MEF2C Alleviates Postoperative Cognitive Dysfunction by Repressing Ferroptosis.

BACKGROUND: Ferroptosis, a form of programmed cell death featured by lipid peroxidation, has been proposed as a potential etiology for postoperative cognitive dysfunction (POCD). Myocyte-specific enhancer factor 2C (MEF2C), a transcription factor expressed in various brain cell types, has been implicated in cognitive disorders. This study sought to ascertain whether MEF2C governs postoperative cognitive capacity by affecting ferroptosis. METHODS: Transcriptomic analysis of public data was used to identify MEF2C as a candidate differentially expressed gene in the hippocampus of POCD mice. The POCD mouse model was established via aseptic laparotomy under isoflurane anesthesia after treatment with recombinant adeno-associated virus 9 (AAV9)-mediated overexpression of MEF2C and/or the glutathione peroxidase 4 (GPX4) inhibitor RSL3. Cognitive performance, Nissl staining, and ferroptosis-related parameters were assessed. Dual-luciferase reporter gene assays and chromatin immunoprecipitation assays were implemented to elucidate the mechanism by which MEF2C transcriptionally activates GPX4. RESULTS: MEF2C mRNA and protein levels decreased in the mouse hippocampus following anesthesia and surgery. MEF2C overexpression ameliorated postoperative memory decline, hindered lipid peroxidation and iron accumulation, and enhanced antioxidant capacity, which were reversed by RSL3. Additionally, MEF2C was found to directly bind to the Gpx4 promoter and activate its transcription. CONCLUSIONS: Our findings suggest that MEF2C may be a promising therapeutic target for POCD through its negative modulation of ferroptosis.

Sirtuin 1 in regulating the p53/glutathione peroxidase 4/gasdermin D axis in acute liver failure.

In this editorial, we comment on the article by Zhou et al. The study reveals the connection between ferroptosis and pyroptosis and the effect of silent information regulator sirtuin 1 (SIRT1) activation in acute liver failure (ALF). ALF is characterized by a sudden and severe liver injury resulting in significant hepatocyte damage, often posing a high risk of mortality. The predominant form of hepatic cell death in ALF involves apoptosis, ferroptosis, autophagy, pyroptosis, and necroptosis. Glutathione peroxidase 4 (GPX4) inhibition sensitizes the cell to ferroptosis and triggers cell death, while Gasdermin D (GSDMD) is a mediator of pyroptosis. The study showed that ferroptosis and pyroptosis in ALF are regulated by blocking the p53/GPX4/GSDMD pathway, bridging the gap between the two processes. The inhibition of p53 elevates the levels of GPX4, reducing the levels of inflammatory and liver injury markers, ferroptotic events, and GSDMD-N protein levels. Reduced p53 expression and increased GPX4 on deletion of GSDMD indicated ferroptosis and pyroptosis interaction. SIRT1 is a NAD-dependent deacetylase, and its activation attenuates liver injury and inflammation, accompanied by reduced ferroptosis and pyroptosis-related proteins in ALF. SIRT1 activation also inhibits the p53/GPX4/GSDMD axis by inducing p53 acetylation, attenuating LPS/D-GalN-induced ALF.

Overview of ferroptosis and pyroptosis in acute liver failure.

In this editorial, we comment on the article by Zhou et al published in a recent issue. We specifically focus on the crucial roles of ferroptosis and pyroptosis in acute liver failure (ALF), a disease with high mortality rates. Ferroptosis is the result of increased intracellular reactive oxygen species due to iron accumulation, glutathione (GSH) depletion, and decreased GSH peroxidase 4 activity, while pyroptosis is a procedural cell death mediated by gasdermin D which initiates a sustained inflammatory process. In this review, we describe the characteristics of ferroptosis and pyroptosis, and discuss the involvement of the two cell death modes in the onset and development of ALF. Furthermore, we summarize several interfering methods from the perspective of ferroptosis and pyroptosis for the alleviation of ALF. These observations might provide new targets and a theoretical basis for the treatment of ALF, which are also crucial for improving the prognosis of patients with ALF.

Mild therapeutic hypothermic protection activates the PI3K/AKT signaling pathway to inhibit TRPM7 and suppress ferroptosis induced by myocardial ischemia­reperfusion injury.

The present study aimed to investigate the role of PI3K­mediated ferroptosis signaling induced by mild therapeutic hypothermia (MTH), which was defined as a temperature of 34˚C, in protecting against myocardial ischemia-reperfusion (I/R) injury (MIRI). To meet this aim, H9C2 cells underwent hypoxia­reperfusion (H/R) and/or MTH. The MTT assay was used to assess cell viability, cytotoxicity was measured using a lactate dehydrogenase cytotoxicity assay, and Annexin V­FITC/PI flow cytometric analysis was used to analyze early and late cell apoptosis. In addition, 84 healthy adult male Sprague­Dawley rats were randomly divided into seven groups (n=12), and underwent I/R and various treatments. Hemodynamics were monitored, and the levels of myocardial injury marker enzymes and oxidative stress markers in myocardial tissue were measured using ELISA. The expression levels of PI3K, AKT, transient receptor potential cation channel subfamily M member 7 (TRPM7), glutathione peroxidase 4 (GPX4) and acyl­CoA synthetase long chain family member 4 (ACSL4) in animals and cells were measured using western blot analysis. These experiments revealed that MTH could effectively reduce myocardial infarct size, improve hemodynamic performance following MIRI and suppress myocardial apoptosis, thereby contributing to the recovery from H/R injury. Mechanistically, MTH was revealed to be able to activate the PI3K/AKT signaling pathway in cells, upregulating GPX4, and downregulating the expression levels of TRPM7 and ACSL4. Treatment with 2­aminoethoxydiphenyl borate (an inhibitor of TRPM7) could further strengthen the myocardial protective effects of MTH, whereas treatment with erastin (promoter of ferroptosis) and wortmannin (inhibitor of PI3K) led to the effective elimination of the myocardial protective effects of MTH. Compared with in the I/R group, the PI3K/AKT activation level and the expression levels of GPX4 were both significantly increased, whereas the expression levels of TRPM7 and ACSL4 were significantly decreased in the I/R + MTH group. Taken together, the results of the present study indicated that MTH may activate the PI3K/AKT signaling pathway to inhibit TRPM7 and suppress ferroptosis induced by MIRI.

ETV4­mediated transcriptional activation of SLC12A5 exacerbates ferroptosis resistance and glucose metabolism reprogramming in breast cancer cells.

Solute carrier family 12 member 5 (SLC12A5) is an oncogene in numerous types of cancer, however its function in breast cancer (BC) remains elusive. ETS translocation variant 4 (ETV4) promotes BC. Therefore, the present study aimed to elucidate the role of SLC12A5 in ferroptosis and glucose metabolism in BC cells as well as to understand the underlying mechanism. Analysis of data from the UALCAN database demonstrated expression levels of SLC12A5 in BC and its association with prognosis. Reverse transcription­quantitative PCR and western blotting were conducted to evaluate the expression levels of SLC12A5 and ETV4 in BC cells. The abilities of BC cells to proliferate, migrate and invade were assessed using Cell Counting Kit­8, colony formation, wound healing and Transwell assays. Thiobarbituric acid reactive substances assay and a C11 BODIPY 581/591 probe were used to evaluate lipid peroxidation. Ferroptosis resistance was evaluated by the measurement of Fe2+ and ferroptosis­related solute carrier family 7a member 11 (SLC7A11), glutathione peroxidase 4 (GPX4), acyl­CoA synthetase long­chain family member 4 (ACSL4) and transferrin receptor 1 (TFR1) protein levels. Glycolysis was assessed via evaluation of extracellular acidification rate, oxygen consumption rate, lactate production and glucose consumption. Finally, luciferase reporter and chromatin immunoprecipitation assay were used to verify the interaction between ETV4 and the SLC12A5 promoter. UALCAN database analysis indicated that SLC12A5 was upregulated in BC tissues and cells and that SLC12A5 elevation indicated a poor prognosis of patients with BC. SLC12A5 knockdown suppressed the BC cell proliferative, migratory and invasive capabilities. Moreover, SLC12A5 knockdown decreased BC cell ferroptosis resistance and glucose metabolism reprogramming. The transcription factor ETV4 was demonstrated to bind to the SLC12A5 promoter and upregulate its transcription. Furthermore, ETV4 overexpression counteracted the suppressive effect of SLC12A5 knockdown on the BC cell proliferative, migratory and invasive abilities, as well as on ferroptosis resistance and glucose metabolism reprogramming. Transcriptional activation of SLC12A5 by ETV4 modulated the migration, invasion, ferroptosis resistance and glucose metabolism reprogramming of BC cells.

Intracellular C5aR1 inhibits ferroptosis in glioblastoma through METTL3-dependent m6A methylation of GPX4.

Glioblastoma (GBM) is the most common primary intracranial malignant tumor. Recent literature suggests that induction of programmed death has become a mainstream cancer treatment strategy, with ferroptosis being the most widely studied mode. Complement C5a receptor 1 (C5aR1) is associated with both tumorigenesis and tumor-related immunity. However, knowledge regarding the role of C5aR1 in GBM progression is limited. In the present study, we observed significant upregulation of C5aR1 in glioma tissue. In addition, C5aR1 expression was found to be closely associated with patient prognosis and survival. Subsequent experimental verification demonstrated that C5aR1 promoted the progression of GBM mainly by suppressing ferroptosis induction, inhibiting the accumulation of lipid peroxides, and stabilizing the expression of the core antiferroptotic factor glutathione peroxidase 4 (GPX4). Aberrant N6-methyladenosine (m6A) modification of GPX4 mRNA contributes significantly to epigenetic tumorigenesis, and here, we report that selective methyltransferase-like 3 (METTL3)-dependent m6A methylation of GPX4 plays a key role in C5AR1 knockdown-induced ferroptosis induction. Mechanistically, ERK1/2 signaling pathway activation increases the METTL3 protein abundance in GBM cells. This activation then increases the stability of METTL3-mediated m6A modifications on GPX4, enabling it to fulfill its transcriptional function. More importantly, in an intracranial xenograft mouse model, PMX205, a C5aR1 inhibitor, promoted alterations in ferroptosis in GBM cells and inhibited GBM progression. In conclusion, our findings suggest that C5aR1 inhibits ferroptosis in GBM cells and promotes MettL3-dependent GPX4 expression through ERK1/2, thereby promoting glioma progression. Our study reveals a novel mechanism by which the intracellular complement receptor C5aR1 suppresses ferroptosis induction and promotes GBM progression. These findings may facilitate the identification of a potential therapeutic target for glioma.

Human-augmented large language model-driven selection of glutathione peroxidase 4 as a candidate blood transcriptional biomarker for circulating erythroid cells.

The identification of optimal candidate genes from large-scale blood transcriptomic data is crucial for developing targeted assays to monitor immune responses. Here, we introduce a novel, optimized large language model (LLM)-based approach for prioritizing candidate biomarkers from blood transcriptional modules. Focusing on module M14.51 from the BloodGen3 repertoire, we implemented a multi-step LLM-driven workflow. Initial high-throughput screening used GPT-4, Claude 3, and Claude 3.5 Sonnet to score and rank the module's constituent genes across six criteria. Top candidates then underwent high-resolution scoring using Consensus GPT, with concurrent manual fact-checking and, when needed, iterative refinement of the scores based on user feedback. Qualitative assessment of literature-based narratives and analysis of reference transcriptome data further refined the selection process. This novel multi-tiered approach consistently identified Glutathione Peroxidase 4 (GPX4) as the top candidate gene for module M14.51. GPX4's role in oxidative stress regulation, its potential as a future drug target, and its expression pattern across diverse cell types supported its selection. The incorporation of reference transcriptome data further validated GPX4 as the most suitable candidate for this module. This study presents an advanced LLM-driven workflow with a novel optimized scoring strategy for candidate gene prioritization, incorporating human-in-the-loop augmentation. The approach identified GPX4 as a key gene in the erythroid cell-associated module M14.51, suggesting its potential utility for biomarker discovery and targeted assay development. By combining AI-driven literature analysis with iterative human expert validation, this method leverages the strengths of both artificial and human intelligence, potentially contributing to the development of biologically relevant and clinically informative targeted assays. Further validation studies are needed to confirm the broader applicability of this human-augmented AI approach.

The role and possible mechanism of the ferroptosis-related SLC7A11/GSH/GPX4 pathway in myocardial ischemia-reperfusion injury.

BACKGROUND: Myocardial ischemia-reperfusion injury (MI/RI) is an unavoidable risk event for acute myocardial infarction, with ferroptosis showing close involvement. We investigated the mechanism of MI/RI inducing myocardial injury by inhibiting the ferroptosis-related SLC7A11/glutathione (GSH)/glutathione peroxidase 4 (GPX4) pathway and activating mitophagy. METHODS: A rat MI/RI model was established, with myocardial infarction area and injury assessed by TTC and H&E staining. Rat cardiomyocytes H9C2 were cultured in vitro, followed by hypoxia/reoxygenation (H/R) modeling and the ferroptosis inhibitor lipoxstatin-1 (Lip-1) treatment, or 3-Methyladenine or rapamycin treatment and overexpression plasmid (oe-SLC7A11) transfection during modeling. Cell viability and death were evaluated by CCK-8 and LDH assays. Mitochondrial morphology was observed by transmission electron microscopy. Mitochondrial membrane potential was detected by fluorescence dye JC-1. Levels of inflammatory factors, reactive oxygen species (ROS), Fe2+, malondialdehyde, lipid peroxidation, GPX4 enzyme activity, glutathione reductase, GSH and glutathione disulfide, and SLC7A11, GPX4, LC3II/I and p62 proteins were determined by ELISA kit, related indicator detection kits and Western blot. RESULTS: The ferroptosis-related SLC7A11/GSH/GPX4 pathway was repressed in MI/RI rat myocardial tissues, inducing myocardial injury. H/R affected GSH synthesis and inhibited GPX4 enzyme activity by down-regulating SLC7A11, thus promoting ferroptosis in cardiomyocytes, which was averted by Lip-1. SLC7A11 overexpression improved H/R-induced cardiomyocyte ferroptosis via the GSH/GPX4 pathway. H/R activated mitophagy in cardiomyocytes. Mitophagy inhibition reversed H/R-induced cellular ferroptosis. Mitophagy activation partially averted SLC7A11 overexpression-improved H/R-induced cardiomyocyte ferroptosis. H/R suppressed the ferroptosis-related SLC7A11/GSH/GPX4 pathway by inducing mitophagy, leading to cardiomyocyte injury. CONCLUSIONS: Increased ROS under H/R conditions triggered cardiomyocyte injury by inducing mitophagy to suppress the ferroptosis-related SLC7A11/GSH/GPX4 signaling pathway activation.

GZMA suppressed GPX4-mediated ferroptosis to improve intestinal mucosal barrier function in inflammatory bowel disease.

BACKGROUND: Our previous study has demonstrated a decreased colonic CD8+CD39+ T cells, enrichment of granzyme A (GZMA), was found in pediatric-onset colitis and inflammatory bowel disease (IBD) characterized by impaired intestinal barrier function. However, the influence of GZMA on intestinal barrier function remains unknown. METHODS: Western blotting(WB), real-time PCR (qPCR), immunofluorescence (IF) and in vitro permeability assay combined with intestinal organoid culture were used to detect the effect of GZMA on intestinal epithelial barrier function in vivo and in vitro. Luciferase, immunoprecipitation (IP) and subcellular fractionation isolation were performed to identify the mechanism through which GZMA modulated intestinal epithelial barrier function. RESULTS: Herein, we, for the first time, demonstrated that CD8+CD39+ T cells promoted intestinal epithelial barrier function through GZMA, leading to induce Occludin(OCLN) and Zonula Occludens-1(ZO-1) expression, which was attributed to enhanced CDX2-mediated cell differentiation caused by increased glutathione peroxidase 4(GPX4)-induced ferroptosis inhibition in vivo and in vitro. Mechanically, GZMA inhibited intestinal epithelial cellular PDE4B activation to trigger cAMP/PKA/CREB cascade signaling to increase CREB nuclear translocation, initiating GPX4 transactivity. In addition, endogenous PKA interacted with CREB, and this interaction was enhanced in response to GZMA. Most importantly, administration of GZMA could alleviate DSS-induced colitis in vivo. CONCLUSION: These findings extended the novel insight of GZMA contributed to intestinal epithelial cell differentiation to improve barrier function, and enhacement of GZMA could be a promising strategy to patients with IBD.

Borna disease virus 1 induces ferroptosis, contributing to lethal encephalitis.

Borna disease virus 1 (BoDV-1) is a neurotropic RNA virus that has been linked to fatal BoDV-1 encephalitis (BVE) in humans. Ferroptosis represents a newly recognized kind of programmed cell death that marked by iron overload and lipid peroxidation. Various viral infections are closely related to ferroptosis. However, the link between BoDV-1 infection and ferroptosis, as well as its role in BVE pathogenesis, remains inadequately understood. Herein, we used primary rat cortical neurons, human microglial HMC3 cells, and Sprague‒Dawley rats as models. BoDV-1 infection induced ferroptosis, as ferroptosis characteristics were detected (iron overload, reactive oxygen species buildup, decreased antioxidant capacity, lipid peroxidation, and mitochondrial damage). Analysis via qRT-PCR and Western blot demonstrated that BoDV-1-induced ferroptosis was mediated through Nrf2/HO-1/SLC7a11/GPX4 antioxidant pathway suppression. Nrf2 downregulation was due to BoDV-1 infection promoting Nrf2 ubiquitination and degradation. Following BoDV-1-induced ferroptosis, the PTGS2/PGE2 signaling pathway was activated, and various intracellular lipid peroxidation products and damage-associated molecular patterns were released, contributing to BVE occurrence and progression. More importantly, inhibiting ferroptosis or the ubiquitin‒proteasome system effectively alleviated BVE. Collectively, these findings demonstrate the interaction between BoDV-1 infection and ferroptosis and reveal BoDV-1-induced ferroptosis as an underlying pathogenic mechanism of BVE.

KLF14 directly downregulates the expression of GPX4 to exert antitumor effects by promoting ferroptosis in cervical cancer.

BACKGROUND: Cervical cancer is the fourth leading cause of cancer-related death among women worldwide, and effective therapeutic strategies for its treatment are limited. Recent studies have indicated that ferroptosis, a form of regulated cell death, is a promising therapeutic strategy. KLF14 has been shown to regulate both cell proliferation and apoptosis in cervical cancer. However, its role in modulating lipid peroxidation and ferroptosis remains largely unexplored and enigmatic. METHODS: SiHa and HeLa cells were transduced with lentiviral vectors to overexpress KLF14. Protein levels were analyzed via western blotting and immunohistochemistry (IHC). LDH assays, calcein-AM/propidium iodide (PI) staining, and generation of cell growth curves using a real-time cell analysis (RTCA) system were used to detect cell damage and proliferation. Cellular ROS, lipid ROS, transmission electron microscopy (TEM), and Fe2+ assays and a xenograft mouse model were used to measure the level of ferroptosis. Proteomics combined with bioinformatics methods was used to screen target genes regulated by KLF14, and CUT&Tag and dual-luciferase assays confirmed the repression of GPX4 by KLF14 via direct binding to its promoter. RESULTS: KLF14 is abnormally expressed in various tumors and downregulated in cervical cancer. Overexpression of KLF14 induced ferroptosis and inhibited cell proliferation in vitro as well as xenograft tumorigenicity in vivo. Mechanistic studies revealed that KLF14 binds to the promoter of GPX4, suppressing its transcriptional activity and thereby decreasing its expression, which contributes to the induction of ferroptosis. Truncation and point mutation analyses of the GPX4 promoter revealed multiple binding sites for KLF14 within the - 1000 bp to + 35 bp region, which are responsible for its inhibitory effect on GPX4 transcription. Additionally, deletion of the zinc finger motif in KLF14 abolished its inhibitory effect on GPX4 promoter activity and cell proliferation. CONCLUSION: Our data revealed a previously unidentified function of KLF14 in promoting ferroptosis, which results in the suppression of cell proliferation. Mechanistically, we revealed a novel regulatory mechanism by which KLF14 targets GPX4. These findings suggest a novel strategy to induce ferroptosis through the targeting of KLF14 in human cervical cancer cells.

Cold plasma irradiation inhibits skin cancer via ferroptosis.

Cold atmospheric plasma (CAP) has been extensively utilized in medical treatment, particularly in cancer therapy. However, the underlying mechanism of CAP in skin cancer treatment remains elusive. In this study, we established a skin cancer model using CAP treatmentin vitro. Also, we established the Xenograft experiment modelin vivo. The results demonstrated that treatment with CAP induced ferroptosis, resulting in a significant reduction in the viability, migration, and invasive capacities of A431 squamous cell carcinoma, a type of skin cancer. Mechanistically, the significant production of reactive oxygen species (ROS) by CAP induces DNA damage, which then activates Ataxia-telangiectasia mutated (ATM) and p53 through acetylation, while simultaneously suppressing the expression of Solute Carrier Family 7 Member 11 (SLC7A11). Consequently, this cascade led to the down-regulation of intracellular Glutathione peroxidase 4 (GPX4), ultimately resulting in ferroptosis. CAP exhibits a favorable impact on skin cancer treatment, suggesting its potential medical application in skin cancer therapy.

ELK4 targets CHMP6 to inhibit ferroptosis and enhance malignant properties of skin cutaneous melanoma cells.

Ferroptosis, a key factor in tumor progression, is poorly understood at the molecular level. This study investigates how ELK4 and CHMP6 regulate skin cutaneous melanoma (SKCM) cell proliferation and ferroptosis. Analysis of TCGA data reveals high expression of ELK4 and CHMP6 in SKCM. Overexpression of ELK4 or CHMP6 enhances cell proliferation, invasion, and migration while reducing ROS and Fe2 + levels. It also increases GPX4 and xCT expression and decreases ACSL4 levels in SKCM cells. The opposite effects are observed with ELK4 or CHMP6 knockdown. ELK4 binds to the CHMP6 promoter, promoting CHMP6 transcription. Knockdown of CHMP6 reverses the oncogenic effects of ELK4 overexpression. In conclusion, ELK4 enhances proliferation, invasion, and migration while inhibiting ferroptosis in SKCM cells by upregulating CHMP6 transcription. This study sheds light on the intricate mechanisms involved in SKCM progression and identifies potential therapeutic targets in melanoma treatment.

Brown adipose tissue-derived exosomes improve polycystic ovary syndrome in mice via STAT3/GPX4 signaling pathway.

Polycystic ovary syndrome (PCOS) is associated with impaired adipose tissue physiology. Elevated brown adipose tissue (BAT) mass or activity has shown potential in the treatment of PCOS. In this study, we aimed to investigate whether BAT-derived exosomes (BAT-Exos), as potential biomarkers of BAT activity, exert similar benefits as BAT in the treatment of PCOS. PCOS was induced in female C57BL/6J mice orally administered 1 mg/kg of letrozole for 21 days. Subsequently, the animals underwent transplantation with BAT or administered BAT-Exos (200 µg) isolated from young healthy mice via the tail vein; healthy female mice were used as controls. The results indicate that BAT-Exos treatment significantly reduced body weight and improved insulin resistance in PCOS mice. In addition, BAT-Exos improved ovulation function by reversing the acyclicity of the estrous cycle, decreasing circulating luteinizing hormone and testosterone, recovering ovarian performance, and improving oocyte quality, leading to a higher pregnancy rate and litter size. Furthermore, western blotting revealed reduced expression of signal transducer and activator of transcription 3 (STAT3) and increased expression of glutathione peroxidase 4 (GPX4) in the ovaries of mice in the BAT-Exos group. To further explore the role of the STAT3/GPX4 signaling pathway in PCOS mice, we treated the mice with an intraperitoneal injection of 5 mg/kg stattic, a STAT3 inhibitor. Consistent with BAT-Exos treatment, the administration of stattic rescued letrozole-induced PCOS phenotypes. These findings suggest that BAT-Exos treatment might be a potential therapeutic strategy for PCOS and that the STAT3/GPX4 signaling pathway is a critical therapeutic target for PCOS.

GPX4 restricts ferroptosis of NKp46+ILC3s to control intestinal inflammation.

Group 3 innate lymphoid cells (ILC3s) are essential for both pathogen defense and tissue homeostasis in the intestine. Dysfunction of ILC3s could lead to increased susceptibility to intestinal inflammation. However, the precise mechanisms governing the maintenance of intestinal ILC3s are yet to be fully elucidated. Here, we demonstrated that ferroptosis is vital for regulating the survival of intestinal ILC3. Ferroptosis-related genes, including GPX4, a key regulator of ferroptosis, were found to be upregulated in intestinal mucosal ILC3s from ulcerative colitis patients. Deletion of GPX4 resulted in a decrease in NKp46+ILC3 cell numbers, impaired production of IL-22 and IL-17A, and exacerbated intestinal inflammation in a T cell-independent manner. Our mechanistic studies revealed that GPX4-mediated ferroptosis in NKp46+ILC3 cells was regulated by the LCN2-p38-ATF4-xCT signaling pathway. Mice lacking LCN2 in ILC3s or administration of a p38 pathway inhibitor exhibited similar phenotypes of ILC3 and colitis to those observed in GPX4 conditional knock-out mice. These observations provide novel insights into therapeutic strategies for intestinal inflammation by modulating ILC3 ferroptosis.

MFSD7C protects hemolysis-induced lung impairments by inhibiting ferroptosis.

Hemolysis drives susceptibility to lung injury and predicts poor outcomes in diseases, such as malaria and sickle cell disease (SCD). However, the underlying pathological mechanism remains elusive. Here, we report that major facilitator superfamily domain containing 7 C (MFSD7C) protects the lung from hemolytic-induced damage by preventing ferroptosis. Mechanistically, MFSD7C deficiency in HuLEC-5A cells leads to mitochondrial dysfunction, lipid remodeling and dysregulation of ACSL4 and GPX4, thereby enhancing lipid peroxidation and promoting ferroptosis. Furthermore, systemic administration of MFSD7C mRNA-loaded nanoparticles effectively prevents lung injury in hemolytic mice, such as HbSS-Townes mice and PHZ-challenged 7 C-/- mice. These findings present the detailed link between hemolytic complications and ferroptosis, providing potential therapeutic targets for patients with hemolytic disorders.

[Sanguinarine induces ferroptosis of colorectal cancer cells by upregulating STUB1 and downregulating GPX4].

OBJECTIVE: To investigate the effect of sanguinarine (SAN) on proliferation and ferroptosis of colorectal cancer cells. METHODS: SW620 and HCT-116 cells treated with different concentrations of SAN were examined for cell viability changes using CCK8 assay to determine the IC50 of SAN in the two cells. The inhibitory effects of SAN on proliferation, invasion and migration of the cells were evaluated using colony-forming assay and Transwell assays. ROS production in the treated cells was analyzed with flow cytometry, and lipid peroxide production was assessed by detecting malondialdehyde (MDA) level. Glutathione (GSH) levels in the cells were detected, and Western blotting was used to detect the expressions of ferroptosis-related proteins STUB1 and GPX4. RESULTS: SAN significantly inhibited the proliferation, invasion and migration of SW620 and HCT-116 cells. SAN treatment significantly promoted ROS production, increased intracellular MDA level, and lowered GSH level in the two cells (P<0.05). Western blotting showed that SAN significantly upregulated the expression of STUB1 and down-regulated the expression of its downstream protein GPX4 (P<0.05). CONCLUSION: SAN induces ferroptosis in colorectal cancer cells by regulating STUB1/GPX4, which may serve as a new therapeutic target for colorectal cancer.

Identification of genes supporting cold resistance of mammalian cells: lessons from a hibernator.

Susceptibility of human cells to cold stress restricts the use of therapeutic hypothermia and long-term preservation of organs at low temperatures. In contrast, cells of mammalian hibernators possess remarkable cold resistance, but little is known about the molecular mechanisms underlying this phenomenon. In this study, we conducted a gain-of-function screening of genes that confer cold resistance to cold-vulnerable human cells using a cDNA library constructed from the Syrian hamster, a mammalian hibernator, and identified Gpx4 as a potent suppressor of cold-induced cell death. Additionally, genetic deletion of or pharmacological inhibition of Gpx4 revealed that Gpx4 is necessary for suppressing lipid peroxidation specifically under cold in hamster cell lines. Genetic disruption of other ferroptosis-suppressing pathways, namely biopterin synthesis and mitochondrial or plasma membrane CoQ reduction pathways, also accelerated cold-induced cell death under Gpx4 dysfunction. Collectively, ferroptosis-suppressing pathways protect the cells of a mammalian hibernator from cold-induced cell death and the augmentation of these pathways renders cold resistance to cells of non-hibernators, including humans.

Hypoxia inducible factor (HIF) 3α prevents COPD by inhibiting alveolar epithelial cell ferroptosis via the HIF-3α-GPx4 axis.

Rationale: COPD patients are largely asymptomatic until the late stages when prognosis is generally poor. In this study, we shifted the focus to pre-COPD and smoking stages, and found enrichment of hypoxia inducible factor (HIF)-3α is in pre-COPD samples. Smoking induced regional tissue hypoxia and emphysema have been found in COPD patients. However, the mechanisms underlying hypoxia especially HIF-3α and COPD have not been investigated. Methods: We performed bulk-RNA sequencing on 36 peripheral lung tissue specimens from non-smokers, smokers, pre-COPD and COPD patients, and using "Mfuzz" algorithm to analysis the dataset dynamically. GSE171541 and EpCAM co-localization analyses were used to explore HIF-3α localization. Further, SftpcCreert2/+R26LSL-Hif3a knock-in mice and small molecular inhibitors in vitro were used to explore the involvement of HIF-3α in the pathophysiology of COPD. Results: Reactive oxygen species (ROS) and hypoxia were enriched in pre-COPD samples, and HIF-3α was downregulated in alveolar epithelial cells in COPD. In vitro experiments using lentivirus transfection, bulk-RNA seq, and RSL3 showed that the activation of the HIF-3α-GPx4 axis inhibited alveolar epithelial cell ferroptosis when treated with cigarettes smoking extracts (CSE). Further results from SftpcCreert2/+R26LSL-Hif3a knock-in mice demonstrated overexpression of HIF-3α inhibited alveolar epithelial cells ferroptosis and prevented the decline of lung function. Conclusion: Hypoxia and oxidation-related damage begins years before the onset of COPD symptoms, suggesting the imbalance and impairment of intracellular homeostatic system. The activation of the HIF-3α-GPx4 axis is a promising treatment target. By leveraging this comprehensive analysis method, more potential targets could be found and enhancing our understanding of the pathogenesis.

Connexin43 overexpression promoted ferroptosis and increased myocardial vulnerability to ischemia-reperfusion injury in type 1 diabetic mice.

Enhancement of Connexin43 (Cx43) and ferroptosis are respectively associated with the exacerbation of myocardial ischemia-reperfusion injury (MIRI) in diabetes. Myocardial vulnerability to ischemic insult has been shown to vary during early and later phases of diabetes in experimental settings. Whether or not Connexin43 (Cx43) and ferroptosis interplay during MIRI in diabetes is unknown. We, thus, aimed to investigate whether or not the content of myocardial Cx43 may be attributable to myocardial vulnerability to MIRI at different stages of diabetes and also to explore the potential interplay between Cx43 and ferroptosis in this pathology. Age-matched control and subgroups of Streptozotocin-induced diabetic mice were subjected to MIRI induced by 30 minutes coronary artery occlusion and 2 hours reperfusion respectively at 1, 2 and 5 weeks of diabetes. Rat cardiac H9C2 cells were exposed to high glucose (HG) for 48h in the absence or presence of Cx43 gene knockdown followed by hypoxia/reoxygenation (HR) respectively for 6 and 12 hours. Post-ischemic myocardial infarct size was reduced in 1 and 2 weeks DM mice concomitant with enhanced GPX4 and reduced cardiac Cx43 and ferroptosis as compared to control. By contrast, cardiac GPX4 was significantly reduced while Cx43 increased at DM 5 weeks (D5w) which was correspondent to significant increases in ferroptosis and myocardial infarction. Post-ischemic cardiac function was improved in 1 and 2 weeks but worsened in 5w DM mice as compared with non-diabetic control. GAP19 (Cx43 inhibitor) significantly attenuated ferroptosis and reduced myocardial infarction in D5w mice. Erastin (ferroptosis activator) reversed the cardioprotective effect of GAP19. In vitro, HR significantly reduced cell viability accompanied with reduced GPX4 but elevated Cx43 expression, MDA production and ferroptosis. Cx43 gene knockdown in H9C2 resulted in a significant increase in GPX4, reduction in MDA and ferroptosis, and subsequently reduced post-hypoxic cell viability. The beneficial effects of Cx43 gene knock-down was minified or eliminated by Erastin. It is concluded that Cx43 overexpression exacerbates MIRI under diabetic conditions via promoting ferroptosis, while its down-regulation at early state of diabetes is attributable to enhanced myocardial tolerance to MIRI.