Contents lists available at ScienceDirect Journal of Neuroimmunology journal homepage: www.elsevier.com/locate/jneuroim Ischemic damage and early inflammatory infiltration are different in the core and penumbra lesions of rat brain after transient focal cerebral ischemia Emőke Horváth a, ⁎ , Adina Huțanu b,c , Liviu Chiriac d , Minodora Dobreanu b,c , Alex Orădan e , Előd-Ernő Nagy f a Department of Pathology, University of Medicine and Pharmacy of Tîrgu Mures, Gh. Marinescu Street 38, Tîrgu Mures, Romania b Center for Advanced Medical and Pharmaceutical Research, University of Medicine and Pharmacy Tîrgu Mures, Gh. Marinescu Street 38, Tîrgu Mures, Romania c Department of Laboratory Medicine, University of Medicine and Pharmacy of Tîrgu Mures, Gh. Marinescu Street 50, Tîrgu Mures, Romania d National Magnetic Resonance Center, Faculty of Physics, "Babes-Bolyai" University, Cluj-Napoca, Mihail Kogalniceanu Street 1, Cluj-Napoca, Romania e Laboratory Animal Facility-Center for Experimental Medicine, Iuliu Hatieganu University of Medicine and Pharmacy, Louis Pasteur Street 6, Cluj-Napoca, Romania f Department of Biochemistry and Environmental Chemistry, University of Medicine and Pharmacy of Tîrgu Mures, Gh. Marinescu Street 38, Tîrgu Mures, Romania ARTICLE INFO Keywords: Early inflammatory infiltrate Digital morphometry M1/M2 macrophages COX-2 FXIII Cerebral ischemia ABSTRACT Clinical and experimental observations emphasize that inflammation is a direct risk factor for stroke. We per- formed a detailed histological and immunohistochemical analysis, assisted by digital morphometry, to compare the representative brain lesions in the ischemic core and penumbra in a rat model. Focal neuronal necrosis and degeneration were significantly more intense in the core, whereas inflammatory infiltration, MPO, CD68, CD3, FXIII, Cox-2, iNOS2, Arg-1 expressions were stronger in the penumbra. Our findings indicate that neuroin- flammation affects the penumbra more than the core and suggest that targeted modulation of the cellular in- filtrate could be exploited to save brain volume. 1. Introduction Stroke is the second leading cause of death globally and can lead to permanent disability (The World Health Organization updates, 2017). In humans, the immediate therapy of stroke focuses on reducing the volume of acute ischemia in order to improve the clinical outcome. Therapeutic options are limited to early recanalization by intravenous thrombolysis with the clot-busting drug, tissue plasminogen activator administered up to 4.5 h after the onset of symptoms, or/and throm- bectomy; both methods have a narrow therapeutic window (Jauch et al., 2013). The parenchymal lesions in acute ischemic stroke can be classified in two, sometimes poorly delimited morphological zones: is- chemic core (irreversibly injured) and penumbra. The penumbra is severely hypoperfused and is at increased risk of being merged into the ischemic core, if it is not reperfused quickly (Alves et al., 2014). Correct estimation of the penumbra size and drastic reduction of its extent is critical in guiding stroke therapy. Thus, a focus of acute stroke intervention should be the early reperfusion of the pe- numbra (Manning et al., 2014; Fuhrer et al., 2017). In stroke, the duration and intensity of the ischemia determines the severity of neural tissue damage, but other factors, independent of circulation, might also exert important effects, especially in the periinfarct area (Wolinski and Glabinski, 2013). Animal models of cerebral ischemia describe a well-established timing of inflammatory events after the brain injury. First, within minutes after the onset of ischemia, resident microglia cells are acti- vated, with the consequential release of reactive oxygen species and proinflammatory cytokines. These mediators induce the activation of cerebrovascular endothelial cells, promoting adhesion and transmi- gration of leukocytes into the injured tissue. There is some controversy regarding the prioritary presence of neutrophils versus macrophages. Some authors state that the first infiltrating cells are neutrophils (from https://doi.org/10.1016/j.jneuroim.2018.08.002 Received 30 April 2018; Received in revised form 2 August 2018; Accepted 4 August 2018 Abbreviations: MMPs, matrix metalloproteinases; PMNs, polymorphonuclear neutrophils; tMCAO, transient middle cerebral artery occlusion; S, sham group; I, ischemic group; MCA, middle cerebral artery; MRI, magnetic resonance imaging; H&E, haematoxylin and eosin; CV, cresyl violet/Nissl stain; CD15, cluster of differentiation antigen 15; MPO, Myeloperoxidase; CD68, cluster of differentiation antigen 68; iNOS2, inducible nitric oxide synthase; Arg1, arginase; CD3, cluster of differentiation antigen 3; FXIII, coagulation factor XIII; MMP-9, matrix metalloproteinase 9; COX-2, Cyclooxygenase-2; DAB, 3,3'-diaminobenzidine; TTDS, total tissue damage score; TBV, total brain volume; IV, ischemic volume; PGE-2, Prostaglandin E2 ⁎ Corresponding author at: Department of Pathology, University of Medicine and Pharmacy of Tîrgu Mures, 540139 Gh. Marinescu 38, Tîrgu Mures, Romania. E-mail address: emoke.horvath@umftgm.ro (E. Horváth). Journal of Neuroimmunology 324 (2018) 35–42 0165-5728/ © 2018 Elsevier B.V. All rights reserved. T