International Journal of Neuroscience, 2014; 124(8): 601–608Copyright © 2014 Informa Healthcare USA, Inc.ISSN: 0020-7454 print / 1543-5245 onlineDOI: 10.3109/00207454.2013.866110

RESEARCH ARTICLE

Streptozotocin-induced diabetes increases amyloidplaque deposition in AD transgenic mice throughmodulating AGEs/RAGE/NF-κB pathway

Xu Wang,1 Song Yu,1 Jiang-Ping Hu,2 Chun-Yan Wang,3 Yue Wang,1 Hai-Xing Liu,1

and Yu-Li Liu4

1Department of Histology and Embryology, Liaoning University of Traditional Chinese Medicine, Shenyang, P.R. China;2Department of Histology and Embryology, Mudanjiang Medical University, Mudanjiang, Mudanjiang, P.R. China;3Medical Research Laboratory, Jilin Medical College, Jilin, P.R. China; 4Acupuncture and Massage College, LiaoningUniversity of Traditional Chinese Medicine, Shenyang, P.R. China

Background: An increasing number of studies have demonstrated of that diabetes mellitus (DM) is associatedwith an increased prevalence of Alzheimer disease (AD), the underlying mechanisms are still obscure. Methods:We developed a streptozotocin (STZ)-induced diabetic AD transgenic mouse model and evaluated the effect ofhyperglycemia on senile plaque formation. Results: Our data showed that administration of STZ increased thelevel of blood glucose and increased the advanced glycation end products (AGEs) in brain tissue, and furtherenhanced the expression levels of the receptor for AGEs (RAGE) and the nuclear factor-kappa B (NF-κB) in thebrain, and accelerated the senile plaque formation in the transgenic mice. Our results showed that STZ-inducedinsulin-deficient hyperglycemia caused the pathophysiology of AD in APP/PS1 transgenic mice by modulatingthe AGEs/RAGE/NF-κB pathway. Conclusions: Our study suggests that there is a close linkage of DM andcerebral amyloidosis in the pathogenesis of AD.

KEYWORDS: diabetes mellitus, Alzheimer’s disease, β-amyloid, advanced glycation end products

Introduction

Alzheimer’s disease (AD) is a devastating and pro-gressive neurodegenerative disease clinically character-ized by progressive cognitive impairment. The majorpathological changes associated with AD are extracellu-lar senile plaques and intracellular neurofibrillary tan-gles (NFTs) in the brain. Diabetes mellitus (DM) isa metabolic disease in which a person has high bloodsugar, either because the body does not produce enoughinsulin or the body does not respond to the insulin thatis produced. Both AD and DM are two of the mostcommon degenerative diseases in the world. Both haveenormous impacts on patients’ quality of life leading to

Received 9 September 2013; revised 23 October 2013; accepted 12 November2013.

Correspondence: Xu Wang, Department of Histology and Embryology,Liaoning University of Traditional Chinese Medicine, Shenyang 110847, P.R.China. Tel: +86-24-31207087. Fax: +86-24-31207073. E-mail:wangxu@lnutcm.edu.cn

growing health care costs especially in the elderly. In-terestingly, recent clinical and epidemiological evidencesuggest that diabetic patients have a 30∼65% increasedrisk for developing AD [1]. Some experimental studieshave focused on insulin dysfunctions (insulin deficiencyor insulin resistance) as possible mechanisms linking ADand DM [2,3]. However, very few studies have exam-ined the direct effects of hyperglycemia, which is one ofthe most serious health conditions in DM patients, onthe development of AD [4]. Therefore, the underlyingmechanisms responsible for the effects of hyperglycemiaon AD pathogenesis remain largely unknown.

The chronic hyperglycemic status has been shown tofavor glycation reactions, leading to the formation of ad-vanced glycation end products (AGEs) [5]. AGEs areconsidered to play a key role in the pathogenesis of thechronic complications of DM. It was reported that theformation and accumulation of AGEs also contributedto diabetic neuropathic processes in DM [6]. Further-more, there is evidence for the involvement of AGEs inthe pathogenesis of AD. Immunohistochemical studies

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showed that AGEs were expressed in the senile plaquesand NFTs [7], and suggested that AGEs may be an im-portant factor in the progression of AD. So it stands toreason that the AGEs levels may be a possible linkingbetween DM and AD.

AGEs are able to bind to their cell surface receptor(RAGE) on neurons, microglia, astrocytes, and brainendothelial cells and trigger the RAGE-dependent NF-kB activation [8]. There is a growing body of evidenceto suggest that RAGE and NF-kB can participate in theamyloid-β precursor protein (APP) processing and Aβ

production, the primary component in the pathophysi-ology of AD [9,10].

The present study employs the APP/PS1 doubletransgenic mice expressing both the gene for humanAPP and presenilin 1 (PS1). Both mutations areassociated with early-onset AD. These mice show anage-dependent increase in the Aβ and in the number ofamyloid plaques in the brain. The experimental diabetesin the APP/PS1 transgenic mouse is induced by usingstreptozotocin (STZ), a naturally occurring chemicalthat has a particular toxicity toward pancreatic β cells.Our results indicate that the systemic hyperglycemiainduced by STZ exacerbates the production of AGEs,increases the expression of RAGE and NF-κB, andfurther accelerates the cerebral amyloidosis in theAPP/PS1 transgenic mice. The present data suggestthat STZ-induced insulin-deficient hyperglycemia canpromote the progression of AD through modulatingAGEs/RAGE/ NF-κB pathway.

Materials and Methods

Transgenic mice

All studies were performed using the male APP/PS1(APPswe/PSEN1dE9) double-transgenic mice. TheAPP/PS1 mice were obtained from the Jackson Labo-ratory (West Grove, PA, USA). The transgenic mousemodel was selected because of the senile plaque forma-tion in the brains of transgenic animals by six to sevenmonths of age. Throughout the experiments, the an-imals were kept in cages in a controlled environment(22–25 ◦C, 50% relative humidity, 12 h light/dark cy-cle) with free access to food and water. All proceduresusing animals were conducted in accordance with thecare and use of medical laboratory animals (Ministry ofHealth PR China, 1998) and the guidelines of the lab-oratory animal ethical standards of Liaoning Universityof Traditional Chinese Medicine.

Induction of diabetes and blood glucosemeasurement

Diabetes was induced in three-month-old APP/PS1 fol-lowing an overnight fast by intraperitoneal injection

of STZ (90 mg/kg, Sigma, dissolved in sodium cit-rate buffer) for consecutive two days. Age-matchedAPP/PS1 mice received citrate buffer only (vehicle con-trol). Mice with nonfasting blood glucose levels higherthan 15 mM/L one week after STZ injection were con-sidered to be diabetic and used in the later experiments[3]. The APP/PS1 mice were analyzed at 20 weeks afterSTZ or vehicle treatment. Blood samples were obtainedby tail prick and blood glucose levels were measured us-ing a portable blood glucose meter (Johnson, m211667,USA).

Immunohistochemistry

After 20 weeks of diabetes, mice were sacrificed by de-capitation. The hippocampus and cerebral cortex weredissected from the left hemisphere and were immedi-ately homogenized for protein extraction. The otherhalf of the brains was fixed in 4% paraformaldehydeand routine paraffin sections (7 μm) were preparedfor morphological analysis. The paraffin sections weredewaxed, rinsed, treated with 3% H2O2 for 10 min.Nonspecific epitopes were then blocked by incubationwith 5% normal goat serum for 30 min, and incubatedovernight with mouse monoclonal anti-β-Amyloid an-tibody (3.0 μg/ml, Sigma), rabbit polyclonal anti-AGEantibody (5.0 μg/ml, Biosynthesis Biotechnology), andrabbit monoclonal anti-NF-κB p65 antibody (1:200,Cell Signaling Technology) antibody at 4 ◦C. After rins-ing in 0.1 M phosphate-buffered saline (PBS) threetimes, sections were incubated with biotinylated goatanti-mouse/rabbit IgG (5.0 μg/ml) for 1 h, followed bystreptavidin peroxidase for 1 h at room temperature. Af-ter rinsing, the sections were treated with 0.025% 3,3-diaminobenzidine for 10 min. The sections were de-hydrated, cleared, covered, and examined under a lightmicroscope equipped with a digital camera (Olympus;AX70U-Photo, Japan).

The quantification of percentage area occupied byplaques and the NF-κB positive cell expression afterimmunostaining were analyzed using the image analy-sis system (Image-Pro Plus 6.0). For quantification, fivesections with the same reference position were selectedfrom each animal (n = 7/group) and processed for assay.In the Aβ staining sections, the area of the plaque, thecortex and hippocampus were quantified (in square mi-crometers). In the NF-κB immunohistochemistry sec-tions, brown nuclei were counted and analyzed in thecortex and hippocampus of APP/PS1 transgenic mice.

Western blotting

Protein lysates preparation and immunoblotting wereperformed as described previously [3]. Cerebral tissueshomogenates were centrifuged at 12 000 g for 30 min at

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4 ◦C. Total extract proteins (30–50 μg) were separatedon 10% SDS-PAGE bis-Tris gels and then transferredonto PVDF membranes. Nonspecific binding sites onthe membrane were blocked by 5% bovine serum albu-min in 0.1% TBS/Tween-20 (TBST) for 1 h, and thenincubated with a primary antibody overnight at 4 ◦C.After washing with TBST, the membranes were incu-bated with horseradish peroxide-conjugated second an-tibody (0.2 μg/ml, Santa Cruz Biotechnology) for 1 hat room temperature. Primary antibodies used includ-ing rabbit polyclonal anti-RAGE antibody (0.5 μg/ml,Abcam), rabbit monoclonal anti-NF-κB p65 antibody(1:500, Cell Signaling Technology), and mouse β-actin(0.04 μg/ml, Santa Cruz Biotechnology). Immunoreac-tive bands were visualized with an enhanced chemilumi-nescence kit (Pierce Biotechnology, Rockford, IL) andChem Doc XRS with Quantity One software (Bio-Rad;Hercules, CA, USA). The bands were scanned and theintensities of the bands were measured using Image-proPlus 6.0 analysis software.

ELISA

Cerebral tissues from transgenic mice were homoge-nized in 7.4 M PBS and centrifuged for 20 min at thespeed of 2000 g remove supernatant. The supernatantswere then loaded onto 48-well plates and the level ofAGEs was determined using an AGE indirect ELISAkit (Cell Biolabs Inc., San Diego, CA, USA) followingthe manufacturer’s instructions. The absorbance wasrecorded at 450 nm using a 48-well plate reader.

Statistical analysis

All values are expressed as mean ± standard deviation(SD). Differences between groups were analyzed by Stu-dent’s t test between STZ-induced diabetic AD modelmice and vehicle controls. All data were analyzed usingSPSS software, and differences were considered signifi-cant at p < 0.05.

Results

Hyperglycemia was induced by the STZtreatment

The body weight and blood glucose of the APP/PS1mice were monitored throughout the study. As shownin Table 1, STZ-treated APP/PS1 mice exhibited a sig-nificantly lower body weight compared with controls(21%), paralleled by a strong increase in blood glucose(259%) at the end of the research.

Table 1. Body weight and blood glucose of STZ-mice andcontrol.

Control STZ

Body weight (g) 26.4 ± 1.34 20.80 ± 1.48∗

Blood glucose (mM) 8.56 ± 0.81 30.74 ± 2.84∗∗

The values are given as mean ± SD.∗p < 0.05 vs. control.∗∗p < 0.01 vs. control.

Hyperglycemia enhances the formation ofAGEs in the APP/PS1 mouse brain

AGEs-positive granules were identified in theperikaryon of cortical neurons (Figure 1A). Therewere far more AGEs-positive granules in corticalneurons of STZ-treated APP/PS1 mice than controls.Furthermore, we measured the level of AGEs in thebrain of STZ-induced diabetic APP/PS1 mice bySandwich ELISA. Data showed that the level of AGEswas markedly increased in STZ-treated APP/PS1 micerelative to controls (0.38 ± 0.12 ng/mg vs. 1.21 ±0.22 ng/mg, p < 0.01) (Figure 1B).

Administration of STZ increases the levels ofRAGE

RAGE is a multiligand receptor of the immunoglobulinsuperfamily of cell surface molecules acting as counter-receptor for AGEs. Thus, we examined the protein levelsof RAGE in the STZ-induced diabetic APP/PS1 mouseand control. As shown in Figure 2, immunoblotting re-sults showed that the protein expression of RAGE wassignificantly increased by 85% in the STZ-treated dia-betic mouse brain, compared with vehicle control (Fig-ure 2B; p < 0.01).

The expression of NF-κB is increased inSTZ-induced APP/PS1 diabetic mice

The engagement of AGEs to RAGE leads to the activa-tion of NF-κB. A large number of data have shown thatNF-κB differently regulates Aβ production and Aβ con-centrations by modulating transactivations of APP andα/β/γ -secretase activity. To investigate the effect of STZtreatment on the NF-κB expression in the APP/PS1transgenic mouse brain, we performed NF-κB immuno-histochemical staining in the cortex and hippocampus ofAPP/PS1 mouse brain. We found that NF-κB was ex-pressed specifically inside the nucleus both in the cor-tex and hippocampus (Figure 3A). Figure 3B showedthat NF-kB positive neurons were higher in cortex of theSTZ-treated APP/PS1 mice compared with untreatedAPP/PS1 mice (p < 0.01). However, there was no

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Figure 1. Hyperglycemia promotes the formation and accumulation of AGEs in the STZ-induceddiabetic APP/PS1 mouse brain. (A) Representative AGEs-immuopositive granules in cortical neuronsfrom the STZ-induced diabetic APP/PS1 mouse and control. (B) The level of AGEs in brain tissue wasmeasured using an ELISA kit. STZ treatment significantly increased the levels of AGEs in APP/PS1transgenic mouse brain. ∗∗p < 0.01 (n = 7).

significant difference in NF-κB expression of hippocam-pus between control group and experimental group(Figure 3C). Consistent with the immunostaining, thewestern blot assay also showed that STZ treatment ofAPP/PS1 transgenic mice significantly increased NF-κBexpression compared to nontreatment mice (Figure 3Dand E; p < 0.05). These data suggest that STZ-induceddiabetes in APP/PS1 transgenic mice increases inflam-matory mediators in the brain.

Administration of STZ enhances senile plaqueformation in APP/PS1 mice

To determine whether hyperglycemia was increasingsenile plaque formation, tissue sections from vehicle-and STZ-treated APP/PS1 mouse brain were stainedwith an antibody against Aβ. As shown in Figure 4,STZ-treated mice displayed the expected numerous,large Aβ-immunoreactive senile plaques in the cortexand hippocampus compared with controls (Figure 4A).Quantification of extracellular Aβ deposits revealed that

Figure 2. Administration of STZ increases the levels of RAGE.(A) Western blots showing the expression levels of RAGE proteinsin APP/PS1 transgenic mouse brain 20 weeks after STZ adminis-tration. β-actin was used as a loading control. (B) STZ adminis-tration leads to a marked increase in the RAGE in the APP/PS1transgenic mouse brain. ∗∗p < 0.01 (n = 7).

the area occupied by senile plaques was significantly in-creased in brains of STZ-treated groups compared tothe vehicle-treated group. In the cortex, the area of se-nile plaques was increased by nearly 34% in the STZ-treated mouse brain (Figure 4B). In the hippocampus,STZ treatment showed significant increase of plaquearea by approximately 54%, as compared with vehicle-treated group (Figure 4C).

Discussion

During the last decade, epidemiological and clinicaldata have suggested that patients with DM have an in-creased risk of developing AD [11]. Some studies havedemonstrated that impaired insulin and insulin signal-ing transduction, which occur in both DM and AD, mayprovide a mechanistic link between the two disorders[12]. Our previous studies have shown that insulin defi-ciency induced impaired insulin signaling including ac-tivation of its downstream GSK3α/β and JNK pathways,and resulted in increased deposition of Aβ and acceler-ated plaque formation in the APP/PS1 transgenic mousebrain [3]. In addition to the impaired insulin action, hy-perglycemia is likely another factor to explain the linkagebetween DM and AD [4]. In the present study, a mousemodel of combined DM and AD was made by inject-ing APP/PS1 transgenic mice with STZ, a known toxinfor pancreatic insulin-producing β cells. We showed thatadministration of STZ markedly increased the level ofblood glucose and caused formation of senile plaques.Deposition of Aβ to form neuritic plaques in the brainis the pathological hallmark of AD [13]. Thus, we usedthis model to assess whether STZ-induced diabetes ex-acerbated cerebral amyloidosis, and to explore the un-derlying mechanisms.

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Figure 3. The expression of NF-κB is increased in STZ-inducedAPP/PS1 diabetic mice. (A) The NF-κB reactive cell was detectedin the cortex and hippocampus from vehicle- and STZ-treatedAPP/PS1 mice. Scale bar = 50 μm. (B) The number of NF-κB positive neurons was significantly increased in STZ treatedAPP/PS1 mice. (C) There were no significant differences in NF-κB positive neurons of the hippocampus between vehicle controlsand STZ-treated mice. (D and E) Western blots showed that STZ-induced diabetes lead to an increase in the expression levels ofNF-κB in the APP/PS1 mice. ∗p < 0.05, ∗∗p < 0.01 (n = 7).

In DM, enhanced blood glucose level leads to theformation of AGEs [14]. AGEs are produced throughthe nonenzymatic glycation as well as oxidation of pro-teins, lipids and nucleic acids. Accumulation of AGEsin cells and tissues is a normal feature of aging, and isaccelerated in AD [15,16]. A large body of evidence hasshown that accelerated AGEs can be also observed inAD and are believed to have a key role in the develop-ment and progression of AD [17]. In AD, several studiesshowed AGEs are localized in the senile plaques, extra-

cellular spaces and further more in NFTs [18,19] andhave been reported to have a direct effect on the Aβ de-position and senile plaque volume [20]. Specifically, for-mation of AGEs accelerates the conversion of Aβ frommonomeric to oligomeric or higher molecular weightforms [21]. Therefore, AGEs could explain many of theneuropathological and biochemical features of AD suchas extensive protein cross-linking, glial induction of ox-idative stress and neuronal cell death [15]. In our animalmodel, we observed that STZ-induced hyperglycemiaresulted in increased levels of AGEs in brain tissues ofAPP/PS1 transgenic mice, suggesting that AGEs forma-tion and accumulation in hyperglycemic state promotedthe formation of senile plaques.

One of the mechanisms of AGEs-mediated Aβ accu-mulation is through their cell surface receptor (RAGE).RAGE is a member of the immunoglobulin superfam-ily of cell surface molecules and engages diverse ligandsrelevant to distinct pathological processes [22]. Stud-ies have demonstrated that RAGE plays a central rolein AD. In AD brain, RAGE is evident in neurons, mi-croglia, astrocytes, and in brain endothelia cells [23].Leu showed that the levels of RAGE proteins were pos-itively correlated with the severity of the AD pathology[24]. RAGE apparently promoted the production of Aβ

in brain and the transport of Aβ from the blood to thebrain that lead to AD progression [9,25]. Furthermore,the engagement of AGEs to RAGE leads to the activa-tion of the NF-κB and triggers intracellular signalingpathways [26,27]. NF-κB has also been implicated indiverse biological processes and extremely detrimentalif its activities are incorrectly regulated [28,29]. NF-κBhas also been implicated in processes of synaptic plas-ticity and memory [30]. The involvement of NF-κB inthe pathophysiology of AD is one of the well-known ex-amples. Some studies have shown that NF-κB can par-ticipate in APP processing and Aβ production, playsan important role in the pathophysiology of AD. It hasbeen reported that NF-κB activates the transcriptionof βAPP, BACE1, and some of the γ -secretase mem-bers and increases protein expression and enzymatic ac-tivities, resulting in enhanced Aβ production [31–33].Consistently, our results showed that STZ-induced dia-betes could increase levels of AGEs, RAGE and NF-κBin APP/PS1 transgenic mice. It is thereafter conceivablethat STZ-induced diabetes can promote the formationand accumulation of senile plaque in AD through theAGEs/RAGE/NF-κB signaling pathway. However, wecannot rule out the possibilities that STZ treatment byitself promotes the formation and accumulation of senileplaque via other mechanisms relevant to AGEs/RAGEaxis.

The roles of RAGE and NF-κB on BACE1 upregu-lation were recently reported in animal models and cul-tured cells. BACE1 is an essential protease that cleaves

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Figure 4. Administration of STZ enhances senile plaque formation in APP/PS1 mice. (A) Twentyweeks after STZ administration, the senile plaques were detected using Aβ immunohistochemistry inthe cortex and hippocampus of transgenic mice. Scale bar = 50 μm. (B and C) Quantification of thepercentage area occupied by plaques in the cortex (B) and hippocampus (C). ∗p < 0.05, ∗∗p < 0.01(n = 7).

APP to generate Aβ peptides, a central component ofneuritic plaques in AD brains [34]. It was reported thatthe expression and activity of BACE1 were increasedin cells overexpressing RAGE and in RAGE-injectedbrains of Tg2576 mice [35]. In addition, the NF-κBactivates the transcription of BACE1 and results in en-hanced Aβ production [31,32]. We have previously re-ported the changes of BACE1 protein expression inour mouse model [3]. So, we supposed that activatedBACE1 could be the substrate of AGEs/RAGE/NF-κBpathway to result in the increased senile plaques in dia-betic APP/PS1 transgenic mice.

In summary, we have shown that hyperglycemia inSTZ-induced diabetic APP/PS1 transgenic mice canenhance the AGEs levels, and consequently promotesthe formation of senile plaques through their bind-

ing with RAGE followed by the activation of NF-κB.Taken together, our results demonstrate that activationof the AGEs/RAGE/NF-κB pathway, as a result of hy-perglycemia, may play vital roles in the formation of se-nile plaque and pathogenesis of AD. Thus, the upreg-ulation of AGEs/RAGE/NF-κB pathway in the diabeticbrain could be one of the novel molecular mechanismsunderlying the development of AD, providing a possiblemechanistic link between DM and AD.

Conclusions

Our data showed that administration of STZ increasedthe level of blood glucose and increased the AGEs inbrain tissue, and further enhanced the expression levels

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Streptozotocin-induced diabetes increases amyloid plaque deposition 607

of the RAGE and the NF-κB in the brain, and acceler-ated the senile plaque formation in the transgenic mice.The present data indicate that STZ-induced insulin-deficient hyperglycemia caused the pathophysiology ofAD in APP/PS1 transgenic mice by modulating theAGEs/RAGE/NF-κB pathway. Our study suggests thatthere is a close linkage of DM and cerebral amyloidosisin the pathogenesis of AD.

Declaration of Interest

The authors declare that they have no conflict of inter-ests. The authors alone are responsible for the contentand writing of this paper.

The study was supported by the Natural Sci-ence Foundation of China (81100810, 81203004, and81100808), the Postdoctoral Science Foundation ofChina (2012M510849), and the Technological AgencyFoundation of Liaoning Province (2011408004).

References

1. Arvanitakis Z, Wilson RS, Bienias JL, Evans DA, Bennett DA.Diabetes mellitus and risk of Alzheimer disease and decline incognitive function. Arch Neurol 2004;61:661–666.

2. Cao D, Lu H, Lewis TL, Li L. Intake of sucrose-sweetened wa-ter induces insulin resistance and exacerbates memory deficitsand amyloidosis in a transgenic mouse model of Alzheimer dis-ease. J Biol Chem 2007;282:36275–36282.

3. Wang X, Zheng W, Xie JW, Wang T, Wang SL, Teng WP, WangZY. Insulin deficiency exacerbates cerebral amyloidosis and be-havioral deficits in an Alzheimer transgenic mouse model. MolNeurodegener 2010;5:46–58.

4. Burdo JR, Chen Q, Calcutt NA, Schubert D. The pathologicalinteraction between diabetes and presymptomatic Alzheimer’sdisease. Neurobiol Aging 2009;30:1910–1917.

5. Podell BK, Ackart DF, Kirk NM, Eck SP, Bell C, BasarabaRJ. Non-diabetic hyperglycemia exacerbates disease severity inMycobacterium tuberculosis infected guinea pigs. PLoS One2012;7:e46824.

6. Wada R, Nishizawa Y, Yagihashi N, Takeuchi M, Ishikawa Y,Yasumura K, Nakano M, Yagihashi S. Effects of OPB-9195,anti-glycation agent, on experimental diabetic neuropathy. EurJ Clin Invest 2001;31:513–520.

7. Sasaki N, Toki S, Chowei H, Saito T, Nakano N, Hayashi Y,Takeuchi M, Makita Z. Immunohistochemical distribution ofthe receptor for advanced glycation end products in neurons andastrocytes in Alzheimer’s disease. Brain Res 2001;888:256–262.

8. Alexiou P, Chatzopoulou M, Pegklidou K, Demopoulos VJ.RAGE: a multi-ligand receptor unveiling novel insights in healthand disease. Curr Med Chem 2010;17:2232–2252.

9. Deane R, Du Yan S, Submamaryan RK, LaRue B, Jovanovic S,Hogg E, Welch D, Manness L, Lin C, Yu J, Zhu H, Ghiso J,Frangione B, Stern A, Schmidt AM, Armstrong DL, ArnoldB, Liliensiek B, Nawroth P, Hofman F, Kindy M, Stern D,Zlokovic B. RAGE mediates amyloid-beta peptide transportacross the blood-brain barrier and accumulation in brain. NatMed 2003;9:907–913.

10. Bourne KZ, Ferrari DC, Lange-Dohna C, Rossner S, WoodTG, Perez-Polo JR. Differential regulation of BACE1 pro-

moter activity by nuclear factor-kappaB in neurons and gliaupon exposure to beta-amyloid peptides. J Neurosci Res2007;85:1194–1204.

11. Xu W, Qiu C, Winblad B, Fratiglioni L. The effect of border-line diabetes on the risk of dementia and Alzheimer’s disease.Diabetes 2007;56:211–216.

12. Kroner Z. The relationship between Alzheimer’s disease and di-abetes. Type 3 diabetes? Altern Med Rev 2009;14:373–379.

13. Ly PT, Wu Y, Zou H, Wang R, Zhou W, Kinoshita A,Zhang M, Yang Y, Cai F, Woodgett J, Song W. Inhibitionof GSK3beta-mediated BACE1 expression reduces Alzheimer-associated phenotypes. J Clin Invest 2012;123:224–235.

14. Yamagishi S, Takeuchi M, Inagaki Y, Nakamura K, ImaizumiT. Role of advanced glycation end products (AGEs) and theirreceptor (RAGE) in the pathogenesis of diabetic microangiopa-thy. Int J Clin Pharmacol Res 2003;23:129–134.

15. Munch G, Deuther-Conrad W, Gasic-Milenkovic J. Glycox-idative stress creates a vicious cycle of neurodegenerationin Alzheimer’s disease–a target for neuroprotective treatmentstrategies? J Neural Transm Suppl 2002;62:303–307.

16. Odetti P, Rossi S, Monacelli F, Poggi A, Cirnigliaro M, FedericiM, Federici A. Advanced glycation end products and bone lossduring aging. Ann N Y Acad Sci 2005;1043:710–717.

17. Shuvaev VV, Laffont I, Serot JM, Fujii J, Taniguchi N, Siest G.Increased protein glycation in cerebrospinal fluid of Alzheimer’sdisease. Neurobiol Aging 2001;22:397–402.

18. Takeuchi M, Yamagishi S. Possible involvement of ad-vanced glycation end-products (AGEs) in the pathogene-sis of Alzheimer’s disease. Curr Pharm Des 2008;14:973–978.

19. Srikanth V, Maczurek A, Phan T, Steele M, Westcott B,Juskiw D, Munch G. Advanced glycation endproducts andtheir receptor RAGE in Alzheimer’s disease. Neurobiol Aging2009;32:763–777.

20. Kuhla B, Loske C, Garcia De Arriba S, Schinzel R, HuberJ, Munch G. Differential effects of “Advanced glycation end-products” and beta-amyloid peptide on glucose utilization andATP levels in the neuronal cell line SH-SY5Y. J Neural Transm2004;111:427–439.

21. Loske C, Gerdemann A, Schepl W, Wycislo M, Schinzel R,Palm D, Riederer P, Munch G. Transition metal-mediated gly-coxidation accelerates cross-linking of beta-amyloid peptide.Eur J Biochem 2000;267:4171–4178.

22. Yan SF, Ramasamy R, Schmidt AM. The RAGE axis: a funda-mental mechanism signaling danger to the vulnerable vascula-ture. Circ Res 2010;106:842–853.

23. Deane R, Bell RD, Sagare A, Zlokovic BV. Clearance ofamyloid-beta peptide across the blood-brain barrier. implicationfor therapies in Alzheimer’s disease. CNS Neurol Disord DrugTargets 2009;8:16–30.

24. Lue LF, Walker DG, Jacobson S, Sabbagh M. Receptor foradvanced glycation end products. its role in Alzheimer’s dis-ease and other neurological diseases. Future Neurol 2009;4:167–177.

25. Yan SD, Bierhaus A, Nawroth PP, Stern DM. RAGEand Alzheimer’s disease. a progression factor for amyloid-beta-induced cellular perturbation? J Alzheimers Dis 2009;16:833–843.

26. Li J, Schmidt AM. Characterization and functional analysis ofthe promoter of RAGE, the receptor for advanced glycation endproducts. J Biol Chem 1997;272:16498–16506.

27. Huttunen HJ, Fages C, Rauvala H. Receptor for advanced gly-cation end products (RAGE)-mediated neurite outgrowth andactivation of NF-kappaB require the cytoplasmic domain of thereceptor but different downstream signaling pathways. J BiolChem 1999;274:19919–19924.

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ealth

care

.com

by

Lin

kopi

ngs

Uni

vers

ity o

n 08

/19/

14Fo

r pe

rson

al u

se o

nly.

608 X. Wang et al.

28. Kabe Y, Ando K, Hirao S, Yoshida M, Handa H. Redox reg-ulation of NF-kappaB activation. distinct redox regulation be-tween the cytoplasm and the nucleus. Antioxid Redox Signal2005;7:395–403.

29. Smale ST. Hierarchies of NF-kappaB target-gene regulation.Nat Immunol 2011;12:689–694.

30. Kaltschmidt B, Ndiaye D, Korte M, Pothion S, Arbibe L, Prul-lage M, Pfeiffer J, Lindecke A, Staiger V, Israel A, KaltschmidtC, Memet S. NF-kappaB regulates spatial memory formationand synaptic plasticity through protein kinase A/CREB signal-ing. Mol Cell Biol 2006;26:2936–2946.

31. Guglielmotto M, Aragno M, Tamagno E, Vercellinatto I,Visentin S, Medana C, Catalano MG, Smith MA, Perry G,Danni O, Boccuzzi G, Tabaton M. AGEs/RAGE complex up-regulates BACE1 via NF-kappaB pathway activation. NeurobiolAging 2010;33:196 e113–127.

32. Chen CH, Zhou W, Liu S, Deng Y, Cai F, Tone M, Tone Y,Tong Y, Song W. Increased NF-kappaB signalling up-regulatesBACE1 expression and its therapeutic potential in Alzheimer’sdisease. Int J Neuropsychopharmacol 2011;1–14.

33. Chami L, Buggia-Prevot V, Duplan E, Delprete D, ChamiM, Peyron JF, Checler F. Nuclear factor-kappaB regulates be-taAPP and beta- and gamma-secretases differently at physiolog-ical and supraphysiological Abeta concentrations. J Biol Chem2012;287:24573–24584.

34. Zhou W, Cai F, Li Y, Yang GS, O’Connor KD, Holt RA, SongW. BACE1 gene promoter single-nucleotide polymorphisms inAlzheimer’s disease. J Mol Neurosci 2010;42:127–133.

35. Cho HJ, Son SM, Jin SM, Hong HS, Shin DH, Kim SJ, Huh K,Mook-Jung I. RAGE regulates BACE1 and Abeta generation viaNFAT1 activation in Alzheimer’s disease animal model. FASEBJ 2009;23:2639–2649.

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