Objective: To achieve a comprehensive method able to distinguish the secretome changes caused by oxidative stress regulation or induction of cell death, and thus be able to monitor indicators of cell death progression in blood.

Background: Molecules secreted by cells play key roles in the regulation of many physiological processes, being therefore good predictors of the cellular physiological state. As a result, the secretome is an important source of potential biomarkers, more likely to be reflected in biological fluids. Oxidative stress is perhaps the most common factor, and the main cause of cell death involved in a large variety of disorders. However, oxidative stress (de)regulation may occurs through diverse mechanisms leading to different responses, which can be reflected in the secretome.

Methods: A cell model was treated with different stimulus of hydrogen peroxide to induce oxidative stress without cell death or to promote cell death. The newly generated secretome spiked with the proper internal standards, was analyzed by a quantitative mass spectrometry approach (SWATH-MS). Additionally, cell viability assays were led to correlate with the MS data.

Results: A large number of proteins and metabolites were quantified between control and mild oxidative stress conditions, from them 119 up- and 115 downregulated proteins, and 519 up- and 921 downregulated metabolite features were highlighted by allowing a clear distinction of the conditions. Since no impact in cellular integrity was detected, these markers can be considered indicators of stress previous to cell death. In addition, a panel of 24 proteins markedly increasing in the cell death condition was detected, being a good indicator of cell death.

Conclusions: An integrative approach was introduced, and successfully applied in identification of oxidative stress biomarkers in cellular secretome, allowing the creation of libraries with proteins and metabolites that can be used to interrogate other samples on routine analysis. To transpose the proposed method to clinical diagnosis, these potential biomarkers will be validated in CSF and plasma from animal subjected to an oxidative stress insult and finally, in plasma of patients with neurodegenerative disorders, where these markers will be used to evaluated the progression of the disease and the respective therapy.

XIII CNC Annual Meeting, 2015, 14th Human Proteome Organization World Congress – HUPO 2015.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/monitoring-oxidative-stress-and-progression-to-cell-death-from-secretome-to-blood-diagnosis/