EXPLORING THE AMYLOID BETA (ABETA) DEPENDENT METABOLIC FINGERPRINT OF HIPPOCAMPAL NEURONS BY TWO-DIMENSIONAL COMPREHENSIVE GAS CHROMATOGRAPHY: POTENTIALS IN DRUG DISCOVERY STUDIES

Two-dimensional comprehensive GC-MS (GC×GC-MS) represent the most advanced GC platform with great potentials in metabolomic studies due to its increased separation power, sensitivity and structured 2D patterns that can be adopted for samples informative features location and cross-comparison. The resulting chromatographic profiles, in fact, processed by suitable chemometrical approaches, enable to contemporarily run profiling and fingerprinting characterization of complex samples [1]. In the present study, we investigate the potentials of a GC×2GC system equipped with a conventional first dimension column (1D - 30 m x 0.25 mm ID) coupled to two second dimension columns of variable lengths (2D FID 1.6 m x 0.1 mm ID and 2DMS 1.8 m x 0.1 mm ID) and parallel dual detection by MS and FID in the metabolite profiling of neurons treated with Amyloid Beta42 Peptides (ABeta42), aiming to investigate the existence of metabolic changes occurring during early stage of Alzheimer Disease (AD) onset. AD is the most common cause of dementia; it is considered the consequence of several cellular degenerative processes that primarily affect brain networks related to memory and cognition, such as the hippocampus. The accumulation of oligomers of Amyloid Beta peptides (ABeta), such as ABeta42 produced by the proteolytic processing of the amyloid precursor protein (APP) is one of the initiating event that triggers the progressive dismantling of the synapses, neuronal circuits and networks. Here, through the use of Microelectrode Arrays (MEAs), we have first characterized the neuronal firing inhibition induced by ABeta42 and then propose to investigate if the same peptides are also able to modify the neuronal metabolite fine print. Afterwards, as also DNA methylation/demethylation has a critical role in the regulation of gene expression, we aim also to understand if it can be related to AD for a possible relationship between these latter, excitability and metabolomic profiles of neurons. In this project, newly designed chemical entities will be tested to investigate the interplay between DNA methylation/demethylation on neuronal excitability and metabolic profile. By this way we will hopefully provide new molecular tools for an improved diagnosis, prognosis, and therapy. Preliminary results emphasize the potentials of GC×2GC-MS in terms of: (a) enhanced information sensitivity, being limited the amount of biologic material available for each sampling, (b) accuracy and reliability of the resultant metabolic fine-print thus enabling reliable and consistent evaluation of drug interaction with living cells and,(c) information potential that is extended to a large number of chemical features thus providing an almost complete picture of metabolic changes undergone.