorylation of microtubule-associated protein tau plays a pivotal role in Alzheimer’s disease (AD). We previously found that O-GlcNAcylation inversely correlates to hyperphosphorylation of tau in AD brain, and downregulation of brain O-GlcNAcylation promotes tau hyperphosphorylation and AD-like neurodegeneration in mice. Objective: Herein we investigated the effect of increasing O-GlcNAcylation by using intermittent dosing with low doses of a potent novel O-GlcNAcase (OGA) inhibitor on AD-like brain changes and cognitive function in a mouse model of sporadic AD (sAD) induced by intracerebroventricular (ICV) injection of streptozotocin (STZ). Methods: STZ was injected into the lateral ventricle of C57BL/6J mice. From the second day, Thiamme2-G (TM2G) or saline, as a vehicle control, was orally administered to the ICV-STZ mice three times per week for five weeks. A separate group of ICV-saline mice treated with saline was used as a baseline control. Behavioral tests, including open field and novel object recognition, were conducted three weeks after the first dose of the TM2G or saline. Protein O-GlcNAcylation, tau hyperphosphorylation, synaptic proteins, and neuroinflammation in the mouse brain were assessed by western blotting. Results: ICV-STZ caused decreased protein O-GlcNAcylation. Enhancement of O-GlcNAcylation to moderate levels by using low-dose OGA inhibitor in ICV-STZ mice prevented STZ-induced body weight loss, rescued cognitive impairments, and restored AD-like pathologies, including hyperphosphorylation of tau and abnormalities in synaptic proteins and neuroinflammation. Conclusion: These findings suggest that moderately increasing protein O-GlcNAcylation by using low doses of OGA inhibitor may be a suitable therapeutic strategy for sAD.

Neurofibrillary tangles (NFTs) made of abnormally hyperphosphorylated tau are a hallmark of Alzheimer’s disease (AD) and related tauopathies. Regional distribution of NFTs is associated with the progression of the disease and has been proposed to be a result of prion-like propagation of misfolded tau. Tau in AD brain is heterogenous and presents in various forms. In the present study, we prepared different tau fractions by sedimentation combined with sarkosyl solubility from AD brains and analyzed their biochemical and pathological properties. We found that tau in oligomeric fraction (O-tau), sarkosyl-insoluble fractions 1 and 2 (SI1-tau and SI2-tau) and monomeric heat-stable fraction (HS-tau) showed differences in truncation, hyperphosphorylation, and resistance to proteinase K. O-tau, SI1-tau, and SI2-tau, but not HS-tau, were hyperphosphorylated at multiple sites and contained SDS- and β-mercaptoethanol–resistant high molecular weight aggregates, which lacked the N-terminal portion of tau. O-tau and SI2-tau displayed more truncation and less hyperphosphorylation than SI1-tau. Resistance to proteinase K was increased from O-tau to SI1-tau to SI2-tau. O-tau and SI1-tau, but not SI2-tau or HS-tau, captured tau from cell lysates and seeded tau aggregation in cultured cells. Heat treatment could not kill the prion-like activity of O-tau to capture normal tau. Hippocampal injection of O-tau into 18-month-old FVB mice induced significant tau aggregation in both ipsilateral and contralateral hippocampi, but SI1-tau only induced tau pathology in the ipsilateral hippocampus, and SI2-tau and HS-tau failed to induce any detectable tau aggregation. These findings suggest that O-tau and SI1-tau have prion-like activities and may serve as seeds to recruit tau and template tau to aggregate, resulting in the propagation of tau pathology. Heterogeneity of tau pathology within AD brain results in different fractions with different biological and prion-like properties, which may pose a major challenge in targeting tau for development of effective therapeutic treatments.

Neurofibrillary tangles of abnormally hyperphosphorylated Tau are a hallmark of Alzheimer's disease (AD) and related tauopathies. Tau is truncated at multiple sites by various proteases in AD brain. Although many studies have reported the effect of truncation on the aggregation of Tau, these studies mostly employed highly artificial conditions, using heparin sulfate or arachidonic acid to induce aggregation. Here, we report for the first time the pathological activities of various truncations of Tau, including site-specific phosphorylation, self-aggregation, binding to hyperphosphorylated and oligomeric Tau isolated from AD brain tissue (AD O-Tau), and aggregation seeded by AD O-Tau. We found that deletion of the first 150 or 230 amino acids (aa) enhanced Tau's site-specific phosphorylation, self-aggregation, and binding to AD O-Tau and aggregation seeded by AD O-Tau, but deletion of the first 50 aa did not produce a significant effect. Deletion of the last 50 aa was found to modulate Tau's site-specific phosphorylation, promote its self-aggregation, and cause it to be captured by and aggregation seeded by AD O-Tau, whereas deletion of the last 20 aa had no such effects. Among the truncated Taus, Tau(151–391) showed the highest pathological activities. AD O-Tau induced aggregation of Tau(151–391) in vitro and in cultured cells. These findings suggest that the first 150 aa and the last 50 aa protect Tau from pathological characteristics and that their deletions facilitate pathological activities. Thus, inhibition of Tau truncation may represent a potential therapeutic approach to suppress Tau pathology in AD and related tauopathies.

Protein phosphatase 2A (PP2A) is the major tau phosphatase. Its activity toward tau is regulated by the methylation of PP2A catalytic subunit (PP2Ac) at Leu309. Protein phosphatase methylesterase-1 (PME-1) demethylates PP2Ac and suppresses its activity. We previously found that glycogen synthase kinase-3β (GSK-3β) suppresses PME-1 expression. However, the underlying molecular mechanism is unknown. In the present study, we analyzed the promoter of PME-1 gene and found that human PME-1 promoter contains two lymphoid enhancer binding factor-1/T-cell factor (LEF1/TCF) cis-elements in which β-catenin serves as a co-activator. β-catenin acted on these two cis-elements and promoted PME-1 expression. GSK-3β phosphorylated β-catenin and suppressed its function in promoting PME-1 expression. Inhibition and activation of GSK-3β by PI3K-AKT pathway promoted and suppressed, respectively, PME-1 expression in primary cultured neurons, SH-SY5Y cells and in the mouse brain. These findings suggest that GSK-3β phosphorylates β-catenin and suppresses its function on PME-1 expression, resulting in an increase of PP2Ac methylation.

Alternative splicing of tau exon 10 generates tau isoforms with three or four microtubule-binding repeats, 3R-tau or 4R-tau, which are under developmental regulation. Dysregulation of tau exon 10 splicing is sufficient to cause neurodegenerative disorders. The RNA-binding Fox3 (Rbfox3), identified as NeuN, regulates RNA processing. However, whether Rbfox3/NeuN regulates tau exon 10 splicing is unknown. In the present study, we found that the developmental expression of 4R-tau coincided with the expression of Rbfox3 in rat brains. Rbfox3 enhanced tau exon 10 inclusion. Tau intron 10 contains UGCAUG, the conservative binding sequence of Rbfox3. Intron 10 of tau pre-mRNA was co-immunoprecipitated by Rbfox3/NeuN. Deletion mutants of the RNA recognition motif (RRM) or three RNA-binding sites of the RRM in Rbfox3/NeuN failed to enhance tau exon 10 inclusion. Rbfox3, specifically expressed in the fetal brain, did not affect tau exon 10 splicing. The level of Rbfox3/NeuN was reduced and was associated with the ratio of 4R-tau/3R-tau in the excitotoxic mouse brains induced by kainic acid. These findings suggest that Rbfox3/NeuN regulates the alternative splicing of tau exon 10 and that decreased Rbfox3/NeuN may lower the ratio of 4R-tau/3R-tau.

Background Recent studies indicated that circulatory factors in blood plasma from young animals can reactivate neurogenesis, restore synaptic plasticity, and improve cognitive function in aged animals. Here, we investigated if young plasma could have a possible therapeutic effect for treatment of Alzheimer’s disease (AD)-like pathologies and cognitive impairment in triple-transgenic AD (3×Tg-AD) mice. Methods We intravenously injected plasma from 2- to 3-month-old C57BL/6 J wild-type mice into 16–17-month-old 3×Tg-AD mice twice a week for 8 weeks. The behavioral tests including open field, novel object recognition, Morris water maze, and reversal Morris water maze were conducted after 4-week plasma injections. The effect of young plasma on tau and Aβ pathologies and on the levels of synaptic proteins and neuroinflammation were assessed by Western blots and immunohistochemical staining. Results Young plasma treatment improved short-term memory in the novel object recognition test and enhanced the spatial learning and memory in Morris water maze test and reversal Morris water maze test. Biochemical studies revealed that young plasma treatment reduced both tau and Aβ pathologies, as well as neuroinflammation in the mouse brain. However, we did not detect any significant changes in levels of synaptic proteins or the dentate gyrus neurogenesis in the mouse brain after the treatment with young plasma. Conclusions These data indicate that young blood plasma not only ameliorates tau and Aβ pathologies but also enhances the cognitive function in 3×Tg-AD mice. These findings suggest that transfusion with young blood plasma could be a potentially effective treatment for AD.

Background Abnormally hyperphosphorylated tau is the major protein of neurofibrillary tangles in Alzheimer's disease. Insulin activates PI3K-AKT signaling and regulates tau phosphorylation. Impaired brain insulin signaling is involved in Alzheimer's disease pathogenesis. However, the effect of peripheral insulin on tau phosphorylation is controversial. Objective In the present study, we determined the effect of peripheral insulin administration on tau phosphorylation in brain. Methods We intraperitoneally injected a super physiological dose of insulin to mice and analyzed PI3K-AKT signaling and tau phosphorylation in brains by western blots. Results We found that peripherally administered insulin activated the PI3K-AKT signaling pathway immediately in the liver, but not in the brain. Tau phosphorylation in the mouse brain was found to be first decreased (15 min) and then increased (30 min and 60 min) after peripheral insulin administration and these changes correlated inversely with body temperature and the level of brain protein O-GlcNAcylation. Maintaining body temperature of mice post peripheral insulin administration prevented the insulin/hypoglycemia-induced tau hyperphosphorylation after peripheral insulin administration. Conclusion These findings suggest that peripheral insulin can induce tau hyperphosphorylation through both hypothermia and downregulation of brain protein O-GlcNAcylation during hypoglycemia.

Background Evidence from clinical studies and basic research has shown a strong correlation between Alzheimer's disease (AD) and type 2 diabetes. Tau, a neuronal microtubule-associated protein, is hyperphosphorylated and aggregated into neurofibrillary tangles in the AD brain. However, the expression of tau in pancreas is under debate. Objective We determined the expression of tau in mouse pancreas. Methods We used western blots, immunoprecipitation, and immunohistochemical staining to analyze pancreatic expression of tau in mice. Results We found that neither total tau nor phosphorylated tau was detectable in the mouse pancreas by western blots. Immunostaining with pan tau antibodies R134d and Tau-5 revealed bright and dense varicosities in the pancreatic islets and the exocrine pancreas. These varicosities were immunoreactive to synapsin 1, a presynaptic marker which can outline autonomic nerve profiles in pancreas, exhibiting complete colocalization with tau. Importantly, endocrine cells in islets did not exhibit specific immunoreactivity to any of pan tau antibodies tested, nor did the exocrine cells. Conclusion In the mouse pancreas, we found that tau is exclusively expressed in autonomic nerve fibers, but there is no detectable expression in endocrine cells in the islet.

Background Alzheimer's disease (AD) is the most common cause of dementia. Studies indicate that neuroinflammation plays an important role in the pathophysiology of AD. High-mobility group box 1 (HMGB1) is an important chromatin protein. It can be secreted by immune cells and passively released from damaged cells to promote inflammation. HMGB1 also can recruit stem cells and promote their proliferation and tissue repairing. However, the role of HMGB1 in the progression of AD is currently unknown. Objective The aims were to investigate the effect of HMGB1 on the AD-related pathologies and cognitive function using 3×Tg-AD mouse model. Methods Female 5-month-old 3×Tg-AD mice were intracerebroventricularly injected with 4.5 μg of HMGB1 or with saline as a control. The levels of interesting protein were assessed by western blots or immunofluorescence. The effect of HMGB1 on the cognitive function was evaluated by one-trial novel object recognition test and Morris water maze. Results Intracerebroventricular injection of recombinant HMGB1 ameliorated cognitive impairment in 5-6-month-old 3×Tg-AD mice. The levels of synapsin 1, synaptophysin, MAP2, NeuN, and phosphorylated CREB were increased in HMGB1-treated 3×Tg-AD mouse brains. HMGB1 decreased intracellular amyloid-β level but did not affect tau phosphorylation. HMGB1 treatment also promoted neurogenesis in the dentate gyrus and increased the level of GFAP in the 3×Tg-AD mouse brains. Conclusion These results reveal a novel function of HMGB1 in enhancing neuroplasticity and improving cognitive function in 3×Tg-AD mice.

Recent pre-clinical and clinical studies suggest that general anesthesia in infants and children may increase the risk of learning disabilities. Currently, there is no treatment for preventing anesthesia-induced neurotoxicity and potential long-term functional impairment. Animal studies have shown that neonatal exposure to anesthesia can induce acute neurotoxicity and long-term behavioral changes that can be detected a few months later. It is currently unknown whether neonatal exposure, especially repeated exposures, to general anesthesia can induce or increase the risk for cognitive impairment during aging. Here, we report that repeated exposures of neonatal mice (P7–9 days old) to anesthesia with sevoflurane (3 h/day for 3 days) led to cognitive impairment that was detectable at the age of 18–19 months, as assessed by using novel object recognition, Morris water maze, and fear conditioning tests. The repeated neonatal exposures to anesthesia did not result in detectable alterations in neurobehavioral development, in tau phosphorylation, or in the levels of synaptic proteins in the aged mouse brains. Importantly, we found that treatment with intranasal insulin prior to anesthesia exposure can prevent mice from anesthesia-induced cognitive impairment. These results suggest that neonatal exposure to general anesthesia could increase the risk for cognitive impairment during aging. This study also supports pre-treatment with intranasal administration of insulin to be a simple, effective approach to prevent infants and children from the increased risk for age-related cognitive impairment induced by neonatal exposure to general anesthesia.

Moderate dietary restriction can ameliorate age-related chronic diseases such as Alzheimer's disease (AD) by increasing the expression of neurotrophic factors and promoting neurogenesis in the brain. Glycogen synthase kinase-3β (GSK-3β) signaling is essential for the coordination of progenitor cell proliferation and differentiation during brain development. The mechanisms by which GSK-3β is involved in dietary restriction-induced neurogenesis and cognitive improvement remain unclear. Six-month-old male 3xTg-AD and wild-type mice were fed on alternate days (intermittent fasting, IF) or ad libitum (AL) for 3 months. GSK-3β activity was regulated by bilaterally infusing lentiviral vectors carrying siRNA targeting GSK-3β into the dentate gyrus region of the hippocampus. Intermittent fasting promoted neuronal differentiation and maturation in the dentate gyrus and ameliorated recognized dysfunction in 3xTg-AD mice. These effects were reversed by siRNA targeting GSK-3β. After intermittent fasting, the insulin and protein kinase A signaling pathways were inhibited, while the adenosine monophosphate-activated protein kinase and brain-derived neurotrophic factor pathways were activated. These findings suggest that intermittent fasting can promote neuronal differentiation and maturation in the hippocampus by activating GSK-3β, thus improving learning and memory.

Sevoflurane and isoflurane are among the most commonly used general anesthetics for children including infants, but their impact on metabolism, especially on blood glucose level, in children is not well understood. We investigated the impacts of anesthesia of neonatal (7-8 days old) and adult (2-3 months old) mice with the inhalational anesthetics 2.5% sevoflurane or 1.5% isoflurane, or the injectable anesthetics propofol (150 mg/kg) or avertin (375 mg/kg), for up to 6 hours. We found that sevoflurane and isoflurane induced severe hypoglycemia in neonatal mice and that this phenomenon was specific to the inhalational anesthetics because the injectable anesthetics propofol and avertin did not induce hypoglycemia. Surprisingly, the inhalational anesthesia induced hyperglycemia instead in adult mice. We also demonstrated that the inhalational anesthesia-induced hypoglycemia was a major cause of death for the neonatal mice receiving intranasal administration of saline prior to anesthesia. These studies revealed severe hypoglycemia in neonatal mice during anesthesia with sevoflurane or isoflurane. If this phenomenon also occurs in human, our findings would warrant closely monitoring blood glucose level and maintaining it in the normal range in infants receiving inhalational anesthesia.

The aim of this study was to investigate the effect of melatonin (MT) and its metabolite N(1)-acetyl-N(2)-formyl-5-methoxykynuramine (AFMK) on Alzheimer-like learning and memory impairment in rats intracerebroventricularly injected with streptozotocin (STZ). The results showed that the escape latency of the STZ group was longer than that of the control (CON), MT, and AFMK groups. Increased levels of hyperphosphorylated tau, neurofilament proteins, and malondialdehyde and decreased superoxide dismutase levels were observed in the brains of the rats from the STZ group compared with the brains of the rats from the CON, MT, AFMK high and low group. These results suggest that exogenous MT and AFMK can improve memory impairment and downregulate AD-like hyperphosphorylation induced by STZ, most likely through their antioxidation function. Meanwhile, we found that an equal dose of AFMK had a stronger effect than that of MT. Our results indicate that MT and its metabolite AFMK represent novel treatment strategies for Alzheimer's disease.

Concerns about the potential neurotoxicity of general anesthesia to the developing brain have been increasing in recent years. Animal studies have shown that neonatal exposure to general anesthesia causes both acute neurotoxicity and behavioral abnormalities later in life. In the present study, we observed over-activation of neuronal apoptosis in the brain of neonatal mice after a single exposure to anesthesia with sevoflurane for 6 hours at the age of 7 days. More importantly, we found that insulin administered through intranasal delivery prior to anesthesia prevented anesthesia-induced over-activation of neuronal apoptosis. This study provides experimental evidence for a potential effective, yet simple, method to prevent anesthesia-induced neurotoxicity in children, especially in infants.

The amyloid cascade hypothesis has been dominating drug discovery for Alzheimer's disease (AD) for the last two decades. The failure of the development of effective drugs for slowing down or reversing the progression of AD warrants the AD field to consider out-of-the-box thinking and therapeutic approaches. We propose the multifactorial hypothesis of AD, emphasizing that AD is caused by multiple etiological factors, which may result in common brain pathology and functional consequences through several separate but integrated molecular pathways. More than one etiological factor and mechanistic pathway may be involved in a single individual with sporadic AD, and different individuals may have different etiological factors, involving different mechanisms/pathways. We urge the recognition of the multifactorial nature of AD and the paradigm shift of AD drug development from a single target to multiple targets, either with the multitarget-directed ligands approach or the cocktail therapy approach. We believe that patient stratification and the use of the precision medicine model will also benefit AD drug discovery.

Impairment of cerebral glucose uptake/metabolism in individuals with Alzheimer's disease (AD) is believed to lead to downregulation of protein O-GlcNAcylation, which contributes to tau pathogenesis through tau hyperphosphorylation. Level of glucose transporter 3 (GLUT3), a neuronal specific glucose transporter, is decreased in AD brain, which may contribute to impaired brain glucose uptake/metabolism. However, what causes the reduction of GLUT3 in AD brain is not fully understood. Here, we report 1) that decrease of GLUT3 is associated with the reduction of protein O-GlcNAcylation in AD brain, 2) that GLUT3 level is negatively correlated with calpain I activation in human brain, 3) that calpain I proteolyzes GLUT3 at the N-terminus in vitro, and 4) that activation of calpain I is negatively correlated with protein O-GlcNAcylation in AD brain. Furthermore, we found that overexpression of GLUT3 enhances protein O-GlcNAcylation in N2a cells. Overexpression of calpain I suppresses protein O-GlcNAcylation in these cells. These findings suggest a novel mechanism by which calpain I overactivation leads to GLUT3 degradation and the consequent down-regulation of protein O-GlcNAcylation in AD brain.

Alzheimer's disease (AD), which accounts for three fourth of all cases of dementia, is a major public health problem in modern society and, yet, there is no effective treatment available that can prevent or inhibit this chronic progressive neurodegenerative disease. A major current drug target is intraneuronal abnormally hyperphosphorylated microtubule-associated protein tau which is a histopathological hallmark of this disease and of a family of neurodegenerative diseases called tauopathies. Areas covered: In this review, the authors discuss a growing number of studies that describe the nature and mechanism of tau pathology and various drug discovery options and most recent developments in tau-based therapeutics. PubMed was used to obtain relevant literature while clinicaltrials.gov site and Google search were employed to obtain the latest information on tau based AD clinical trials. Expert opinion: In authors' opinion, loss of neuronal connectivity leads to the hyperphosphorylation of tau and is thus a key therapeutic target. Rescue of neuronal connectivity loss and hyperphosphorylation of tau are most promising approaches. Consequently, tau immunotherapy has a high therapeutic potential.

O-GlcNAcylation is a post-translational modification of proteins. Protein kinase A (PKA)-cAMP response element binding protein (CREB) signaling plays critical roles in multiple biological processes. Isoforms α and β of PKA catalytic subunit (PKAc) and CREB are modified by O-GlcNAcylation. In the present study, we determined the role of O-GlcNAcylation in PKAc isoform-specific CREB signaling. We found that up-regulation of O-GlcNAcylation enhanced CREB phosphorylation, but suppressed CREB expression in exogenous PKAc isoform-unspecific manner. PKAc isoforms affected exogenous expression of OGT or OGA and protein O-GlcNAcylation differently. Up-regulation of O-GlcNAcylation did not significantly affect net PKAcα-CREB signaling, but enhanced PKAcβ-CREB signaling. The role of O-GlcNAcylation in PKA-CREB signaling was desensitized by insulin treatment. This study suggests a role of O-GlcNAcylation in PKA-CREB signaling by affecting phosphorylation of CREB in a PKAc isoform-specific manner.

Hyperphosphorylation of tau and imbalanced expression of 3R-tau and 4R-tau as a result of dysregulation of tau exon 10 splicing are believed to be pivotal to the pathogenesis of tau pathology, but the molecular mechanism leading to the pathologic tau formation in Alzheimer’s disease (AD) brain is not fully understood. In the present study, we found that casein kinase 1ε (CK1ε) was increased significantly in AD brains. Overexpression of CK1ε in cultured cells led to increased tau phosphorylation at many sites. Moreover, we found that CK1ε suppressed tau exon 10 inclusion. Levels of CK1ε were positively correlated to tau phosphorylation, 3R-tau expression and tau pathology, and negatively correlated to 4R-tau in AD brains. Overexpression of CK1ε in the mouse hippocampus increased tau phosphorylation and impaired spontaneous alternation behavior. These data suggest that CK1ε is involved in the regulation of tau phosphorylation, the alternative splicing of tau exon 10, and cognitive performance. Up-regulation of CK1ε might contribute to tau pathology by hyperphosphorylating tau and by dysregulating the alternative splicing of tau exon 10 in AD.

O-GlcNAcylation is a common posttranslational modification of nucleocytoplasmic proteins with β-N-acetylglucosamine (GlcNAc) and regulates numerous biological processes. By using mouse models of cerebral ischemia induced by permanent and transient middle cerebral artery occlusion (MCAO), we observed an initial elevation (~1.7-fold, 1–4 hours after ischemia) and then decline of O-GlcNAcylation during cerebral ischemia. We found that moderate increase (

Hyperphosphorylation and aggregation of the neuronal protein tau are responsible for neurodegenerative diseases called tauopathies. Dysregulation of the alternative splicing of tau exon 10 results in alterations of the ratio of two tau isoforms, 3R-tau and 4R-tau, which have been seen in several tauopathies. Transactive response DNA-binding protein of 43 kDa (TDP-43) is involved in the regulation of RNA processing, including splicing. Cytoplasmic aggregation of TDP-43 has been observed in the brains of individuals with chronic traumatic encephalopathy or Alzheimer's disease, diseases in which neurofibrillary tangles of hyperphosphorylated tau are hallmarks. Here, we investigated the role of TDP-43 in tau exon 10 splicing. We found that TDP-43 promoted tau exon 10 inclusion, which increased production of the 4R-tau isoform. Moreover, TDP-43 could bind to intron 9 of tau pre-mRNA. Deletion of the TDP-43 N or C terminus promoted its cytoplasmic aggregation and abolished or diminished TDP-43-promoted tau exon 10 inclusion. Several TDP-43 mutations associated with amyotrophic lateral sclerosis or frontotemporal lobar degeneration with ubiquitin inclusions promoted tau exon 10 inclusion more effectively than wild-type TDP-43 but did not affect TDP-43 cytoplasmic aggregation in cultured cells. The ratio of 3R-tau/4R-tau was decreased in transgenic mouse brains expressing human TDP-43 and increased in the brains expressing the disease-causing mutation TDP-43M337V, in which cytoplasmic TDP-43 was increased. These findings suggest that TDP-43 promotes tau exon 10 inclusion and 4R-tau expression and that disease-related changes of TDP-43, truncations and mutations, affect its function in tau exon 10 splicing, possibly because of TDP-43 mislocalization to the cytoplasm.

Alternative splicing of tau exon 10 generates tau isoforms with three or four microtubule-binding repeats, 3R-tau and 4R-tau, which is equally expressed in adult human brain. Imbalanced expression in 3R-tau and 4R-tau has been found in several sporadic and inherited tauopathies, suggesting that dysregulation of tau exon 10 is sufficient to cause neurodegenerative diseases. We previously reported that Dyrk1A, which is overexpressed in Down syndrome brains, regulates alternative splicing of exogenous tau exon 10. In the present study, we investigated the regulation of endogenous tau exon 10 splicing by Dyrk1A. We found that inhibition of Dyrk1A enhanced tau exon 10 inclusion, leading to an increase in 4R-tau/3R-tau ratio in differentiated-human neuronal progenitors and in the neonatal rat brains. Accompanied with overexpression of Dyrk1A, 3R-tau was increased and 4R-tau was decreased in the neonatal brains of Ts65Dn mice, a model of Down syndrome. Treatment with Dyrk1A inhibitor, green tea flavonol epigallocatechin-gallate (EGCG), from gestation to adulthood suppressed 3R-tau expression and rescued anxiety and memory deficits in Ts65Dn mouse brains. Thus, Dyrk1A might be an ideal therapeutic target for Alzheimer’s disease, especially for Down syndrome and EGCG which inhibits Dyrk1A may have potential effect on the treatment or prevention of this disease.

Glycogen synthase kinase-3β (GSK-3β) is the major tau kinase. Its phosphorylation at Ser9 suppresses the activity. In Alzheimer's disease (AD) brain, GSK-3β is truncated at the C terminus by overactivated calpain I, leading to an increase in its activity. However, the effect of truncation on its phosphorylation is unknown. We found here that in AD brain and in cultured cells, C-terminally truncated GSK-3β is less phosphorylated at Ser9 than the full-length enzyme. The truncation promotes GSK-3β nuclear translocation and enhances its interaction with protein phosphatase 2A (PP2A), leading to dephosphorylation. Thus, the truncation of GSK-3β may enhance its activity through Ser9 dephosphorylation by PP2A. Our findings shed new light on the role of calpain-GSK-3β-PP2A in tau pathogenesis of AD.

In the brains of individuals with Alzheimer's disease (AD) and chronic traumatic encephalopathy, tau pathology is accompanied usually by intracellular aggregation of transactive response DNA-binding protein 43 (TDP-43). However, the role of TDP-43 in tau pathogenesis is not understood. Here, we investigated the role of TDP-43 in tau expression in vitro and in vivo. We found that TDP-43 suppressed tau expression by promoting its mRNA instability through the UG repeats of its 3΄-untranslated region (3΄-UTR). The C-terminal region of TDP-43 was required for this function. Neurodegenerative diseases-causing TDP-43 mutations affected tau mRNA instability differentially, in that some promoted and others did not significantly affect tau mRNA instability. The expression levels of tau and TDP-43 were inverse in the frontal cortex and the cerebellum. Accompanied with cytoplasmic accumulation of TDP-43, tau expression was elevated in TDP-43M337V transgenic mouse brains. The level of TDP-43, which is decreased in AD brains, was found to correlate negatively with the tau level in human brain. Our findings indicate that TDP-43 suppresses tau expression by promoting the instability of its mRNA. Down-regulation of TDP-43 may be involved in the tau pathology in AD and related neurodegenerative disorders.

General anesthesia increases the risk for cognitive impairment and Alzheimer's disease (AD) in vulnerable individuals such as the elderly. We previously reported that prior administration of insulin through intranasal delivery can prevent the anesthesia-induced cognitive impairment and biochemical changes in the brain. However, little is known about the underlying molecular mechanisms. Here, we report that general anesthesia resulted in downregulation of mammalian/mechanistic target of rapamycin (mTOR) and eukaryotic elongation factor 2 (eEF2) in the brain along with reduction of presynaptic proteins and brain-derived neurotrophic factor and cognitive impairment in aged mice. Prior administration of intranasal insulin prevented these anesthesia-induced changes. These results suggest the involvement of the mTOR-eEF2 signaling pathway in the anesthesia-induced brain changes and cognitive impairment and in the prevention of these changes with insulin. Correlation analyses and the use of eEF2 kinase inhibitor further support our conclusions. These studies shed light on the molecular mechanism by which anesthesia and insulin could act on synaptic proteins and cognitive function.

Children, after multiple exposures to general anesthesia, appear to be at an increased risk of developing learning disabilities. Almost all general anesthetics-including sevoflurane, which is commonly used for children-are potentially neurotoxic to the developing brain. Anesthesia exposure during development might also be associated with behavioral deficiencies later in life. To date, there is no treatment to prevent anesthesia-induced neurotoxicity and behavioral changes. In this study, we anesthetized 7-day-old neonatal mice with sevoflurane for 3 h per day for three consecutive days and found that the anesthesia led to mild behavioral abnormalities later in life that were detectable by using the novel object recognition test, Morris water maze, and fear conditioning test. Biochemical and immunohistochemical studies indicate that anesthesia induced a decrease in brain levels of postsynaptic density 95 (PSD95), a postsynaptic marker, and marked activation of neuronal apoptosis in neonatal mice. Importantly, insulin administered through intranasal delivery prior to anesthesia was found to prevent the anesthesia-induced long-term behavioral abnormalities, reduction of PSD95, and activation of neuronal apoptosis. These findings suggest that intranasal insulin administration could be an effective approach to prevent the increased risk of neurotoxicity and chronic damage caused by anesthesia in the developing brain.

Type 2 diabetes mellitus (T2DM) and Alzheimer's disease (AD) are both serious global health problems with high prevalence. These two diseases have some common features, including risk factors, age-associated disease onsets, insulin resistance, impaired glucose metabolism, deregulation of O-GlcNAcylation, chronic oxidative stress, and inflammation. Some of these features, such as insulin resistance, impaired glucose metabolism, and deregulation of O-GlcNAcylation, may serve as molecular links between T2DM and AD. Research on these molecular links is reviewed and discussed in this chapter. Understanding of these molecular links will help uncover the disease mechanisms and design therapeutic strategies to prevent and treat these two devastating diseases.

Alzheimer's disease (AD) is characterized by the presence of extracellular amyloid β plaques and intraneuronal neurofibrillary tangles of hyperphosphorylated microtubule-associated protein tau in the brain. Aggregation of transactive response DNA-binding protein of 43 kDa (TDP-43) in the neuronal cytoplasm is another feature of AD. However, how TDP-43 is associated with AD pathogenesis is unknown. Here, we found that casein kinase 1e (CK1e) phosphorylated TDP-43 at Ser403/404 and Ser409/410. In AD brains, the level of CK1e was dramatically increased and positively correlated with the phosphorylation of TDP-43 at Ser403/404 and Ser409/410. Overexpression of CK1e promoted its cytoplasmic aggregation and suppressed TDP-43-promoted tau mRNA instability and tau exon 10 inclusion, leading to an increase of tau and 3R-tau expressions. Levels of CK1e and TDP-43 phosphorylation were positively correlated with the levels of total tau and 3R-tau in human brains. Furthermore, we observed, in pilot immunohistochemical studies, that the severe tau pathology was accompanied by robust TDP-43 pathology and a high level of CK1e. Taken together, our findings suggest that the elevation of CK1e in AD brain may phosphorylate TDP-43, promote its cytoplasmic aggregation and suppress its function in tau mRNA processing, leading to acceleration/exacerbation of tau pathology. Thus, the elevation of CK1e may link TDP-43 to tau pathogenesis in AD brain.