Methyltransferase-like 3 silenced inhibited the ferroptosis development via regulating the glutathione peroxidase 4 levels in the intracerebral hemorrhage progression

ABSTRACT This study examined the effects of methyltransferase-like 3 (METTL3) on ferroptosis during intracerebral hemorrhage (ICH) progression. The brain microvascular endothelial cells (BMVECs) were stimulated with oxygen and glucose deprivation (OGD) and hemin to establish an ICH model. Cell viability was tested using a CCK8 assay. The levels of Fe2+, glutathione, reactive oxygen species, LPO, and MDA were determined using the corresponding commercial kits. Cell death was analyzed using TUNEL and propidium iodide staining. The correlation between METTL3 and glutathione peroxidase 4 (GPX4) was analyzed using Spearman’s correlation test and further confirmed using the CHIP assay. Western blotting and RT-qPCR were performed to measure the relative expression levels. Mice were injected with 0.2 units collagenase IV to establish an ICH model in vivo. We found that the Fe2+, reactive oxygen species, LPO, and MDA levels were enhanced, and glutathione was depleted in OGD/H-treated BMVECs as well as in ICH mice. Additionally, cell viability and SLC7A11 protein levels decreased, and cell death and TFR1 protein levels increased in OGD/H-treated BMVECs. METTL3 silencing relieves OGD/H-induced injury in BMVECs. In addition, METTL3 was significantly negatively related to GPX4, which was further confirmed by the CHIP assay. Silencing of METTL3 decreased the N6-methyladenosine levels of GPX4 and increased its mRNA levels of GPX4. GPX4 knockdown neutralized the role of METTL3 in OGD/H-treated BMVECs. These results implied that ferroptosis occurred in the ODG/H-treated BMVECs and ICH mouse models. METTL3 silencing effectively suppressed ferroptosis by regulating N6-methyladenosine and mRNA levels of GPX4. Graphical abstract

[1]  Zhiwei Huang,et al.  Naringenin alleviates myocardial ischemia/reperfusion injury by regulating the nuclear factor-erythroid factor 2-related factor 2 (Nrf2) /System xc-/ glutathione peroxidase 4 (GPX4) axis to inhibit ferroptosis , 2021, Bioengineered.

[2]  Shuyuan Liu,et al.  METTL3-mediated m6A modification of Bcl-2 mRNA promotes non-small cell lung cancer progression , 2021, Oncology reports.

[3]  Qiang Wang,et al.  Bioinformatics Analysis Identifies Potential Ferroptosis Key Genes in the Pathogenesis of Intracerebral Hemorrhage , 2021, Frontiers in Neuroscience.

[4]  N. Rabelo,et al.  Intracerebral Hemorrhage and Ferroptosis: Something Else that STICH Should Know? , 2021, World neurosurgery.

[5]  S. Kyuwa,et al.  Capsid integrity RT-qPCR for the selective detection of intact SARS-CoV-2 in wastewater , 2021, Science of The Total Environment.

[6]  Ji Wang,et al.  Metformin induces Ferroptosis by inhibiting UFMylation of SLC7A11 in breast cancer , 2021, Journal of Experimental & Clinical Cancer Research.

[7]  Hong Wang,et al.  Baicalin Inhibits Ferroptosis in Intracerebral Hemorrhage , 2021, Frontiers in Pharmacology.

[8]  Junjie Chen,et al.  mTORC1 couples cyst(e)ine availability with GPX4 protein synthesis and ferroptosis regulation , 2021, Nature Communications.

[9]  S. Yue,et al.  Legumain promotes tubular ferroptosis by facilitating chaperone-mediated autophagy of GPX4 in AKI , 2021, Cell death & disease.

[10]  Bing Chen,et al.  Long non-coding RNA H19 protects against intracerebral hemorrhage injuries via regulating microRNA-106b-5p/acyl-CoA synthetase long chain family member 4 axis , 2021, Bioengineered.

[11]  Yanjun Che,et al.  Baicalin suppresses autophagy-dependent ferroptosis in early brain injury after subarachnoid hemorrhage , 2021, Bioengineered.

[12]  Zhengzheng Wang,et al.  HBXIP drives metabolic reprogramming in hepatocellular carcinoma cells via METTL3‐mediated m6A modification of HIF‐1α , 2020, Journal of cellular physiology.

[13]  V. Yong,et al.  Neuroinflammation in intracerebral haemorrhage: immunotherapies with potential for translation , 2020, The Lancet Neurology.

[14]  Gaiqing Wang,et al.  Ferroptosis, a Regulated Neuronal Cell Death Type After Intracerebral Hemorrhage , 2020, Frontiers in Cellular Neuroscience.

[15]  Zhonghua Ma,et al.  N6‐methyladenosine (m6A) RNA modification in cancer stem cells , 2020, Stem cells.

[16]  D. Tang,et al.  Oxidative Damage and Antioxidant Defense in Ferroptosis , 2020, Frontiers in Cell and Developmental Biology.

[17]  Xiang Song,et al.  Role of GPX4-Mediated Ferroptosis in the Sensitivity of Triple Negative Breast Cancer Cells to Gefitinib , 2020, Frontiers in Oncology.

[18]  V. Anggono,et al.  Altered Expression of the m6A Methyltransferase METTL3 in Alzheimer’s Disease , 2020, eNeuro.

[19]  Yongsheng Li,et al.  The interaction between ferroptosis and lipid metabolism in cancer , 2020, Signal Transduction and Targeted Therapy.

[20]  Dan Han,et al.  Loss of Cardiac Ferritin H Facilitates Cardiomyopathy via Slc7a11-Mediated Ferroptosis , 2020, Circulation research.

[21]  B. Stockwell,et al.  Transferrin Receptor Is a Specific Ferroptosis Marker , 2020, Cell reports.

[22]  Meng Gao,et al.  NDP-MSH binding melanocortin-1 receptor ameliorates neuroinflammation and BBB disruption through CREB/Nr4a1/NF-κB pathway after intracerebral hemorrhage in mice , 2019, Journal of Neuroinflammation.

[23]  Zi-Zhen Zhang,et al.  METTL3-mediated N6-methyladenosine modification is critical for epithelial-mesenchymal transition and metastasis of gastric cancer , 2019, Molecular Cancer.

[24]  Q. Ding,et al.  METTL3-mediated m6A modification of HDGF mRNA promotes gastric cancer progression and has prognostic significance , 2019, Gut.

[25]  X. Yang,et al.  MiR-27a promotes the autophagy and apoptosis of IL-1β treated-articular chondrocytes in osteoarthritis through PI3K/AKT/mTOR signaling , 2019, Aging.

[26]  D. Lin,et al.  METTL3 facilitates tumor progression via an m6A-IGF2BP2-dependent mechanism in colorectal carcinoma , 2019, Molecular cancer.

[27]  Jung-Shin Lee,et al.  Rapid method for chromatin immunoprecipitation (ChIP) assay in a dimorphic fungus, Candida albicans , 2019, Journal of Microbiology.

[28]  M. Conrad,et al.  Role of GPX4 in ferroptosis and its pharmacological implication. , 2019, Free radical biology & medicine.

[29]  Fan Zhang,et al.  Rnf112 deletion protects brain against intracerebral hemorrhage (ICH) in mice by inhibiting TLR-4/NF-κB pathway. , 2018, Biochemical and biophysical research communications.

[30]  J. Qiu,et al.  N 6-methyladenosine modification and METTL3 modulate enterovirus 71 replication , 2018, Nucleic acids research.

[31]  Zongxi Sun,et al.  Panax notoginseng Saponins Protect Cerebral Microvascular Endothelial Cells against Oxygen-Glucose Deprivation/Reperfusion-Induced Barrier Dysfunction via Activation of PI3K/Akt/Nrf2 Antioxidant Signaling Pathway , 2018, Molecules.

[32]  Wei Li,et al.  METTL3-mediated m6A modification is required for cerebellar development , 2018, PLoS biology.

[33]  G. Lip,et al.  Anticoagulation Resumption After Intracerebral Hemorrhage , 2018, Current Atherosclerosis Reports.

[34]  Z. Shao,et al.  Melatonin resists oxidative stress‐induced apoptosis in nucleus pulposus cells , 2018, Life sciences.

[35]  K. Somasundaram,et al.  Essential role of METTL3-mediated m6A modification in glioma stem-like cells maintenance and radioresistance , 2018, Oncogene.

[36]  C. Weimar,et al.  Epidemiology, Prognosis and Prevention of Non-Traumatic Intracerebral Hemorrhage. , 2017, Current pharmaceutical design.

[37]  Samie R. Jaffrey,et al.  m6A RNA methylation promotes XIST-mediated transcriptional repression , 2016, Nature.

[38]  A. Walch,et al.  Inactivation of the ferroptosis regulator Gpx4 triggers acute renal failure in mice , 2014, Nature Cell Biology.

[39]  S. Lorkowski,et al.  Highly Efficient Transfection of Human THP-1 Macrophages by Nucleofection , 2014, Journal of visualized experiments : JoVE.

[40]  M. R. Lamprecht,et al.  Ferroptosis: An Iron-Dependent Form of Nonapoptotic Cell Death , 2012, Cell.

[41]  T. Hoppe,et al.  Western blot analysis of the autophagosomal membrane protein LGG-1/LC3 in Caenorhabditis elegans. , 2019, Methods in enzymology.