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Dr. Ginevra Giacomello

Dissertation: Metabolic alteration of circulating monocytes during neuroinflammation: method development and in vitro studies in Alzheimer’s disease and multiple sclerosis

This thesis focuses on the study of the reprogramming of the immunometabolism of infiltrating monocytes during neuroinflammation. Three main results have been achieved. First, the development and validation of a method for analysing changes in the major metabolic pathways of cells. To do so, the analytical challenges presented by the high polarity of the targeted metabolites and the chelating properties of citrate and phosphates have been addressed and overcome. The developed method allows the simultaneous analysis of key-role metabolites with HPLC-ESI-QQQ-MS. It is a suitable tool for future studies to quickly, reliably, and reproducibly investigate the metabolism of cells in different conditions or under various stimulations. The proof of concept of this method is shown by the results of the following two points. Second, in vitro elucidation of monocyte metabolic alterations when infiltrating into the CSF from the bloodstream. These results provide an insight on the immunometabolism rewiring from the production of amino acids (serine and glycine) in normal conditions towards a more active TCA cycle and production of glutamine when stimulated with CSF. This knowledge gives a valuable overview of the effects caused by the compartment change of monocytes and provides the basis for understanding the metabolic adaptation of infiltrating immune cells under different conditions. Third, the display of the significant variations in the metabolism reprogramming of infiltrating monocytes in three study cases: in healthy individuals, in patients with AD and with MScl. These outcomes elucidate how a healthy donor CSF affects monocytes compared to that of patients. Moreover, it distinguishes the alterations caused by a disease more correlated to neurodegeneration, such as AD, from those caused by a disease primarily neuroinflammatory like MScl. The results presented in this thesis are of relevance for a more in-depth understanding of the metabolic fate of monocytes in CSF and to build solid bases for further studies of their role in neuroinflammation. It is one step forward in elucidating the complexity of alterations that occur in our CNS in the case of neurodegenerative or neuroinflammatory diseases.