MicroRNA‐181a restricts human γδ T cell differentiation by targeting Map3k2 and Notch2

γδ T cells are a conserved population of lymphocytes that contributes to anti‐tumor responses through its overt type 1 inflammatory and cytotoxic properties. We have previously shown that human γδ T cells acquire this profile upon stimulation with IL‐2 or IL‐15, in a differentiation process dependent on MAPK/ERK signaling. Here, we identify microRNA‐181a as a key modulator of human γδ T cell differentiation. We observe that miR‐181a is highly expressed in patients with prostate cancer and that this pattern associates with lower expression of NKG2D, a critical mediator of cancer surveillance. Interestingly, miR‐181a expression negatively correlates with an activated type 1 effector profile obtained from in vitro differentiated γδ T cells and miR‐181a overexpression restricts their levels of NKG2D and TNF‐α. Upon in silico analysis, we identify two miR‐181a candidate targets, Map3k2 and Notch2, which we validate via overexpression coupled with luciferase assays. These results reveal a novel role for miR‐181a as critical regulator of human γδ T cell differentiation and highlight its potential for manipulation of γδ T cells in next‐generation immunotherapies.     

Osteonecrosis of the Jaw Developed in Mice: DISEASE VARIANTS REGULATED BY γδ T CELLS IN ORAL MUCOSAL BARRIER IMMUNITY*

Osteonecrosis of the jaw (ONJ), an uncommon co-morbidity in patients treated with bisphosphonates (BP), occurs in the segment of jawbone interfacing oral mucosa. This study aimed to investigate a role of oral mucosal barrier γδ T cells in the pathogenesis of ONJ. Female C57Bl/6J (B6) mice received a bolus zoledronate intravenous injection (ZOL, 540 μg/kg), and their maxillary left first molars were extracted 1 week later. ZOL-treated mice (WT ZOL) delayed oral wound healing with patent open wounds 4 weeks after tooth extraction with characteristic oral epithelial hyperplasia. γδ T cells appeared within the tooth extraction site and hyperplastic epithelium in WT ZOL mice. In ZOL-treated γδ T cell null (Tcrd^−/− ZOL) mice, the tooth extraction open wound progressively closed; however, histological ONJ-like lesions were identified in 75 and 60% of WT ZOL and Tcrd^−/− ZOL mice, respectively. Although the bone exposure phenotype of ONJ was predominantly observed in WT ZOL mice, Tcrd^−/− ZOL mice developed the pustule/fistula disease phenotype. We further addressed the role of γδ T cells from human peripheral blood (h-γδ T cells). When co-cultured with ZOL-pretreated human osteoclasts in vitro, h-γδ T cells exhibited rapid expansion and robust IFN-γ secretion. When h-γδ T cells were injected into ZOL-treated immunodeficient (Rag2^−/− ZOL) mice, the oral epithelial hyperplasia developed. However, Rag2^−/− ZOL mice did not develop osteonecrosis. The results indicate that γδ T cells are unlikely to influence the core osteonecrosis mechanism; however, they may serve as a critical modifier contributing to the different oral mucosal disease variations of ONJ.     

Induction of γδT cells from HSC‐enriched BMCs co‐cultured with iPSC‐derived thymic epithelial cells

T cells bearing γδ antigen receptors have been investigated as potential treatments for several diseases, including malignant tumours. However, the clinical application of γδT cells has been hampered by their relatively low abundance in vivo and the technical difficulty of inducing their differentiation from hematopoietic stem cells (HSCs) in vitro. Here, we describe a novel method for generating mouse γδT cells by co‐culturing HSC‐enriched bone marrow cells (HSC‐eBMCs) with induced thymic epithelial cells (iTECs) derived from induced pluripotent stem cells (iPSCs). We used BMCs from CD45.1 congenic C57BL/6 mice to distinguish them from iPSCs, which expressed CD45.2. We showed that HSC‐eBMCs and iTECs cultured with IL‐2 + IL‐7 for up to 21 days induced CD45.1⁺ γδT cells that expressed a broad repertoire of Vγ and Vδ T‐cell receptors. Notably, the induced lymphocytes contained few or no αβT cells, NK1.1⁺ natural killer cells, or B220⁺ B cells. Adoptive transfer of the induced γδT cells to leukemia‐bearing mice significantly reduced tumour growth and prolonged mouse survival with no obvious side effects, such as tumorigenesis and autoimmune diseases. This new method suggests that it could also be used to produce human γδT cells for clinical applications.     

p87 and p101 Subunits Are Distinct Regulators Determining Class IB Phosphoinositide 3-Kinase (PI3K) Specificity

Class I_B phosphoinositide 3-kinase γ (PI3Kγ) comprises a single catalytic p110γ subunit, which binds to two non-catalytic subunits, p87 or p101, and controls a plethora of fundamental cellular responses. The non-catalytic subunits are assumed to be redundant adaptors for Gβγ enabling G-protein-coupled receptor-mediated regulation of PI3Kγ. Growing experimental data provide contradictory evidence. To elucidate the roles of the non-catalytic subunits in determining the specificity of PI3Kγ, we tested the impact of p87 and p101 in heterodimeric p87-p110γ and p101-p110γ complexes on the modulation of PI3Kγ activity in vitro and in living cells. RT-PCR, biochemical, and imaging data provide four lines of evidence: (i) specific expression patterns of p87 and p101, (ii) up-regulation of p101, providing the basis to consider p87 as a protein forming a constitutively and p101 as a protein forming an inducibly expressed PI3Kγ, (iii) differences in basal and stimulated enzymatic activities, and (iv) differences in complex stability, all indicating apparent diversity within class I_B PI3Kγ. In conclusion, expression and activities of PI3Kγ are modified differently by p87 and p101 in vitro and in living cells, arguing for specific regulatory roles of the non-catalytic subunits in the differentiation of PI3Kγ signaling pathways.     

Characterization of a Novel α-Conotoxin from Conus textile That Selectively Targets α6/α3β2β3 Nicotinic Acetylcholine Receptors

α6β2 Nicotinic acetylcholine receptors (nAChRs) expressed by dopaminergic neurons in the CNS are potential therapeutic targets for the treatment of several neuropsychiatric diseases, including nicotine addiction and Parkinson disease. However, recent studies indicate that the α6 subunit can also associate with the β4 subunit to form α6β4 nAChRs that are difficult to pharmacologically distinguish from α6β2, α3β4, and α3β2 subtypes. The current study characterized a novel 16-amino acid α-conotoxin (α-CTx) TxIB from Conus textile whose sequence is GCCSDPPCRNKHPDLC-amide as deduced from gene cloning. The peptide and an analog with an additional C-terminal glycine were chemically synthesized and tested on rat nAChRs heterologously expressed in Xenopus laevis oocytes. α-CTx TxIB blocked α6/α3β2β3 nAChR with an IC₅₀ of 28 nm. In contrast, the peptide showed little or no block of other tested subtypes at concentrations up to 10 μm. The three-dimensional solution structure of α-CTx TxIB was determined using NMR spectroscopy. α-CTx TxIB represents a uniquely selective ligand for probing the structure and function of α6β2 nAChRs.     

Isolation and Ex Vivo Culture of Vδ1+CD4+γδ T Cells,an Extrathymic αβT-cell Progenitor

The thymus, the primary organ for the generation of αβ T cells and backbone of the adaptive immune system in vertebrates, has long been considered as the only source of αβT cells. Yet, thymic involution begins early in life leading to a drastically reduced output of naïve αβT cells into the periphery. Nevertheless, even centenarians can build immunity against newly acquired pathogens. Recent research suggests extrathymic αβT cell development, however our understanding of pathways that may compensate for thymic loss of function are still rudimental. γδ T cells are innate lymphocytes that constitute the main T-cell subset in the tissues. We recently ascribed a so far unappreciated outstanding function to a γδ T cell subset by showing that the scarce entity of CD4⁺ Vδ1⁺γδ T cells can transdifferentiate into αβT cells in inflammatory conditions. Here, we provide the protocol for the isolation of this progenitor from peripheral blood and its subsequent cultivation. Vδ1 cells are positively enriched from PBMCs of healthy human donors using magnetic beads, followed by a second step wherein we target the scarce fraction of CD4⁺ cells with a further magnetic labeling technique. The magnetic force of the second labeling exceeds the one of the first magnetic label, and thus allows the efficient, quantitative and specific positive isolation of the population of interest. We then introduce the technique and culture condition required for cloning and efficiently expanding the cells and for identification of the generated clones by FACS analysis. Thus, we provide a detailed protocol for the purification, culture and ex vivo expansion of CD4⁺ Vδ1⁺γδ T cells. This knowledge is prerequisite for studies that relate to this αβT cell progenitor`s biology and for those who aim to identify the molecular triggers that are involved in its transdifferentiation.     

Key Residues in the Nicotinic Acetylcholine Receptor β2 Subunit Contribute to α-Conotoxin LvIA Binding

α-Conotoxin LvIA (α-CTx LvIA) is a small peptide from the venom of the carnivorous marine gastropod Conus lividus and is the most selective inhibitor of α3β2 nicotinic acetylcholine receptors (nAChRs) known to date. It can distinguish the α3β2 nAChR subtype from the α6β2* (* indicates the other subunit) and α3β4 nAChR subtypes. In this study, we performed mutational studies to assess the influence of residues of the β2 subunit versus those of the β4 subunit on the binding of α-CTx LvIA. Although two β2 mutations, α3β2[F119Q] and α3β2[T59K], strongly enhanced the affinity of LvIA, the β2 mutation α3β2[V111I] substantially reduced the binding of LvIA. Increased activity of LvIA was also observed when the β2-T59L mutant was combined with the α3 subunit. There were no significant difference in inhibition of α3β2[T59I], α3β2[Q34A], and α3β2[K79A] nAChRs when compared with wild-type α3β2 nAChR. α-CTx LvIA displayed slower off-rate kinetics at α3β2[F119Q] and α3β2[T59K] than at the wild-type receptor, with the latter mutant having the most pronounced effect. Taken together, these data provide evidence that the β2 subunit contributes to α-CTx LvIA binding and selectivity. The results demonstrate that Val¹¹¹ is critical and facilitates LvIA binding; this position has not previously been identified as important to binding of other 4/7 framework α-conotoxins. Thr⁵⁹ and Phe¹¹⁹ of the β2 subunit appear to interfere with LvIA binding, and their replacement by the corresponding residues of the β4 subunit leads to increased affinity.     

Recruitment of the γ-Tubulin Ring Complex to Drosophila Salt-stripped Centrosome Scaffolds

Extracting isolated Drosophila centrosomes with 2 M KI generates salt-resistant scaffolds that lack the centrosomal proteins CP190, CP60, centrosomin, and γ-tubulin. To clarify the role of these proteins in microtubule nucleation by centrosomes and to identify additional centrosome components required for nucleation, we have developed an in vitro complementation assay for centrosome function. Centrosome aster formation is reconstituted when these inactive, salt-stripped centrosome scaffolds are supplemented with a soluble fraction of a Drosophila embryo extract. The CP60 and CP190 can be removed from this extract without effect, whereas removing the γ-tubulin destroys the complementing activity. Consistent with these results, we find no evidence that these three proteins form a complex together. Instead, γ-tubulin is found in two distinct protein complexes of 240,000 and ∼3,000,000 D. The larger complex, which is analogous to the Xenopus γ-tubulin ring complex (γTuRC) (Zheng, Y., M.L. Wong, B. Alberts, and T. Mitchison. 1995. Nature. 378:578–583), is necessary but not sufficient for complementation. An additional factor found in the extract is required. These results provide the first evidence that the γTuRC is required for microtubule nucleation at the centrosome.     

Characterization and structural analyses of a novel glycosyltransferase acting on the β-1,2-glucosidic linkages

The IALB_1185 protein, which is encoded in the gene cluster for endo-β-1,2-glucanase homologs in the genome of Ignavibacterium album, is a glycoside hydrolase family (GH) 35 protein. However, most known GH35 enzymes are β-galactosidases, which is inconsistent with the components of this gene cluster. Thus, IALB_1185 is expected to possess novel enzymatic properties. Here, we showed using recombinant IALB_1185 that this protein has glycosyltransferase activity toward β-1,2-glucooligosaccharides, and that the kinetic parameters for β-1,2-glucooligosaccharides are not within the ranges for general GH enzymes. When various aryl- and alkyl-glucosides were used as acceptors, glycosyltransfer products derived from these acceptors were subsequently detected. Kinetic analysis further revealed that the enzyme has wide aglycone specificity regardless of the anomer, and that the β-1,2-linked glucose dimer sophorose is an appropriate donor. In the complex of wild-type IALB_1185 with sophorose, the electron density of sophorose was clearly observed at subsites −1 and +1, whereas in the E343Q mutant–sophorose complex, the electron density of sophorose was clearly observed at subsites +1 and +2. This observation suggests that binding at subsites −1 and +2 competes through Glu102, which is consistent with the preference for sophorose as a donor and unsuitability of β-1,2-glucooligosaccharides as acceptors. A pliable hydrophobic pocket that can accommodate various aglycone moieties was also observed in the complex structures with various glucosides. Overall, our biochemical and structural data are indicative of a novel enzymatic reaction. We propose that IALB_1185 be redefined β-1,2-glucooligosaccharide:d-glucoside β-d-glucosyltransferase as a systematic name and β-1,2-glucosyltransferase as an accepted name.     

A Role for the αvβ3 Integrin in the Transmigration of Monocytes

The β2 integrins and intercellular adhesion molecule-1 (ICAM-1) are important for monocyte migration through inflammatory endothelium. Here we demonstrate that the integrin αvβ3 is also a key player in this process. In an in vitro transendothelial migration assay, monocytes lacking β3 integrins revealed weak migratory ability, whereas monocytes expressing β3 integrins engaged in stronger migration. This migration could be partially blocked by antibodies against the integrin chains αL, β2, αv, or IAP, a protein functionally associated with αvβ3 integrin. Transfection of β3 integrin chain cDNA into monocytes lacking β3 integrins resulted in expression of the αvβ3 integrin and conferred on these cells an enhanced ability to transmigrate through cell monolayers expressing ICAM-1. These monocytes also engaged in αLβ2-dependent locomotion on recombinant ICAM-1 which was enhanced by αvβ3 integrin occupancy. Antibodies against IAP were able to revert this αvβ3 integrin-dependent cell locomotion to control levels. Finally, adhesion assays revealed that occupancy of αvβ3 integrin could decrease monocyte binding to ICAM-1.In conclusion, we show that αvβ3 integrin modulates αLβ2 integrin-dependent monocyte adhesion to and migration on ICAM-1. This could represent a novel mechanism to promote monocyte motility on vascular ICAM-1 and initiate subsequent transendothelial migration.     

Moieties of Complement iC3b Recognized by the I-domain of Integrin αXβ2

Complement fragment iC3b serves as a major opsonin for facilitating phagocytosis via its interaction with complement receptors CR3 and CR4, also known by their leukocyte integrin family names, αMβ2 and αXβ2, respectively. Although there is general agreement that iC3b binds to the αM and αX I-domains of the respective β2-integrins, much less is known regarding the regions of iC3b contributing to the αX I-domain binding. In this study, using recombinant αX I-domain, as well as recombinant fragments of iC3b as candidate binding partners, we have identified two distinct binding moieties of iC3b for the αX I-domain. They are the C3 convertase-generated N-terminal segment of the C3b α’-chain (α’NT) and the factor I cleavage-generated N-terminal segment in the CUBf region of α-chain. Additionally, we have found that the CUBf segment is a novel binding moiety of iC3b for the αM I-domain. The CUBf segment shows about a 2-fold higher binding activity than the α’NT for αX I-domain. We also have shown the involvement of crucial acidic residues on the iC3b side of the interface and basic residues on the I-domain side.     

Glycogen synthase kinase 3β promotes osteosarcoma invasion and migration via regulating PTEN and phosphorylation of focal adhesion kinase

Aim: Typical features of human osteosarcoma are highly invasive and migratory capacities. Our study aimed to investigate the roles of glycogen synthase kinase 3β (GSK3β) in human osteosarcoma metastasis.Methods: GSK3β expressions in clinical osteosarcoma tissues with or without metastasis were examined by immunohistochemical staining. The expressions of GSK3β, p-GSK3β^Ser9, and p-GSK3β^Tyr216 in human osteoblast cells (hFOB1.19) and human osteosarcoma cells (MG63, SaOS-2, and U2-OS) were detected by Western blotting. The GSK3β activity was measured by non-radio isotopic in vitro kinase assay. Migration and invasion abilities of MG-63 cells treated with small-molecular GSK3β inhibitors were respectively examined by monolayer-based wound-healing assay and transwell assay. The mRNA expressions of GSK3β, matrix metalloproteinase-2 (MMP-2), MMP-9, phosphatase with tensin homology (PTEN), and focal adhesion kinase (FAK) were detected after siRNA transfection for 72 h. Meanwhile, protein expressions of GSK3β, FAK, p-FAK^Y397, PTEN, MMP-2, and MMP-9 were measured by Western blotting.Results: Clinical osteosarcoma tissues with metastasis showed higher GSK3β expressions. MG63 and U2-OS cells that were easy to occur metastasis showed significantly higher expressions and activities of GSK3β than SaOS-2 cells. Inhibition of GSK3β with small-molecular GSK3β inhibitors in MG63 cells significantly attenuated cell migration and invasion. These effects were associated with reduced expressions of MMP-2 and MMP-9. Moreover, increased PTEN and decreased p-FAK^Y397 expressions were observed following GSK3β knockdown by siRNA transfection. Conclusion: GSK3β might promote osteosarcoma invasion and migration via pathways associated with PTEN and phosphorylation of FAK.     

Recombinant disintegrin targets α(v) β(3) integrin and leads to mediator production

Integrin αvβ3 is most likely the foremost modulator of angiogenesis among all known integrins. Recombinant disintegrin DisBa-01, originally obtained from snake venom glands, binds to αvβ3, thereby significantly inhibiting adhesion and generating in vivo anti-metastatic ability. However, its function in mediator production is not clear. Here, we observed that the mediators VEGF-A, IL-8, and TGF-β are not produced by human umbilical vein endothelial cells (HUVEC cell line) or monocyte/macrophage cells (SC cell line) when cells adhered to vitronectin. However, when exposed to DisBa-01, HUVECs produced higher levels of TGF-β, and SC cells produced higher levels of VEGF-A. Nonetheless, HUVECs also showed an enhancement of apoptosis after losing adherence when exposed to disintegrin, which is a characteristic of anoikis. We propose that disintegrin DisBa-01 could be used to modulate integrin αvβ3 functions.     

High temperature promotes amyloid β-protein production and γ-secretase complex formation via Hsp90

Alzheimer''s disease (AD) is characterized by neuronal loss and accumulation of β-amyloid-protein (Aβ) in the brain parenchyma. Sleep impairment is associated with AD and affects about 25–40% of patients in the mild-to-moderate stages of the disease. Sleep deprivation leads to increased Aβ production; however, its mechanism remains largely unknown. We hypothesized that the increase in core body temperature induced by sleep deprivation may promote Aβ production. Here, we report temperature-dependent regulation of Aβ production. We found that an increase in temperature, from 37 °C to 39 °C, significantly increased Aβ production in amyloid precursor protein-overexpressing cells. We also found that high temperature (39 °C) significantly increased the expression levels of heat shock protein 90 (Hsp90) and the C-terminal fragment of presenilin 1 (PS1-CTF) and promoted γ-secretase complex formation. Interestingly, Hsp90 was associated with the components of the premature γ-secretase complex, anterior pharynx-defective-1 (APH-1), and nicastrin (NCT) but was not associated with PS1-CTF or presenilin enhancer-2. Hsp90 knockdown abolished the increased level of Aβ production and the increased formation of the γ-secretase complex at high temperature in culture. Furthermore, with in vivo experiments, we observed increases in the levels of Hsp90, PS1-CTF, NCT, and the γ-secretase complex in the cortex of mice housed at higher room temperature (30 °C) compared with those housed at standard room temperature (23 °C). Our results suggest that high temperature regulates Aβ production by modulating γ-secretase complex formation through the binding of Hsp90 to NCT/APH-1.     

A Conformational Change of the γ Subunit Indirectly Regulates the Activity of Cyanobacterial F1-ATPase

The central shaft of the catalytic core of ATP synthase, the γ subunit consists of a coiled-coil structure of N- and C-terminal α-helices, and a globular domain. The γ subunit of cyanobacterial and chloroplast ATP synthase has a unique 30–40-amino acid insertion within the globular domain. We recently prepared the insertion-removed α₃β₃γ complex of cyanobacterial ATP synthase (Sunamura, E., Konno, H., Imashimizu-Kobayashi, M., and Hisabori, T. (2010) Plant Cell Physiol. 51, 855–865). Although the insertion is thought to be located in the periphery of the complex and far from catalytic sites, the mutant complex shows a remarkable increase in ATP hydrolysis activity due to a reduced tendency to lapse into ADP inhibition. We postulated that removal of the insertion affects the activity via a conformational change of two central α-helices in γ. To examine this hypothesis, we prepared a mutant complex that can lock the relative position of two central α-helices to each other by way of a disulfide bond formation. The mutant obtained showed a significant change in ATP hydrolysis activity caused by this restriction. The highly active locked complex was insensitive to N-dimethyldodecylamine-N-oxide, suggesting that the complex is resistant to ADP inhibition. In addition, the lock affected ϵ inhibition. In contrast, the change in activity caused by removal of the γ insertion was independent from the conformational restriction of the central axis component. These results imply that the global conformational change of the γ subunit indirectly regulates complex activity by changing both ADP inhibition and ϵ inhibition.     

c-Abl kinase-mediated phosphorylation of γ-tubulin promotes γ-tubulin ring complexes assembly and microtubule nucleation

Cytoskeletal microtubules (MTs) are nucleated from γ-tubulin ring complexes (γTuRCs) located at MT organizing centers (MTOCs), such as the centrosome. However, the exact regulatory mechanism of γTuRC assembly is not fully understood. Here, we showed that the nonreceptor tyrosine kinase c-Abl was associated with and phosphorylated γ-tubulin, the essential component of the γTuRC, mainly on the Y443 residue by in vivo (immunofluorescence and immunoprecipitation) or in vitro (surface plasmon resonance) detection. We further demonstrated that phosphorylation deficiency significantly impaired γTuRC assembly, centrosome construction, and MT nucleation. c-Abl/Arg deletion and γ-tubulin Y443F mutation resulted in an abnormal morphology and compromised spindle function during mitosis, eventually causing uneven chromosome segregation. Our findings reveal that γTuRC assembly and nucleation function are regulated by Abl kinase-mediated γ-tubulin phosphorylation, revealing a fundamental mechanism that contributes to the maintenance of MT function.     

The thymic cortical epithelium determines the TCR repertoire of IL-17-producing γδT cells

The thymus provides a specialized microenvironment in which distinct subsets of thymic epithelial cells (TECs) support T-cell development. Here, we describe the significance of cortical TECs (cTECs) in T-cell development, using a newly established mouse model of cTEC deficiency. The deficiency of mature cTECs caused a massive loss of thymic cellularity and impaired the development of αβT cells and invariant natural killer T cells. Unexpectedly, the differentiation of certain γδT-cell subpopulations—interleukin-17-producing Vγ4 and Vγ6 cells—was strongly dysregulated, resulting in the perturbation of γδT-mediated inflammatory responses in peripheral tissues. These findings show that cTECs contribute to the shaping of the TCR repertoire, not only of “conventional” αβT cells but also of inflammatory “innate” γδT cells.     

Hyperphosphorylation of Tau Induced by Naturally Secreted Amyloid-β at Nanomolar Concentrations Is Modulated by Insulin-dependent Akt-GSK3β Signaling Pathway

Alzheimer disease (AD) is neuropathologically characterized by the formation of senile plaques from amyloid-β (Aβ) and neurofibrillary tangles composed of phosphorylated Tau. Although there is growing evidence for the pathogenic role of soluble Aβ species in AD, the major question of how Aβ induces hyperphosphorylation of Tau remains unanswered. To address this question, we here developed a novel cell coculture system to assess the effect of extracellular Aβ at physiologically relevant levels naturally secreted from donor cells on the phosphorylation of Tau in recipient cells. Using this assay, we demonstrated that physiologically relevant levels of secreted Aβ are sufficient to cause hyperphosphorylation of Tau in recipient N2a cells expressing human Tau and in primary culture neurons. This hyperphosphorylation of Tau is inhibited by blocking Aβ production in donor cells. The expression of familial AD-linked PSEN1 mutants and APP ΔE693 mutant that induce the production of oligomeric Aβ in donor cells results in a similar hyperphosphorylation of Tau in recipient cells. The mechanism underlying the Aβ-induced Tau hyperphosphorylation is mediated by the impaired insulin signal transduction because we demonstrated that the phosphorylation of Akt and GSK3β upon insulin stimulation is less activated under this condition. Treating cells with the insulin-sensitizing drug rosiglitazone, a peroxisome proliferator-activated receptor γ agonist, attenuates the Aβ-dependent hyperphosphorylation of Tau. These findings suggest that the disturbed insulin signaling cascade may be implicated in the pathways through which soluble Aβ induces Tau phosphorylation and further support the notion that correcting insulin signal dysregulation in AD may offer a potential therapeutic approach.