NCOA4 requires a [3Fe-4S] to sense and maintain the iron homeostasis.

NCOA4 is a selective cargo receptor for ferritinophagy, the autophagic turnover of ferritin (FTH), a process critical for regulating intracellular iron bioavailability. However, how ferritinophagy flux is controlled through NCOA4 in iron-dependent processes needs to be better understood. Here, we show that the C-terminal FTH-binding domain of NCOA4 harbors a [3Fe-4S]-binding site with a stoichiometry of approximately one labile [3Fe-4S] cluster per NCOA4 monomer. By analyzing the interaction between NCOA4 and HERC2 ubiquitin ligase or NCOA4 and FTH, we demonstrate that NCOA4 regulates ferritinophagy by sensing the intracellular iron-sulfur cluster levels. Under iron-repletion conditions, HERC2 recognizes and recruits holo-NCOA4 as a substrate for polyubiquitination and degradation, favoring ferritin iron storage. Under iron-depletion conditions, NCOA4 exists in the form of apo-protein and binds ferritin to promote the occurrence of ferritinophagy and release iron. Thus, we identify an iron-sulfur cluster [3Fe-4S] as a critical cofactor in determining the fate of NCOA4 in favoring iron storage in ferritin or iron release via ferritinophagy and provide a dual mechanism for selective interaction between HERC2 and [3Fe-4S]-NCOA4 for proteasomal degradation or between ferritin and apo-NCOA4 for ferritinophagy in the control of iron homeostasis.

SS-31 efficacy in a mouse model of Friedreich ataxia by upregulation of frataxin expression.

Friedreich ataxia (FRDA) is a serious hereditary neurodegenerative disease, mostly accompanied with hypertrophic cardiomyopathy, caused by the reduced expression of frataxin (FXN). However, there is still no effective treatment. Our previous studies have shown that SS-31, a mitochondrion-targeted peptide, is capable to upregulate the expression of FXN and improve the mitochondrial function in cells derived from FRDA patients. To further explore the potential of SS-31, we used the GAA expansion-based models, including Y47 and YG8R (Fxn KIKO) mice, primary neurons and macrophages from the mice and cells derived from FRDA patients. After once-daily intraperitoneal injection of 1 mg/kg SS-31 for 1 month, we observed the significant improvement of motor function. The vacuolation in dorsal root ganglia, lesions in dentate nuclei and the lost thickness of myelin sheath of spinal cord were all repaired after SS-31 treatment. In addition, the hypertrophic cardiomyocytes and disarrayed abnormal Purkinje cells were dramatically reduced. Interestingly, we found that SS-31 treatment upregulated FXN expression not only at the translational levels as observed in cell culture but also at mRNA levels in vivo. Consequently, mitochondrial morphology and function were greatly improved in all tested tissues. Importantly, our data provided additional evidence that the maintenance of the therapeutic benefits needed continuous drug administration. Taken together, our findings have demonstrated the effectiveness of SS-31 treatment through the upregulation of FXN in vivo and offer guidance of the potential usage in the clinical application for FRDA.

Rescuers from the Other Shore: Intercellular Mitochondrial Transfer and Its Implications in Central Nervous System Injury and Diseases.

As the powerhouse and core of cellular metabolism and survival, mitochondria are the essential organelle in mammalian cells and maintain cellular homeostasis by changing their content and morphology to meet demands through mitochondrial quality control. It has been observed that mitochondria can move between cells under physiological and pathophysiological conditions, which provides a novel strategy for preserving mitochondrial homeostasis and also a therapeutic target for applications in clinical settings. Therefore, in this review, we will summarize currently known mechanisms of intercellular mitochondrial transfer, including modes, triggers, and functions. Due to the highly demanded energy and indispensable intercellular linkages of the central nervous system (CNS), we highlight the mitochondrial transfer in CNS. We also discuss future application possibilities and difficulties that need to be addressed in the treatment of CNS injury and diseases. This clarification should shed light on its potential clinical applications as a promising therapeutic target in neurological diseases. Intercellular mitochondrial transfer maintains the homeostasis of central nervous system (CNS), and its alteration is related to several neurological diseases. Supplementing exogenous mitochondrial donor cells and mitochondria, or utilizing some medications to regulate the process of transfer might mitigate the disease and injury.

Ironing out macrophages in atherosclerosis.

The most common cause of death worldwide is atherosclerosis and related cardiovascular disorders. Macrophages are important players in the pathogenesis of atherosclerosis and perform critical functions in iron homeostasis due to recycling iron by phagocytosis of senescent red blood cells and regulating iron availability in the tissue microenvironment. With the growth of research on the "iron hypothesis" of atherosclerosis, macrophage iron has gradually become a hotspot in the refined iron hypothesis. Macrophages with the M1, M2, M(Hb), Mox, and other phenotypes have been defined with different iron-handling capabilities related to the immune function and immunometabolism of macrophages, which influence the progression of atherosclerosis. In this review, we focus on macrophage iron and its effects on the development of atherosclerosis. We also cover the contradictory discoveries and propose a possible explanation. Finally, pharmaceutical modulation of macrophage iron is discussed as a promising target for atherosclerosis therapy.

Oridonin attenuates the progression of atherosclerosis by inhibiting NLRP3 and activating Nrf2 in apolipoprotein E-deficient mice.

Oridonin, a well-known traditional Chinese herbal medicinal product isolated from Isodon rubescens (Hemsl.) H.Hara, has many potential properties, including anti-inflammatory and antioxidant activities. However, there is no evidence whether oridonin have a protective effect on atherosclerosis. This study focused on the effects of oridonin on oxidative stress and inflammation generated from atherosclerosis. The therapeutic effect on atherosclerosis was evaluated by intraperitoneal injection of oridonin in a high-fat fed ApoE-/- mouse model. We isolated mouse peritoneal macrophages and detected the effect of oridonin on oxidized low-density lipoprotein-induced lipid deposition. Oil red O staining, Masson's staining, dihydroethidium fluorescence staining, immunohistochemical staining, western blotting analysis, immunofluorescence, enzyme-linked immunosorbent assay and quantitative real-time PCR were used to evaluate the effect on atherosclerosis and explore the mechanisms. Oridonin treatment significantly alleviated the progression of atherosclerosis, reduced macrophage infiltration and stabilized plaques. Oridonin could significantly inhibit inflammation associated with NLRP3 activation. Oridonin significantly reduced oxidative stress by blocking Nrf2 ubiquitination and degradation. We also found that oridonin could prevent the formation of foam cells by increasing lipid efflux protein and reducing lipid uptake protein in macrophages. Oridonin has a protective effect on atherosclerosis in ApoE-/- mice, which may be related to the inhibition of NLRP3 and the stabilization of Nrf2. Therefore, oridonin may be a potential therapeutic agent for atherosclerosis.

Cur@SF NPs alleviate Friedreich's ataxia in a mouse model through synergistic iron chelation and antioxidation.

Abnormal iron metabolism, mitochondrial dysfunction and the derived oxidative damage are the main pathogeneses of Friedrich's ataxia (FRDA), a single-gene inherited recessive neurodegenerative disease characterized by progressive cerebellar and sensory ataxia. This disease is caused by frataxin (FXN) mutation, which reduces FXN expression and impairs iron sulfur cluster biogenesis. To date, there is no effective therapy to treat this condition. Curcumin is proposed harboring excellent ability to resist oxidative stress through Nrf2 activation and its newly found ability to chelate iron. However, its limitation is its poor water solubility and permeability. Here, we synthesized slow-release nanoparticles (NPs) by loading curcumin (Cur) into silk fibroin (SF) to form NPs with an average size of 150 nm (Cur@SF NPs), which exhibited satisfactory therapeutic effects on the improvement of FRDA manifestation in lymphoblasts (1 µM) derived from FRDA patients and in YG8R mice (150 mg/kg/5 days). Cur@SF NPs not only removed iron from the heart and diminished oxidative stress in general but also potentiate iron-sulfur cluster biogenesis, which compensates FXN deficiency to improve the morphology and function of mitochondria. Cur@SF NPs showed a significant advantage in neuron and myocardial function, thereby improving FRDA mouse behavior scores. These data encourage us to propose that Cur@SF NPs are a promising therapeutic compound in the application of FRDA disease.

Iron regulatory protein 2 modulates the switch from aerobic glycolysis to oxidative phosphorylation in mouse embryonic fibroblasts.

The importance of the role of iron regulatory proteins (IRPs) in mitochondrial iron homeostasis and function has been raised. To understand how an IRP affects mitochondrial function, we used globally Irp2-depleted mouse embryonic fibroblasts (MEFs) and found that Irp2 ablation significantly induced the expression of both hypoxia-inducible factor subunits, Hif1α and Hif2α. The increase of Hif1α up-regulated its targeted genes, enhancing glycolysis, and the increase of Hif2α down-regulated the expression of iron-sulfur cluster (Fe-S) biogenesis-related and electron transport chain (ETC)-related genes, weakening mitochondrial respiration. Inhibition of Hif1α by genetic knockdown or a specific inhibitor prevented Hif1α-targeted gene expression, leading to decreased aerobic glycolysis. Inhibition of Hif2α by genetic knockdown or selective disruption of the heterodimerization of Hif2α and Hif1ß restored the mitochondrial ETC and coupled oxidative phosphorylation (OXPHOS) by enhancing Fe-S biogenesis and increasing ETC-related gene expression. Our results indicate that Irp2 modulates the metabolic switch from aerobic glycolysis to OXPHOS that is mediated by Hif1α and Hif2α in MEFs.

Iron overload contributes to general anaesthesia-induced neurotoxicity and cognitive deficits.

BACKGROUND: Increasing evidence suggests that multiple or long-time exposure to general anaesthesia (GA) could be detrimental to cognitive development in young subjects and might also contribute to accelerated neurodegeneration in the elderly. Iron is essential for normal neuronal function, and excess iron in the brain is implicated in several neurodegenerative diseases. However, the role of iron in GA-induced neurotoxicity and cognitive deficits remains elusive. METHODS: We used the primary hippocampal neurons and rodents including young rats and aged mice to examine whether GA impacted iron metabolism and whether the impact contributed to neuronal outcomes. In addition, a pharmacological suppression of iron metabolism was performed to explore the molecular mechanism underlying GA-mediated iron overload in the brain. RESULTS: Our results demonstrated that GA, induced by intravenous ketamine or inhalational sevoflurane, disturbed iron homeostasis and caused iron overload in both in vitro hippocampal neuron culture and in vivo hippocampus. Interestingly, ketamine- or sevoflurane-induced cognitive deficits, very likely, resulted from a novel iron-dependent regulated cell death, ferroptosis. Notably, iron chelator deferiprone attenuated the GA-induced mitochondrial dysfunction, ferroptosis, and further cognitive deficits. Moreover, we found that GA-induced iron overload was activated by NMDAR-RASD1 signalling via DMT1 action in the brain. CONCLUSION: We conclude that disturbed iron metabolism may be involved in the pathogenesis of GA-induced neurotoxicity and cognitive deficits. Our study provides new vision for consideration in GA-associated neurological disorders.

Ferritin is secreted via 2 distinct nonclassical vesicular pathways.

Ferritin turnover plays a major role in tissue iron homeostasis, and ferritin malfunction is associated with impaired iron homeostasis and neurodegenerative diseases. In most eukaryotes, ferritin is considered an intracellular protein that stores iron in a nontoxic and bioavailable form. In insects, ferritin is a classically secreted protein and plays a major role in systemic iron distribution. Mammalian ferritin lacks the signal peptide for classical endoplasmic reticulum-Golgi secretion but is found in serum and is secreted via a nonclassical lysosomal secretion pathway. This study applied bioinformatics and biochemical tools, alongside a protein trafficking mouse models, to characterize the mechanisms of ferritin secretion. Ferritin trafficking via the classical secretion pathway was ruled out, and a 2:1 distribution of intracellular ferritin between membrane-bound compartments and the cytosol was observed, suggesting a role for ferritin in the vesicular compartments of the cell. Focusing on nonclassical secretion, we analyzed mouse models of impaired endolysosomal trafficking and found that ferritin secretion was decreased by a BLOC-1 mutation but increased by BLOC-2, BLOC-3, and Rab27A mutations of the cellular trafficking machinery, suggesting multiple export routes. A 13-amino-acid motif unique to ferritins that lack the secretion signal peptide was identified on the BC-loop of both subunits and plays a role in the regulation of ferritin secretion. Finally, we provide evidence that secretion of iron-rich ferritin was mediated via the multivesicular body-exosome pathway. These results enhance our understanding of the mechanism of ferritin secretion, which is an important piece in the puzzle of tissue iron homeostasis.

Iron Pathophysiology in Friedreich's Ataxia.

Friedreich's ataxia (FRDA) is a degenerative disease that affects both the central and the peripheral nervous systems and non-neural tissues including, mainly, heart, and endocrine pancreas. It is an autosomal recessive disease caused by a GAA triplet-repeat localized within an Alu sequence element in intron 1 of frataxin (FXN) gene, which encodes a mitochondrial protein FXN. This protein is essential for mitochondrial function by the involvement of iron-sulfur cluster biogenesis. The effects of its deficiency also include disruption of cellular, particularly mitochondrial, iron homeostasis, i.e., relatively more iron accumulated in mitochondria and less iron presented in cytosol. Though iron toxicity is commonly thought to be mediated via Fenton reaction, oxidative stress seems not to be the main problem to result in detrimental effects on cell survival, particularly neuron survival. Therefore, the basic research on FXN function is urgently demanded to understand the disease. This chapter focuses on the outcome of FXN expression, regulation, and function in cellular or animal models of FRDA and on iron pathophysiology in the affected tissues. Finally, therapeutic strategies based on the control of iron toxicity and iron cellular redistribution are considered. The combination of multiple therapeutic targets including iron, oxidative stress, mitochondrial function, and FXN regulation is also proposed.

NLRP3 inflammasome-dependent pyroptosis is proposed to be involved in the mechanism of age-dependent isoflurane-induced cognitive impairment.

Wang Z et al. recently published a paper, titled "Critical role of NLRP3-caspase-1 pathway in age-dependent isoflurane-induced microglial inflammatory response and cognitive impairment". The finding in this paper is consistent with our previous study on NLRP3-caspase-1 pathway. Here, we propose that NLRP3 inflammasome-dependent pyroptosis may be involved in the mechanism of age-dependent isoflurane-induced cognitive impairment and discuss that inhibiting NLRP3 inflammasome activation with a novel inhibitor MCC950 may ameliorate age-dependent isoflurane-induced neuro-inflammation.

Tuning the Dipole-Dipole Interaction in a Quantum Gas with a Rotating Magnetic Field.

We demonstrate the tuning of the magnetic dipole-dipole interaction (DDI) within a dysprosium Bose-Einstein condensate by rapidly rotating the orientation of the atomic dipoles. The tunability of the dipolar mean-field energy manifests as a modified gas aspect ratio after time-of-flight expansion. We demonstrate that both the magnitude and the sign of the DDI can be tuned using this technique. In particular, we show that a magic rotation angle exists at which the mean-field DDI can be eliminated, and at this angle, we observe that the expansion dynamics of the condensate is close to that predicted for a nondipolar gas. The ability to tune the strength of the DDI opens new avenues toward the creation of exotic soliton and vortex states as well as unusual quantum lattice phases and Weyl superfluids.

The N-Terminus of Iron-Sulfur Cluster Assembly Factor ISD11 Is Crucial for Subcellular Targeting and Interaction with l-Cysteine Desulfurase NFS1.

Assembly of iron-sulfur (FeS) clusters is an important process in living cells. The initial sulfur mobilization step for FeS cluster biosynthesis is catalyzed by l-cysteine desulfurase NFS1, a reaction that is localized in mitochondria in humans. In humans, the function of NFS1 depends on the ISD11 protein, which is required to stabilize its structure. The NFS1/ISD11 complex further interacts with scaffold protein ISCU and regulator protein frataxin, thereby forming a quaternary complex for FeS cluster formation. It has been suggested that the role of ISD11 is not restricted to its role in stabilizing the structure of NFS1, because studies of single-amino acid variants of ISD11 additionally demonstrated its importance for the correct assembly of the quaternary complex. In this study, we are focusing on the N-terminal region of ISD11 to determine the role of N-terminal amino acids in the formation of the complex with NFS1 and to reveal the mitochondrial targeting sequence for subcellular localization. Our in vitro studies with the purified proteins and in vivo studies in a cellular system show that the first 10 N-terminal amino acids of ISD11 are indispensable for the activity of NFS1 and especially the conserved "LYR" motif is essential for the role of ISD11 in forming a stable and active complex with NFS1.

Optimizing depth-of-field extension in optical sectioning microscopy techniques using a fast focus-tunable lens.

Volume imaging based on a fast focus-tunable lens (FTL) allows three-dimensional (3D) observation within milliseconds by extending the depth-of-field (DOF) with sub-micrometer transverse resolution on optical sectioning microscopes. However, the previously published DOF extensions were neither axially uniform nor fit with theoretical prediction. In this work, complete theoretical treatments of focus extension with confocal and various multiphoton microscopes are established to correctly explain the previous results. Moreover, by correctly placing the FTL and properly adjusting incident beam diameter, a uniform DOF is achieved in which the actual extension nicely agrees with the theory. Our work not only provides a theoretical platform for volumetric imaging with FTL but also demonstrates the optimized imaging condition.

Protective effect of melatonin on the development of abdominal aortic aneurysm in a rat model.

BACKGROUND: Oxidative injury, inflammation, and apoptosis are involved in the progression of abdominal aortic aneurysm (AAA). Melatonin (MLT) has been reported with an effective antioxidant activity. The objective of the present study was to investigate whether MLT could suppress the development of AAA. METHODS: The AAA model was introduced by intraluminal perfusion of elastase in rats. All rats were divided into three groups as follows: (1) sham; (2) AAA + vehicle; and (3) AAA + MLT. Daily administration of MLT (10 mg/kg/d) or vehicle started 3 d before the perfusion and continued for 28 d after perfusion. An ultrasound system was applied to measure the dilation of the aorta. Histologic assays were performed to evaluate the structure, morphology, and apoptotic cells of the aortas; biochemical assays to determine the levels of proteins and lipid peroxide, activities of superoxide dismutase and NADPH oxidases, and cell viability; dihydroethidium fluorescence staining and flow cytometry to detect the presence of reactive oxygen species, and/or cell apoptosis; and electron microscopy to observe the ultrastructure of mitochondria. Cell lines A7R5 and RAW 264.7 were used for in vitro experiments. RESULTS: MLT treatment inhibited dilation of the aorta very likely through its antioxidant property; significantly reduced the levels of lipid peroxide, activities of NADPH oxidases, and content of reactive oxygen species; remarkably inhibited NF-κB signaling pathway and activities of matrix metalloproteinases triggered by elastase perfusion. As a result, the mitochondrion-dependent apoptosis was suppressed, cellular energy (ATP) supply was recovered, and mitochondrial morphology remained intact. CONCLUSIONS: Our results demonstrate the beneficial effects of MLT on inhibition of AAA formation, suggesting that MLT could be a potential agent for prevention of the development of human AAA.

TGFß-activated Kinase 1 (TAK1) Inhibition by 5Z-7-Oxozeaenol Attenuates Early Brain Injury after Experimental Subarachnoid Hemorrhage.

Accumulating evidence suggests that activation of mitogen-activated protein kinases (MAPKs) and nuclear factor NF-κB exacerbates early brain injury (EBI) following subarachnoid hemorrhage (SAH) by provoking proapoptotic and proinflammatory cellular signaling. Here we evaluate the role of TGFß-activated kinase 1 (TAK1), a critical regulator of the NF-κB and MAPK pathways, in early brain injury following SAH. Although the expression level of TAK1 did not present significant alternation in the basal temporal lobe after SAH, the expression of phosphorylated TAK1 (Thr-187, p-TAK1) showed a substantial increase 24 h post-SAH. Intracerebroventricular injection of a selective TAK1 inhibitor (10 min post-SAH), 5Z-7-oxozeaenol (OZ), significantly reduced the levels of TAK1 and p-TAK1 at 24 h post-SAH. Involvement of MAPKs and NF-κB signaling pathways was revealed that OZ inhibited SAH-induced phosphorylation of p38 and JNK, the nuclear translocation of NF-κB p65, and degradation of IκBα. Furthermore, OZ administration diminished the SAH-induced apoptosis and EBI. As a result, neurological deficits caused by SAH were reversed. Our findings suggest that TAK1 inhibition confers marked neuroprotection against EBI following SAH. Therefore, TAK1 might be a promising new molecular target for the treatment of SAH.

Phosphorylation of Akt by SC79 Prevents Iron Accumulation and Ameliorates Early Brain Injury in a Model of Experimental Subarachnoid Hemorrhage.

Previous studies have demonstrated that activation of Akt may alleviate early brain injury (EBI) following subarachnoid hemorrhage (SAH). This study is undertaken to determine whether iron metabolism is involved in the beneficial effect of Akt activation after SAH. Therefore, we used a novel molecule, SC79, to activate Akt in an experimental Sprague-Dawley rat model of SAH. Rats were randomly divided into four groups as follows: sham, SAH, SAH + vehicle, SAH + SC79. The results confirmed that SC79 effectively enhanced the defense against oxidative stress and alleviated EBI in the temporal lobe after SAH. Interestingly, we found that phosphorylation of Akt by SC79 reduced cell surface transferrin receptor-mediated iron uptake and promoted ferroportin-mediated iron transport after SAH. As a result, SC79 administration diminished the iron content in the brain tissue. Moreover, the impaired Fe-S cluster biogenesis was recovered and loss of the activities of the Fe-S cluster-containing enzymes were regained, indicating that injured mitochondrial functions are restored to healthy levels. These findings suggest that disrupted iron homeostasis could contribute to EBI and Akt activation may regulate iron metabolism to relieve iron toxicity, further protecting neurons from EBI after SAH.

Human glutaredoxin 3 can bind and effectively transfer [4Fe-4S] cluster to apo-iron regulatory protein 1.

Glutaredoxin 3 (GLRX3) is a member of monothiol glutaredoxins with a CGFS active site that has been demonstrated to function in cellular iron sensing and trafficking via its bound iron-sulfur cluster. Human GLRX3 has been shown to form a dimer that binds two bridging [2Fe-2S] clusters with glutathione (GSH) as a ligand, assembling a compound 2GLRX3-2[2Fe-2S]-4GSH. Each iron of the iron-sulfur clusters is bound to the thiols of the cysteines, one of which is from the active site of GLRX3, the other from the noncovalently bound GSH. Here, we show that the recombinant human GLRX3 isolated anaerobically from Escherichia coli can incorporate [4Fe-4S] cluster in the absence of GSH, revealed by spectral and enzymatic analysis. [4Fe-4S] cluster-containing GLRX3 is competent for converting iron regulatory protein 1 (apo-IRP1) into aconitase within 30 min, via intact iron-sulfur cluster transfer. These in vitro studies suggest that human GLRX3 is important for cytosolic Fe-S protein maturation.

Low expression of ferroxidases is implicated in the iron retention in human atherosclerotic plaques.

The effect of iron on the progress of atherosclerosis is still controversial. To explore the relationship between atherosclerotic plaques and iron metabolism and how iron is accumulated in plaque macrophages, we performed Oil red O staining to detect the lipid of the atherosclerotic plaques, enzyme-linked immunosorbent assay to detect the intracellular lipids (total cholesterol, free cholesterol) and serum hepcidin, Western-blot to examine the iron-related proteins, immunohistochemical and immunofluorescence assays to localize ferroportin 1 in macrophages. The contents of serum iron and transferrin saturation were measured. The results confrimed that atherosclerotic plaques were all lipid-rich. Compared to normal vessel wall, atherosclerotic plaques had significantly higher levels of ferritin and ferroportin 1. Strikingly, we found the much lower levels of ferroxidases ceruloplasmin and hephaestin in plaque tissue than the normal vessel, while the content of serum hepcidin, iron and transferrin saturation were similar in these two groups. The novel finding suggests that the inability of ferrous iron to be oxidized into ferric iron might be a potential mechanism for iron retention in plaques.

Blockage of mitochondrial calcium uniporter prevents iron accumulation in a model of experimental subarachnoid hemorrhage.

Previous studies have shown that iron accumulation is involved in the pathogenesis of brain injury following subarachnoid hemorrhage (SAH) and chelation of iron reduced mortality and oxidative DNA damage. We previously reported that blockage of mitochondrial calcium uniporter (MCU) provided benefit in the early brain injury after experimental SAH. This study was undertaken to identify whether blockage of MCU could ameliorate iron accumulation-associated brain injury following SAH. Therefore, we used two reagents ruthenium red (RR) and spermine (Sper) to inhibit MCU. Sprague-Dawley (SD) rats were randomly divided into four groups including sham, SAH, SAH+RR, and SAH+Sper. Biochemical analysis and histological assays were performed. The results confirmed the iron accumulation in temporal lobe after SAH. Interestingly, blockage of MCU dramatically reduced the iron accumulation in this area. The mechanism was revealed that inhibition of MCU reversed the down-regulation of iron regulatory protein (IRP) 1/2 and increase of ferritin. Iron-sulfur cluster dependent-aconitase activity was partially conserved when MCU was blocked. In consistence with this and previous report, ROS levels were notably reduced and ATP supply was rescued; levels of cleaved caspase-3 dropped; and integrity of neurons in temporal lobe was protected. Taken together, our results indicated that blockage of MCU could alleviate iron accumulation and the associated injury following SAH. These findings suggest that the alteration of calcium and iron homeostasis be coupled and MCU be considered to be a therapeutic target for patients suffering from SAH.