Potential convergence of olfactory dysfunction in Parkinson's disease and COVID-19: The role of neuroinflammation.

Parkinson's disease (PD) is a prevalent neurodegenerative disorder that affects 7-10 million individuals worldwide. A common early symptom of PD is olfactory dysfunction (OD), and more than 90% of PD patients suffer from OD. Recent studies have highlighted a high incidence of OD in patients with SARS-CoV-2 infection. This review investigates the potential convergence of OD in PD and COVID-19, particularly focusing on the mechanisms by which neuroinflammation contributes to OD and neurological events. Starting from our fundamental understanding of the olfactory bulb, we summarize the clinical features of OD and pathological features of the olfactory bulb from clinical cases and autopsy reports in PD patients. We then examine SARS-CoV-2-induced olfactory bulb neuropathology and OD and emphasize the SARS-CoV-2-induced neuroinflammatory cascades potentially leading to PD manifestations. By activating microglia and astrocytes, as well as facilitating the aggregation of α-synuclein, SARS-CoV-2 could contribute to the onset or exacerbation of PD. We also discuss the possible contributions of NF-κB, the NLRP3 inflammasome, and the JAK/STAT, p38 MAPK, TLR4, IL-6/JAK2/STAT3 and cGAS-STING signaling pathways. Although olfactory dysfunction in patients with COVID-19 may be reversible, it is challenging to restore OD in patients with PD. With the emergence of new SARS-CoV-2 variants and the recurrence of infections, we call for continued attention to the intersection between PD and SARS-CoV-2 infection, especially from the perspective of OD.

Yeast Prion Protein Sup35 Initiates α-Synuclein Pathology in Parkinson's Disease.

Sinus infection of Saccharomyces cerevisiae accelerates the aggregation of α-synuclein (α-syn) in A53T mice, which was caused by prion protein Sup35. Sup35 promotes α-syn aggregation in vitro and in vivo and leads to Parkinson's disease (PD)-like motor impairment in wildtype mice, suggesting that the yeast Sup35 triggers α-syn pathology in PD.

Ndfip1 protected dopaminergic neurons via regulating mitochondrial function and ferroptosis in Parkinson's disease.

Increasing evidence has shown that mitochondrial dysfunction and iron accumulation contribute to the pathogenesis of Parkinson's disease (PD). Nedd4 family interacting protein 1 (Ndfip1) is an adaptor protein of the Nedd4 E3 ubiquitin ligases. We have previously reported that Ndfip1 showed a neuroprotective effect in cell models of PD. However, whether Ndfip1 could protect dopaminergic neurons in PD animal models in vivo and the possible mechanisms are not known. Here, our results showed that the expression of Ndfip1 decreased in the substantia nigra (SN) of 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced PD mouse model. Overexpression of Ndfip1 could improve MPTP-induced motor dysfunction significantly and antagonize the loss of dopaminergic neurons in the SN of MPTP-induced mice. Further study showed that overexpression of Ndfip1 might protect against MPTP-induced neurotoxicity through regulation of voltage-dependent anion-selective channel (VDAC). In addition, we observed the downregulation of Ndfip1 and upregulation of VDAC1/2 in 1-methyl-4-phenylpyridinium ion (MPP+)-induced SH-SY5Y cells. Furthermore, high expression of Ndfip1 in SH-SY5Y cells inhibited MPP+-induced increase of VDAC1/2 and restored MPP+-induced mitochondrial dysfunction. Furthermore, Ndfip1 prevented MPP+-induced increase in the expression of long-chain acyl-CoA synthetase 4 (ACSL4), suggesting the possible role of Ndfip1 in regulating ferroptosis. Our results provide new evidence for the neuroprotective effect of Ndfip1 on dopaminergic neurons in PD animal models and provide promising targets for the treatment of iron-related diseases, including PD.

Characterization of graded 6-Hydroxydopamine unilateral lesion in medial forebrain bundle of mice.

Parkinson's disease (PD) is the second most common age-related neurodegenerative disease, with a progressive loss of dopaminergic cells and fibers. The purpose of this study was to use different doses of 6-hydroxydopamine (6-OHDA) injection into the medial forebrain bundle (MFB) of mice to mimic the different stages of the disease and to characterize in detail their motor and non-motor behavior, as well as neuropathological features in the nigrostriatal pathway. MFB were injected with 0.5 µg, 1 µg, 2 µg of 6-OHDA using a brain stereotaxic technique. 6-OHDA induced mitochondrial damage dose-dependently, as well as substantia nigra pars compacta (SNpc) tyrosine hydroxylase-positive (TH+) cell loss and striatal TH fiber loss. Activation of astrocytes and microglia in the SNpc and striatum were consistently observed at 7 weeks, suggesting a long-term glial response in the nigrostriatal system. Even with a partial or complete denervation of the nigrostriatal pathway, 6-OHDA did not cause anxiety, although depression-like behavior appeared. Certain gait disturbances were observed in 0.5 µg 6-OHDA lesioned mice, and more extensive in 1 µg group. Despite the loss of more neurons from 2 µg 6-OHDA, there was no further impairment in behaviors compared to 1 µg 6-OHDA. Our data have implications that 1 µg 6-OHDA was necessary and sufficient to induce motor and non-motor symptoms in mice, thus a valuable mouse tool to explore disease progression and new treatment in PD.

Contrasting Iron Metabolism in Undifferentiated Versus Differentiated MO3.13 Oligodendrocytes via IL-1ß-Induced Iron Regulatory Protein 1.

Parkinson's disease (PD) is a prevalent neurodegenerative disorder characterized by the loss of dopaminergic neurons and the accumulation of iron in the substantia nigra. While iron accumulation and inflammation are implicated in PD pathogenesis, their impact on oligodendrocytes, the brain's myelin-forming cells, remains elusive. This study investigated the influence of interleukin-1ß (IL-1ß), an elevated proinflammatory cytokine in PD, on iron-related proteins in MO3.13 oligodendrocytes. We found that IL-1ß treatment in undifferentiated MO3.13 oligodendrocytes increased iron regulatory protein 1 and transferrin receptor 1 (TfR1) expression while decreasing ferroportin 1 (FPN1) expression. Consequently, iron uptake was enhanced, and iron release was reduced, leading to intracellular iron accumulation. Conversely, IL-1ß treatment in differentiated MO3.13 oligodendrocytes exhibited the opposite effect, with decreased TfR1 expression, increased FPN1 expression, and reduced iron uptake. These findings suggest that IL-1ß-induced dysregulation of iron metabolism in oligodendrocytes may contribute to the pathological processes observed in PD. IL-1ß can increase the iron content in undifferentiated oligodendrocytes, thus facilitating the differentiation of undifferentiated MO3.13 oligodendrocytes. In differentiated oligodendrocytes, IL-1ß may facilitate iron release, providing a potential source of iron for neighboring dopaminergic neurons, thereby initiating a cascade of deleterious events. This study provides valuable insights into the intricate interplay between inflammation, abnormal iron accumulation, and oligodendrocyte dysfunction in PD. Targeting the IL-1ß-mediated alterations in iron metabolism may hold therapeutic potential for mitigating neurodegeneration and preserving dopaminergic function in PD.

Mitochondrial iron dyshomeostasis and its potential as a therapeutic target for Parkinson's disease.

Abnormal iron accumulation has been implicated in the etiology of Parkinson's disease (PD). Understanding how iron damages dopaminergic neurons in the substantia nigra (SN) of PD is particularly important for developing targeted neurotherapeutic strategies for the disease. However, it is still not fully understood how excess iron contributes to the neurodegeneration of dopaminergic neurons in PD. There has been increased attention on mitochondrial iron dyshomeostasis, iron-induced mitochondrial dysfunction and ferroptosis in PD. Therefore, this review begins with a brief introduction to describe cellular iron metabolism and the dysregulation of iron metabolism in PD. Then we provide an update on how iron is delivered to mitochondria and induces the damage of dopaminergic neurons in PD. In addition, we also summarize new research progress on iron-dependent ferroptosis in PD and mitochondria-localized proteins involved in ferroptosis. This will provide new insight into potential therapeutic strategies targeting mitochondrial iron dysfunction.

Cell senescence induced by toxic interaction between α-synuclein and iron precedes nigral dopaminergic neuron loss in a mouse model of Parkinson's disease.

Cell senescence has been implicated in the pathology of Parkinson's disease (PD). Both abnormal α-synuclein aggregation and iron deposition are suggested to be the triggers, facilitators, and aggravators during the development of PD. In this study, we investigated the involvement of α-synuclein and iron in the process of cell senescence in a mouse model of PD. In order to overexpress α-syn-A53T in the substantia nigra pars compacta (SNpc), human α-syn-A53T was microinjected into both sides of the SNpc in mice. We found that overexpression of α-syn-A53T for one week induced significant pro-inflammatory senescence-associated secretory phenotype (SASP), increased cell senescence-related proteins (ß-gal, p16, p21, H2A.X and γ-H2A.X), mitochondrial dysfunction accompanied by dysregulation of iron-related proteins (L-ferritin, H-ferritin, DMT1, IRP1 and IRP2) in the SNpc. In contrast, significant loss of nigral dopaminergic neurons and motor dysfunction were only observed after overexpression of α-syn-A53T for 4 weeks. In PC12 cells stably overexpressing α-syn-A53T, iron overload (ferric ammonium citrate, FAC, 100 µM) not only increased the level of reactive oxygen species (ROS), p16 and p21, but also exacerbated the processes of oxidative stress and cell senescence signalling induced by α-syn-A53T overexpression. Interestingly, reducing the iron level with deferoxamine (DFO) or knockdown of transferrin receptor 1 (TfR1) significantly improved both the phenotypes and dysregulated proteins of cell senescence induced by α-syn-A53T overexpression. All these evidence highlights the toxic interaction between iron and α-synuclein inducing cell senescence, which precedes nigral dopaminergic neuronal loss in PD. Further investigation on cell senescence may yield new therapeutic agents for the prevention or treatment of PD.

Gut microbiota-induced CXCL1 elevation triggers early neuroinflammation in the substantia nigra of Parkinsonian mice.

Gut microbiota disturbance and systemic inflammation have been implicated in the degeneration of dopaminergic neurons in Parkinson's disease (PD). How the alteration of gut microbiota results in neuropathological events in PD remains elusive. In this study, we explored whether and how environmental insults caused early neuropathological events in the substantia nigra (SN) of a PD mouse model. Aged (12-month-old) mice were orally administered rotenone (6.25 mg·kg-1·d-1) 5 days per week for 2 months. We demonstrated that oral administration of rotenone to ageing mice was sufficient to establish a PD mouse model and that microglial activation and iron deposition selectively appeared in the SN of the mice prior to loss of motor coordination and dopaminergic neurons, and these events could be fully blocked by microglial elimination with a PLX5622-formulated diet. 16 S rDNA sequencing analysis showed that the gut microbiota in rotenone-treated mice was altered, and mice receiving faecal microbial transplantation (FMT) from ageing mice treated with rotenone for 2 months exhibited the same pathology in the SN. We demonstrated that C-X-C motif chemokine ligand-1 (CXCL1) was an essential molecule, as intravenous injection of CXCL1 mimicked almost all the pathology in serum and SN induced by oral rotenone and FMT. Using metabolomics and transcriptomics analyses, we identified the PPAR pathway as a key pathway involved in rotenone-induced neuronal damage. Inhibition of the PPARγ pathway was consistent in the above models, whereas its activation by linoleic acid (60 mg·kg-1·d-1, i.g. for 1 week) could block these pathological events in mice intravenously injected with CXCL1. Altogether, these results reveal that the altered gut microbiota resulted in neuroinflammation and iron deposition occurring early in the SN of ageing mice with oral administration of rotenone, much earlier than motor symptoms and dopaminergic neuron loss. We found that CXCL1 plays a crucial role in this process, possibly via PPARγ signalling inhibition. This study may pave the way for understanding the "brain-gut-microbiota" molecular regulatory networks in PD pathogenesis. The aged C57BL/6 male mice with rotenone intragastric administration showed altered gut microbiota, which caused systemic inflammation, PPARγ signalling inhibition and neuroinflammation, brain iron deposition and ferroptosis, and eventually dopaminergic neurodegeneration in PD.

Ferritin Is Secreted from Primary Cultured Astrocyte in Response to Iron Treatment via TRPML1-Mediated Exocytosis.

Impaired iron homeostasis has been proven to be one of the critical contributors to the pathology of Parkinson's disease (PD). Ferritin is considered an intracellular protein responsible for storing cytosolic iron. Recent studies have found that ferritin can be secreted from cells independent of the classical endoplasmic reticulum-Golgi system. However, the precise mechanisms underlying the secretion of ferritin in the brain were not elucidated. In the present study, we demonstrated that the primary cultured astrocytes do have the ability to secrete ferritin, which is enhanced by iron treatment. Increased ferritin secretion was accompanied by increased protein expression of ferritin response to iron stimulation. Further study showed that iron-induced expression and secretion of ferritin could be inhibited by CQ or 3-MA pretreatment. In addition, the knockdown of transient receptor potential mucolipin 1 (TRPML1) antagonized iron-induced ferritin secretion, accompanied by further increased intracellular protein levels of ferritin. Further study demonstrated that ferritin colocalized with LAMP1 in iron-treated astrocytes. On the contrary, ras-associated protein 27a (Rab27a) knockdown further enhanced iron-induced ferritin secretion and decreased intracellular protein levels of ferritin. Furthermore, we also showed that the secretory autophagy protein tripartite motif containing 16 (TRIM16) and sec22b decreased in iron-treated astrocytes. These results suggested that astrocytes might secrete ferritin via TRPML1-mediated exocytosis. This provides new evidence for the mechanisms underlying the secretion of ferritin in primary cultured astrocytes under a high iron environment.

Dopaminergic neurodegeneration in the substantia nigra is associated with olfactory dysfunction in mice models of Parkinson's disease.

Olfactory dysfunction represents a prodromal stage in Parkinson's disease (PD). However, the mechanisms underlying hyposmia are not specified yet. In this study, we first observed an early olfactory dysfunction in mice with intragastric rotenone administration, consistent with dopaminergic neurons loss and α-synuclein pathology in the olfactory bulb. However, a much severer olfactory dysfunction was observed without severer pathology in olfactory bulb when the loss of dopaminergic neurons in the substantia nigra occurred. Then, we established the mice models by intrastriatal α-synuclein preformed fibrils injection and demonstrated the performance in the olfactory discrimination test was correlated to the loss of dopaminergic neurons in the substantia nigra, without any changes in the olfactory bulb analyzed by RNA-sequence. In mice with intranasal ferric ammonium citrate administration, we observed olfactory dysfunction when dopaminergic neurodegeneration in substantia nigra occurred and was restored when dopaminergic neurons were rescued. Finally we demonstrated that chemogenetic inhibition of dopaminergic neurons in the substantia nigra was sufficient to cause hyposmia and motor incoordination. Taken together, this study shows a direct relationship between nigral dopaminergic neurodegeneration and olfactory dysfunction in PD models and put forward the understandings that olfactory dysfunction represents the early stage of neurodegeneration in PD progression.

TFEB regulates cellular labile iron and prevents ferroptosis in a TfR1-dependent manner.

Autophagy is a major clearance pathway for misfolded α-synuclein which promotes ferroptosis through NCOA4-mediated ferritin degradation. The regulation of these two processes to achieve improved neuroprotection in Parkinson's disease (PD) must be elucidated. Transcription factor EB (TFEB) is a master regulator of both autophagy and lysosome biogenesis, and lysosomes are important cellular iron storage organelles; however, the role of TFEB in ferroptosis and iron metabolism remains unclear. In this study, TFEB overexpression promoted the clearance of misfolded α-synuclein and prevented ferroptosis and iron overload. TFEB overexpression up-regulated transferrin receptor 1 (TfR1) synthesis and increased the localization of TfR1 in the lysosome, facilitating lysosomal iron import and transient lysosomal iron storage. TFEB overexpression increased the levels of cellular iron-safe storage proteins (both ferritin light and heavy chains). These functions in iron metabolism maintain the cellular labile iron at a low level and electrical activity, even under iron overload conditions. Notably, lower levels of cellular labile iron and the upregulation of ferritin light and heavy chains were reversed after TfR1 knockdown in cells overexpressing TFEB, indicating that TFEB regulates cellular labile iron and suppresses ferroptosis in a TfR1 dependent manner. Taken together, this evidence of the regulation of iron metabolism enriches our understanding of the function of TFEB. In addition, TFEB overexpression protects against ferroptosis and iron overload and provides a new direction and perspective for autophagy regulation in PD.

Heme oxygenase-1: The roles of both good and evil in neurodegenerative diseases.

Heme oxygenase-1 (HO-1) is the only way for cells to decompose heme. It can cleave heme to produce carbon monoxide (CO), ferrous iron (Fe2+ ), and biliverdin (BV). BV is reduced to bilirubin (BR) by biliverdin reductase(BVR). In previous studies, HO-1 was considered to have protective effects because of its anti-inflammatory, anti-apoptosis, and antiproliferation functions. However, emerging experimental studies have found that the metabolites derived from HO-1 can cause increase iin intracellular oxidative stress, mitochondrial damage, iron death, and autophagy. Because of its particularity, it is very meaningful to understand its exact mechanism. In this review, we summarized the protective and toxic effects of HO-1, its potential mechanism, its role in neurodegenerative diseases and related drug research. This knowledge may be beneficial to the development of new therapies for neurodegenerative diseases and is crucial to the development of new therapeutic strategies and biomarkers.

Alpha-synuclein Affects Certain Iron Transporters of BV2 Microglia Cell through its ferric reductase activity.

Alpha-synuclein (α-syn) is a major component of lewy bodies, which is biomarker of Parkinson's disease (PD). It accumulates in substantia nigra pars compacts (SNpc) to form insoluble aggregates and cause neurotoxicity, which is often accompanied by iron deposition. In this study, we compared the iron reductase activity between monomeric α-syn (M-α-syn) and oligomeric α-syn (O-α-syn), investigated the effect of α-syn on iron metabolism of BV2 microglia cells as well. We found that α-syn had ferric reductase activity, and O-α-syn had stronger enzyme activity than M-α-syn. M-α-syn upregulated iron uptake protein, divalent metal transporter1 (DMT1) expression and iron influx, but did not regulate iron release protein, ferroportin1 (FPN1) expression and iron efflux. O-α-syn elevated the expression of both DMT1 and FPN1, thus increased the iron influx and efflux in BV2 microglial cells, but the expressions of iron regulatory protein1 and hypoxia inducible factor2α have no significant change. Moreover, both M-α-syn and O-α-syn could increase the mRNA expressions of TNF-α and IL-1ß in BV2 microglia cells. Taken together, our data suggest that both types of α-syn can activate microglia, which leads to increased expressions of pro-inflammatory factors. α-syn can affect DMT1 and FPN1 expressions in BV2 microglia cells, which might be through its ferric reductase activity.

Chemogenetic and optogenetic stimulation of zona incerta GABAergic neurons ameliorates motor impairment in Parkinson's disease.

Parkinson's disease (PD) is characterized by the degeneration of dopaminergic neurons in the substantia nigra and leads to progressive motor dysfunction. While studies have focused on the basal ganglia network, recent evidence suggests neuronal systems outside the basal ganglia are also related to PD pathogenesis. The zona incerta (ZI) is a predominantly inhibitory subthalamic region for global behavioral modulation. This study investigates the role of GABAergic neurons in the ZI in a mouse model of 6-hydroxydopamine (6-OHDA)-induced PD. First, we found a decrease in GABA-positive neurons in the ZI, and then the mice used chemogenetic/optogenetic stimulation to activate or inhibit GABAergic neurons. The motor performance of PD mice was significantly improved by chemogenetic/optogenetic activation of GABAergic neurons, and repeated chemogenetic activation of ZI GABAergic neurons increased the dopamine content in the striatum. Our work identifies the role of ZI GABAergic neurons in regulating motor behaviors in 6-OHDA-lesioned PD model mice.

[A glass micropipette vacuum technique of cerebrospinal fluid sampling in C57BL/6 mice].

The purpose of this study was to establish a suitable method for extracting cerebrospinal fluid (CSF) from C57BL/6 mice. A patch clamp electrode puller was used to draw a glass micropipette, and a brain stereotaxic device was used to fix the mouse's head at an angle of 135° from the body. Under a stereoscopic microscope, the skin and muscle tissue on the back of the mouse's head were separated, and the dura mater at the cerebellomedullary cistern was exposed. The glass micropipette (with an angle of 20° to 30° from the dura mater) was used to puncture at a point 1 mm inboard of Y-shaped dorsal vertebral artery for CSF sampling. After the first extraction, the glass micropipette was connected with a 1 mL sterile syringe to form a negative pressure device for the second extraction. The results showed that the successful rate of CSF extraction was 83.33% (30/36). Average CSF extraction amount was (7.16 ± 0.43) µL per mouse. In addition, C57BL/6 mice were given intranasally ferric ammonium citrate (FAC) to establish a model of brain iron accumulation, and the CSF extraction technique established in the present study was used for sampling. The results showed that iron content in the CSF from the normal saline control group was not detected, while the iron content in the CSF from FAC-treated group was (76.24 ± 38.53) µmol/L, and the difference was significant. These results suggest that glass micropipette vacuum technique of CSF sampling established in the present study has the advantages of simplicity, high success rate, large extraction volume, and low bleeding rate, and is suitable for the research on C57BL/6 mouse neurological disease models.

Intraperitoneal injection of iron dextran induces peripheral iron overload and mild neurodegeneration in the nigrostriatal system in C57BL/6 mice.

AIMS: Elevated iron levels in the affected areas of brain are linked to several neurodegenerative diseases including Parkinson's disease (PD). This study investigated the influence of peripheral iron overload in peripheral tissues, as well as its entry into the brain regions on lysosomal functions. The survival of dopaminergic neurons in the nigrostriatal system and motor coordination were also investigated. MAIN METHODS: An intraperitoneal injection of iron dextran (FeDx) mouse model was established. Western blot was used to detect iron deposition and lysosomal functions in the liver, spleen, hippocampal (HC), striatum (STR), substantia nigra (SN) and olfactory bulb (OB). Iron in serum and cerebrospinal fluid (CSF) was determined by an iron assay kit. Immunofluorescence and immunohistochemical staining were applied to detect dopaminergic neurons and fibers. Motor behavior was evaluated by gait analysis. KEY FINDINGS: Iron was deposited consistently in the liver and spleen, and serum iron was elevated. While iron deposition occurred late in the HC, STR and SN, without apparently affecting CSF iron levels. Although cathepsin B (CTSB), cathepsin D (CTSD), glucocerebrosidase (GCase) and lysosome integrated membrane protein 2 (LIMP-2) protein levels were dramatically up-regulated in the liver and spleen, they were almost unchanged in the brain regions. However, CTSB was up-regulated in acute iron-overloaded OB and primary cultured astrocytes. The number of dopaminergic neurons in the SN remained unchanged, and mice did not exhibit significant motor incoordination. SIGNIFICANCE: Intraperitoneal injection of FeDx in mice induces largely peripheral iron overload while not necessarily sufficient to cause severe disruption of the nigrostriatal system.

4-Aminopyridine Protects Nigral Dopaminergic Neurons in the MPTP Mouse Model of Parkinson's Disease.

Various pharmacological blockers targeting K+ channel have been identified to be related to the treatment of Parkinson's disease (PD). Previous studies showed that 4-Aminopyridine (4-AP), a wide-spectrum K+ channel blocker, was able to attenuate apomorphine-induced rotation in parkinsonism rats, indicating the possible beneficial effects in attenuation of PD motor symptoms. However, it is unclear whether 4-AP exhibits neuroprotective effects against the neurodegeneration of substantia nigra (SN)-striatum system in PD. In this study, the 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mouse model was employed to evaluate the neuroprotective effects of 4-AP. Results showed that 4-AP inhibited MPTP-induced dopaminergic neuronal loss in the SN as well as dopamine depletion in the striatum. Behavior indexes of open field test and rotarod test confirmed that 4-AP attenuated MPTP-induced motor deficits. We also showed that 4-AP treatment could significantly attenuate the MPTP-induced increase in malonaldehyde (MDA) levels and decrease in superoxide dismutase (SOD) levels. Additionally, MPTP significantly reduced the Bcl-2 expression and promoted the Caspase-3 activation; 4-AP protected dopaminergic neurons against MPTP-induced neurotoxicity by reversing these changes. These results indicate that 4-AP exerts a neuroprotective effect on dopaminergic neurons against MPTP by decreasing oxidative stress and apoptosis. This provides a promising therapeutic target for the treatment of PD.

Iron Overload Causes Ferroptosis But Not Apoptosis in MO3.13 Oligodendrocytes.

Oligodendrocytes are the most iron-rich cells in the brain. Studies have shown that oligodendrocytes are very sensitive to oxidative stress, and iron overload is more likely to cause damage to oligodendrocytes. The purpose of this experiment was to investigate the damaging effect and mechanism of ferric ammonium citrate (FAC) on MO3.13 oligodendrocytes. In FAC treatment group, the intracellular iron concentration and intracellular reactive oxygen species were increased. There were no obvious changes in nucleus and chromatin, but increased mitochondrial membrane density, decreased mitochondrial cristae and mitochondrial length were observed. Glutathione peroxidase 4 (GPX4) expression was decreased, but the ratio of Bcl-2/Bax protein levels and cleaved caspase-3 expression did not change. Moreover, the iron chelator deferoxamine (DFO) and the ferroptosis inhibitor ferrostatin-1(Fer-1) could inhibit the upregulation of GPX4, which indicating that DFO and Fer-1 could inhibit ferroptosis in MO3.13 oligodendrocytes induced by iron overload. Furthermore, the phosphorylation level of p53 was not changed, while the ratio of protein expressions of p-Erk1/2/Erk1/2 were markedly increased. Taken together, our data suggest that iron overload induces ferroptosis but not apoptosis in oligodendrocytes. The mechanism may be related to mitogen-activated protein kinase pathway activation rather than p53 pathway activation.