Please wait a minute...
 Home  About the Journal Editorial Board Aims & Scope Peer Review Policy Subscription Contact us
Early Edition  //  Current Issue  //  Open Special Issues  //  Archives  //  Most Read  //  Most Downloaded  //  Most Cited
Aging and disease    2020, Vol. 11 Issue (3) : 547-558     DOI: 10.14336/AD.2019.0715
Orginal Article |
Neuroprotective Effects of Endogenous Secretory Receptor for Advanced Glycation End-products in Brain Ischemia
Shimizu Yu1,2, Harashima Ai1, Munesue Seiichi1, Oishi Masahiro1,2, Hattori Tsuyoshi3, Hori Osamu3, Kitao Yasuko3, Yamamoto Hiroshi1,4, Leerach Nontaphat1, Nakada Mitsutoshi2, Yamamoto Yasuhiko1,*, Hayashi Yasuhiko2,5
1Department of Biochemistry and Molecular Vascular Biology,
2Department of Neurosurgery and
3Department of Neuroanatomy, Kanazawa University Graduate School of Medical Sciences, Kanazawa 920-8641, Japan.
4Komatsu University, Komatsu, Ishikawa 923-8511, Japan.
5Department of Neurosurgery, Kanazawa Medical University, Uchinada 920-0293, Japan.
Download: PDF(847 KB)   HTML
Export: BibTeX | EndNote | Reference Manager | ProCite | RefWorks    

The receptor for advanced glycation end-products (RAGE) is expressed on human brain endothelial cells (HBEC) and is implicated in neuronal cell death after ischemia. We report that endogenous secretory RAGE (esRAGE) is a splicing variant form of RAGE that functions as a decoy against ischemia-induced neuronal cell damage. This study demonstrated that esRAGE was associated with heparan sulphate proteoglycans on HBEC. The parabiotic experiments between human esRAGE overexpressing transgenic (Tg), RAGE knockout (KO), and wild-type (WT) mice revealed a significant neuronal cell damage in the CA1 region of the WT side of parabiotic WT→WT mice, but not of Tg→WT mice, 7 days after bilateral common carotid artery occlusion. Human esRAGE was detected around the CA1 neurons in the WT side of the parabiotic Tg→WT pair, but not in the KO side of the Tg→KO pair. To elucidate the dynamic transfer of esRAGE into the brain, we used the blood-brain barrier (BBB) system (PharmaCo-Cell) with or without RAGE knockdown in endothelial cells. A RAGE-dependent transfer of esRAGE was demonstrated from the vascular to the brain side. These findings suggested that esRAGE is associated with heparan sulphate proteoglycans and is transferred into the brain via BBB to exert its neuroprotective effects in ischemia.

Keywords Receptor for advanced glycation end-products (RAGE)      endogenous secretory RAGE (esRAGE)      parabiosis      delayed neuronal cell damage      blood-brain barrier     
Corresponding Authors: Yamamoto Yasuhiko   
About author:

These authors shared first-authorship.

Just Accepted Date: 26 July 2019   Issue Date: 13 May 2020
E-mail this article
E-mail Alert
Articles by authors
Shimizu Yu
Harashima Ai
Munesue Seiichi
Oishi Masahiro
Hattori Tsuyoshi
Hori Osamu
Kitao Yasuko
Yamamoto Hiroshi
Leerach Nontaphat
Nakada Mitsutoshi
Yamamoto Yasuhiko
Hayashi Yasuhiko
Cite this article:   
Shimizu Yu,Harashima Ai,Munesue Seiichi, et al. Neuroprotective Effects of Endogenous Secretory Receptor for Advanced Glycation End-products in Brain Ischemia[J]. Aging and disease, 2020, 11(3): 547-558.
URL:     OR
Figure 1.  Parabiosis and BCCAO. A) Parabiosis was done between wild-type (WT) and WT mice (WT→WT), esRAGE transgenic and WT mice (Tg→WT), and esRAGE transgenic and RAGE knockout (KO) mice (Tg→KO); gray-colored and right-side mice underwent BCCAO and further analyses. B) Experimental timeline. C, Laser-Doppler flowmetry data for evaluating the cerebral cortical microperfusion. Baseline, baseline data (100%); BCCAO, data at 1 min during the occlusion; After, data at 30 min after BCCAO. Values are mean ± SD. D) Hematoxylin-eosin stain of the hippocampus. E) Human esRAGE levels in the sera of WT sides of WT→WT and Tg→WT pairs and non-parabiosed esRAGE Tg mice (n = 4-8). ND, not detected. Values are mean ± SEM.
Figure 2.  Neuronal cell damage. A) HE and Nissl stains of the hippocampal CA1 region of WT sides of WT→WT and Tg→WT pairs and of KO side of Tg→KO pair after 7 days of BCCAO (left panel). Surviving neuron numbers per area in hippocampal CA1 region were counted in WT sides of WT→WT and Tg→WT pairs and in KO side of Tg→KO pair with or without BCCAO (right panel) (n = 4-8). Values are mean ± SEM. B) TUNEL stain. Green signals indicate apoptotic cells of WT sides of WT→WT and Tg→WT pairs and of KO side of Tg→KO pair after 7 days of BCCAO (left panel). Apoptosis cell numbers per total cell numbers were counted in WT sides of WT→WT and Tg→WT pairs and in KO side of Tg→KO pair with or without BCCAO (right panel) (n = 4-8). Values are mean ± SEM.
Figure 3.  Immunohistochemical detection of human esRAGE. A) Immunohistochemical study for the detection of human esRAGE (green signals). Hippocampal vasculatures and CA1 regions of WT side of WT→WT and esRAGE Tg (Tg)→WT pairs, the Tg side of the WT→Tg pair, and the RAGE knockout (KO) side of the Tg→KO pair without BCCAO. B) Immunostaining for NeuN (a neuronal marker, green) and human esRAGE (red) (left panel) as well as GFAP (a glial marker, green) and human esRAGE (red) (right panel) in brain cortex and hippocampus of WT sides of Tg→WT pair. Blue signals indicate nuclei [4',6-diamidino-2-phenylindole (DAPI) stain].
Figure 4.  Quantitative detection of human esRAGE A) Human esRAGE contents in the brain parenchyma of WT side of WT→WT pair, WT side of Tg→WT pair, KO side of Tg→KO pair, and Tg side of WT→Tg pair (n = 3). B) Serum human esRAGE concentrations in Tg side of Tg→WT pair, Tg side of Tg→KO pair, WT side of Tg→WT pair, and KO side of Tg→KO pair (n = 3). C, Serum mouse sRAGE concentrations in KO and WT mice (n = 3). ND, not detected; ns, not significant. Values are mean ± SEM.
Figure 5.  Association of esRAGE with endothelial cells. A and B) Immunofluorescence studies of human brain endothelial cells (HBEC) in culture. Green, Red and blue signals indicate esRAGE, heparin sulfate and nucleus (DAPI), respectively. Bar, 50 µm. C) Human esRAGE levels in culture media of HBEC (n = 4). esRAGE, 1 µg/ml; Heparin, 0.1 IU/ml; heparitinase, 1 mU/ml. Values are mean ± SEM. C) Serum levels of human esRAGE in the esRAGE Tg mice with or without heparin injection (n = 4-8). Values are mean ± SEM.
Figure 6.  Transfer of esRAGE through BBB. A) In vitro (BBB) model system composed of primary cultures of monkey brain capillary endothelial cells coupled with rat pericytes and astrocytes. Recombinant esRAGE (20 µg/ml) was added to the upper (vessel side) chambers of the model and transferred esRAGE level was quantified in the lower (brain side) chambers. Endothelial cells were treated with scrambled (control) or RAGE shRNA vectors (knockdown). The integrity of the in vitro primate BBB was unaffected by RAGE knockdown, assessed by high trans-endothelial electrical resistance (TEER) (n = 5). B) Human esRAGE levels in brain side were quantified (n = 5). Control, scrambled vector-treated; RAGE knockdown, RAGE shRNA vectors-treated. Values are mean ± SEM.
[1] Moskowitz MA, Lo EH, Iadecola C, et al. (2010). The science of stroke: mechanisms in search of treatments. Neuron, 29: 181-98.
[2] Prabhakaran S, Ruff I and Bernstein RA (2015). Acute stroke intervention: A systematic review. JAMA, 313: 1451-1462.
[3] Pulsinelli WA, Levy DE, Duffy TE, et al. (1982). Regional cerebral blood flow and glucose metabolism following transient forebrain ischemia. Ann Neurol, 11: 499-502.
[4] Tajiri S, Oyadomari S, Yano S, et al. (2004). Ischemia-induced neuronal cell death is mediated by the endoplasmic reticulum stress pathway involving CHOP. Cell Death and Differentiation, 11: 403-415.
[5] Kirino T (1982). Delayed neuronal death in the gerbil hippocampus following ischemia. Brain Res, 6: 57-69.
[6] Wang WP, Iyo AH, Miguel-Hidalgo J, et al. (2006). Agmatine protects against cell damage induced by NMDA and glutamate in cultured hippocampal neurons. Brain Res, 21: 210-216.
[7] Ma L1, Carter RJ, Morton AJ, et al. (2003). RAGE is expressed in pyramidal cells of the hippocampus following moderate hypoxic-ischemic brain injury in rats. Brain Res, 966: 167-174.
[8] Koyama R, Tao K, Sasaki T, et al. (2012). GABAergic excitation after febrile seizures induces ectopic granule cells and adult epilepsy. Nat Med, 18: 1271-1278.
[9] Lein ES, Zhao X, Gage FH, et al. (2004). Defining a molecular atlas of the hippocampus using DNA microarrays and high-throughput in situ hybridization. J Neurosci, 24: 3879-3889.
[10] Quillinan N, Grewal H, Deng G, et al. (2015). Region-specific role for GluN2B-containing NMDA receptors in injury to Purkinje cells and CA1 neurons following global cerebral ischemia. Neuroscience, 284: 555-565.
[11] Dihné M, Grommes C, Lutzenburg M, et al. (2002). Different mechanisms of secondary neuronal damage in thalamic nuclei after focal cerebral ischemia in rats. Stroke, 33: 3006-3011.
[12] Siesjö BK, Bengtsson F (1989). Calcium fluxes, calcium antagonists, and calcium-related pathology in brain ischemia, hypoglycemia, and spreading depression: a unifying hypothesis. J Cereb Blood Flow Metab, 9: 127-140.
[13] Chen YJ, Nguyen HM, Maezawa I, et al. (2016). The potassium channel kca3.1 constitutes a pharmacological target for neuroinflammation associated with ischemia/reperfusion stroke. J Cereb Blood Flow Metab, 36: 2146-2161.
[14] Love S (1999). Oxidative stress in brain ischemia. Brain Pathol, 9: 119-131.
[15] Kumar R, Bukowski MJ, Wider JM, et al. (2016). Mitochondrial dynamics following global cerebral ischemia. Mol Cell Neurosci, 76: 68-75.
[16] Kamide T, Kitao Y, Takeichi T, et al. (2012). RAGE mediates vascular injury and inflammation after global cerebral ischemia. Neurochem Int, 60: 220-228.
[17] Sajjad Muhammad, Waleed Barakat, Stoyan Stoyanov, et al. (2008). The HMGB1 Receptor RAGE Mediates Ischemic Brain Damage. J Neurosci, 28: 12023-12031.
[18] Qiu J, Nishimura M, Wang Y, et al. (2008). Early release of HMGB-1 from neurons after the onset of brain ischemia. J Cereb Blood Flow Metab, 28:927-938.
[19] Kim Y, Kim C, Son SM, et al. (2016). The novel RAGE interactor PRAK is associated with autophagy signaling in Alzheimer's disease pathogenesis. Mol Neurodegener, 11: 4.
[20] Myint KM, Yamamoto Y, Doi T, et al. (2006). RAGE control of diabetic nephropathy in a mouse model: effects of RAGE gene disruption and administration of low-molecular weight heparin. Diabetes, 55: 2510-2522.
[21] Bucciarelli LG, Wendt T, Qu W, et al. (2002) RAGE blockade stabilizes established atherosclerosis in diabetic apolipoprotein E-null mice. Circulation, 106: 2827-2835.
[22] Yamamoto Y, Yamamoto H (2012). Controlling the receptor for advanced glycation end-products to conquer diabetic vascular complications. J Diabetes Investig, 3: 107-114.
[23] Yan SD, Chen X, Fu J, et al. (1996). RAGE and amyloid-beta peptide neurotoxicity in Alzheimer's disease. Nature, 382: 685-691.
[24] Yamamoto Y, Kato I, Doi T, et al. (2001). Development and prevention of advanced diabetic nephropathy in RAGE-overexpressing mice. J Clin Invest, 108: 261-268.
[25] Tanaka N, Yonekura H, Yamagishi S, et al. (2000). The receptor for advanced glycation end products is induced by the glycation products themselves and tumor necrosis factor-α through nuclear factor-κB, and by 17β-estradiol through Sp-1 in human vascular endothelial cells. J Biol Chem, 275: 25781-25790.
[26] Yonekura H, Yamamoto Y, Sakurai S, et al. (2003). Novel splice variants of the receptor for advanced glycation end-products expressed in human vascular endothelial cells and pericytes, and their putative roles in diabetes-induced vascular injury. Biochem J, 370: 1097-1109.
[27] Yamagishi S, Matsui T (2010). Soluble form of a receptor for advanced glycation end products (sRAGE) as a biomarker. Front Biosci (Elite Ed), 2: 1184-1195.
[28] Sugihara T, Munesue S, Yamamoto Yet al. (2012). Endogenous secretory receptor for advanced glycation end-products inhibits amyloid-β1-42 uptake into mouse brain. J Alzheimers Dis, 28: 709-720.
[29] Yamamoto Y, Doi T, Kato I, et al. (2005). Receptor for advanced glycation end products is a promising target of diabetic nephropathy. Ann N Y Acad Sci, 1043: 562-566.
[30] Muhammad S, Barakat W, Stoyanov S, et al. (2008). The HMGB1 receptor RAGE mediates ischemic brain damage. J Neurosci, 28: 12023-12031.
[31] Hassid BG, Nair MN, Ducruet AF, et al. (2009). Neuronal RAGE expression modulates severity of injury following transient focal cerebral ischemia. J Clin Neurosci, 16: 302-306.
[32] Nakatsuji H, Kishida K, Sekimoto R, et al. (2014). Tracing the movement of adiponectin in a parabiosis model of wild-type and adiponectin-knockout mice. FEBS Open Bio, 4: 276-282.
[33] Sakurai S, Yamamoto Y, Tamei H, et al. (2006). Development of an ELISA for esRAGE and its application to type 1 diabetic patients. Diabetes Res Clin Pract, 73(2): 158-165.
[34] Takeuchi A, Yamamoto Y, Tsuneyama K, et al. (2007). Endogenous secretory receptor for advanced glycation endproducts as a novel prognostic marker in chondrosarcoma. Cancer, 109(12): 2532-2540.
[35] Cheng C, Tsuneyama K, Kominami R, et al. (2005). Expression profiling of endogenous secretory receptor for advanced glycation end products in human organs. Mod Pathol, 18(10): 1385-1396.
[36] Labat-Moleur F, Guillermet C, Lorimier P, et al. (1998). TUNEL apoptotic cell detection in tissue sections: critical evaluation and improvement. J Histochem Cytochem, 46: 327-334.
[37] Shimamura M, Zhou P, Casolla B, et al. (2013). Prostaglandin E2 type 1 receptors contribute to neuronal apoptosis after transient forebrain ischemia. J Cereb Blood Flow Metab, 33: 1207-1214.
[38] Kurinami H, Shimamura M, Ma T, et al. (2014). Prohibitin viral gene transfer protects hippocampal CA1 neurons from ischemia and ameliorates postischemic hippocampal dysfunction. Stroke, 45: 1131-1138.
[39] Sasaki T, Kitagawa K, Omura-Matsuoka E, et al. (2007). The phosphodiesterase inhibitor rolipram promotes survival of newborn hippocampal neurons after ischemia. Stroke, 38: 1597-1605.
[40] Cheng Y, Deshmukh M, D'Costa A, et al. (1998). Caspase inhibitor affords neuroprotection with delayed administration in a rat model of neonatal hypoxic-ischemic brain injury. J Clin Invest, 101: 1992-1999.
[41] Bohara M, Kambe Y, Nagayama T, et al. (2014). C-type natriuretic peptide modulates permeability of the blood-brain barrier. J Cereb Blood Flow Metab, 34: 589-596.
[42] Yamamoto Y, Mingkun Liang, Seiichi Munesue, et al. (2019). Vascular RAGE transports oxytocin into the brain to elicit its maternal bonding behaviour in mice. Commun Biol, 2: 76.
[43] Srinivasan B, Kolli AR, Esch MB, et al. (2015). TEER measurement techniques for in vitro barrier model systems. J Lab Autom, 20: 107-126.
[44] Gardner TW, Lieth E, Khin SA, et al. (1997). Astrocytes increase barrier properties and ZO-1 expression in retinal vascular endothelial cells. Invest Ophthalmol Vis Sci, 38: 2423-2427.
[45] Monden M, Koyama H, Otsuka Y, et al. (2013). Receptor for advanced glycation end products regulates adipocyte hypertrophy and insulin sensitivity in mice: involvement of Toll-like receptor 2. Diabetes, 62: 478-489.
[46] Shichita T, Sugiyama Y, Ooboshi H, et al. (2009). Pivotal role of cerebral interleukin-17-producing gammadeltaT cells in the delayed phase of ischemic brain injury. Nat Med, 15: 946-950.
[47] Shichita T, Hasegawa E, Kimura A, et al. (2012). Peroxiredoxin family proteins are key initiators of post-ischemic inflammation in the brain. Nat Med, 18: 911-917.
[48] Tang SC, Yeh SJ, Tsai LK, et al. (2016). Cleaved but not endogenous secretory RAGE is associated with outcome in acute ischemic stroke. Neurology, 86: 270-276.
[49] Mizumoto S, Takahashi J, Sugahara K, et al. (2012). Receptor for advanced glycation end products (RAGE) functions as receptor for specific sulfated glycosaminoglycans, and anti-RAGE antibody or sulfated glycosaminoglycans delivered in vivo inhibit pulmonary metastasis of tumor cells. J Biol Chem, 287: 18985-18994.
[50] Deane R, Du Yan S, Submamaryan RK, et al. (2003). RAGE mediates amyloid-beta peptide transport across the blood-brain barrier and accumulation in brain. Nat Med, 9: 907-913.
[1] Yongming Jia, Na Wang, Yingbo Zhang, Di Xue, Haoming Lou, Xuewei Liu. Alteration in the Function and Expression of SLC and ABC Transporters in the Neurovascular Unit in Alzheimer’s Disease and the Clinical Significance[J]. Aging and disease, 2020, 11(2): 390-404.
[2] Zhang Can, Brandon Nicole R., Koper Kerryann, Tang Pei, Xu Yan, Dou Huanyu. Invasion of Peripheral Immune Cells into Brain Parenchyma after Cardiac Arrest and Resuscitation[J]. Aging and disease, 2018, 9(3): 412-425.
[3] Ren Changhong, Li Ning, Wang Brian, Yang Yong, Gao Jinhuan, Li Sijie, Ding Yuchuan, Jin Kunlin, Ji Xunming. Limb Ischemic Perconditioning Attenuates Blood-Brain Barrier Disruption by Inhibiting Activity of MMP-9 and Occludin Degradation after Focal Cerebral Ischemia[J]. Aging and disease, 2015, 6(6): 406-417.
Full text



Copyright © 2014 Aging and Disease, All Rights Reserved.
Address: Aging and Disease Editorial Office 3400 Camp Bowie Boulevard Fort Worth, TX76106 USA
Fax: (817) 735-0408 E-mail:
Powered by Beijing Magtech Co. Ltd