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Iron is an essential molecule for biological processes, but its accumulation can lead to oxidative stress and cellular death. Due to its oxidative effects, iron accumulation is implicated in the process of aging and neurodegenerative diseases. However, the mechanism for this increase in iron with aging, and whether this increase is localized to specific cellular compartment(s), are not known. Here, we measured the levels of iron in different tissues of aged mice, and demonstrate that while cytosolic non-heme iron is increased in the liver and muscle tissue, only the aged brain exhibits an increase in both the cytosolic and mitochondrial non-heme iron. This increase in brain iron is associated with elevated levels of local hepcidin mRNA and protein in the brain. We also demonstrate that the increase in hepcidin is associated with increased ubiquitination and reduced levels of the only iron exporter, feroportin-1 (FPN1). Overall, our studies provide a potential mechanism for iron accumulation in the brain through increased local expression of hepcidin, and subsequent iron accumulation due to decreased iron export. Additionally, our data support that aging is associated with mitochondrial and cytosolic iron accumulation only in the brain and not in other tissues.

Chronic loss of Augmenter of Liver Regeneration (ALR) results in mitochondrial myopathy with cataracts; however, the mechanism for this disorder remains unclear. Here, we demonstrate that loss of ALR, a principal component of the MIA40/ALR protein import pathway, results in impaired cytosolic Fe/S cluster biogenesis in mammalian cells. Mechanistically, MIA40/ALR facilitates the mitochondrial import of ATP-binding cassette (ABC)-B8, an inner mitochondrial membrane protein required for cytoplasmic Fe/S cluster maturation, through physical interaction with ABCB8. Downregulation of ALR impairs mitochondrial ABCB8 import, reduces cytoplasmic Fe/S cluster maturation, and increases cellular iron through the iron regulatory protein-iron response element system. Our finding thus provides a mechanistic link between MIA40/ALR import machinery and cytosolic Fe/S cluster maturation through the mitochondrial import of ABCB8, and offers a potential explanation for the pathology seen in patients with ALR mutations.

Chronic loss of Augmenter of Liver Regeneration (ALR) results in mitochondrial myopathy with cataracts, however, the mechanism for this disorder remains unclear. Here, we demonstrate that loss of ALR, a principal component of the MIA40/ALR protein import pathway, results in impaired cytosolic Fe/S cluster biogenesis in mammalian cells. Mechanistically, MIA40/ALR facilitates the mitochondrial import of ATP binding cassette (ABC)-B8, an inner mitochondrial membrane protein required for cytoplasmic Fe/S cluster maturation, through physical interaction with ABCB8. Downregulation of ALR impairs mitochondrial ABCB8 import, reduces cytoplasmic Fe/S cluster maturation, and increases cellular iron through the iron regulatory protein-iron response element system. Our finding provides a mechanistic link between MIA40/ALR import machinery and cytosolic Fe/S cluster maturation through the mitochondrial import of ABCB8, and offers a potential explanation for the pathology seen in patients with ALR mutations.

Introduction: A loss of function mutation in the mRNA binding protein ZFP36L2 has been associated with congenital heart diseases in humans, although the mechanism remains unclear. Objectives: We sought to elucidate the role of ZFP36L2 in the heart using mice with cardiac-specific ZFP36L2 deletion. Results: Cardiac-specific Zfp36l2 knockout (cs-L2KO) female mice exhibit a dramatic phenotype in which the majority of mice die after repeated pregnancies due to heart failure. We performed RNA-seq in H9c2 cells with ZFP36L2 KD, and found the expression of multiple mTOR pathway genes were altered, including marked repression of Redd1 and Sesn2 . Consistent with the RNA-seq data, the activity of mTORC1 was increased in cs-L2KO mouse hearts and in ZFP36L2 KD H9c2 cells. Loss of TSC2 mitigated mTORC1 hyperactivity in ZFP36L2 KD cells, indicating that ZFP36L2 regulates mTORC1 activity through the TSC2 complex. Overexpression of REDD1 rescued increased mTORC1 activity with ZFP36L2 KD, suggesting that ZFP36L2 regulates mTORC1, in-part through REDD1. Under total amino acid starvation, we observed higher p-S6K T389 levels in ZFP36L2 KD cells, and SESN2 overexpression mitigated p-S6K T389 , indicating loss of ZFP36L2 also regulates mTORC1 through amino acid sensing. p53 is a transcriptional activator of both REDD1 and SESN2, and we found that ZFP36L2 was required to maintain p53 levels by directly destabilizing MDM2 mRNA. Accordingly, administration of Nutlin3, an MDM2 inhibitor, reversed the reduction in p53, REDD1 and SESN2 protein levels, and prevented the increase in S6K phosphorylation in response to ZFP36L2 KD. Next, we treated pregnant mice with rapamycin, and cs-L2KO mice treated with rapamycin displayed significantly improved cardiac function after delivery. Finally, we analyzed human heart samples from patients with peri-partum cardiomyopathy and found ZFP36L2 protein to be significantly decreased and p-S6K T389 levels increased, compared to healthy controls. Conclusions: Our studies demonstrate that ZFP36L2 regulates mTORC1 activity through modifying p53 stability, and that the disruption of this pathway induces peri-partum cardiomyopathy. Inhibition of mTORC1 hyperactivity with rapamycin can be a potential therapy for this disease.

Introduction: Sirtuins are NAD+ dependent deacetylases and critical regulators of energy metabolism and response to oxidative stress. Sirtuin2 (SIRT2) is a cytoplasmic member of the sirtuin family, and has been shown to regulate cellular iron homeostasis through deacetylation of nuclear factor erythroid-derived 2-related factor 2 (NRF2). However, whether SIRT2-NRF2 pathway is involved in the development of heart failure remains unknown. Methods and results: To investigate the functional role of SIRT2 in the response to cardiac stress, SIRT2 knockout (KO) mice and their littermate controls were subjected to pressure overload by transverse aortic constriction (TAC). SIRT2 KO had normal appearance and cardiovascular parameters at baseline. However, in response to TAC, Sirt2 -/- mice displayed resistance to the pathological hypertrophic response, whereas wild type (WT) mice developed cardiac hypertrophy and heart failure. In addition, SIRT2 KO mice displayed less cardiac damage after /reperfusion injury. SIRT2 knockdown in neonatal rat cardiomyocytes (NRCM) reduced reactive oxygen species (ROS) production and cell death after H2O2 treatment. Since cellular oxidative stress is one of major contributor of cardiac dysfunction caused by both I/R injury and pressure overload, we examined whether NRF2 is associated with SIRT2-mediated cardiac response to oxidative stress. Levels of NRF2 was upregulated in NRCM with SIRT2 knockdown and treated with H2O2 compared to wild type (WT) cells. Moreover, NRF2 is translocated into the nucleus and its anti-oxidant target proteins are upregulated in NRCM with SIRT2 knockdown. SIRT2 was also found to bind and deacetylate NRF2 directly as determined by co-immunoprecipitation studies. This led to a reduction of its nuclear translocation and transcriptional activity. Finally, knockdown of both SIRT2 and NRF2 diminished the effects of SIRT2 knockdown on ROS production and cellular damage. Conclusion: These results indicate that SIRT2 contributes to pressure overload and I/R injury induced heart impairment in mice, and promotes oxidative stress injury in cardiomyocytes via deacetylating NRF2 and altering its activity.

Introduction: A loss of function mutation in the mRNA binding protein ZFP36L2 has been associated with congenital heart diseases in humans, although the mechanism remains unclear. Objectives: We sought to elucidate the role of ZFP36L2 in the heart using mice with cardiac-specific ZFP36L2 deletion. Results: Cardiac-specific Zfp36l2 knockout (cs-L2KO) female mice exhibit a dramatic phenotype in which the majority of mice die after repeated pregnancies due to heart failure. We performed RNA-seq in H9c2 cells with ZFP36L2 KD, and found the expression of multiple mTOR pathway genes were altered, including marked repression of Redd1 and Sesn2 . Consistent with the RNA-seq data, the activity of mTORC1 was increased in cs-L2KO mouse hearts and in ZFP36L2 KD H9c2 cells. Loss of TSC2 mitigated mTORC1 hyperactivity in ZFP36L2 KD cells, indicating that ZFP36L2 regulates mTORC1 activity through the TSC2 complex. Overexpression of REDD1 rescued increased mTORC1 activity with ZFP36L2 KD, suggesting that ZFP36L2 regulates mTORC1, in-part through REDD1. Under total amino acid starvation, we observed higher p-S6K T389 levels in ZFP36L2 KD cells, and SESN2 overexpression mitigated p-S6K T389 , indicating loss of ZFP36L2 also regulates mTORC1 through amino acid sensing. p53 is a transcriptional activator of both REDD1 and SESN2, and we found that ZFP36L2 was required to maintain p53 levels by directly destabilizing MDM2 mRNA. Accordingly, administration of Nutlin3, an MDM2 inhibitor, reversed the reduction in p53, REDD1 and SESN2 protein levels, and prevented the increase in S6K phosphorylation in response to ZFP36L2 KD. Next, we treated pregnant mice with rapamycin, and cs-L2KO mice treated with rapamycin displayed significantly improved cardiac function after delivery. Finally, we analyzed human heart samples from patients with peri-partum cardiomyopathy and found ZFP36L2 protein to be significantly decreased and p-S6K T389 levels increased, compared to healthy controls. Conclusions: Our studies demonstrate that ZFP36L2 regulates mTORC1 activity through modifying p53 stability, and that the disruption of this pathway induces peri-partum cardiomyopathy. Inhibition of mTORC1 hyperactivity with rapamycin can be a potential therapy for this disease.

In this study, polyethylene glycol (PEG) with hydroxyapatite (HA), with the incorporation of physical gold nanoparticles (AuNPs), was created and equipped through a surface coating technique in order to form PEG-HA-AuNP nanocomposites. The surface morphology and chemical composition were characterized using scanning electron microscopy (SEM), atomic force microscopy (AFM), UV–Vis spectroscopy (UV–Vis), Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), and contact angle assessment. The effects of PEG-HA-AuNP nanocomposites on the biocompatibility and biological activity of MC3T3-E1 osteoblast cells, endothelial cells (EC), macrophages (RAW 264.7), and human mesenchymal stem cells (MSCs), as well as the guiding of osteogenic differentiation, were estimated through the use of an in vitro assay. Moreover, the anti-inflammatory, biocompatibility, and endothelialization capacities were further assessed through in vivo evaluation. The PEG-HA-AuNP nanocomposites showed superior biological properties and biocompatibility capacity for cell behavior in both MC3T3-E1 cells and MSCs. These biological events surrounding the cells could be associated with the activation of adhesion, proliferation, migration, and differentiation processes on the PEG-HA-AuNP nanocomposites. Indeed, the induction of the osteogenic differentiation of MSCs by PEG-HA-AuNP nanocomposites and enhanced mineralization activity were also evidenced in this study. Moreover, from the in vivo assay, we further found that PEG-HA-AuNP nanocomposites not only facilitate the anti-immune response, as well as reducing CD86 expression, but also facilitate the endothelialization ability, as well as promoting CD31 expression, when implanted into rats subcutaneously for a period of 1 month. The current research illustrates the potential of PEG-HA-AuNP nanocomposites when used in combination with MSCs for the regeneration of bone tissue, with their nanotopography being employed as an applicable surface modification approach for the fabrication of biomaterials.

SIRT2 is a cytoplasmic sirtuin that plays a role in various cellular processes, including tumorigenesis, metabolism, and inflammation. Since these processes require iron, we hypothesized that SIRT2 directly regulates cellular iron homeostasis. Here, we have demonstrated that SIRT2 depletion results in a decrease in cellular iron levels both in vitro and in vivo. Mechanistically, we determined that SIRT2 maintains cellular iron levels by binding to and deacetylating nuclear factor erythroid-derived 2-related factor 2 (NRF2) on lysines 506 and 508, leading to a reduction in total and nuclear NRF2 levels. The reduction in nuclear NRF2 leads to reduced ferroportin 1 (FPN1) expression, which in turn results in decreased cellular iron export. Finally, we observed that Sirt2 deletion reduced cell viability in response to iron deficiency. Moreover, livers from Sirt2-/- mice had decreased iron levels, while this effect was reversed in Sirt2-/- Nrf2-/- double-KO mice. Taken together, our results uncover a link between sirtuin proteins and direct control over cellular iron homeostasis via regulation of NRF2 deacetylation and stability.

Excess cellular iron increases reactive oxygen species (ROS) production and causes cellular damage. Mitochondria are the major site of iron metabolism and ROS production; however, few studies have investigated the role of mitochondrial iron in the development of cardiac disorders, such as ischemic heart disease or cardiomyopathy (CM). We observe increased mitochondrial iron in mice after ischemia/reperfusion (I/R) and in human hearts with ischemic CM, and hypothesize that decreasing mitochondrial iron protects against I/R damage and the development of CM. Reducing mitochondrial iron genetically through cardiac‐specific overexpression of a mitochondrial iron export protein or pharmacologically using a mitochondria‐permeable iron chelator protects mice against I/R injury. Furthermore, decreasing mitochondrial iron protects the murine hearts in a model of spontaneous CM with mitochondrial iron accumulation. Reduced mitochondrial ROS that is independent of alterations in the electron transport chain's ROS producing capacity contributes to the protective effects. Overall, our findings suggest that mitochondrial iron contributes to cardiac ischemic damage, and may be a novel therapeutic target against ischemic heart disease.

The role of posttranscriptional metabolic gene regulatory programs in diabetes is not well understood. Here, we show that the RNA-binding protein tristetraprolin (TTP) is reduced in the livers of diabetic mice and humans and is transcriptionally induced in response to insulin treatment in murine livers in vitro and in vivo. Liver-specific Ttp-KO (lsTtp-KO) mice challenged with high-fat diet (HFD) have improved glucose tolerance and peripheral insulin sensitivity compared with littermate controls. Analysis of secreted hepatic factors demonstrated that fibroblast growth factor 21 (FGF21) is posttranscriptionally repressed by TTP. Consistent with increased FGF21, lsTtp-KO mice fed HFD have increased brown fat activation, peripheral tissue glucose uptake, and adiponectin production compared with littermate controls. Downregulation of hepatic Fgf21 via an adeno-associated virus-driven shRNA in mice fed HFD reverses the insulin-sensitizing effects of hepatic Ttp deletion. Thus, hepatic TTP posttranscriptionally regulates systemic insulin sensitivity in diabetes through liver-derived FGF21.

Cells respond to iron deficiency by activating iron-regulatory proteins to increase cellular iron uptake and availability. However, it is not clear how cells adapt to conditions when cellular iron uptake does not fully match iron demand. Here, we show that the mRNA-binding protein tristetraprolin (TTP) is induced by iron deficiency and degrades mRNAs of mitochondrial Fe/S-cluster-containing proteins, specifically Ndufs1 in complex I and Uqcrfs1 in complex III, to match the decrease in Fe/S-cluster availability. In the absence of TTP, Uqcrfs1 levels are not decreased in iron deficiency, resulting in nonfunctional complex III, electron leakage, and oxidative damage. Mice with deletion of Ttp display cardiac dysfunction with iron deficiency, demonstrating that TTP is necessary for maintaining cardiac function in the setting of low cellular iron. Altogether, our results describe a pathway that is activated in iron deficiency to regulate mitochondrial function to match the availability of Fe/S clusters.

The prevalence of obesity is rising globally, and the United States has one of the highest obesity rates in the world: ≈17% of the young and >33% of adults are obese.1 Obesity is associated with chronic low-grade systemic inflammation, which is considered a critical underlying factor in the development of insulin resistance (IR).2 IR is a major risk factor for type 2 diabetes mellitus (T2DM) and cardiovascular disease.3 In the development of obesity, white adipose tissue, particularly the abdominal adipose tissue, is the key site that mediates systemic inflammation and IR, though other organs, such as skeletal muscle and liver, have also been implicated.4 Adipose tissue is a highly vascularized organ where every adipocyte is connected to at least one capillary.5 To maintain normal adipose tissue function, the proper signaling between adipocytes and endothelial cells (ECs) from the surrounding vasculature is important.6 There is a growing body of evidence suggesting that EC dysfunction contributes to the pathogenesis of atherosclerosis, obesity, and T2DM.7,8 Therefore, it is of key interest to further study the role of the crosstalk between adipose tissue ECs and adipocytes in obesity-associated IR and to identify potential therapeutic targets for novel interventions. Recently, several reports suggested that microRNAs (miRs) are important mediators of the development of inflammation and IR in obese adipose tissue.9 Subsequently, numerous studies explored targeting specific miRs in diabetic complications to mitigate the pathological sequela of T2DM.9 Given these points, using miRs to modulate adipocyte–EC axis in adipose tissue may offer new tools to combat the growing epidemic of obesity and its associated comorbidities. Article, see p 810 Over the past 2 decades, several studies elucidated the underlying molecular mechanisms linking inflammation to obesity-associated IR. Hotamisligil et al10 was the first to demonstrate …

Background Sex‐related differences in cardiac function and iron metabolism exist in humans and experimental animals. Male patients and preclinical animal models are more susceptible to cardiomyopathies and heart failure. However, whether similar differences are seen in iron‐overload cardiomyopathy is poorly understood. Methods and Results Male and female wild‐type and hemojuvelin‐null mice were injected and fed with a high‐iron diet, respectively, to develop secondary iron overload and genetic hemochromatosis. Female mice were completely protected from iron‐overload cardiomyopathy, whereas iron overload resulted in marked diastolic dysfunction in male iron‐overloaded mice based on echocardiographic and invasive pressure‐volume analyses. Female mice demonstrated a marked suppression of iron‐mediated oxidative stress and a lack of myocardial fibrosis despite an equivalent degree of myocardial iron deposition. Ovariectomized female mice with iron overload exhibited essential pathophysiological features of iron‐overload cardiomyopathy showing distinct diastolic and systolic dysfunction, severe myocardial fibrosis, increased myocardial oxidative stress, and increased expression of cardiac disease markers. Ovariectomy prevented iron‐induced upregulation of ferritin, decreased myocardial SERCA2a levels, and increased NCX1 levels. 17β‐Estradiol therapy rescued the iron‐overload cardiomyopathy in male wild‐type mice. The responses in wild‐type and hemojuvelin‐null female mice were remarkably similar, highlighting a conserved mechanism of sex‐dependent protection from iron‐overload‐mediated cardiac injury. Conclusions Male and female mice respond differently to iron‐overload‐mediated effects on heart structure and function, and females are markedly protected from iron‐overload cardiomyopathy. Ovariectomy in female mice exacerbated iron‐induced myocardial injury and precipitated severe cardiac dysfunction during iron‐overload conditions, whereas 17β‐estradiol therapy was protective in male iron‐overloaded mice.

Iron chelation rather than supplementation would be beneficial to patients with heart diseases. More effort should be devoted to the development of iron chelators that could target mitochondrial iron. Iron is an essential trace mineral for normal mammalian physiology. It is incorporated into iron/sulfur (Fe/S) clusters that serve as important cofactors for many enzymes, including the mitochondrial respiratory chain, and it gives blood and muscle their signature red color through its presence in heme.1 Because of its involvement in many important cellular processes, dysregulation of iron often causes disease. There are 2 broad categories of diseases associated with iron dysregulation—iron overload and iron deficiency. Each category can be further subdivided based on whether the changes in iron occur at the cellular or at the systemic level. Iron is critical for developmental processes, and studies have linked severe systemic iron deficiency to impaired brain development in children.2 However, most of these studies were conducted in developing countries where iron deficiency tends to be more severe. In developed countries, the major disorder associated with iron deficiency is anemia. Although systemic iron deficiency and anemia may have been used interchangeably, these 2 terms refer to different conditions. Systemic iron deficiency refers to low amounts of iron in circulation because of limited iron absorption or excess iron loss. Anemia is defined as low hemoglobin content within red blood cells and can be caused by low systemic iron, mutations in hemoglobin genes, vitamin B12 deficiency, or underlying chronic diseases. Limited oxygen-carrying capacity secondary to anemia can cause fatigue; however, it generally does not cause mortality or major damage to vital organs. Except for high-output heart failure (HF) associated with severe anemia, iron deficiency and anemia are not associated with any other major cardiac disorders. Cellular iron deficiency is a rare phenomenon, and except …

Objectives The goal of this study was to characterize the regulation of heme and non-heme iron in human failing hearts. Background Iron is an essential molecule for cellular physiology, but in excess it facilitates oxidative stress. Mitochondria are the key regulators of iron homeostasis through heme and iron-sulfur cluster synthesis. Because mitochondrial function is depressed in failing hearts and iron accumulation can lead to oxidative stress, we hypothesized that iron regulation may also be impaired in heart failure (HF). Methods We measured mitochondrial and cytosolic heme and non-heme iron levels in failing human hearts retrieved during cardiac transplantation surgery. In addition, we examined the expression of genes regulating cellular iron homeostasis, the heme biosynthetic pathway, and micro-RNAs that may potentially target iron regulatory networks. Results Although cytosolic non-heme iron levels were reduced in HF, mitochondrial iron content was maintained. Moreover, we observed a significant increase in heme levels in failing hearts, with corresponding feedback inhibition of the heme synthetic enzymes and no change in heme degradation. The rate-limiting enzyme in heme synthesis, delta-aminolevulinic acid synthase 2 (ALAS2), was significantly upregulated in HF. Overexpression of ALAS2 in H9c2 cardiac myoblasts resulted in increased heme levels, and hypoxia and erythropoietin treatment increased heme production through upregulation of ALAS2. Finally, increased heme levels in cardiac myoblasts were associated with excess production of reactive oxygen species and cell death, suggesting a maladaptive role for increased heme in HF. Conclusions Despite global mitochondrial dysfunction, heme levels are maintained above baseline in human failing hearts.

The erbB family of receptor tyrosine kinases are known to play important roles in normal epithelial development and epithelial neoplasia. Considerable evidence also suggests that signaling through the epidermal growth factor receptor (EGFR) plays an important role in multistage skin carcinogenesis in mice; however, less is known about the role of erbB2. In this study, to further examine the role of both erbB2 and EGFR in epithelial carcinogenesis, we examined the effect of a dual erbB2/EGFR tyrosine kinase inhibitor, GW2974, given in the diet on skin tumor promotion during two-stage carcinogenesis in wild-type and BK5.erbB2 mice. In BK5.erbB2 mice, erbB2 is overexpressed in the basal layer of epidermis and leads to heightened sensitivity to skin tumor development. GW2974 effectively inhibited skin tumor promotion by 12-O-tetradecanoylphorbol-13-acetate in wild-type and BK5.erbB2 mice, although a more marked effect was seen in BK5.erbB2 mice. In addition, this inhibitory effect was reversible when GW2974 treatment was withdrawn. GW2974 inhibited 12-O-tetradecanoylphorbol-13-acetate–induced epidermal hyperproliferation, which correlated with reduced activation of both the EGFR and erbB2. These results support the hypothesis that both the EGFR and erbB2 play an important role in the development of skin tumors during two-stage skin carcinogenesis, especially during the tumor promotion stage. Furthermore, the marked sensitivity of BK5.erbB2 mice to the inhibitory effects of GW2974 during tumor promotion suggest greater efficacy for this compound when erbB2 is overexpressed or amplified as an early event in the carcinogenic process. Cancer Prev Res; 3(8); 940–52. ©2010 AACR.

Background Heme is an essential iron‐containing molecule for cardiovascular physiology, but in excess it may increase oxidative stress. Failing human hearts have increased heme levels, with upregulation of the rate‐limiting enzyme in heme synthesis, δ‐aminolevulinic acid synthase 2 ( ALAS 2), which is normally not expressed in cardiomyocytes. We hypothesized that increased heme accumulation (through cardiac overexpression of ALAS 2) leads to increased oxidative stress and cell death in the heart. Methods and Results We first showed that ALAS 2 and heme levels are increased in the hearts of mice subjected to coronary ligation. To determine the causative role of increased heme in the development of heart failure, we generated transgenic mice with cardiac‐specific overexpression of ALAS 2. While ALAS 2 transgenic mice have normal cardiac function at baseline, their hearts display increased heme content, higher oxidative stress, exacerbated cell death, and worsened cardiac function after coronary ligation compared to nontransgenic littermates. We confirmed in cultured cardiomyoblasts that the increased oxidative stress and cell death observed with ALAS 2 overexpression is mediated by increased heme accumulation. Furthermore, knockdown of ALAS 2 in cultured cardiomyoblasts exposed to hypoxia reversed the increases in heme content and cell death. Administration of the mitochondrial antioxidant MitoTempo to ALAS 2‐overexpressing cardiomyoblasts normalized the elevated oxidative stress and cell death levels to baseline, indicating that the effects of increased ALAS 2 and heme are through elevated mitochondrial oxidative stress. The clinical relevance of these findings was supported by the finding of increased ALAS 2 induction and heme accumulation in failing human hearts from patients with ischemic cardiomyopathy compared to nonischemic cardiomyopathy. Conclusions Heme accumulation is detrimental to cardiac function under ischemic conditions, and reducing heme in the heart may be a novel approach for protection against the development of heart failure.

Heme is an essential molecule for living aerobic organisms and is involved in a remarkable array of diverse biological processes. In the cardiovascular system, heme plays a major role in gas exchange, mitochondrial energy production, antioxidant defense, and signal transduction. Although heme, as

Introduction: Iron is essential for the activity of several cellular proteins, but excess free iron can cause cellular damage through production of reactive oxygen species (ROS). Iron accumulation in mitochondria, the major site of cellular iron homeostasis, leads to cardiomyopathy. However, it is not known whether a reduction in baseline mitochondrial (as opposed to cytosolic) iron can protect against ischemia-reperfusion (I/R) injury in the heart. We hypothesized that since mitochondria are the major site of iron homeostasis and that mitochondrial iron can lead to oxidative damage, a reduction in mitochondrial iron at baseline would be sufficient to protect against I/R injury. Results: Transgenic (TG) mice with cardiomyocyte-specific overexpression of the mitochondrial iron export protein ATP-binding cassette (ABC)-B8 had significantly lower mitochondrial iron in the heart than nontransgenic (NTG) littermates at baseline, but their cardiac function and the expression of key antioxidant systems were similar to NTG littermates. In response to I/R, TG mice displayed significantly less apoptosis and lipid peroxidation products and better preserved cardiac function than NTG littermates, suggesting that a reduction in mitochondrial iron protects against I/R injury. To confirm these results, we next took a pharmacological approach to assess the effects of a reduction in mitochondrial vs cytosolic iron on the response to I/R using 2,2’-bipyridyl (BPD, a mitochondria-accessible iron chelator) and deferoxamine (DFO, an iron chelator that can only reduce cytosolic iron). Treating rat cardiomyoblast H9C2 cells with BPD but not DFO significantly lowered chelatable mitochondrial iron and protected against H 2 O 2 induced cell death, and pretreatment with BPD but not DFO protected mice against I/R injury and reduced ROS production, suggesting that a reduction in baseline mitochondrial, but not cytosolic, iron is sufficient to protect against I/R injury. Conclusions: Our findings demonstrate that selective reduction in mitochondrial iron is protective in I/R injury. Thus, targeting mitochondrial iron with selective iron chelators may provide a novel approach for treatment of ischemic heart disease.

Nature Communications 8: Article number: 14095 (2017); Published: 24 January 2017; Updated: 30 August 2017. The authors inadvertently omitted Eltyeb Abdelwahid, who contributed to the generation of animal models and their initial evaluation, from the author list. This has now been corrected in both the PDF and HTML versions of the Article.

Patients with type 2 diabetes often present with cardiovascular complications; however, it is not clear how diabetes promotes cardiac dysfunction. In murine models, deletion of the gene encoding aryl hydrocarbon nuclear translocator (ARNT, also known as HIF1β) in the liver or pancreas leads to a diabetic phenotype; however, the role of ARNT in cardiac metabolism is unknown. Here, we determined that cardiac-specific deletion of Arnt in adult mice results in rapid development of cardiomyopathy (CM) that is characterized by accumulation of lipid droplets. Compared with hearts from ARNT-expressing mice, ex vivo analysis of ARNT-deficient hearts revealed a 2-fold increase in fatty acid (FA) oxidation as well as a substantial increase in the expression of PPARα and its target genes. Furthermore, deletion of both Arnt and Ppara preserved cardiac function, improved survival, and completely reversed the FA accumulation phenotype, indicating that PPARα mediates the detrimental effects of Arnt deletion in the heart. Finally, we determined that ARNT directly regulates Ppara expression by binding to its promoter and forming a complex with HIF2α. Together, these findings suggest that ARNT is a critical regulator of myocardial FA metabolism and that its deletion leads to CM and an increase in triglyceride accumulation through PPARα.

Disorders of iron homeostasis are very common, yet the molecular mechanisms of iron regulation remain understudied. Over 20 years have passed since the first characterization of iron-regulatory proteins (IRP) as mediators of cellular iron-deficiency response in mammals through iron acquisition. However, little is known about other mechanisms necessary for adaptation to low-iron states. In this review, we present recent evidence that establishes the existence of a new iron-regulatory pathway aimed at iron conservation and optimization of iron use through suppression of nonessential iron-consuming processes. Moreover, we discuss the possible links between iron homeostasis and energy metabolism uncovered by studies of iron-deficiency response.