Pinning down phosphorylated tau and tauopathies

Neurofibrillary tangles (NFTs) are prominent neuronal lesions in a large subset of neurodegenerative diseases, including Alzheimer's disease (AD). NFTs are mainly composed of insoluble Tau that is hyperphosphorylated on many serine or threonine residues preceding proline (pSer/Thr-Pro). Tau hyperphosphorylation abolishes its biological function to bind microtubules and promotes microtubule assembly and precedes neurodegeneration. Not much is known about how tau is further regulated following phosphorylation. Notably, we have recently shown that phosphorylated Ser/Thr-Pro motifs exist in two distinct conformations. The conversion between two conformations in some proteins is catalyzed by the prolyl isomerase Pin1. Pin1 binds to tau phosphorylated specifically on the Thr231-Pro site and probably catalyzes cis/trans isomerization of pSer/Thr-Pro motif(s), thereby inducing conformational changes in tau. Such conformational changes can directly restore the ability of phosphorylated Tau to bind microtubules and promote microtubule assembly and/or facilitate tau dephosphorylation by its phosphatase PP2A, as PP2A activity is conformation-specific. Furthermore, Pin1 expression inversely correlates with the predicted neuronal vulnerability in normally aged brain and also with actual neurofibrillary degeneration in AD brain. Moreover, deletion of the gene encoding Pin1 in mice causes progressive age-dependent neuropathy characterized by motor and behavioral deficits, tau hyperphosphorylation, tau filament formation and neuronal degeneration. Distinct from all other mouse models where transgenic overexpression of specific proteins elicits tau-related pathologies, Pin1 is the first protein whose depletion causes age-dependent neurodegeneration and tau pathologies. Thus, Pin1 is pivotal in maintaining normal neuronal function and preventing age-dependent neurodegeneration. This could represent a promising interventive target to prevent neurodegenerative diseases.     

Interaction of tau isoforms with Alzheimer's disease abnormally hyperphosphorylated tau and in vitro phosphorylation into the disease-like protein

The microtubule-associated protein tau is a family of six isoforms that becomes abnormally hyperphosphorylated and accumulates in neurons undergoing neurodegeneration in the brains of patients with Alzheimer disease (AD). We investigated the isoform-specific interaction of normal tau with AD hyperphosphorylated tau (AD P-tau). We found that the binding of AD P-tau to normal human recombinant tau was tau4L > tau4S > tau4 and tau3L > tau3S > tau3, and that its binding to tau4L was greater than to tau3L. AD P-tau also inhibited the assembly of microtubules promoted by each tau isoform and caused disassembly when added to preassembled microtubules. This inhibition and depolymerization of microtubules by the AD P-tau corresponded directly to the degree of its interaction with the different tau isoforms. In vitro hyperphosphorylation of recombinant tau (P-tau) conferred AD P-tau-like characteristics. Like AD P-tau, P-tau interacted with and sequestered normal tau and inhibited microtubule assembly. These studies suggest that the AD P-tau interacts preferentially with the tau isoforms that have the amino-terminal inserts and four microtubule binding domain repeats and that hyperphosphorylation of tau appears to be sufficient to acquire AD P-tau characteristics. Thus, lack of amino-terminal inserts and extra microtubule binding domain repeat in fetal human brain might be protective from Alzheimer's neurofibrillary degeneration.     

Inverse and distinct modulation of tau-dependent neurodegeneration by presenilin 1 and amyloid-beta in cultured cortical neurons: evidence that tau phosphorylation is the limiting factor in amyloid-beta-induced cell death

Alzheimer's disease (AD) is characterized by massive neuron loss in distinct brain regions, extracellular accumulations of the amyloid precursor protein-fragment amyloid-beta (A beta) and intracellular tau fibrils containing hyperphosphorylated tau. Experimental evidence suggests a relation between presenilin (PS) mutations, A beta formation, and tau phosphorylation in triggering cell death; however, how A beta and PS affect tau-dependent degeneration is unknown. Using herpes simplex virus 1-mediated gene-transfer of fluorescent-tagged tau constructs in primary cortical neurons, we demonstrate that tau expression exerts a neurotoxic effect that is increased with a construct mimicking disease-like hyperphosphorylation [pseudohyperphosphorylated (PHP) tau]. Live imaging revealed that PHP tau expression is associated with increased perikarya suggesting the development of a 'ballooned' phenotype as a specific feature of tau-mediated cell death. Transgenic expression of PS1 suppressed tau-induced neurodegeneration. In contrast, A beta amplified degeneration in the presence of wt tau but not of PHP tau. The data indicate that PS1 and A beta inversely modulate tau-dependent neurodegeneration at distinct steps. They indicate that the mode by which PHP tau causes neurotoxicity is downstream of A beta and that tau phosphorylation is the limiting factor in A beta-induced cell death. Suppression of tau expression or inhibition of tau phosphorylation at disease-relevant sites may provide an effective therapeutic strategy to prevent neurodegeneration in Alzheimer's disease.     

MARKing tau for tangles and toxicity

In healthy neurons, tau proteins regulate microtubule function in the axon. In the brains of individuals with Alzheimer's disease, tau is hyperphosphorylated and aggregated into intraneuronal deposits called neurofibrillary tangles (NFTs). Hyperphosporylation dislodges tau from the microtubule surface, potentially resulting in compromised axonal integrity and the accumulation of toxic tau peptides. Recent biochemical and animal model studies have re-evaluated tau phosphorylation and other aspects of neurofibrillar pathology. The results indicate that phosphorylation of tau's microtubule-binding domain by the protein kinase MARK primes tau for hyperphosphorylation by the kinases GSK-3 and Cdk5, which in turn triggers the aggregation of tau into filaments and tangles. Toxic consequences for the neuron might be exacerbated by tangle formation but are already evident during the early steps of the process.     

Alzheimer neurofibrillary degeneration: significance, etiopathogenesis, therapeutics and prevention

Alzheimer disease (AD) is multi-factorial and heterogeneous. Independent of the aetiology, this disease is characterized clinically by chronic and progressive dementia and histopathologically by neurofibrillary degeneration of abnormally hyperphosphorylated tau seen as intraneuronal neurofibrillary tangles, neuropil threads and dystrophic neurites, and by neuritic (senile) plaques of beta-amyloid. The neurofibrillary degeneration is apparently required for the clinical expression of AD, and in related tauopathies it leads to dementia in the absence of amyloid plaques. While normal tau promotes assembly and stabilizes microtubules, the abnormally hyperphosphorylated tau sequesters normal tau, MAP1 and MAP2, and disrupts microtubules. The abnormal hyperphosphorylation of tau also promotes its self-assembly into tangles of paired helical and or straight filaments. Tau is phosphorylated by a number of protein kinases. Glycogen synthase kinase-3 (GSK-3) and cyclin dependent protein kinase 5 (cdk5) are among the kinases most implicated in the abnormal hyperphosphorylation of tau. Among the phosphatases which regulate the phosphorylation of tau, protein phosphatase-2A (PP-2A), the activity of which is down-regulated in AD brain, is by far the major enzyme. The inhibition of abnormal hyperphosphorylation of tau is one of the most promising therapeutic targets for the development of disease modifying drugs. A great advantage of inhibiting neurofibrillary degeneration is that it can be monitored by evaluating the levels of total tau and tau phosphorylated at various known abnormally hyperphosphorylated sites in the cerebrospinal fluid of patients, obtained by lumbar puncture. There are at least five subgroups of AD, each is probably caused by a different etiopathogenic mechanism. The AD subgroup identification of patients can help increase the success of clinical trials and the development of specific and potent disease modifying drugs.     

Overactivation of glycogen synthase kinase-3 by inhibition of phosphoinositol-3 kinase and protein kinase C leads to hyperphosphorylation of tau and impairment of spatial memory

Neurofibrillary tangles (NFTs) consisting of the hyperphosphorylated microtubule-associated protein tau are a defining pathological characteristic of Alzheimer's disease (AD). Hyperphosphorylation of tau is hypothesized to impair the microtubule stabilizing function of tau, leading to the formation of paired helical filaments and neuronal death. Glycogen synthase kinase-3 (GSK-3) has been shown to be one of several kinases that mediate tau hyperphosphorylation in vitro. However, molecular mechanisms underlying overactivation of GSK-3 and its potential linkage to AD-like pathologies in vivo remain unclear. Here, we demonstrate that injection of wortmannin (a specific inhibitor of phosphoinositol-3 kinase) or GF-109203X (a specific inhibitor of protein kinase C) into the left ventricle of rat brains leads to overactivation of GSK-3, hyperphosphorylation of tau at Ser 396/404/199/202 and, most significantly, impaired spatial memory. The effects of wortmannin and GF-109203X are additive. Significantly, specific inhibition of GSK-3 activity by LiCl prevents hyperphosphorylation of tau, and spatial memory impairment resulting from PI3K and PKC inhibition. These results indicate that in vivo inhibition of phosphoinositol-3 kinase and protein kinase C results in overactivation of GSK-3 and tau hyperphosphorylation and support a direct role of GSK-3 in the formation of AD-like cognitive deficits.     

Cdk5 is involved in NFT-like tauopathy induced by transient cerebral ischemia in female rats

Although neurofibrillary tangle (NFT) formation is a central event in both familial and sporadic Alzheimer's disease (AD), neither cellular origin nor functional consequence of the NFTs are fully understood. This largely is due to the lack of available in vivo models for neurofibrillary degeneration (NFD). NFTs have only been identified in transgenic mice, bearing a transgene for a rare hereditary neurodegenerative disease, frontotemporal dementia with Parkinsonism linked to chromosome 17 (FTDP17). Epidemiological evidence suggests a much higher occurrence of dementia in stroke patients. This may represent the underlying cause of the pathogenesis of sporadic AD, which accounts for the majority of AD cases. We examined pathological markers of AD in a rodent stroke model. Here we show that after transient cerebral ischemia, hyperphosphorylated tau accumulates in neurons of the cerebral cortex in the ischemic area, forms filaments similar to those present in human neurodegenerative tauopathies and colocalizes with markers of apoptosis. As a potential underlying mechanism, we were able to determine that transient ischemia induced tau hyperphosphorylation and NFT-like conformations are associated with aberrant activation of cyclin dependent kinase 5 (Cdk5) and can be rescued by delivery of a potent, but non-specific cyclin dependent kinase inhibitor, roscovitine to the brain. Our study further indicates that accumulation of p35 and its calpain-mediated cleavage product, p25 may account for the deregulation of Cdk5 induced by transient ischemia. We conclude that Cdk5 may be the principal protein kinase responsible for tau hyperphosphorylation and may be a hallmark of the tauopathies in this stroke model.     

Epitope mapping of mAbs AT8 and Tau5 directed against hyperphosphorylated regions of the human tau protein

Post-mortem diagnosis of Alzheimer's disease relies on high numbers of senile plaques and neurofibrillary tangles (NFTs) stained in distinct brain areas. NFTs mostly consist of hyperphosphorylated versions of the microtubule attached tau protein (PHF-tau) with more than 30 serine and threonine phosphorylation sites identified so far. Characterization of hyperphosphorylated tau regions and the hope to develop robust assays for early AD diagnosis relies mostly on phosphorylation-dependent monoclonal antibodies (mAbs) recognizing only disease-specific phosphorylation patterns. Here, we report that anti-PHF-tau mAb AT8 recognizes an epitope doubly phosphorylated at serine 202 and threonine 205, which was not influenced by a third phosphate group at serine 199. But mAb AT8 was cross-reactive to two doubly phosphorylated motifs containing either serines 199 and 202 or serines 205 and 208 of the human tau sequence. The epitope of anti-tau mAb Tau5 was mapped to the human tau sequence 218-225, which is not phosphorylated in vivo.     

Structural and functional implications of tau hyperphosphorylation: information from phosphorylation-mimicking mutated tau proteins

Abnormal tau-immunoreactive filaments are a hallmark of tauopathies, including Alzheimer's disease (AD). A higher phosphorylation ("hyperphosphorylation") state of tau protein may represent a critical event. To determine the potential role of tau hyperphosphorylation in these disorders, mutated tau proteins were produced where serine/threonine residues known to be highly phosphorylated in tau filaments isolated from AD patients were substituted for glutamate to simulate a paired helical filament (PHF)-like tau hyperphosphorylation. We demonstrate that, like hyperphosphorylation, glutamate substitutions induce compact structure elements and SDS-resistant conformational domains in tau protein. Hyperphosphorylation-mimicking glutamate-mutated tau proteins display a complete functional loss in its ability to promote microtubule nucleation which can partially be overcome by addition of the osmolyte trimethylamine N-oxide (TMAO), which is similar to phosphorylated tau. In addition, glutamate-mutated tau proteins fail to interact with the dominant brain protein phosphatase 2A isoform ABalphaC, and exhibit a reduced ability to assemble into filaments. Interestingly, wild-type tau and phosphorylation-mimicking tau similarly bind to microtubules when added alone, but the mutated tau is almost completely displaced from the microtubule surface by equimolar concentrations of wild-type tau. The data indicate that glutamate-mutated tau proteins provide a useful model for analyzing the functional consequences of tau hyperphosphorylation. They suggest that several mechanisms contribute to the abnormal tau accumulation observed during tauopathies, in particular a selective displacement of hyperphosphorylated tau from microtubules, a functional loss in promoting microtubule nucleation, and a failure to interact with phosphatases.     

IL-5 blocks apoptosis and tau hyperphosphorylation induced by Aβ<Subscript>25–35</Subscript> peptide in PC12 cells

The primary features of Alzheimer’s disease (AD) are extracellular amyloid plaques consisting mainly of deposits of amyloid β (Aβ) peptides and intracellular neurofibrillary tangles (NFTs). Sets of evidence suggest that interleukin-5 (IL-5) is involved in the pathogenesis of AD. Herein, we investigated the protective role of IL-5 in PC12 cells, to provide new insights into understanding this disease. Western blot was employed to assess the protein levels of Bax and phospho-tau as well as phospho-JAK2; MTT assay was performed to decipher cell viability. Treatment of IL-5 decreased Aβ_25–35-induced tau phosphorylation and apoptosis, effects blunted by JAK2 inhibition. IL-5 prevents Aβ_25–35-evoked tau protein hyperphosphorylation and apoptosis through JAK2 signaling.     

Linking alterations in tau phosphorylation and cleavage during neuronal apoptosis

Neurofibrillary tangles (NFTs) are classic lesions of Alzheimer's disease. NFTs are bundles of abnormally phosphorylated tau, the paired helical filaments. The initiating mechanisms of NFTs and their role in neuronal loss are still unknown. Accumulating evidence supports a role for the activation of proteolytic enzymes, caspases, in neuronal death observed in brains of patients with Alzheimer's disease. Alterations in tau phosphorylation and tau cleavage by caspases have been previously reported in neuronal apoptosis. However, the links between the alterations in tau phosphorylation and its proteolytic cleavage have not yet been documented. Here, we show that, during staurosporine-induced neuronal apoptosis, tau first undergoes transient hyperphosphorylation, which is followed by dephosphorylation and cleavage. This cleavage generated a 10-kDa fragment in addition to the 17- and 50-kDa tau fragments previously reported. Prior tau dephosphorylation by a glycogen synthase kinase-3beta inhibitor, lithium, enhanced tau cleavage and sensitized neurons to staurosporine-induced apoptosis. Caspase inhibition prevented tau cleavage without reversing changes in tau phosphorylation linked to apoptosis. Furthermore, the microtubule depolymerizing agent, colchicine, induced tau dephosphorylation and caspase-independent tau cleavage and degradation. Both phenomena were blocked by inhibiting protein phosphatase 2A (PP2A) by okadaic acid. These experiments indicate that tau dephosphorylation precedes and is required for its cleavage and degradation. We propose that the absence of cleavage and degradation of hyperphosphorylated tau (due to PP2A inhibition) may lead to its accumulation in degenerating neurons. This mechanism may contribute to the aggregation of hyperphosphorylated tau into paired helical filaments in Alzheimer's disease where reduced PP2A activity has been reported.     

饥饿对小鼠脑中tau蛋白磷酸化和O-GlcNAc糖基化的影响

为了探讨大脑中葡萄糖摄取和代谢障碍在阿尔茨海默病(Alzheimer$sdisease,AD)神经退行性病变中的作用,将昆明种小鼠进行饥饿和再喂食处理,并使用多种磷酸化tau蛋白特异性的抗体和蛋白O-GlcNAc糖基化特异性抗体进行检测,观察饥饿及恢复喂养后不同时间点大脑皮质中tau蛋白糖基化及多个位点磷酸化的变化.结果显示:饥饿处理引起小鼠大脑皮质中总蛋白和tau蛋白的O-GlcNAc糖基化水平降低,同时tau蛋白磷酸化水平升高,饥饿引起的tauO-GlcNAc糖基化和磷酸化改变均在恢复进食后逆转成正常水平.该研究结果提示:大脑中tau蛋白的磷酸化和O-GlcNAc糖基化之间存在相互调节,脑中葡萄糖代谢障碍可能通过下调tau蛋白O-GlcNAc糖基化水平使tau蛋白产生异常过度磷酸化,进而促发AD的病理进程.这一结果为在早期阶段通过逆转tau蛋白异常过度磷酸化治疗AD成为可能提供了实验基础.     

Anisomycin过度激活丝裂原活化蛋白激酶使tau发生过度磷酸化

阿尔茨海默病(Alzheimer's disease,AD)的病理特征之一是神经元内存在神经原纤维缠结(neurofibrillary tangles,NFTs),后者是由过度磷酸化的微管相关蛋白tau形成的双股螺旋细丝(paired helical filaments,PHFs)构成.为了探讨丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)在微管相关蛋白tau磷酸化中的作用及机制,本实验用0.1 μg/mL、0.2 μg/mL和0.4μg/mL三种不同浓度的MAPK激动剂anisomycin处理小鼠成神经瘤细胞株(mouse neuroblastoma cells,N2a),检测MAPK活性的变化及其与tau蛋白多个AD相关位点过度磷酸化的关系,并检测糖原合酶激酶-3(glycogen synthase kinase-3,GSK-3)和蛋白激酶A(protein kinase A,PKA)的活性变化.结果显示,anisomycin以剂量依赖的方式激活MAPK活性,但免疫印迹结果显示tau蛋白的Ser-198/199/202位点和Ser-396/404位点的过度磷酸化只在anisomycin浓度为0.4 μg/mL时出现,三种浓度的anisomycin均未引起tau蛋白Ser-214位点磷酸化的改变;同时,GSK-3活性在anisomycin为0.1 μg/mL时没有明显变化,当anisomycin浓度升高到0.2 μg/mL和0.4 μg/mL时出现明显增高,而PKA的活性没有明显的改变.使用GSK-3的特异性抑制剂氯化锂(LiCl)则完全阻断MAPK被过度激活导致的tau蛋白磷酸化水平的增高,而同时MAPK活性不受影响.以上结果提示:过度激活MAPK可以导致tau蛋白Ser-198/199/202和Ser-396/404位点过度磷酸化,其机制可能涉及MAPK激活GSK-3的间接作用.     

Hyperphosphorylation of tau induces local polyproline II helix

Alzheimer's disease is characterized by two protein precipitates, extracellular amyloid plaques and intracellular neurofibrillary tangles (NFTs). The primary constituent of NFTs is a hyperphosphorylated form of the microtubule-binding protein tau. Hyperphosphorylation of tau on over 30 residues, primarily within proline-rich sequences, is associated with conformational changes whose nature is poorly defined. Peptides derived from the proline-rich region of tau (residues 174-242) were synthesized, and the conformations were analyzed for the nonphosphorylated and phosphorylated peptides. CD and NMR data indicate that phosphorylation of serine and threonine residues in proline-rich sequences induces a conformational change to a type II polyproline helix. The largest phosphorylation-dependent conformational changes observed by CD were for tau peptides incorporating residues 174-183 or residues 229-238. Phosphoserine and phosphothreonine residues exhibited ordered values of (3)J(alphaN) (3.1-6.2 Hz; mean = 4.7 Hz) compared to nonphosphorylated serine and threonine. Phosphorylation of a tau peptide consisting of tau residues 196-209 resulted in the disruption of a nascent alpha-helix. These results suggest that global reorganization of tau may occur upon hyperphosphorylation of proline-rich sequences in tau.     

Neuroglobin attenuates Alzheimer-like tau hyperphosphorylation by activating Akt signaling

Neuroglobin (Ngb) is a recently identified member of hemoglobin family, distributed mainly in central and peripheral nervous systems. Recent studies suggest that Ngb can protect neural cells from β-amyloid-induced toxicity in Alzheimer disease (AD). Hyperphosphorylation of tau is another characterized pathological hallmark in the AD brains; however, it is not reported whether Ngb also affects tau phosphorylation. In this study, we found that the level of Ngb was significantly reduced in Tg2576 mice (a recognized mouse model of AD) and TgMAPt mice, and the level of Ngb was negatively correlated with tau phosphorylation. Over-expression of Ngb attenuates tau hyperphosphorylation at multiple AD-related sites induced by up-regulation of glycogen synthase kinase-3β (GSK-3β), a crucial tau kinase. While Ngb activates Akt and thus inhibits GSK-3β, simultaneously inhibition of Akt abolishes the effects of Ngb on GSK-3β inhibition and tau hyperphosphorylation. Our data indicate that Ngb may attenuate tau hyperphosphorylation through activating Akt signaling pathway, implying a therapeutic target for AD.     

Death-associated protein kinase 1 has a critical role in aberrant tau protein regulation and function

The presence of tangles composed of phosphorylated tau is one of the neuropathological hallmarks of Alzheimer''s disease (AD). Tau, a microtubule (MT)-associated protein, accumulates in AD potentially as a result of posttranslational modifications, such as hyperphosphorylation and conformational changes. However, it has not been fully understood how tau accumulation and phosphorylation are deregulated. In the present study, we identified a novel role of death-associated protein kinase 1 (DAPK1) in the regulation of the tau protein. We found that hippocampal DAPK1 expression is markedly increased in the brains of AD patients compared with age-matched normal subjects. DAPK1 overexpression increased tau protein stability and phosphorylation at multiple AD-related sites. In contrast, inhibition of DAPK1 by overexpression of a DAPK1 kinase-deficient mutant or by genetic knockout significantly decreased tau protein stability and abolished its phosphorylation in cell cultures and in mice. Mechanistically, DAPK1-enhanced tau protein stability was mediated by Ser71 phosphorylation of Pin1, a prolyl isomerase known to regulate tau protein stability, phosphorylation, and tau-related pathologies. In addition, inhibition of DAPK1 kinase activity significantly increased the assembly of MTs and accelerated nerve growth factor-mediated neurite outgrowth. Given that DAPK1 has been genetically linked to late onset AD, these results suggest that DAPK1 is a novel regulator of tau protein abundance, and that DAPK1 upregulation might contribute to tau-related pathologies in AD. Therefore, we offer that DAPK1 might be a novel therapeutic target for treating human AD and other tau-related pathologies.     

Acetylcholine receptors and tau phosphorylation

Alzheimer's disease (AD) is characterized by the presence, in the brain of the patients, of two aberrant structures: intracellular neurofibrillary tangles (NFTs), containing an abnormal hyperphosphorylated form of tau protein, and extracellular senile plaques (SPs), mainly composed by fibrillar amyloid beta peptide. Another feature of AD is the neurodegeneration and dysfunction of basal forebrain cholinergic system. A possible connection among those AD characteristics could occur. Thus, the purpose of this short review is to summarize the involvement of nicotinic (nAChR) and muscarinic (mAChR) receptors on tau phosphorylation, in a direct way, or through the previous interaction of some of these receptors with amyloid beta. Several studies have demonstrated that nAChR activation results in a significantly increase of tau phosphorylation, whereas mAChR activation, may prevent tau phosphorylation.     

Ⅱ型糖尿病大鼠海马Alzheimer病样Tau蛋白过度磷酸化修饰及机制探讨

Tau蛋白过度磷酸化是Alzheimer病 (Alzheimer′s disease, AD) 的一个重要特征.本研究检测了Ⅱ型糖尿病大鼠海马tau蛋白磷酸化水平,对其形成机制进行探讨. 以同龄正常Wistar大鼠作为对照,高脂高蛋白高糖饮食加小剂量链脲佐菌素（streptozotocin,STZ）注射诱导造Ⅱ型糖尿病模型（T2DM组）.放免法检测血浆胰岛素;葡萄糖氧化酶法检测血浆葡萄糖;蛋白质印迹技术检测各组大鼠海马内总tau蛋白、tau蛋白上部分位点磷酸化、神经细胞膜上胰岛素受体及葡萄糖转运子3（glucose transport 3,GLUT3）水平;表面等离子共振技术（surface plasmon resonance, SPR）检测细胞膜上胰岛素受体与血浆胰岛素结合力;γ32-P标记的ATP和特异性底物肽检测海马内胰岛素信号传导系统中的关键酶糖原合酶激酶-3β（glycogen synthase kinase-3β, GSK-3β）活性.结果显示,T2DM组血浆血糖、血浆胰岛素及运用HOMA-IR公式计算的胰岛素抵抗指数显著高于对照组.蛋白质印迹结果显示两组大鼠海马回总tau蛋白水平无差异;T2DM组中tau蛋白在Ser199、Thr212、Ser214、Thr217、Ser396及Ser422位点上的磷酸化水平均显著高于对照组;T2DM组海马神经细胞膜上胰岛素受体水平及与胰岛素结合的功能均显著低于对照组;GSK-3β活性检测结果显示,T2DM组大鼠模型海马回中GSK-3β活性明显增高.研究结果表明,Ⅱ型糖尿病中由于胰岛素抵抗导致GSK-3β激活从而出现AD样tau蛋白的过度磷酸化,葡萄糖代谢紊乱也可能在tau蛋白的过度磷酸化起一定作用.     

Developmental regulation of tau phosphorylation, tau kinases, and tau phosphatases

Tau is a neuronal microtubule-associated protein. Its hyperphosphorylation plays a critical role in Alzheimer disease (AD). Expression and phosphorylation of tau are regulated developmentally, but its dynamic regulation and the responsible kinases or phosphatases remain elusive. Here, we studied the developmental regulation of tau in rats during development from embryonic day 15 through the age of 24 months. We found that tau expression increased sharply during the embryonic stage and then became relatively stable, whereas tau phosphorylation was much higher in developing brain than in mature brain. However, the extent of tau phosphorylation at seven of the 14 sites studied was much less in developing brain than in AD brain. Tau phosphorylation during development matched the period of active neurite outgrowth in general. Tau phosphorylation at various sites had different topographic distributions. Several tau kinases appeared to regulate tau phosphorylation collectively at overlapping sites, and the decrease of overall tau phosphorylation in adult brain might be due to the higher levels of tau phosphatases in mature brain. These studies provide new insight into the developmental regulation of site-specific tau phosphorylation and identify the likely sites required for the abnormal hyperphosphorylation of tau in AD.