Abstract
Stroke is one of the leading causes of death and permanent disability in the elderly. However, most of the experimental studies on stroke are based on young animals, and we hypothesised that age can substantially affect the stroke response. The two-vessel occlusion model of global ischemia by occluding the common carotid arteries for 15 min at 40 mmHg of blood pressure was carried out in 3- and 18-month-old male Sprague–Dawley rats. The adhesion molecules E- and P-selectin, cell adhesion molecules (CAMs), both intercellular (ICAM-1) and vascular (VCAM-1), as well as glial fibrillary acidic protein (GFAP), and cleaved caspase-3 were measured at 48 h after ischemia in the cerebral cortex and hippocampus using Western blot, qPCR and immunofluorescence techniques. Diametric expression of GFAP and a different morphological pattern of caspase-3 labelling, although no changes in the cell number, were observed in the neurons of young and old animals. Expression of E-selectin and CAMs was also modified in an age- and ischemia/reperfusion-dependent manner. The hippocampus and cerebral cortex had similar response patterns for most of the markers studied. Our data suggest that old and young animals present different time-courses of neuroinflammation and apoptosis after ischemic damage. On the other hand, these results suggest that neuroinflammation is dependent on age rather than on the different vulnerability described for the hippocampus and cerebral cortex. These differences should be taken into account in searching for therapeutic targets.
Similar content being viewed by others
Abbreviations
- BBB:
-
Blood–brain barrier
- BSA:
-
Bovine serum albumin
- CA:
-
Cornu Ammonis
- CAM:
-
Cellular adhesion molecules
- DABCO:
-
1,4-Diazabicyclo(2.2.2)octane
- DAPI:
-
4′,6-Diamidino-2-phenylindole
- GFAP:
-
Glial fibrillary acidic protein
- I/R:
-
Ischemia/reperfusion
- ICAM:
-
Intercellular adhesion molecule
- MCAO:
-
Middle cerebral artery occlusion
- MRI:
-
Magnetic resonance imaging
- PFA:
-
Paraformaldehyde
- PBST:
-
Buffer sodium phosphate with Triton X-100
- TBST:
-
Tris-buffered saline 50 mM with Tween-20 0.2 %
- VCAM:
-
Vascular adhesion molecule
References
Amor S, Puentes F, Baker D, van der Valk P (2010) Inflammation in neurodegenerative diseases. Immunology 129:154–169
Anthony DC, Bolton SJ, Fearn S, Perry VH (1997) Age-related effects of interleukin-1 beta on polymorphonuclear neutrophil-dependent increases in blood–brain barrier permeability in rats. Brain 120(Pt 3):435–444
Anyanwu EC (2007) Neurochemical changes in the aging process: implications in medication in the elderly. Sci World J 7:1603–1610
Arumugam TV, Phillips TM, Cheng A, Morrell CH, Mattson MP, Wan R (2010) Age and energy intake interact to modify cell stress pathways and stroke outcome. Ann Neurol 67:41–52
Ayuso MI, Garcia-Bonilla L, Martin ME, Salinas M (2010) Assessment of protein expression levels after transient global cerebral ischemia using an antibody microarray analysis. Neurochem Res 35:1239–1247
Back T (1998) Pathophysiology of the ischemic penumbra—revision of a concept. Cell Mol Neurobiol 18:621–638
Badan I, Platt D, Kessler C, Popa-Wagner A (2003) Temporal dynamics of degenerative and regenerative events associated with cerebral ischemia in aged rats. Gerontology 49:356–365
Bala K, Tripathy BC, Sharma D (2006) Neuroprotective and anti-ageing effects of curcumin in aged rat brain regions. Biogerontology 7:81–89
Barkalow FJ, Goodman MJ, Gerritsen ME, Mayadas TN (1996) Brain endothelium lack one of two pathways of P-selectin-mediated neutrophil adhesion. Blood 88:4585–4593
Bendel O, Alkass K, Bueters T, von Euler M, von Euler G (2005) Reproducible loss of CA1 neurons following carotid artery occlusion combined with halothane-induced hypotension. Brain Res 1033:135–142
Blamire AM, Anthony DC, Rajagopalan B, Sibson NR, Perry VH, Styles P (2000) Interleukin-1beta-induced changes in blood–brain barrier permeability, apparent diffusion coefficient, and cerebral blood volume in the rat brain: a magnetic resonance study. J Neurosci 20:8153–8159
Buga AM, Di Napoli M, Popa-Wagner A (2013) Preclinical models of stroke in aged animals with or without comorbidities: role of neuroinflammation. Biogerontology 14:651–662
Busch SA, Silver J (2007) The role of extracellular matrix in CNS regeneration. Curr Opin Neurobiol 17:120–127
Cacheaux LP, Ivens S, David Y, Lakhter AJ, Bar-Klein G, Shapira M, Heinemann U, Friedman A, Kaufer D (2009) Transcriptome profiling reveals TGF-beta signaling involvement in epileptogenesis. J Neurosci 29:8927–8935
Candelario-Jalil E (2009) Injury and repair mechanisms in ischemic stroke: considerations for the development of novel neurotherapeutics. Curr Opin Investig Drugs 10:644–654
Colangelo AM, Alberghina L, Papa M (2014) Astrogliosis as a therapeutic target for neurodegenerative diseases. Neurosci Lett
Collins TC, Petersen NJ, Menke TJ, Souchek J, Foster W, Ashton CM (2003) Short-term, intermediate-term, and long-term mortality in patients hospitalized for stroke. J Clin Epidemiol 56:81–87
Dijkhuizen RM, Knollema S, van der Worp HB, Ter Horst GJ, De Wildt DJ, Berkelbach van der Sprenkel JW, Tulleken KA, Nicolay K (1998) Dynamics of cerebral tissue injury and perfusion after temporary hypoxia-ischemia in the rat: evidence for region-specific sensitivity and delayed damage. Stroke 29:695–704
Donnan GA, Fisher M, Macleod M, Davis SM (2008) Stroke. Lancet 371:1612–1623
Durukan A, Tatlisumak T (2007) Acute ischemic stroke: overview of major experimental rodent models, pathophysiology, and therapy of focal cerebral ischemia. Pharmacol Biochem Behav 87:179–197
Duverger D, MacKenzie ET (1988) The quantification of cerebral infarction following focal ischemia in the rat: influence of strain, arterial pressure, blood glucose concentration, and age. J Cereb Blood Flow Metab 8:449–461
Dziennis S, Mader S, Akiyoshi K, Ren X, Ayala P, Burrows GG, Vandenbark AA, Herson PS, Hurn PD, Offner HA (2011) Therapy with recombinant T-cell receptor ligand reduces infarct size and infiltrating inflammatory cells in brain after middle cerebral artery occlusion in mice. Metab Brain Dis 26:123–133
Dziewulska D (1997) Age-dependent changes in astroglial reactivity in human ischemic stroke. Immunohistochemical study. Folia Neuropathol 35:99–106
Eckle VS, Buchmann A, Bursch W, Schulte-Hermann R, Schwarz M (2004) Immunohistochemical detection of activated caspases in apoptotic hepatocytes in rat liver. Toxicol Pathol 32:9–15
Fan W, Dai Y, Xu H, Zhu X, Cai P, Wang L, Sun C, Hu C, Zheng P, Zhao BQ (2014) Caspase-3 modulates regenerative response after stroke. Stem Cells 32:473–486
Fricker M, Vilalta A, Tolkovsky AM, Brown GC (2013) Caspase inhibitors protect neurons by enabling selective necroptosis of inflamed microglia. J Biol Chem 288:9145–9152
Gelderblom M, Leypoldt F, Steinbach K, Behrens D, Choe CU, Siler DA, Arumugam TV, Orthey E, Gerloff C, Tolosa E, Magnus T (2009) Temporal and spatial dynamics of cerebral immune cell accumulation in stroke. Stroke 40:1849–1857
Ginsberg MD, Pulsinelli WA (1994) The ischemic penumbra, injury thresholds, and the therapeutic window for acute stroke. Ann Neurol 36:553–554
Gotsch U, Jager U, Dominis M, Vestweber D (1994) Expression of P-selectin on endothelial cells is upregulated by LPS and TNF-alpha in vivo. Cell Adhes Commun 2:7–14
He Z, Meschia JF, Brott TG, Dickson DW, McKinney M (2006) Aging is neuroprotective during global ischemia but leads to increased caspase-3 and apoptotic activity in hippocampal neurons. Curr Neurovasc Res 3:181–186
Hossmann KA (1994) Viability thresholds and the penumbra of focal ischemia. Ann Neurol 36:557–565
Ivens S, Kaufer D, Flores LP, Bechmann I, Zumsteg D, Tomkins O, Seiffert E, Heinemann U, Friedman A (2007) TGF-beta receptor-mediated albumin uptake into astrocytes is involved in neocortical epileptogenesis. Brain 130:535–547
Jin R, Yang G, Li G (2010) Inflammatory mechanisms in ischemic stroke: role of inflammatory cells. J Leukoc Biol 87:779–789
Kadhim HJ, Duchateau J, Sebire G (2008) Cytokines and brain injury: invited review. J Intensive Care Med 23:236–249
Kirino T, Tamura A, Sano K (1985) Selective vulnerability of the hippocampus to ischemia—reversible and irreversible types of ischemic cell damage. Prog Brain Res 63:39–58
Kriz J, Lalancette-Hebert M (2009) Inflammation, plasticity and real-time imaging after cerebral ischemia. Acta Neuropathol 117:497–509
Lakhan SE, Kirchgessner A, Hofer M (2009) Inflammatory mechanisms in ischemic stroke: therapeutic approaches. J Transl Med 7:97
Lalonde CC, Mielke JG (2014) Selective vulnerability of hippocampal sub-fields to oxygen-glucose deprivation is a function of animal age. Brain Res 1543:271–279
Li Y, Powers C, Jiang N, Chopp M (1998) Intact, injured, necrotic and apoptotic cells after focal cerebral ischemia in the rat. J Neurol Sci 156:119–132
Li C, Zhao R, Gao K, Wei Z, Yin MY, Lau LT, Chui D, Hoi Yu AC (2011) Astrocytes: implications for neuroinflammatory pathogenesis of Alzheimer’s disease. Curr Alzheimer Res 8:67–80
Lipton P (1999) Ischemic cell death in brain neurons. Physiol Rev 79:1431–1568
Liu F, McCullough LD (2011) Middle cerebral artery occlusion model in rodents: methods and potential pitfalls. J Biomed Biotechnol 2011:464701
Liu F, McCullough LD (2012) Interactions between age, sex, and hormones in experimental ischemic stroke. Neurochem Int 61:1255–1265
Liu F, Benashski SE, Persky R, Xu Y, Li J, McCullough LD (2012) Age-related changes in AMP-activated protein kinase after stroke. Age (Dordr) 34:157–168
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) Method. Methods 25:402–408
Llorente IL, Burgin TC, Perez-Rodriguez D, Martinez-Villayandre B, Perez-Garcia CC, Fernandez-Lopez A (2013) Unfolded protein response to global ischemia following 48 h of reperfusion in the rat brain: the effect of age and meloxicam. J Neurochem 127(5):701–710
McIntosh CT, Warnock JN (2013) Side-specific characterization of aortic valve endothelial cell adhesion molecules under cyclic strain. J Heart Valve Dis 22:631–639
Mehta SL, Manhas N, Raghubir R (2007) Molecular targets in cerebral ischemia for developing novel therapeutics. Brain Res Rev 54:34–66
Meisel C, Schwab JM, Prass K, Meisel A, Dirnagl U (2005) Central nervous system injury-induced immune deficiency syndrome. Nat Rev Neurosci 6:775–786
Montori S, Dos Anjos S, Rios-Granja MA, Perez-Garcia CC, Fernandez-Lopez A, Martinez-Villayandre B (2010a) AMPA receptor downregulation induced by ischaemia/reperfusion is attenuated by age and blocked by meloxicam. Neuropathol Appl Neurobiol 36:436–447
Montori S, Martinez-Villayandre B, Dos-Anjos S, Llorente IL, Burgin TC, Fernandez-Lopez A (2010b) Age-dependent modifications in the mRNA levels of the rat excitatory amino acid transporters (EAATs) at 48 hour reperfusion following global ischemia. Brain Res 1358:11–19
Montori S, Dos-Anjos S, Martinez-Villayandre B, Regueiro-Purrinos MM, Gonzalo-Orden JM, Ruano D, Fernandez-Lopez A (2010c) Age and meloxicam attenuate the ischemia/reperfusion-induced down-regulation in the NMDA receptor genes. Neurochem Int 56:878–885
Nedergaard M, Gjedde A, Diemer NH (1986) Focal ischemia of the rat brain: autoradiographic determination of cerebral glucose utilization, glucose content, and blood flow. J Cereb Blood Flow Metab 6:414–424
Perry VH, Anthony DC, Bolton SJ, Brown HC (1997) The blood–brain barrier and the inflammatory response. Mol Med Today 3:335–341
Petri B, Phillipson M, Kubes P (2008) The physiology of leukocyte recruitment: an in vivo perspective. J Immunol 180:6439–6446
Popa-Wagner A, Dinca I, Yalikun S, Walker L, Kroemer H, Kessler C (2006) Accelerated delimitation of the infarct zone by capillary-derived nestin-positive cells in aged rats. Curr Neurovasc Res 3:3–13
Popa-Wagner A, Badan I, Walker L, Groppa S, Patrana N, Kessler C (2007) Accelerated infarct development, cytogenesis and apoptosis following transient cerebral ischemia in aged rats. Acta Neuropathol 113:277–293
Rami A, Bechmann I, Stehle JH (2008) Exploiting endogenous anti-apoptotic proteins for novel therapeutic strategies in cerebral ischemia. Prog Neurobiol 85:273–296
Rojas JI, Zurru MC, Romano M, Patrucco L, Cristiano E (2007) Acute ischemic stroke and transient ischemic attack in the very old-risk factor profile and stroke subtype between patients older than 80 years and patients aged less than 80 years. Eur J Neurol 14:895–899
Rolls A, Shechter R, Schwartz M (2009) The bright side of the glial scar in CNS repair. Nat Rev Neurosci 10:235–241
Rosamond W, Flegal K, Furie K, Go A, Greenlund K, Haase N, Hailpern SM, Ho M, Howard V, Kissela B, Kittner S, Lloyd-Jones D, McDermott M, Meigs J, Moy C, Nichol G, O’Donnell C, Roger V, Sorlie P, Steinberger J, Thom T, Wilson M, Hong Y, American Heart Association Statistics Committee and Stroke Statistics Subcommittee (2008) Heart disease and stroke statistics–2008 update: a report from the American Heart Association Statistics Committee and Stroke Statistics Subcommittee. Circulation 117:e25–e146
Salas A, Shimaoka M, Phan U, Kim M, Springer TA (2006) Transition from rolling to firm adhesion can be mimicked by extension of integrin alphaLbeta2 in an intermediate affinity state. J Biol Chem 281:10876–10882
Schilling M, Besselmann M, Leonhard C, Mueller M, Ringelstein EB, Kiefer R (2003) Microglial activation precedes and predominates over macrophage infiltration in transient focal cerebral ischemia: a study in green fluorescent protein transgenic bone marrow chimeric mice. Exp Neurol 183:25–33
Sinha N, Baquer NZ, Sharma D (2005) Anti-lipidperoxidative role of exogenous dehydroepiendrosterone (DHEA) administration in normal ageing rat brain. Indian J Exp Biol 43:420–424
Sofroniew MV, Vinters HV (2010) Astrocytes: biology and pathology. Acta Neuropathol 119:7–35
Stanimirovic D, Satoh K (2000) Inflammatory mediators of cerebral endothelium: a role in ischemic brain inflammation. Brain Pathol 10:113–126
Stevens SL, Bao J, Hollis J, Lessov NS, Clark WM, Stenzel-Poore MP (2002) The use of flow cytometry to evaluate temporal changes in inflammatory cells following focal cerebral ischemia in mice. Brain Res 932:110–119
Sughrue ME, Mehra A, Connolly ES Jr, D’Ambrosio AL (2004) Anti-adhesion molecule strategies as potential neuroprotective agents in cerebral ischemia: a critical review of the literature. Inflamm Res 53:497–508
Tamura A, Graham DI, McCulloch J, Teasdale GM (1981) Focal cerebral ischaemia in the rat: 2. Regional cerebral blood flow determined by [14C]iodoantipyrine autoradiography following middle cerebral artery occlusion. J Cereb Blood Flow Metab 1:61–69
Tanaka R, Komine-Kobayashi M, Mochizuki H, Yamada M, Furuya T, Migita M, Shimada T, Mizuno Y, Urabe T (2003) Migration of enhanced green fluorescent protein expressing bone marrow-derived microglia/macrophage into the mouse brain following permanent focal ischemia. Neuroscience 117:531–539
Taylor S, Wakem M, Dijkman G, Alsarraj M, Nguyen M (2010) A practical approach to RT-qPCR-Publishing data that conform to the MIQE guidelines. Methods 50:S1–S5
Wang Q, Tang XN, Yenari MA (2007) The inflammatory response in stroke. J Neuroimmunol 184:53–68
Wang N, Zhang Y, Wu L, Wang Y, Cao Y, He L, Li X, Zhao J (2013) Puerarin protected the brain from cerebral ischemia injury via astrocyte apoptosis inhibition. Neuropharmacology 79C:282–289
Wasserman JK, Yang H, Schlichter LC (2008) Glial responses, neuron death and lesion resolution after intracerebral hemorrhage in young vs. aged rats. Eur J Neurosci 28:1316–1328
World Health Organization (WHO) (2011) The top 10 causes of death. Fact sheet number 310. WHO, Geneva
Xu XJ, Plesan A, Yu W, Hao JX, Wiesenfeld-Hallin Z (2001) Possible impact of genetic differences on the development of neuropathic pain-like behaviors after unilateral sciatic nerve ischemic injury in rats. Pain 89:135–145
Yilmaz G, Granger DN (2008) Cell adhesion molecules and ischemic stroke. Neurol Res 30:783–793
Yilmaz G, Arumugam TV, Stokes KY, Granger DN (2006) Role of T lymphocytes and interferon-gamma in ischemic stroke. Circulation 113:2105–2112
Zarow C, Vinters HV, Ellis WG, Weiner MW, Mungas D, White L, Chui HC (2005) Correlates of hippocampal neuron number in Alzheimer’s disease and ischemic vascular dementia. Ann Neurol 57:896–903
Zhang R, Chopp M, Zhang Z, Jiang N, Powers C (1998) The expression of P- and E-selectins in three models of middle cerebral artery occlusion. Brain Res 785:207–214
Zhang M, Li WB, Geng JX, Li QJ, Sun XC, Xian XH, Qi J, Li SQ (2007) The upregulation of glial glutamate transporter-1 participates in the induction of brain ischemic tolerance in rats. J Cereb Blood Flow Metab 27:1352–1368
Acknowledgments
We wish to thank Marta Fernandez Caso from the University of Leon for technical support and personal help. This study was supported by Junta of Castilla of León (LE184A12-2). Diego Pérez Rodríguez is granted by Junta de Castilla y León (EDU/346/2013)
Conflict of interest
The authors declare that they have no conflict of interests.
Author information
Authors and Affiliations
Corresponding author
About this article
Cite this article
Anuncibay-Soto, B., Pérez-Rodríguez, D., Llorente, I.L. et al. Age-dependent modifications in vascular adhesion molecules and apoptosis after 48-h reperfusion in a rat global cerebral ischemia model. AGE 36, 9703 (2014). https://doi.org/10.1007/s11357-014-9703-7
Received:
Accepted:
Published:
DOI: https://doi.org/10.1007/s11357-014-9703-7