Bioactive molecules isolated from olive pomace extract protect murine cortex neurons from NMDA-mediated cell death

Changes in intracellular calcium concentration are crucial events during signal transduction processes involved in several cellular physiological functions. Moreover, even a limited dysregulation in intracellular calcium homeostasis can promotes the onset and the progression of neuropathological processes. We have shown previously that the olive pomace extract, obtained in condition of high pressure and high temperature (HPTE), contains bioactive molecules able to prevent calcium-induced cell damages on human neuroblastoma (SK-N-BE) and mouse brain endothelioma (bEnd5) cell lines, operating through the regulation of the cell dynamics that involve the calcium ions. In this thesis, we report the effect of HPTE on primary cell culture of murine cortical neurons used as a cellular model. Specifically, in order to induce a cytotoxic influx of Ca2+ through the opening of the NMDA (N-Methyl-D-Aspartic-Acid) receptor (NMDAR), we treated neurons with NMDA in the presence or in the absence of HPTE for 24 hours. We demonstrate that HPTE 1) significantly reduces the cytotoxic effect of NMDA, evaluated as percentage of cell death, and 2) operates preventing the increase of the intracellular calcium concentration mediated by the NMDAR. To identify the chemical nature of the active factors in the olive pomace extract, we have fractionated HPTE by RP-HPLC, followed by the assay of the cell protective effect. Successively, on the fractions containing the bioactivity and using mass spectrometry techniques, we established that the relevant molecules could be derivatives of the epigallocatechin or of the loganin. The ultimate aim of this work is to provide new information about the structural and functional specificity of molecules present in the HPTE in order to propose new possible strategies for therapeutic interventions in neurodegenerative diseases.