METTL14 通过调控 GPX4 的 m6A 修饰促进骨关节炎软骨细胞铁死亡。

METTL14 promotes chondrocyte ferroptosis in osteoarthritis via m6A modification of GPX4.

机构信息

Tianjin University, Tianjin, China.

Specialized Orthopedics Construction Office, Tianjin Nankai Hospital, Tianjin, China.

出版信息

Int J Rheum Dis. 2024 Aug;27(8):e15297. doi: 10.1111/1756-185X.15297.

DOI:10.1111/1756-185X.15297

PMID:39175261

Abstract

BACKGROUND

Ferroptosis is caused by iron-dependent peroxidation of membrane phospholipids and chondrocyte ferroptosis contributes to osteoarthritis (OA) progression. Glutathione peroxidase 4 (GPX4) plays a master role in blocking ferroptosis. N6-methyladenosine (m6A) is an epigenetic modification among mRNA post-transcriptional modifications. This study investigated the effect of methyltransferase-like 14 (METTL14), the key component of the m6A methyltransferase, on chondrocyte ferroptosis via m6A modification.

METHODS

An OA rat model was established through an intra-articular injection of monosodium iodoacetate in the right knee. OA cartilages in rat models were used for gene expression analysis. Primary mouse chondrocytes or ADTC5 cells were stimulated with IL-1β or erastin. The m6A RNA methylation quantification kit was used to measure m6A level. The effect of METTL14 and GPX4 on ECM degradation and ferroptosis was investigated through western blotting, fluorescence immunostaining, propidium iodide staining, and commercially available kits. The mechanism of METTL14 action was explored through MeRIP-qPCR assays.

RESULTS

METTL14 and m6A expression was upregulated in osteoarthritic cartilages and IL-1β-induced chondrocytes. METTL14 depletion repressed the IL-1β or erastin-stimulated ECM degradation and ferroptosis in mouse chondrocytes. METTL14 inhibited GPX4 gene through m6A methylation modification. GPX4 knockdown reversed the si-METTL14-mediated protection in IL-1β-induced chondrocytes.

CONCLUSION

METTL14 depletion inhibits ferroptosis and ECM degradation by suppressing GPX4 mRNA m6A modification in injured chondrocytes.

摘要

背景

铁死亡是由膜磷脂的铁依赖性过氧化引起的，软骨细胞铁死亡导致骨关节炎（OA）进展。谷胱甘肽过氧化物酶 4（GPX4）在阻断铁死亡中起主要作用。N6-甲基腺苷（m6A）是 mRNA 转录后修饰中的一种表观遗传修饰。本研究通过 m6A 修饰研究了甲基转移酶样 14（METTL14），即 m6A 甲基转移酶的关键组成部分，对软骨细胞铁死亡的影响。

方法

通过向右侧膝关节内注射单碘乙酸钠建立 OA 大鼠模型。使用 OA 大鼠模型中的软骨进行基因表达分析。原代小鼠软骨细胞或 ADTC5 细胞用 IL-1β或 erastin 刺激。使用 m6A RNA 甲基化定量试剂盒测量 m6A 水平。通过 Western blot、荧光免疫染色、碘化丙啶染色和市售试剂盒研究 METTL14 和 GPX4 对 ECM 降解和铁死亡的影响。通过 MeRIP-qPCR 测定探索 METTL14 作用的机制。

结果

METTL14 和 m6A 在骨关节炎软骨和 IL-1β 诱导的软骨细胞中表达上调。METTL14 耗竭抑制了小鼠软骨细胞中 IL-1β 或 erastin 刺激的 ECM 降解和铁死亡。METTL14 通过 m6A 甲基化修饰抑制 GPX4 基因。GPX4 敲低逆转了 si-METTL14 在 IL-1β 诱导的软骨细胞中介导的保护作用。