Abstract Iron has emerged as a significant cause of neurotoxicity in several neurodegenerative conditions, including Alzheimers disease (AD), Parkinsons disease (PD), sporadic Creutzfeldt-Jakob disease (sCJD), and others. In some cases, the underlying cause of iron mis-metabolism is known, while in others, our understanding is, at best, incomplete. Recent evidence implicating key proteins involved in the pathogenesis of AD, PD, and sCJD in cellular iron metabolism suggests that imbalance of brain iron homeostasis associated with these disorders is a direct consequence of disease pathogenesis. A complete understanding of the molecular events leading to this phenotype is lacking partly because of the complex regulation of iron homeostasis within the brain. Since systemic organs and the brain share several iron regulatory mechanisms and iron-modulating proteins, dysfunction of a specific pathway or selective absence of iron-modulating protein(s) in systemic organs has provided important insights into the maintenance of iron homeostasis within the brain. Here, we review recent information on the regulation of iron uptake and utilization in systemic organs and within the complex environment of the brain, with particular emphasis on the underlying mechanisms leading to brain iron mis-metabolism in specific neurodegenerative conditions. Mouse models that have been instrumental in understanding systemic and brain disorders associated with iron mis-metabolism are also described, followed by current therapeutic strategies which are aimed at restoring brain iron homeostasis in different neurodegenerative conditions. We conclude by highlighting important gaps in our understanding of brain iron metabolism and mis-metabolism, particularly in the context of neurodegenerative disorders. Antioxid. Redox Signal. 00, 000-000.
Oxygen sensing transcription factor HIF-1 is activated due to accumulation of regulatory subunit HIF-1? by posttranslational stability mechanism during hypoxia or by several other stimuli even in normoxia. HIF-1? is also regulated by NF-kB mediated transcription mechanism. Reactive oxygen species (ROS) act as an important regulator of HIF-1 either by affecting prolyl hydroxylase activity, the critical determinant of HIF-1? stabilization or by activating NF-kB to promote HIF-1? transcription. Insulin is known to activate HIF-1 by a ROS dependent mechanism but the molecular mechanism of HIF-1? regulation is not known so far. Here we show that insulin regulates HIF-1? by a novel transcriptional mechanism by a ROS-sensitive activation of Sp1 in 3T3-L1 preadipocyte. Insulin shows little effect on HIF-1? protein stability, but increases HIF-1? promoter activity. Mutation analyses, electrophoretic mobility shift assay and chromatin immunoprecipitation assay confirm the role of Sp1 in HIF-1? transcription. We further demonstrate that insulin-induced ROS generation initiates signaling pathway involving phosphatidylinositol 3-kinase and protein kinase C for Sp1 mediated HIF-1? transcription. In summary, we reveal that insulin regulates HIF-1? by a novel transcriptional mechanism involving Sp1.
A large program was conducted by the Government of India to study the prevalence and profile of chronic hepatitis B virus (HBV) infection and its risk factors in pregnant women attending a tertiary care hospital in India. From September 2004 to December 2008 consecutive pregnant women attending the antenatal clinic were screened and those found positive for HBsAg were enrolled. Healthy non-pregnant women of child-bearing age, who presented for blood donation during the same period, served as controls. Women with symptoms of liver disease or those aware of their HBsAg status were excluded. Of the 20,104 pregnant women screened, 224 (1.1%) and of the 658 controls, 8 (1.2%) were HBsAg positive (P?=?ns). Previous blood transfusions and surgery were significant risk factors for infection with HBV. Of the women who were HBsAg positive, the ALT levels were normal in 54% of the women and HBV DNA levels were above 2,000 ?IU/ml in 71% of women. The median HBV DNA levels were higher in women who were HBeAg positive compared to the HBeAg negative group. The most common HBV genotype was D (84%) followed by A?+?D and A (8% each). In conclusion, the prevalence of HBsAg positivity among asymptomatic pregnant women in North India is 1.1% with 71% having high HBV DNA levels. These women may have a high risk of transmitting infection to their newborns.
This study shows that the morphogenic regulator EFG1 level affects the drug susceptibilities of Candida albicans when grown on solid growth media. The Deltaefg1 mutant showed sensitivity particularly to those drugs that target ergosterol or its metabolism. Efg1p disruption showed a gene-dosage effect on drug susceptibilities and resulted in enhanced susceptibility to drugs in the homozygous mutant as compared with the wild type, heterozygous and revertant strains. The enhanced sensitivity to drugs was independent of the status of ATP-binding cassette and MFS multidrug efflux pumps of C. albicans. The Deltaefg1 mutant displayed increased membrane fluidity that coincided with the downregulation of ERG11 and upregulation of OLE1 and ERG3, leading to enhanced passive diffusion of drugs. Interestingly, Deltaefg1 mutant cells displayed enhanced levels of endogenous ROS levels. Notably, the higher levels of ROS in the Deltaefg1 mutant could be reversed by the addition of antioxidants. However, the restoration of ROS levels did not reverse the drug sensitivities of the Deltaefg1 mutant. Taken together, we, for the first time, establish a new role to EFG1 in affecting the drug susceptibilities of C. albicans cells, independent of ROS and known drug efflux mechanisms.
Imbalance of iron homeostasis has been reported in sporadic Creutzfeldt-Jakob-disease (sCJD) affected human and scrapie infected animal brains, but the contribution of this phenotype to disease associated neurotoxicity is unclear.
Glutathione (GSH) depletion is often detected in chronic pathological conditions like hepatitis C infection, alcohol consumption or xenobiotic assault with simultaneous reactive oxygen species (ROS) generation and hepatic iron overload. However, relation between GSH depletion and regulators of iron homeostasis is not clear so far. To determine that hepatic HepG2 cells were treated with GSH synthesis inhibitor butathione sulfoximine (BSO) and a dual regulation of ceruloplasmin (Cp) that involves in hepatic iron release was detected unlike other iron homeostasis regulators. BSO treatment that caused marginal GSH deficiency increased Cp synthesis due to increased transcription mediated by activator protein (AP)-1-binding site. In higher GSH deficiency (> 40 %) with increased ROS generation, Cp expression was decreased due to promotion of Cp mRNA decay mediated by 3untranslated region (3UTR) as found by transfecting chimera of chloramphenicol acetyl transferase (CAT) gene with Cp 3UTR. RNA gel shift assay showed significant reduction in 3UTR binding protein complex in similar condition. Decreased CAT expression and RNA-protein complex binding are reversed by pretreatment with antioxidant N-acetyl cysteine suggesting 3UTR binding protein complex is redox-sensitive. This unique and opposite regulation of Cp provides a mechanism of hepatic iron-deposition during glutathione deficiency detected in chronic pathological conditions.
Ceruloplasmin (Cp), a copper-containing protein, plays a significant role in body iron homeostasis as aceruloplasminemia patients and Cp knock-out mice exhibit iron overload in several tissues including liver and brain. Several other functions as oxidant, as antioxidant, and in nitric oxide metabolism are also attributed to Cp. Despite its role in iron oxidation and other biological oxidation reactions the regulation of Cp by reactive oxygen species (ROS) remains unexplored. Cp is synthesized in liver as a secretory protein and predominantly as a glycosylphosphatidylinositol-anchored membrane-bound form in astroglia. In this study we demonstrated that Cp expression is decreased by an mRNA decay mechanism in response to extracellular (H2O2) or intracellular oxidative stress (by mitochondrial chain blockers rotenone or antimycin A) in both hepatic and astroglial cells. The promotion of Cp mRNA decay is conferred by its 3-untranslated region (UTR). When chloramphenicol acetyltransferase (CAT) gene was transfected as a chimera with Cp 3-UTR in hepatic or astroglial cells, in response to either H2O2, rotenone, or antimycin A, the expression of CAT transcript was decreased, whereas expression of a 3-UTR-less CAT transcript remained unaffected. RNA gel shift assay showed significant reduction in 3-UTR-binding protein complex by ROS in both cell types that was reversed by the antioxidant N-acetylcysteine suggesting that ROS affects RNA-protein complex formation to promote Cp mRNA decay. Our finding is not only the first demonstration of regulation of Cp by ROS by a novel post-transcriptional mechanism but also provides a mechanism of iron deposition in neurodegenerative diseases.
Intracellular pathogens employ several strategies for iron acquisition from host macrophages for survival and growth, whereas macrophage resists infection by actively sequestering iron. Here, we show that instead of allowing macrophage to sequester iron, protozoan parasite Leishmania donovani (LD) uses a novel strategy to manipulate iron uptake mechanisms of the host and utilizes the taken up iron for its intracellular growth. To do so, intracellular LD directly scavenges iron from labile iron pool of macrophages. Depleted labile iron pool activates iron sensors iron-regulatory proteins IRP1 and IRP2. IRPs then bind to iron-responsive elements present in the 3 UTR of iron uptake gene transferrin receptor 1 by a post-transcriptional mRNA stability mechanism. Increased iron-responsive element-IRP interaction and transferrin receptor 1 expressions in spleen-derived macrophages from LD-infected mice confirm that LD employs similar mechanism to acquire iron during infection into mammalian hosts. Increased intracellular LD growth by holo-transferrin supplementation and inhibited growth by iron chelator treatment confirm the significance of this modulated iron uptake pathway of host in favour of the parasite.
Hepatic iron is known to regulate insulin signaling pathways and to influence insulin sensitivity in insulin resistance (IR) patients. However, the role of insulin on hepatic iron homeostasis remains unexplored. Here, we report that insulin promotes transferrin-bound iron uptake but shows no influence on non transferrin-bound iron uptake in human hepatic HepG2 cells. As a mechanism we detected increased transferrin receptor-1 (TfR1) expression both at protein and mRNA levels. Unaltered stability of protein and transcript of TfR1 suggested the regulation at transcriptional level that was confirmed by promoter activity. Involvement of transcription factor hypoxia inducible factor-1 (HIF-1) was shown by mutational analyses of the TfR1 promoter region and by electrophoretic mobility shift assay. When HepG2 cells were transfected with specific siRNA targeted to 3UTR of HIF-1?, the regulatory subunit of HIF-1; insulin-induced TfR1 expression and iron uptake were inhibited. Transfection of cDNA expressing stable form of HIF-1? reversed the increased TfR1 expression and iron uptake. These results suggest a novel role of insulin in hepatic iron uptake by a HIF-1 dependent transcriptional regulation of TfR1.
Recent evidence established a crucial role for mammalian oxygen sensing transcription factor hypoxia inducible factor-1 (HIF-1) in innate immunity against intracellular pathogens. In response to most of these pathogens host phagocytes increase transcription of HIF-1?, the regulatory component of HIF-1 to express various effector molecules against invaders. Leishmania donovani (LD), a protozoan parasite and the causative agent of fatal visceral leishmaniasis resides in macrophages within mammalian host. The mechanism of HIF-1 activation or its role in determining the fate of LD in infected macrophages is still not known. To determine that J774 macrophages were infected with LD and about four-fold increase in HIF-1 activity and HIF-1? expression were detected. A strong increase in HIF-1? expression and nuclear localization was also detected in LD-infected J774 cells, peritoneal macrophages and spleen derived macrophages of LD-infected BALB/c mice. A two-fold increase in HIF-1? mRNA was detected in LD-infected macrophages suggesting involvement of a transcriptional mechanism that was confirmed by promoter activity. We further revealed that LD also induced HIF-1? expression by depleting host cellular iron pool to affect prolyl hydroxylase activity resulting in to stabilization of HIF-1?. To determine the role of HIF-1 on intracellular LD, cells were transfected with HIF-1? siRNA to attenuate its expression and then infected with LD. Although, initial infection rate of LD in HIF-1? attenuated cells was not affected but intracellular growth of LD was significantly inhibited; while, over-expression of stabilized form of HIF-1? promoted intracellular growth of LD in host macrophage. Our results strongly suggest that LD activates HIF-1 by dual mechanism for its survival advantage within macrophage.
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