OXIDATIVE STRESS-INDUCED DOWNREGULATION OF EXTRACELLULAR MATRIX COMPONENTS AND RECEPTOR TYROSINE KINASES IN HUMAN NEUROBLASTOMA CELLS

Oxidative stress is a central pathological mechanism in neurodegenerative diseases, yet systematic characterization of secretome alterations under oxidative conditions remains limited. We established an in vitro model to investigate how oxidative challenge modulates extracellular protein profiles in human neuronal cells. SH-SY5Y neuroblastoma cells were exposed to 150 microM hydrogen peroxide for 24 hours under serum-free conditions to mimic pathological oxidative stress. Conditioned media were collected and analyzed using mass spectrometry-based proteomics. Differentially expressed proteins were subjected to Gene Ontology functional enrichment and Reactome pathway analysis using g:Profiler. Proteomic profiling identified 2,029 proteins with significantly reduced abundance following oxidative treatment, indicating broad suppression of secreted and extracellular proteins. Functional enrichment analyses revealed marked downregulation of biological processes critical for neuronal maintenance, including extracellular matrix organization, calcium ion binding, and cell adhesion. Notably, integrin-interacting proteins (ACTN1, ACTN2, ACTN4, COMP), heparin-binding molecules (FBLN1, FBLN2, FBN1), and key structural extracellular matrix components such as collagens (COL1A1, COL3A1), fibronectin (FN1), laminins (LAMA4, LAMA5), von Willebrand factor (VWF) were strongly reduced. In parallel, several transmembrane receptor tyrosine kinases involved in neurotrophic signaling, including EGFR, EPHB2, ALK, BMPR2, showed decreased abundance. These findings demonstrate that oxidative stress induces coordinated suppression of extracellular matrix integrity and receptor-mediated signaling pathways, suggesting an early molecular signature.

This work was supported by Spanish Ministry of Science and Innovation and Agencia Estatal de Investigación plus FEDER Funds through grants PID2023-149767OB-I00 (FJM) and ‘Unidad de Excelencia María de Maeztu’ CEX2024-001431-M funded by MICIU/AEI/10.13039/501100011033 and by ’ERDF A way of making Europe’.