Peptides analogous to tethered ligands liberated by activated protein C exert neuroprotective effects in glutamate-induced excitotoxicity

Abstract

Haemostatic proteinases may appear in brain tissue after injury and under inflammation as a result of the blood-brain barrier disruption. Serine proteinases regulate cell functions through G-protein-coupled transmembrane protease-activated receptors (PARs). Proteinases cleave only one peptide bond of receptor exodomain, which results in the formation of a new N-terminus (“tethered ligand”) that can specifically interact with the second extracellular loop of the receptor and activate it. Two types of receptors (EPCR and PAR1) are necessary for the cytoprotective effect of activated protein C (APC) on endothelial cells and neurons. APC activates PAR-1 and controls gene expression of proinflammatory and proapoptotic factors. APC exerts a protective effect in stressed neurons and hypoxic brain endothelium, modulates the activity of endothelial cell genes involved in apoptosis, and stabilizes the endothelial barrier. We suppose that the peptides analogous to the PAR1 tethered ligand released by APC may have a neuroprotective effect similar to that of APC. We have simulated ischemic brain damage using a model of glutamate excitotoxicity on the primary culture of neonatal rat hippocampal neurons. It was shown that NPNDKYEPF-amide (peptide 9) and NPNDKYEPFWE (peptide 11) more effectively reduced the level of apoptosis during neuronal excitotoxicity in comparison with APC, while the influence of these peptides on the number of living and necrotic cells was analogous to that of APC. The findings suggest that the protective effect of the peptides analogous to the PAR1 tethered ligand is comparable to the protective effect of APC under glutamate excitotoxicity. Investigation of the mechanisms of PAR1 agonist peptides action and development of their shortened versions with high neuroprotective activity may be a relevant approach to the search of novel neuroprotective drugs for treating neurodegenerative diseases and strokes.