M6A Methyltransferase METTL3 Modulates Traumatic Brain Injury by Targeting Ferroptosis.

Background: Traumatic brain injury (TBI) is a disease caused by external forces that damage brain structure and function. After TBI, iron accumulation and reactive oxygen species (ROS) increase lipid peroxidation, promoting ferroptosis. Methyltransferase-like 3 (METTL3) inhibits ferroptosis by modulating related signaling pathways. This study investigates the effects of METTL3 on neuronal ferroptosis in TBI, offering new insights and potential therapies.

Methods: TBI mouse and neuron cell models were established and treated with METTL3 overexpression. The Morris Water Maze (MWM) test evaluated cognitive function. Histological staining of brain tissues was conducted to assess brain injury, nuclear pyknosis, and iron accumulation. The activation of neurons, microglia, and astrocytes were detected using immunofluorescence staining. Neuron cell proliferation was measured using the Cell Counting Kit 8 (CCK-8). Quantitative PCR (qPCR) and western blot detected the mRNA and protein expression. Ferroptosis was assessed by measuring the accumulation of iron, malondialdehyde (MDA), superoxide dismutase (SOD), and ROS. The quantification of the N6-methyladenosine (m6A) RNA methylation levels in cells was quantified using the m6A-ELISA assay. Methylated RNA immunoprecipitation (MeRIP) assays were conducted to analyze the m6A modification on GPX4 mRNA. The interaction between YTHDF2 and GPX4 mRNA was measured using RNA pulldown and RNA immunoprecipitation (RIP) assays.

Results: METTL3 expression was downregulated in TBI-injured brain tissues. Overexpression of METTL3 improved cognitive function and brain recovery while simultaneously reducing ferroptosis and neuroinflammation. METTL3 overexpression upregulated GPX4 expression both in vitro and in vivo. Further studies indicated that m6A reader protein YTHDF2 binds to GPX4 mRNA, consequently mediating the METTL3-regulated m6A enrichment and RNA stability of GPX4. Knockdown of GPX4 and treatment with ferroptosis inducer abolished the protective effects of METTL3 on neurons.

Conclusion: METTL3 exhibits anti-ferroptosis properties and promotes brain injury recovery after TBI by regulating the m6A modification and RNA stability of GPX4.