Search bioRxiv⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.64898/2026.06.15.732494

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

Pourhadi, M.·Ranxhi, B.·Sukaria, S. P.·Hussein, F. H.·Todi, S. V.·LeWitt, P. A.·Tsou, W.-L.
View original source ↗Export metadata ↓

Abstract

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

Explore related subjects

neurosciencePreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Pourhadi, M., Ranxhi, B., Sukaria, S. P., Hussein, F. H., Todi, S. V., LeWitt, P. A., Tsou, W.-L.. 2026-06-16. Chronic alpha-Synuclein Over-Expression and Ceruloplasmin Challenge Promote Distinct Iron and Redox Responses in M17 Cells. https://doi.org/10.64898/2026.06.15.732494

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

A Mechanistic Account of Dedifferentiation in the Aging Visual System

To uncover the mechanisms of age-related dedifferentiation, we simulated neural degradation in a computational model of the ventral visual stream. Severing connections or adding noise, whether in inferotemporal cortex (IT) or upstream, reduced IT differentiation, indicating that dedifferentiation need not originate locally. Degrading peripheral V1, which preferentially drove place-selective IT neurons, selectively reduced place differentiation, a candidate mechanism for the stronger place dedifferentiation reported in aging.

neuroscience↗
bioRxiv2026

Reusing pipettes in Patch-seq: a foundation for automated multimodal profiling

Patch-seq is a technique for single-cell multimodal characterization. It combines whole-cell electrophysiology, single-cell transcriptomics, and morphological reconstruction in single neurons. Patch-seq morphologies are a critical bridge among cell types, genes, physiology, and large-scale connectivity platforms such as EM connectomes and single-cell-resolution projectomes. However, patch-seq experimental complexity and operator dependence limit throughput. Automation could enable scalable Patch-seq but requires procedural adaptations, including detergent-based pipette cleaning, robotic nucleus extraction and collection. The impact of these automations on Patch-seq quality has not been systematically evaluated. Here, we show that each of these modifications preserves or improves key Patch-seq outcomes. Automated nucleus extraction performs comparably to, or better than, expert manual operation, preserving key RNA and morphological metrics. Detergent-cleaned, reused pipettes supported a viable sealing success rate across multiple cleaning cycles and preserved electrophysiological quality and transcriptomic yield without introducing excess contamination. Sonication effectively allows collection of nuclear material without damaging RNA and supports subsequent pipette cleaning and reuse. Enhanced automated sealing and break-in procedures increased throughput regardless of brain region, cell type or species. Together, these results validate the core procedural adaptations required for automated Patch-seq and establish a foundation for high-throughput, standardized multimodal single-cell characterization.

neuroscience↗
bioRxiv2026

Development of direction selectivity in the Xenopus tadpole optic tectum and midbrain tegmentum

Detecting motion direction is a fundamental function performed by direction selective (DS) neurons. DS neurons in the optic tectum (superior colliculus in mammals) enable the tracking of moving objects. DS neurons of the accessory optic system support reflexive eye / head movements that stabilize gaze during self-motion. By developmental stage 48 (10 days postfertilization), Xenopus tadpoles display visually guided behaviors that require both types of DS neurons. Our previous work has shown that these two regions are not only functionally distinct but also follow different developmental programs. Here we characterized and compared how direction selectivity develops in both visual processing centers. Using in vivo electrophysiological recordings during moving bar visual stimulation, we measured DS synaptic inputs and spiking outputs in both regions across three key developmental stages of the retinotectal projection. Tegmental neurons were direction selective ab initio, and both input and output tuning remained stable across development. Tectal neurons were observed to receive DS inputs early, and their input selectivity also remained stable across development. However, their spiking output grew substantially more DS at the later developmental stage. This was due at least in part to a naturally occurring decrease in intrinsic excitability which preferentially dampened overall spike rates, but especially in response to the non-preferred direction of motion, thereby increasing direction selectivity. These results reveal distinct developmental trajectories in parallel visual pathways. Tegmental direction selectivity is established early and stays stable, whereas tectal output tuning sharpens during development through a decrease in intrinsic excitability.

neuroscience↗