Chronic alpha-Synuclein Over-Expression and Ceruloplasmin Challenge Promote Distinct Iron and Redox Responses in M17 Cells

BackgroundWhile -synuclein (-syn) accumulation and iron dysregulation are hallmarks of Parkinsons Disease, the adaptations that enable neuronal survival under chronic protein stress remain unclear. Here, we investigated how -syn overexpression and ceruloplasmin (Cp)-mediated iron modulation alters iron and redox homeostasis. MethodsWe utilized human BE(2)-M17 neuroblastoma cell lines stably expressing different levels of -syn to examine the interplay between -syn, Ceruloplasmin (Cp)-mediated iron modulation, and the cellular response to oxidative stress. Analyses included Western blotting, immunofluorescence staining, soluble/insoluble fractionation, glutathione, reactive oxygen species (ROS) and hydrogen peroxide (H2O2) quantification, lipid peroxidation, ferrous iron, and cell viability. ResultsOur data suggest an unexpected relationship between chronic -syn expression and cellular redox regulation. Despite carrying a greater -syn burden, cells with higher -syn expression exhibit lower basal ROS, H2O2, and lipid peroxidation compared to control cells. These changes are not accompanied by activation of canonical antioxidant pathways suggesting that the reduced oxidative profile arises through alternative mechanisms. Besides, -syn over-expressing cells display significant remodeling of iron-handling pathways, including altered expression of ferritin heavy chain, transferrin receptor, and ferroportin, suggesting that chronically high -syn levels are associated with changes in iron homeostasis. In addition, this phenotype is not maintained following Cp overexpression. Although Cp reduces Fe{superscript 2} levels, it also induces substantial increases in ROS and H2O2 without corresponding changes in GPX4, glutathione, or related antioxidant systems. Thus, the reduced basal oxidative profile observed in -syn-over-expressing cells does not reflect enhanced canonical antioxidant capacity. Instead, chronically high -syn levels appear to be associated with adaptive remodeling of iron and redox pathways that become sensitive to oxidative imbalance. ConclusionChronic -syn over-expression promotes adaptive remodeling of iron and redox homeostasis, associated with reduced basal oxidative stress but increased sensitivity to Cp-mediated perturbation. These data link -syn burden to iron metabolism and stress-dependent vulnerability in synucleinopathies. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/732494v1_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@9ab785org.highwire.dtl.DTLVardef@186a200org.highwire.dtl.DTLVardef@1f5c13forg.highwire.dtl.DTLVardef@162249e_HPS_FORMAT_FIGEXP M_FIG C_FIG