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Theranostics 2024; 14(7):2687-2705. doi:10.7150/thno.94072 This issue Cite

Research Paper

Abnormal mitochondrial iron metabolism damages alveolar type II epithelial cells involved in bleomycin-induced pulmonary fibrosis

Min Shao¹, Haipeng Cheng², Xiaohong Li², Yujia Qiu¹, Yunna Zhang¹, Yanfen Chang¹, Jiafeng Fu¹, Mengxia Shen¹, Xinxin Xu¹, Dandan Feng¹, Yang Han¹, ShaoJie Yue³, Yan Zhou^1✉, Ziqiang Luo^1,4✉

1. Department of Physiology, Xiangya School of Medicine, Central South University, Changsha, Hunan 410078, China.
2. Department of Pathology, The Second Xiangya Hospital, Central South University, Changsha, Hunan, 410000, China.
3. Department of Pediatrics, Xiangya Hospital, Central South University, Changsha, Hunan, 410013, China.
4. Hunan Key Laboratory of Organ Fibrosis, Changsha, Hunan, 410013, China.

✉ Corresponding authors: Dr. Yan Zhou or Prof. Ziqiang Luo, Department of Physiology, Xiangya Medical School, Central South University, Changsha, Hunan 410078, China. Tel.: +86-731-82355051; E-mail: zhouyanxyedu.cn or luoziqiangedu.cn.

Abstract

Rationale: Pulmonary fibrosis is a chronic progressive lung disease with limited therapeutic options. We previously revealed that there is iron deposition in alveolar epithelial type II cell (AECII) in pulmonary fibrosis, which can be prevented by the iron chelator deferoxamine. However, iron in the cytoplasm and the mitochondria has two relatively independent roles and regulatory systems. In this study, we aimed to investigate the role of mitochondrial iron deposition in AECII injury and pulmonary fibrosis, and to find potential therapeutic strategies.

Methods: BLM-treated mice, MLE-12 cells, and primary AECII were employed to establish the mouse pulmonary fibrosis model and epithelial cells injury model, respectively. Mitochondrial transplantation, siRNA and plasmid transfection, western blotting (WB), quantitative real-time polymerase chain reaction (RT-qPCR), polymerase chain reaction (PCR), immunofluorescence, immunoprecipitation (IP), MitoSOX staining, JC-1 staining, oxygen consumption rate (OCR) measurement, and Cell Counting Kit-8 (CCK8) assay were utilized to elucidate the role of mitochondrial iron deposition in cell and lung fibrosis and determine its mechanism.

Results: This study showed that prominent mitochondrial iron deposition occurs within AECII in bleomycin (BLM)-induced pulmonary fibrosis mouse model and in BLM-treated MLE-12 epithelial cells. Further, the study revealed that healthy mitochondria rescue BLM-damaged AECII mitochondrial iron deposition and cell damage loss. Mitoferrin-2 (MFRN2) is the main transporter that regulates mitochondrial iron metabolism by transferring cytosolic iron into mitochondria, which is upregulated in BLM-treated MLE-12 epithelial cells. Direct overexpression of MFRN2 causes mitochondrial iron deposition and cell damage. In this study, decreased ubiquitination of the ubiquitin ligase F-box/LRR-repeat protein 5 (FBXL5) degraded iron-reactive element-binding protein 2 (IREB2) and promoted MFRN2 expression as well as mitochondrial iron deposition in damaged AECII. Activation of the prostaglandin E2 receptor EP4 subtype (EP4) receptor signaling pathway counteracted mitochondrial iron deposition by downregulating IREB2-MFRN2 signaling through upregulation of FBXL5. This intervention not only reduced mitochondrial iron content but also preserved mitochondrial function and protected against AECII damage after BLM treatment.

Conclusion: Our findings highlight the unexplored roles, mechanisms, and regulatory approaches of abnormal mitochondrial iron metabolism of AECII in pulmonary fibrosis. Therefore, this study deepens the understanding of the mechanisms underlying pulmonary fibrosis and offers a promising strategy for developing effective therapeutic interventions using the EP4 receptor activator.

Keywords: pulmonary fibrosis, type II alveolar epithelial cells, mitochondrial iron deposition, mitoferrin-2, F-box/LRR-repeat protein 5

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