NeuroEPO Preserves Neurons from Glutamate-Induced Excitotoxicity

Article type: Research Article

Authors: Garzón, Fernando^{a; b} | Coimbra, Débora^a | Parcerisas, Antoni^{a; f; g} | Rodriguez, Yamila^{c; d} | García, Julio Cesar^{c; e} | Soriano, Eduardo^{a; f; g; h} | Rama, Ramón^{a; *}

Affiliations: [a] Department of Cell Biology, Physiology and Immunology, Faculty of Biology, University of Barcelona, Spain | [b] Department of Animal Health, University of Nariño, Colombia | [c] Department of Histology, Institute of Preclinical and Basic Sciences, University of Medical Sciences, Havana, Cuba | [d] Center of Molecular Immunology (CIM), Havana, Cuba | [e] National Center for Animals Breeding (Cenpalab), Havana, Cuba | [f] Centro de Investigacion Biomedica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), ISCIII, Madrid, Spain | [g] Vall d’Hebron Institute of Research, Barcelona, Spain | [h] Institucio Catalana de Recerca i Estudis Avancats (ICREA), Barcelona, Spain

Correspondence: [*] Correspondence to: Dr. Ramón Rama, Department Biology Cell, Physiology and Immunology, Faculty of Biology, University of Barcelona. Avenida Diagonal, 643, 08028 Barcelona, Spain. E-mail: [email protected].

Abstract: Many experimental studies show that erythropoietin (EPO) has a neuroprotective action in the brain. EPO in acute and chronic neurological disorders, particularly in stroke, traumatic brain injury, Alzheimer’s disease, Parkinson’s disease, and amyotrophic lateral sclerosis, has neuroprotective effects. We previously reported the neuroprotective effect of NeuroEPO, a low sialic form of EPO, against oxidative stress induced by glutamate excitotoxicity. In this paper, we analyze the effect of NeuroEPO against apoptosis induced by glutamate excitotoxicity in primary neuronal cultures obtained from the forebrains of Wistar rat embryos after 17 days of gestation. Excitotoxicity was induced after nine days of in vitro culture by treatment with a culture medium containing 100μM glutamate for 15 min. To withdraw glutamate, a new medium containing 100 ng NeuroEPO/mL was added. Apoptosis was analyzed after 24 h. Images obtained by phase contrast microscopy show that neurons treated with glutamate exhibit cell body shrinkage, loss of dendrites that do not make contact with neighboring cells, and that NeuroEPO was able to preserve the morphological characteristics of the control. Immunocytochemistry images show that the culture is essentially pure in neurons; that glutamate causes cell mortality, and that this is partially avoided when the culture medium is supplemented with NeuroEPO. Activation of intrinsic apoptotic pathways was analyzed. The decreases in Bcl-2/Bax ratio, increase in the release of cytochrome c, and in the expression and activity of caspase-3 observed in cells treated with glutamate, were restored by NeuroEPO. The results from this study show that NeuroEPO protects cortical neurons from glutamate-induced apoptosis via upregulation of Bcl-2 and inhibit glutamate-induced activation of caspase-3.

Keywords: Apoptosis, erythropoietin, excitotoxicity, neurodegenerative diseases, NeuroEPO, neuroprotection

DOI: 10.3233/JAD-180668

Journal: Journal of Alzheimer's Disease, vol. 65, no. 4, pp. 1469-1483, 2018