Fluorescent Carbon Dots for Sensitive and Rapid Monitoring of Intracellular Ferrous Ion

Although iron is an essential constituent for almost all living organisms, iron dyshomeostasis at a cellular level may trigger oxidative stress and neuronal damage. Hence, there are numerous reported carbon dots (CDs) that have been synthesized and applied to determine intracellular iron ions. However, among reported CDs focused to detect Fe3+ ions, only a few CDs have been designed to specifically determine Fe2+ ions over Fe3+ ions for monitoring of intracellular Fe2+ ions. We have developed the nitrogen-doped CDs (NCDs) for fluorescence turn-off detection of Fe2+ at cellular level. The as-synthesized NCDs exhibit a strong blue fluorescence and low cytotoxicity, acting as fluorescence probes to detect Fe2+ as low as 0.702 µM in aqueous solution within 2 min and visualize intracellular Fe2+ in the concentration range from 0 to 500 µM within 20 min. The as-prepared NCDs possess some advantages such as high biocompatibility, strong fluorescence properties, selectivity, and rapidity for intracellular Fe2+ monitoring, making NCDs an excellent nanoprobe for biosensing of intracellular ferrous ions.

The “Biological Weapons” of Ehrlichia chaffeensis: Novel Molecules and Mechanisms to Subjugate Host Cells

Ehrlichia chaffeensis is an obligatory intracellular bacterium that causes human monocytic ehrlichiosis, an emerging, potentially fatal tick-borne infectious disease. The bacterium enters human cells via the binding of its unique outer-membrane invasin EtpE to the cognate receptor DNase X on the host-cell plasma membrane; this triggers actin polymerization and filopodia formation at the site of E. chaffeensis binding, and blocks activation of phagocyte NADPH oxidase that catalyzes the generation of microbicidal reactive oxygen species. Subsequently, the bacterium replicates by hijacking/dysregulating host-cell functions using Type IV secretion effectors. For example, the Ehrlichia translocated factor (Etf)-1 enters mitochondria and inhibits mitochondria-mediated apoptosis of host cells. Etf-1 also induces autophagy mediated by the small GTPase RAB5, the result being the liberation of catabolites for proliferation inside host cells. Moreover, Etf-2 competes with the RAB5 GTPase-activating protein, for binding to RAB5-GTP on the surface of E. chaffeensis inclusions, which blocks GTP hydrolysis and consequently prevents the fusion of inclusions with host-cell lysosomes. Etf-3 binds ferritin light chain to induce ferritinophagy to obtain intracellular iron. To enable E. chaffeensis to rapidly adapt to the host environment and proliferate, the bacterium must acquire host membrane cholesterol and glycerophospholipids for the purpose of producing large amounts of its own membrane. Future studies on the arsenal of unique Ehrlichia molecules and their interplay with host-cell components will undoubtedly advance our understanding of the molecular mechanisms of obligatory intracellular infection and may identify hitherto unrecognized signaling pathways of human hosts. Such data could be exploited for development of treatment and control measures for ehrlichiosis as well as other ailments that potentially could involve the same host-cell signaling pathways that are appropriated by E. chaffeensis.

Identification of Ferroptosis-Related Gene Prognostic Signature and HSF1 for Reversing Doxorubicin and Gemcitabine Resistance in Uterine Carcinosarcoma

Purpose. Iron metabolism and ferroptosis play crucial roles in the pathogenesis of cancer. In this study, we aim to study the role of ferroptosis-related genes (FRGs) in uterine carcinosarcoma (UCS) and identify potential target for UCS. Methods. Prognostic differentially expressed FRGs were identified of in the TCGA cohort. Integrated analysis, cox regression, and the least absolute shrinkage and selection operator (LASSO) methods of FRGs were performed to construct a multigene signature prognostic model. Moreover, a dataset from Gene Expression Omnibus (GEO) served as an external validation. HSF1 was knockdown in MES-SA and FU-MMT-1 cells, and cell viability, lipid ROS, and intracellular iron level were detected when combined with doxorubicin or gemcitabine. Result. Five FRGs were selected to construct a prognostic model of UCS. The group with high-risk signature score exhibited obviously lower overall survival (OS) than the group with low risk signature score in both TCGA and validated GEO cohorts. Multivariate Cox regression analysis further indicated that the risk score was an independent factor for the prognosis of UCS patients. The high-risk group of UCS has a higher sensitivity in the treatment of doxorubicin and gemcitabine. Knocking down of HSF1 in MES-SA and FU-MMT-1 cells was more sensitive to doxorubicin and gemcitabine via increasing ferroptosis. Conclusions. The five FRGs risk signature prognostic model having a superior and drug sensitivity predictive performance for OS in UCS, and HSF1 is a potential marker sensitive to doxorubicin and gemcitabine in UCS patients.

Abnormal Iron and Lipid Metabolism Mediated Ferroptosis in Kidney Diseases and Its Therapeutic Potential

Ferroptosis is a newly identified form of regulated cell death driven by iron-dependent phospholipid peroxidation and oxidative stress. Ferroptosis has distinct biological and morphology characteristics, such as shrunken mitochondria when compared to other known regulated cell deaths. The regulation of ferroptosis includes different molecular mechanisms and multiple cellular metabolic pathways, including glutathione/glutathione peroxidase 4(GPX4) signaling pathways, which are involved in the amino acid metabolism and the activation of GPX4; iron metabolic signaling pathways, which are involved in the regulation of iron import/export and the storage/release of intracellular iron through iron-regulatory proteins (IRPs), and lipid metabolic signaling pathways, which are involved in the metabolism of unsaturated fatty acids in cell membranes. Ferroptosis plays an essential role in the pathology of various kidneys diseases, including acute kidney injury (AKI), chronic kidney disease (CKD), autosomal dominant polycystic kidney disease (ADPKD), and renal cell carcinoma (RCC). Targeting ferroptosis with its inducers/initiators and inhibitors can modulate the progression of kidney diseases in animal models. In this review, we discuss the characteristics of ferroptosis and the ferroptosis-based mechanisms, highlighting the potential role of the main ferroptosis-associated metabolic pathways in the treatment and prevention of various kidney diseases.

Esophageal Cancer Stem-like Cells Resist Ferroptosis-Induced Cell Death by Active Hsp27-GPX4 Pathway

Cancer stem cells (CSCs), a subpopulation of cancer cells responsible for tumor initiation and treatment failure, are more susceptible to ferroptosis-inducing agents than bulk cancer cells. However, regulatory pathways controlling ferroptosis, which can selectively induce CSC death, are not fully understood. Here, we demonstrate that the CSCs of esophageal squamous carcinoma cells enriched by spheroid culture have increased intracellular iron levels and lipid peroxidation, thereby increasing exposure to several products of lipid peroxidation, such as MDA and 4-HNE. However, CSCs do not reduce cell viability until glutathione is depleted by erastin treatment. Mechanistic studies revealed that damage from elevated lipid peroxidation is avoided through the activation of Hsp27, which upregulates GPX4 and thereby rescues CSCs from ferroptosis-induced cell death. Our results also revealed a correlation between phospho-Hsp27 and GPX4 expression levels and poor prognosis in patients with esophageal cancer. Together, these data indicate that targeting Hsp27 or GPX4 to block this intrinsic protective mechanism against ferroptosis is a potential treatment strategy for eradicating CSC in esophageal squamous cell carcinoma.

Vitamin B6 Alleviates Lipopolysaccharide-induced Myocardial Injury by Ferroptosis and Apoptosis Regulation

Vitamin B6 (VitB6) is a water-soluble vitamin and includes pyridoxine, pyridoxal, pyridoxamine, and their phosphorylated forms. In the current study, we demonstrated that VitB6 could improve lipopolysaccharide (LPS)–induced myocardial injury. We demonstrated that VitB6 can suppress LPS-induced oxidative stress and lipid peroxidation that lead to ferroptosis and apoptosis in vivo and in vitro. Moreover, we found that VitB6 can regulate the expression of iron regulatory proteins, maintaining intracellular iron homeostasis. To confirm that VitB6 could inhibit LPS-induced ferroptosis and apoptosis, we pretreated mice with ferrostatin-1 (Fer-1) and emricasan that efficiently mimicked VitB6 pharmacological effects. This improved the survival rate of mice challenged with a high LPS dose. In addition, VitB6 regulated the expression of LPS-induced apoptosis-related proteins and iron regulatory proteins. It mediated the expression of Nrf2, transcription factor NF-E2–related factor 2, which promoted the expression of antioxidant enzymes and restrained LPS-induced ferroptosis and apoptosis. Overall, our results indicated that VitB6 can be used on novel therapies to relieve LPS-induced myocardial injury.

Exploring the Molecular Mechanisms of Action of Samoan Medicinal Plants via Chemical Genetic Analyses

<p>Natural products are a robust source of drug leads, and medicinal plants have been the source of natural products that are important pharmaceuticals in modern medicine. Samoan medicinal plants have been investigated in the past, but their potential as a source of new drug leads has not been realized. I hypothesized that determining the mechanism of action of Samoan medicinal plant extracts would provide insight into their pharmaceutical potential. The work described herein was carried out on 11 Samoan medicinal plants, from which 22 extracts were prepared. A bioactivity rate of 68% was determined when 15 of the 22 extracts inhibited the growth of yeast (Saccharomyces cerevisiae). The medicinal plant Psychotria insularum was the most potent, thus genome-wide analyses were completed using the haploid deletion mutant library of S. cerevisiae. Yeast strains deficient in iron transport were hypersensitive to the P. insularum aqueous extract. Further investigations showed that exogenous iron supplementation rescued the growth defect induced by P. insularum extracts, suggesting that P. insularum reduced intracellular iron. Fittingly, yeast cells treated with P. insularum extracts contained less intracellular iron than control cells. Paraxodically, the expression levels of iron transporter proteins were upregulated upon extract treatment. When we investigated iron-requiring cellular processes, we found that yeast cells treated with P. insularum extracts exhibited a respiratory deficient phenotype and reduced heme synthesis, indicative of an impaired cellular iron status. These findings suggested that P. insularum reduced bioavailable iron leading to the induction of the low iron response, and indeed the extracts of P. insularum were shown to chelate iron via the iron-chelating CAS assay. To translate results from yeast to mammalian cells, we treated primary murine macrophages with P. insularum extracts and detected an anti-inflammatory response, which we found to correlate with reduced activity of the iron-requiring aconitase enzyme. We further determined using pooled diploid mutant genetic analyses that the extracts of P. insularum did not have a genetic target. To identify the compound mediating the iron chelation mechanism, bioassay-guided isolation was conducted. Fractionation of the crude aqueous extract of P. insularum produced a relatively pure fraction that NMR and the acid-butanol assay identified as a condensed tannin. Together, these results indicate a relationship between iron chelation, a condensed tannin and inflammation. Further, we established an iron chelation mechanism of action by which P. insularum extracts are used to treat inflammation-associated symptoms in traditional Samoan medicine. Lastly, the findings presented here substantiate the reliability of plants with ethnobotanical background as sources for bioactive natural products.</p>

Exploring the Molecular Mechanisms of Action of Samoan Medicinal Plants via Chemical Genetic Analyses

<p>Natural products are a robust source of drug leads, and medicinal plants have been the source of natural products that are important pharmaceuticals in modern medicine. Samoan medicinal plants have been investigated in the past, but their potential as a source of new drug leads has not been realized. I hypothesized that determining the mechanism of action of Samoan medicinal plant extracts would provide insight into their pharmaceutical potential. The work described herein was carried out on 11 Samoan medicinal plants, from which 22 extracts were prepared. A bioactivity rate of 68% was determined when 15 of the 22 extracts inhibited the growth of yeast (Saccharomyces cerevisiae). The medicinal plant Psychotria insularum was the most potent, thus genome-wide analyses were completed using the haploid deletion mutant library of S. cerevisiae. Yeast strains deficient in iron transport were hypersensitive to the P. insularum aqueous extract. Further investigations showed that exogenous iron supplementation rescued the growth defect induced by P. insularum extracts, suggesting that P. insularum reduced intracellular iron. Fittingly, yeast cells treated with P. insularum extracts contained less intracellular iron than control cells. Paraxodically, the expression levels of iron transporter proteins were upregulated upon extract treatment. When we investigated iron-requiring cellular processes, we found that yeast cells treated with P. insularum extracts exhibited a respiratory deficient phenotype and reduced heme synthesis, indicative of an impaired cellular iron status. These findings suggested that P. insularum reduced bioavailable iron leading to the induction of the low iron response, and indeed the extracts of P. insularum were shown to chelate iron via the iron-chelating CAS assay. To translate results from yeast to mammalian cells, we treated primary murine macrophages with P. insularum extracts and detected an anti-inflammatory response, which we found to correlate with reduced activity of the iron-requiring aconitase enzyme. We further determined using pooled diploid mutant genetic analyses that the extracts of P. insularum did not have a genetic target. To identify the compound mediating the iron chelation mechanism, bioassay-guided isolation was conducted. Fractionation of the crude aqueous extract of P. insularum produced a relatively pure fraction that NMR and the acid-butanol assay identified as a condensed tannin. Together, these results indicate a relationship between iron chelation, a condensed tannin and inflammation. Further, we established an iron chelation mechanism of action by which P. insularum extracts are used to treat inflammation-associated symptoms in traditional Samoan medicine. Lastly, the findings presented here substantiate the reliability of plants with ethnobotanical background as sources for bioactive natural products.</p>

Iron, Heme Synthesis and Erythropoietic Porphyrias: A Complex Interplay

Erythropoietic porphyrias are caused by enzymatic dysfunctions in the heme biosynthetic pathway, resulting in porphyrins accumulation in red blood cells. The porphyrins deposition in tissues, including the skin, leads to photosensitivity that is present in all erythropoietic porphyrias. In the bone marrow, heme synthesis is mainly controlled by intracellular labile iron by post-transcriptional regulation: translation of ALAS2 mRNA, the first and rate-limiting enzyme of the pathway, is inhibited when iron availability is low. Moreover, it has been shown that the expression of ferrochelatase (FECH, an iron-sulfur cluster enzyme that inserts iron into protoporphyrin IX to form heme), is regulated by intracellular iron level. Accordingly, there is accumulating evidence that iron status can mitigate disease expression in patients with erythropoietic porphyrias. This article will review the available clinical data on how iron status can modify the symptoms of erythropoietic porphyrias. We will then review the modulation of heme biosynthesis pathway by iron availability in the erythron and its role in erythropoietic porphyrias physiopathology. Finally, we will summarize what is known of FECH interactions with other proteins involved in iron metabolism in the mitochondria.

Ferroptosis as a Major Factor and Therapeutic Target for Neuroinflammation in Parkinson’s Disease

Mounting evidence suggests that ferroptosis is not just a consequence but also a fundamental contributor to the development and progression of Parkinson’s disease (PD). Ferroptosis is characterized as iron-dependent regulated cell death caused by excessive lipid peroxidation, leading to plasma membrane rupture, release of damage-associated molecular patterns, and neuroinflammation. Due to the crucial role of intracellular iron in mediating the production of reactive oxygen species and the formation of lipid peroxides, ferroptosis is intimately controlled by regulators involved in many aspects of iron metabolism, including iron uptake, storage and export, and by pathways constituting the antioxidant systems. Translational and transcriptional regulation of iron homeostasis and redox status provide an integrated network to determine the sensitivity of ferroptosis. We herein review recent advances related to ferroptosis, ranging from fundamental mechanistic discoveries and cutting-edge preclinical animal studies, to clinical trials in PD and the regulation of neuroinflammation via ferroptosis pathways. Elucidating the roles of ferroptosis in the survival of dopaminergic neurons and microglial activity can enhance our understanding of the pathogenesis of PD and provide opportunities for the development of novel prevention and treatment strategies.