Liu Chang, Qiu Siyu, Li Wen, Zhao Liping, He Yingzi, Zeng Xiangli
Department of Otolaryngology-Head and Neck Surgery, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, Guangdong 510630, China.
ENT institute and Department of Otorhinolaryngology, Eye & ENT Hospital, Fudan University, Shanghai 200031, China.
Chin Med J (Engl). 2025 Jul 10. doi: 10.1097/CM9.0000000000003633.
Sensorineural hearing loss is characterized by irreversible hair cell (HC) degeneration. Ferroptosis, which is marked by the accumulation of reactive oxygen species and elevated levels of lipid peroxidation products, has been shown to contribute to drug-mediated auditory impairment. This study aimed to elucidate the role of mixed-lineage leukemia 1 (MLL1) in HC survival in the auditory system.
The HEI-OC1 auditory cell line and postnatal cochlear explants were evaluated using MM-102, a specific MLL1 histone methyltransferase inhibitor. Western blot, quantitative polymerase chain reaction, electron microscopy, and immunofluorescence were used to elucidate the role of MLL1 in regulating ferroptosis in HC injury. RNA sequencing (RNA-seq) was used to analyze the molecular mechanisms of MLL1 intervention in HC injury from an epigenetic perspective.
Our findings demonstrated that immunofluorescence staining revealed crucial role of MM-102 in promoting intracellular accumulation of lipid peroxides and ferrous ions. Subsequent analysis showed MLL1 downregulation-induced mitochondrial dysfunction and endoplasmic reticulum (ER) stress, with transmission electron microscopy imaging confirming ultrastructural alterations in mitochondria and ER. Mechanistic investigations identified the PERK-eIF2α-Atf4-Chop signaling axis as the regulatory pathway, evidenced by Western blot quantification of phosphorylated PERK (p-PERK), Atf4, and Chop levels. RNA-seq analysis revealed 741 differentially expressed genes (335 upregulated, and 406 downregulated). KEGG pathway analysis specifically highlighted significant enrichment of the PI3K/Akt-Lrp1 pathway, with corresponding activation patterns of phospho (p)-Akt and Lrp1 confirmed through Western blot analysis.
MLL1 downregulation initiates ferroptosis in cochlear HCs. This process is intrinsically associated with the activation of mitochondrial dysfunction and ER stress. The study highlights the importance of MLL1 in HC survival, suggesting its potential as a therapeutic target for treating hearing loss.
感音神经性听力损失的特征是毛细胞（HC）发生不可逆的退化。铁死亡以活性氧的积累和脂质过氧化产物水平升高为特征，已被证明与药物介导的听觉损伤有关。本研究旨在阐明混合谱系白血病1（MLL1）在听觉系统中毛细胞存活中的作用。
使用特异性MLL1组蛋白甲基转移酶抑制剂MM-102对HEI-OC1听觉细胞系和出生后耳蜗外植体进行评估。采用蛋白质免疫印迹法、定量聚合酶链反应、电子显微镜和免疫荧光法来阐明MLL1在调节毛细胞损伤中的铁死亡作用。RNA测序（RNA-seq）用于从表观遗传学角度分析MLL1干预毛细胞损伤的分子机制。
我们的研究结果表明，免疫荧光染色显示MM-102在促进细胞内脂质过氧化物和亚铁离子积累方面起关键作用。随后的分析表明，MLL1下调会导致线粒体功能障碍和内质网（ER）应激，透射电子显微镜成像证实了线粒体和内质网的超微结构改变。机制研究确定PERK-eIF2α-Atf4-Chop信号轴为调节途径，蛋白质免疫印迹法定量磷酸化PERK（p-PERK）、Atf4和Chop水平证明了这一点。RNA-seq分析揭示了741个差异表达基因（335个上调，406个下调）。KEGG通路分析特别突出了PI3K/Akt-Lrp1通路的显著富集，通过蛋白质免疫印迹分析证实了磷酸化（p）-Akt和Lrp1的相应激活模式。
MLL1下调会引发耳蜗毛细胞的铁死亡。这一过程与线粒体功能障碍和内质网应激的激活内在相关。该研究突出了MLL1在毛细胞存活中的重要性，表明其作为治疗听力损失的治疗靶点的潜力。
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