Mechanisms of Piezo1-mediated microglial ferroptosis in inhibiting spinal cord injury repair_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

WANG Zesen ^1,2 , HUANG Jiahui ^1,2 , LIANG Bintao ^1,2 , ZHOU Shishuai ^1,2 ,  GUAN Jianzhong ^1,2

1. Department of Orthopaedics, the First Affiliated Hospital of Bengbu Medical College, Bengbu Anhui, 233000, P. R. China;
2. Key Laboratory of Anhui Province for Tissue Transplantation, Bengbu Anhui, 233000, P. R. China;

Corresponding?author：

GUAN Jianzhong, Email: Guanjianzhong@bbmc.edu.cn

Keywords：

Spinal cord injury; Piezo1; microglia; ferroptosis; neuroinflammation; mice

DOI：

10.7507/1002-1892.202602018

Video：

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Objective To investigate the mechanism of the mechanosensitive ion channel Piezo1 in microglial ferroptosis following spinal cord injury (SCI), and to assess the effects of Piezo1 inhibition on ameliorating the injury microenvironment and promoting neurological functional recovery. Methods Primary microglia cells were extracted from neonatal 1-2 days C57BL/6 mice and divided into control group, Yoda1 (Piezo1 agonist) group, and Yoda1+GsMTx4 (Piezo1 inhibitor) group. Live/dead cell staining, reactive oxygen species (ROS) fluorescence staining, 5, 5’, 6, 6’-tetrachloro-1, 1’, 3, 3’-tetraethylbenzimidazolylcarbocyanine iodide (JC-1) mitochondrial membrane potential detection, and transmission electron microscopy were utilized to assess microglial ferroptosis and mitochondrial functional characteristics. SPF female C57BL/6 mice aged 6 to 8 weeks were used to detect the expression of Piezo1 at different time points after SCI by Western blot, and the two time points with no significant change and the most significant change in Piezo1 expression after SCI were selected for subsequent experiments. T₈, T₉ SCI models were established by modified Allen’s method, and were divided into sham operation group, injury group, and injury+shPiezo1 group (Piezo1-targeted interfering virus AAV-shPiezo1 was injected in situ to knock down the expression of Piezo1 14 days before modeling). Colocalization of Piezo1 with microglial markers purinergic receptor P2Y12 (P2ry12), and the expressions of glutathione peroxidase 4 (GPX4) and acyl coenzyme A synthetase long chain member 4 (ACSL4) were observed by immunofluorescence staining. Basso Mouse Scale (BMS) score was used to assess hindlimb motor function in mice. The level of ROS was detected by dihydroethidium (DHE) staining; the content of malondialdehyde (MDA) was detected by MDA kit; the levels of tumor necrosis factor α (TNF-α) and interleukin 10 (IL-10) were detected by ELISA assay; the pathological morphology of spinal cord was observed by HE staining. Results In vitro experiments showed that compared with the control group, the Yoda1 group had typical ultrastructural changes of ferroptosis, such as increased microglial cell death, enhanced ROS fluorescence, mitochondrial membrane potential depolarization, mitochondrial shrinkage and mitochondrial cristae breakage (all P<0.05), while the GsMTx4 group could partially reverse the above effects (P<0.05). In vivo experiments demonstrated that the expression of Piezo1 in spinal cord tissue was up-regulated sequentially after SCI, and reached the peak on the 7th day after SCI (P<0.05), and it was mainly localized in P2ry12-positive microglia. Compared with the injury group, in the injury+shPiezo1 group, the expression of ferroptosis core protein GPX4 in microglia was increased, the expression of ACSL4 was decreased, the levels of ROS and MDA in spinal cord tissue were decreased (P<0.05), the level of pro-inflammatory factor TNF-α was decreased, and the level of anti-inflammatory factor IL-10 was increased (P<0.05). In addition, the BMS score was significantly higher than that of the injury group (P<0.05) from the 14th day after operation, and the spinal cord tissue structure was relatively well preserved, and the cavity area was reduced. Conclusion SCI activates the Piezo1 channel in microglia, triggering mitochondrial dysfunction and mediating cellular ferroptosis, thereby aggravating secondary neuroinflammation. Targeted inhibition of Piezo1 effectively blocks the ferroptosis process, ameliorates the immune microenvironment, and promotes tissue repair and locomotor functional recovery after SCI.

Citation： WANG Zesen, HUANG Jiahui, LIANG Bintao, ZHOU Shishuai, GUAN Jianzhong. Mechanisms of Piezo1-mediated microglial ferroptosis in inhibiting spinal cord injury repair. Chinese Journal of Reparative and Reconstructive Surgery, 2026, 40(7): 1099-1109. doi: 10.7507/1002-1892.202602018 Copy

1.	江琪, 戚超, 孫月榮, 等. 清除脊髓中小膠質細胞對小鼠脊髓損傷后神經修復影響的實驗研究. 中國修復重建外科雜志, 2025, 39(6): 754-761.
2.	Morrison D, Pinpin C, Lee A, et al. Acute immunological phenotypes in individuals with traumatic spinal cord injury. J Neurotrauma, 2025, 42(23-24): 2149-2162.
3.	Brockie S, Zhou C, Fehlings MG. Resident immune responses to spinal cord injury: role of astrocytes and microglia. Neural Regen Res, 2024, 19(8): 1678-1685.
4.	Xu Y, Geng Y, Wang H, et al. Cyclic helix B peptide alleviates proinflammatory cell death and improves functional recovery after traumatic spinal cord injury. Redox Biol, 2023, 64: 102767. doi: 10.1016/j.redox.2023.102767.
5.	Zhang H, Ni W, Yu G, et al. 3, 4-Dimethoxychalcone, a caloric restriction mimetic, enhances TFEB-mediated autophagy and alleviates pyroptosis and necroptosis after spinal cord injury. Theranostics, 2023, 13(2): 810-832.
6.	Feng X, Chen X, Zaeem M, et al. Sesamol attenuates neuroinflammation by regulating the AMPK/SIRT1/NF-κB signaling pathway after spinal cord injury in mice. Oxid Med Cell Longev, 2022, 2022: 8010670. doi: 10.1155/2022/8010670.
7.	Li Q, Song Q, Pei H, et al. Emerging mechanisms of ferroptosis and its implications in lung cancer. Chin Med J (Engl), 2024, 137(7): 818-829.
8.	Yi Y, Jia P, Xie P, et al. Beyond oxidative stress: Ferroptosis as a novel orchestrator in neurodegenerative disorders. Front Immunol, 2025, 16: 1683876. doi: 10.3389/fimmu.2025.1683876.
9.	Ryan F, Blex C, Ngo TD, et al. Ferroptosis inhibitor improves outcome after early and delayed treatment in mild spinal cord injury. Acta Neuropathol, 2024, 147(1): 106. doi: 10.1007/s00401-024-02758-2.
10.	Yao S, Pang M, Wang Y, et al. Mesenchymal stem cell attenuates spinal cord injury by inhibiting mitochondrial quality control-associated neuronal ferroptosis. Redox Biol, 2023, 67: 102871. doi: 10.1016/j.redox.2023.102871.
11.	Zheng Z, Couture D, Adams F, et al. Attenuated tissue damage with mechanical tissue resuscitation in a pig model of spinal cord injury. J Neurotrauma, 2024, 41(9-10): 1020-1029.
12.	Du Y, Xu B, Li Q, et al. The role of mechanically sensitive ion channel Piezo1 in bone remodeling. Front Bioeng Biotechnol, 2024, 12: 1342149. doi: 10.3389/fbioe.2024.1342149.
13.	Yin Q, Zang G, Li N, et al. Agonist-induced Piezo1 activation promote mitochondrial-dependent apoptosis in vascular smooth muscle cells. BMC Cardiovasc Disord, 2022, 22(1): 287. doi: 10.1186/s12872-022-02726-2.
14.	Xu H, Chen X, Luo S, et al. Cardiomyocyte-specific Piezo1 deficiency mitigates ischemia-reperfusion injury by preserving mitochondrial homeostasis. Redox Biol, 2025, 79: 103471. doi: 10.1016/j.redox.2024.103471.
15.	Fan Y, Chen Z, Pathak JL, et al. Differential regulation of adhesion and phagocytosis of resting and activated microglia by dopamine. Front Cell Neurosci, 2018, 12: 309. doi: 10.3389/fncel.2018.00309.
16.	Fu P, Tang R, Yu Z, et al. Rho-associated kinase inhibitors promote microglial uptake via the ERK signaling pathway. Neurosci Bull, 2016, 32(1): 83-91.
17.	Xiao CL, Lai HT, Zhou JJ, et al. Nrf2 signaling pathway: Focus on oxidative stress in spinal cord injury. Mol Neurobiol, 2025, 62(2): 2230-2249.
18.	Li C, Wu Z, Zhou L, et al. Temporal and spatial cellular and molecular pathological alterations with single-cell resolution in the adult spinal cord after injury. Signal Transduct Target Ther, 2022, 7(1): 65. doi: 10.1038/s41392-022-00885-4.
19.	Timofeeva AV, Akhmetzyanova ER, Rizvanov AA, et al. Interaction of microglia with the microenvironment in spinal cord injury. Neuroscience, 2025, 565: 594-603.
20.	Xu L, Wang J, Ding Y, et al. Current Knowledge of Microglia in Traumatic Spinal Cord Injury. Front Neurol, 2022, 12: 796704. doi: 10.3389/fneur.2021.796704.
21.	Cai L, Fan Q, Pang R, et al. Microglia programmed cell death in neurodegenerative diseases and CNS injury. Apoptosis, 2025, 30(1-2): 446-465.
22.	Zheng Q, Wang D, Lin R, et al. Pyroptosis, ferroptosis, and autophagy in spinal cord injury: regulatory mechanisms and therapeutic targets. Neural Regen Res, 2025, 20(10): 2787-2806.
23.	Yu B, Zhang Y, Yang Y, et al. Neonatal-inspired reprogramming of microglial pan-programmed cell death enhances regeneration in adult spinal cord injury. Research (Wash D C), 2025, 8: 0759. doi: 10.34133/research.0759.
24.	Hayta E, Elden H. Acute spinal cord injury: A review of pathophysiology and potential of non-steroidal anti-inflammatory drugs for pharmacological intervention. J Chem Neuroanat, 2018, 87: 25-31.
25.	Jin C, Zhu R, Wang ZW, et al. Dynamic changes in mechanical properties of the adult rat spinal cord after injury. Acta Biomater, 2023, 155: 436-448.
26.	Ikiz ED, Hascup ER, Bae C, et al. Microglial Piezo1 mechanosensitive channel as a therapeutic target in Alzheimer’s disease. Front Cell Neurosci, 2024, 18: 1423410. doi: 10.3389/fncel.2024.1423410.
27.	Seblani M, Decherchi P, Brezun JM. Edema after CNS trauma: A focus on spinal cord injury. Int J Mol Sci, 2023, 24(8): 7159. doi: 10.3390/ijms24087159.
28.	Qi M, Liu R, Zhang F, et al. Roles of mechanosensitive ion channel PIEZO1 in the pathogenesis of brain injury after experimental intracerebral hemorrhage. Neuropharmacology, 2024, 251: 109896. doi: 10.1016/j.neuropharm.2024.109896.
29.	Xie D, Ma Y, Gao C, et al. Piezo1 activation on microglial cells exacerbates demyelination in sepsis by influencing the CCL25/GRP78 pathway. Int Immunopharmacol, 2024, 142(Pt A): 113045. doi: 10.1016/j.intimp.2024.113045.
30.	Dixon SJ, Olzmann JA. The cell biology of ferroptosis. Nat Rev Mol Cell Biol, 2024, 25(6): 424-442.
31.	Feng Z, Min L, Chen H, et al. Iron overload in the motor cortex induces neuronal ferroptosis following spinal cord injury. Redox Biol, 2021, 43: 101984. doi: 10.1016/j.redox.2021.101984.
32.	Michelucci A, Catacuzzeno L. Piezo1, the new actor in cell volume regulation. Pflugers Arch, 2024, 476(7): 1023-1039.
33.	Carraro M, Bernardi P. The mitochondrial permeability transition pore in Ca²⁺ homeostasis. Cell Calcium, 2023, 111: 102719. doi: 10.1016/j.ceca.2023.102719.
34.	Li L, Cao Y, Zhang X, et al. Injectable ROS homeostasis protective hydrogel inhibiting microglial ferroptosis through the Nrf2/Slc7a11/Gpx4 to alleviate neuropathic pain and promote spinal cord injury repair. Redox Biol, 2025, 86: 103816. doi: 10.1016/j.redox.2025.103816.
35.	Chu B, Kon N, Chen D, et al. ALOX12 is required for p53-mediated tumour suppression through a distinct ferroptosis pathway. Nat Cell Biol, 2019, 21(5): 579-591.
36.	Hegdekar N, Sarkar C, Bustos S, et al. Inhibition of autophagy in microglia and macrophages exacerbates innate immune responses and worsens brain injury outcomes. Autophagy, 2023, 19(7): 2026-2044.
37.	Zhu K, Zhu X, Yu J, et al. Effects of HMGB1/RAGE/cathespin B inhibitors on alleviating hippocampal injury by regulating microglial pyroptosis and caspase activation in neonatal hypoxic-ischemic brain damage. J Neurochem, 2023, 167(3): 410-426.
38.	Diao J, Jia Y, Dai E, et al. Ferroptotic therapy in cancer: benefits, side effects, and risks. Mol Cancer, 2024, 23(1): 89. doi: 10.1186/s12943-024-01999-9.
39.	Stockwell BR. Ferroptosis turns 10: Emerging mechanisms, physiological functions, and therapeutic applications. Cell, 2022, 185(14): 2401-2421.
40.	Brooks NAH, Riar I, Klegeris A. Mitochondrial damage-associated molecular patterns: Neuroimmunomodulators in central nervous system pathophysiology. Neural Regen Res, 2026, 21(4): 1322-1338.
41.	Baral H, Kaundal RK. Novel insights into neuroinflammatory mechanisms in traumatic brain injury: Focus on pattern recognition receptors as therapeutic targets. Cytokine Growth Factor Rev, 2025, 83: 18-34.

1. 江琪, 戚超, 孫月榮, 等. 清除脊髓中小膠質細胞對小鼠脊髓損傷后神經修復影響的實驗研究. 中國修復重建外科雜志, 2025, 39(6): 754-761.
2. Morrison D, Pinpin C, Lee A, et al. Acute immunological phenotypes in individuals with traumatic spinal cord injury. J Neurotrauma, 2025, 42(23-24): 2149-2162.
3. Brockie S, Zhou C, Fehlings MG. Resident immune responses to spinal cord injury: role of astrocytes and microglia. Neural Regen Res, 2024, 19(8): 1678-1685.
4. Xu Y, Geng Y, Wang H, et al. Cyclic helix B peptide alleviates proinflammatory cell death and improves functional recovery after traumatic spinal cord injury. Redox Biol, 2023, 64: 102767. doi: 10.1016/j.redox.2023.102767.
5. Zhang H, Ni W, Yu G, et al. 3, 4-Dimethoxychalcone, a caloric restriction mimetic, enhances TFEB-mediated autophagy and alleviates pyroptosis and necroptosis after spinal cord injury. Theranostics, 2023, 13(2): 810-832.
6. Feng X, Chen X, Zaeem M, et al. Sesamol attenuates neuroinflammation by regulating the AMPK/SIRT1/NF-κB signaling pathway after spinal cord injury in mice. Oxid Med Cell Longev, 2022, 2022: 8010670. doi: 10.1155/2022/8010670.
7. Li Q, Song Q, Pei H, et al. Emerging mechanisms of ferroptosis and its implications in lung cancer. Chin Med J (Engl), 2024, 137(7): 818-829.
8. Yi Y, Jia P, Xie P, et al. Beyond oxidative stress: Ferroptosis as a novel orchestrator in neurodegenerative disorders. Front Immunol, 2025, 16: 1683876. doi: 10.3389/fimmu.2025.1683876.
9. Ryan F, Blex C, Ngo TD, et al. Ferroptosis inhibitor improves outcome after early and delayed treatment in mild spinal cord injury. Acta Neuropathol, 2024, 147(1): 106. doi: 10.1007/s00401-024-02758-2.
10. Yao S, Pang M, Wang Y, et al. Mesenchymal stem cell attenuates spinal cord injury by inhibiting mitochondrial quality control-associated neuronal ferroptosis. Redox Biol, 2023, 67: 102871. doi: 10.1016/j.redox.2023.102871.
11. Zheng Z, Couture D, Adams F, et al. Attenuated tissue damage with mechanical tissue resuscitation in a pig model of spinal cord injury. J Neurotrauma, 2024, 41(9-10): 1020-1029.
12. Du Y, Xu B, Li Q, et al. The role of mechanically sensitive ion channel Piezo1 in bone remodeling. Front Bioeng Biotechnol, 2024, 12: 1342149. doi: 10.3389/fbioe.2024.1342149.
13. Yin Q, Zang G, Li N, et al. Agonist-induced Piezo1 activation promote mitochondrial-dependent apoptosis in vascular smooth muscle cells. BMC Cardiovasc Disord, 2022, 22(1): 287. doi: 10.1186/s12872-022-02726-2.
14. Xu H, Chen X, Luo S, et al. Cardiomyocyte-specific Piezo1 deficiency mitigates ischemia-reperfusion injury by preserving mitochondrial homeostasis. Redox Biol, 2025, 79: 103471. doi: 10.1016/j.redox.2024.103471.
15. Fan Y, Chen Z, Pathak JL, et al. Differential regulation of adhesion and phagocytosis of resting and activated microglia by dopamine. Front Cell Neurosci, 2018, 12: 309. doi: 10.3389/fncel.2018.00309.
16. Fu P, Tang R, Yu Z, et al. Rho-associated kinase inhibitors promote microglial uptake via the ERK signaling pathway. Neurosci Bull, 2016, 32(1): 83-91.
17. Xiao CL, Lai HT, Zhou JJ, et al. Nrf2 signaling pathway: Focus on oxidative stress in spinal cord injury. Mol Neurobiol, 2025, 62(2): 2230-2249.
18. Li C, Wu Z, Zhou L, et al. Temporal and spatial cellular and molecular pathological alterations with single-cell resolution in the adult spinal cord after injury. Signal Transduct Target Ther, 2022, 7(1): 65. doi: 10.1038/s41392-022-00885-4.
19. Timofeeva AV, Akhmetzyanova ER, Rizvanov AA, et al. Interaction of microglia with the microenvironment in spinal cord injury. Neuroscience, 2025, 565: 594-603.
20. Xu L, Wang J, Ding Y, et al. Current Knowledge of Microglia in Traumatic Spinal Cord Injury. Front Neurol, 2022, 12: 796704. doi: 10.3389/fneur.2021.796704.
21. Cai L, Fan Q, Pang R, et al. Microglia programmed cell death in neurodegenerative diseases and CNS injury. Apoptosis, 2025, 30(1-2): 446-465.
22. Zheng Q, Wang D, Lin R, et al. Pyroptosis, ferroptosis, and autophagy in spinal cord injury: regulatory mechanisms and therapeutic targets. Neural Regen Res, 2025, 20(10): 2787-2806.
23. Yu B, Zhang Y, Yang Y, et al. Neonatal-inspired reprogramming of microglial pan-programmed cell death enhances regeneration in adult spinal cord injury. Research (Wash D C), 2025, 8: 0759. doi: 10.34133/research.0759.
24. Hayta E, Elden H. Acute spinal cord injury: A review of pathophysiology and potential of non-steroidal anti-inflammatory drugs for pharmacological intervention. J Chem Neuroanat, 2018, 87: 25-31.
25. Jin C, Zhu R, Wang ZW, et al. Dynamic changes in mechanical properties of the adult rat spinal cord after injury. Acta Biomater, 2023, 155: 436-448.
26. Ikiz ED, Hascup ER, Bae C, et al. Microglial Piezo1 mechanosensitive channel as a therapeutic target in Alzheimer’s disease. Front Cell Neurosci, 2024, 18: 1423410. doi: 10.3389/fncel.2024.1423410.
27. Seblani M, Decherchi P, Brezun JM. Edema after CNS trauma: A focus on spinal cord injury. Int J Mol Sci, 2023, 24(8): 7159. doi: 10.3390/ijms24087159.
28. Qi M, Liu R, Zhang F, et al. Roles of mechanosensitive ion channel PIEZO1 in the pathogenesis of brain injury after experimental intracerebral hemorrhage. Neuropharmacology, 2024, 251: 109896. doi: 10.1016/j.neuropharm.2024.109896.
29. Xie D, Ma Y, Gao C, et al. Piezo1 activation on microglial cells exacerbates demyelination in sepsis by influencing the CCL25/GRP78 pathway. Int Immunopharmacol, 2024, 142(Pt A): 113045. doi: 10.1016/j.intimp.2024.113045.
30. Dixon SJ, Olzmann JA. The cell biology of ferroptosis. Nat Rev Mol Cell Biol, 2024, 25(6): 424-442.
31. Feng Z, Min L, Chen H, et al. Iron overload in the motor cortex induces neuronal ferroptosis following spinal cord injury. Redox Biol, 2021, 43: 101984. doi: 10.1016/j.redox.2021.101984.
32. Michelucci A, Catacuzzeno L. Piezo1, the new actor in cell volume regulation. Pflugers Arch, 2024, 476(7): 1023-1039.
33. Carraro M, Bernardi P. The mitochondrial permeability transition pore in Ca²⁺ homeostasis. Cell Calcium, 2023, 111: 102719. doi: 10.1016/j.ceca.2023.102719.
34. Li L, Cao Y, Zhang X, et al. Injectable ROS homeostasis protective hydrogel inhibiting microglial ferroptosis through the Nrf2/Slc7a11/Gpx4 to alleviate neuropathic pain and promote spinal cord injury repair. Redox Biol, 2025, 86: 103816. doi: 10.1016/j.redox.2025.103816.
35. Chu B, Kon N, Chen D, et al. ALOX12 is required for p53-mediated tumour suppression through a distinct ferroptosis pathway. Nat Cell Biol, 2019, 21(5): 579-591.
36. Hegdekar N, Sarkar C, Bustos S, et al. Inhibition of autophagy in microglia and macrophages exacerbates innate immune responses and worsens brain injury outcomes. Autophagy, 2023, 19(7): 2026-2044.
37. Zhu K, Zhu X, Yu J, et al. Effects of HMGB1/RAGE/cathespin B inhibitors on alleviating hippocampal injury by regulating microglial pyroptosis and caspase activation in neonatal hypoxic-ischemic brain damage. J Neurochem, 2023, 167(3): 410-426.
38. Diao J, Jia Y, Dai E, et al. Ferroptotic therapy in cancer: benefits, side effects, and risks. Mol Cancer, 2024, 23(1): 89. doi: 10.1186/s12943-024-01999-9.
39. Stockwell BR. Ferroptosis turns 10: Emerging mechanisms, physiological functions, and therapeutic applications. Cell, 2022, 185(14): 2401-2421.
40. Brooks NAH, Riar I, Klegeris A. Mitochondrial damage-associated molecular patterns: Neuroimmunomodulators in central nervous system pathophysiology. Neural Regen Res, 2026, 21(4): 1322-1338.
41. Baral H, Kaundal RK. Novel insights into neuroinflammatory mechanisms in traumatic brain injury: Focus on pattern recognition receptors as therapeutic targets. Cytokine Growth Factor Rev, 2025, 83: 18-34.

Chinese Journal of Reparative and Reconstructive Surgery

Mechanisms of Piezo1-mediated microglial ferroptosis in inhibiting spinal cord injury repair

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