Effects of mild hypothermia on neurological function in rats with spinal cord injury based on adenosine monophosphate activated protein kinase/Nod-like receptor protein 3 pathway_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

 PAN Jianzhu ¹ , YANG Zhong ²

1. Trauma Center, Tianjin Fifth Central Hospital, Tianjin, 300450, P. R. China;
2. Department of Orthopedics, Tianjin Fifth Central Hospital, Tianjin, 300450, P. R. China;

Corresponding?author：

PAN Jianzhu, Email: mt6jsp@163.com

Keywords：

Spinal cord injury; mild hypothermia; adenosine monophosphate activated protein kinase; Nod-like receptor protein 3; neurological function

DOI：

10.7507/1002-1892.202507019

Video：

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Objective To investigate the effect of mild hypothermia on neurological function in rats with spinal cord injury (SCI) based on the adenosine monophosphate activated protein kinase (AMPK)/Nod-like receptor protein 3 (NLRP3) pathway. Methods The rat model of SCI was established by Allen’s method. The 7-8 weeks old SPF male SD rats were randomly divided into SCI group, mild hypothermia group (SCI+mild hypothermia treatment), Compound C group (SCI+mild hypothermia treatment+20 mg/kg AMPK/NLRP3 pathway inhibitor Compound C), and normal rats with laminectomy as sham-operation group. Basso, Beattie, and Bresnahan (BBB) score was used to evaluate the motor ability of rats at 1, 3, 7, 14 days after treatment. After 14 days, the rats were sacrificed, and the spinal cord histopathological morphology was observed by HE staining, the neuronal apoptosis in spinal cord tissue was detected by TUNEL assay, and the serum levels of interleukin 2 (IL-2), IL-6, transforming growth factor β₁ (TGF-β₁), malondialdehyde (MDA), and superoxide dismutase (SOD) were detected by ELISA. The expression of AMPK/NLRP3 pathway protein, including B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), and cleaved Caspase-9 were detected by Western blot. Results At 1 day after treatment, the rats in SCI group, mild hypothermia group, and Compound C group did not recover their motor ability; at 3, 7, and 14 days, the BBB score of SCI group was significantly lower than that of SCI group (P<0.05), the BBB score of mild hypothermia group was significantly higher than that of SCI group (P<0.05), and the BBB score of Compound C group was significantly lower than that of mild hypothermia group (P<0.05). Compared with the sham-operation group, the SCI group displayed obvious pathological changes in the spinal cord tissue, with disordered tissue architecture, inflammatory infiltration, and blurred interstitial boundaries. The neuronal apoptosis rate, Bax/Bcl-2 ratio, cleaved Caspase-9 expression, NLRP3 protein expression, serum IL-2, IL-6, and MDA levels were elevated, whereas serum TGF-β₁, SOD levels, and spinal cord phosphorylation AMPK/AMPK protein expression significantly decreased (P<0.05). Compared with the SCI group, the above phenomena significantly improved in the mild hypothermia group (P<0.05). Conclusion Subnormal hypothermia can attenuate neurological dysfunction after spinal cord injury in rats, potentially by activating the AMPK/NLRP3 signaling pathway.

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2.	Liu W, Luo F, Wu H, et al. Noggin protein can induce the differentiation of rat bone marrow mesenchymal stem cells to neurons and repair spinal cord injury. Discov Med, 2023, 35(179): 956-964.
3.	Hu X, Xu W, Ren Y, et al. Spinal cord injury: molecular mechanisms and therapeutic interventions. Signal Transduct Target Ther, 2023, 8(1): 245. doi: 10.1038/s41392-023-01477-6.
4.	李均, 徐藝, 冉陽, 等. 漢黃芩素調節AMPK/NLRP3信號通路對H/R誘導的心肌細胞凋亡的影響. 現代生物醫學進展, 2024, 24(17): 3211-3216,3225.
5.	Ye JX, Wang M, Wang RY, et al. Hydroxysafflor yellow A inhibits hypoxia/reoxygenation-induced cardiomyocyte injury via regulating the AMPK/NLRP3 inflammasome pathway. Int Immunopharmacol, 2020, 82: 106316. doi: 10.1016/j.intimp.2020.106316.
6.	Wang J, Wang T, Zhang H, et al. Effect of mild hypothermia vs normothermia cardiopulmonary bypass on postoperative bleeding in patients undergoing coronary artery bypass grafting: protocol of a multi-center, randomized, controlled trial. BMC Surg, 2024, 24(1): 323. doi: 10.1186/s12893-024-02634-6.
7.	Fu D, Chen C, He L, et al. Protective effect of mild hypothermia on spinal cord ischemia-induced delayed paralysis and spinal cord injury. Neurochem Res, 2022, 47(5): 1212-1225.
8.	梅繼林, 楊志新, 李曉寧. 夾脊電針對急性脊髓損傷大鼠脊髓組織NOD樣受體熱蛋白結構域相關蛋白3介導的細胞焦亡的影響. 針刺研究, 2024, 49(2): 110-118.
9.	鄧宏哲, 陳昆, 李鵬, 等. 人參皂苷Rg1通過調控AMPK/NLRP3通路介導的細胞焦亡抑制大鼠肺纖維化研究. 中草藥, 2023, 54(3): 841-848.
10.	王力航, 姚書眈, 湯倩, 等. 亞低溫對SCI大鼠神經細胞凋亡、線粒體呼吸及TNF-α水平的影響. 中國老年學雜志, 2023, 43(9): 2220-2224.
11.	田珂, 趙瑞, 張昆龍, 等. 前扣帶回皮質參與小鼠脊髓損傷后疼痛和認知障礙共病. 神經解剖學雜志, 2025, 41(1): 9-17.
12.	朱強, 胡國良, 李全春, 等. 鼠神經生長因子、亞低溫聯合甲強龍在急性脊髓損傷中的應用及對氧化應激水平的影響. 中國現代醫學雜志, 2021, 31(23): 79-83.
13.	Liu X, Zhang Y, Wang Y, et al. Inflammatory response to spinal cord injury and its treatment. World Neurosurg, 2021, 155: 19-31.
14.	Chalif JI, Chavarro VS, Mensah E, et al. Epidural spinal cord stimulation for spinal cord injury in humans: A systematic review. J Clin Med, 2024, 13(4): 1090. doi: 10.3390/jcm13041090.
15.	Xiao Q, Liu Y, Zhang X, et al. Mild hypothermia ameliorates hepatic ischemia reperfusion injury by inducing RBM3 expression. Apoptosis, 2022, 27(11-12): 899-912.
16.	Ma Y, He Z, Wang J, et al. Mild hypothermia promotes neuronal differentiation of human neural stem cells via RBM3-SOX11 signaling pathway. iScience, 2024, 27(4): 109435. doi: 10.1016/j.isci.2024.109435.
17.	Yao S, Wang L, Chen Q, et al. The effect of mild hypothermia plus rutin on the treatment of spinal cord injury and inflammatory factors by repressing TGF-β/smad pathway. Acta Cir Bras, 2021, 36(3): e360307. doi: 10.1590/ACB360307.
18.	Chen MH, Fu QM. The roles of AMPK in revascularization. Cardiol Res Pract, 2020, 2020: 4028635. doi: 10.1155/2020/4028635.
19.	Zhang J, Huang L, Shi X, et al. Metformin protects against myocardial ischemia-reperfusion injury and cell pyroptosis via AMPK/NLRP3 inflammasome pathway. Aging (Albany NY), 2020, 12(23): 24270-24287.
20.	Zhou Z, Liu Z, Zhang C, et al. Mild hypothermia alleviates early brain injury after subarachnoid hemorrhage via suppressing pyroptosis through AMPK/NLRP3 inflammasome pathway in rats. Brain Res Bull, 2023, 193: 72-83.
21.	Guo R, Gao S, Feng Y, et al. Ulinastatin attenuates spinal cord injury by targeting AMPK/NLRP3 signaling pathway. J Chem Neuroanat, 2022, 125: 102145. doi: 10.1016/j.jchemneu.2022.102145.
22.	Cheng Z, Tu J, Wang K, et al. Wogonin alleviates NLRP3 inflammasome activation after cerebral ischemia-reperfusion injury by regulating AMPK/SIRT1. Brain Res Bull, 2024, 207: 110886. doi: 10.1016/j.brainresbull.2024.110886.
23.	Luo X, Bao X, Weng X, et al. The protective effect of quercetin on macrophage pyroptosis via TLR2/Myd88/NF-κB and ROS/AMPK pathway. Life Sci, 2022, 291: 120064. doi: 10.1016/j.lfs.2021.120064.
24.	He LL, Wang YC, Ai YT, et al. Qiangji decoction alleviates neurodegenerative changes and hippocampal neuron apoptosis induced by D-galactose via regulating AMPK/SIRT1/NF-κB signaling pathway. Front Pharmacol, 2021, 12: 735812. doi: 10.3389/fphar.2021.735812.
25.	Song H, Ding Z, Chen J, et al. The AMPK-SIRT1-FoxO1-NF-κB signaling pathway participates in hesperetin-mediated neuroprotective effects against traumatic brain injury via the NLRP3 inflammasome. Immunopharmacol Immunotoxicol, 2022, 44(6): 970-983.

1. Husain MA, Smith R, Sorge RE, et al. Inhibition of the RNA regulator HuR mitigates spinal cord injury by potently suppressing post-injury neuroinflammation. FASEB J, 2025, 39(9): e70588. doi: 10.1096/fj.202500236R.
2. Liu W, Luo F, Wu H, et al. Noggin protein can induce the differentiation of rat bone marrow mesenchymal stem cells to neurons and repair spinal cord injury. Discov Med, 2023, 35(179): 956-964.
3. Hu X, Xu W, Ren Y, et al. Spinal cord injury: molecular mechanisms and therapeutic interventions. Signal Transduct Target Ther, 2023, 8(1): 245. doi: 10.1038/s41392-023-01477-6.
4. 李均, 徐藝, 冉陽, 等. 漢黃芩素調節AMPK/NLRP3信號通路對H/R誘導的心肌細胞凋亡的影響. 現代生物醫學進展, 2024, 24(17): 3211-3216,3225.
5. Ye JX, Wang M, Wang RY, et al. Hydroxysafflor yellow A inhibits hypoxia/reoxygenation-induced cardiomyocyte injury via regulating the AMPK/NLRP3 inflammasome pathway. Int Immunopharmacol, 2020, 82: 106316. doi: 10.1016/j.intimp.2020.106316.
6. Wang J, Wang T, Zhang H, et al. Effect of mild hypothermia vs normothermia cardiopulmonary bypass on postoperative bleeding in patients undergoing coronary artery bypass grafting: protocol of a multi-center, randomized, controlled trial. BMC Surg, 2024, 24(1): 323. doi: 10.1186/s12893-024-02634-6.
7. Fu D, Chen C, He L, et al. Protective effect of mild hypothermia on spinal cord ischemia-induced delayed paralysis and spinal cord injury. Neurochem Res, 2022, 47(5): 1212-1225.
8. 梅繼林, 楊志新, 李曉寧. 夾脊電針對急性脊髓損傷大鼠脊髓組織NOD樣受體熱蛋白結構域相關蛋白3介導的細胞焦亡的影響. 針刺研究, 2024, 49(2): 110-118.
9. 鄧宏哲, 陳昆, 李鵬, 等. 人參皂苷Rg1通過調控AMPK/NLRP3通路介導的細胞焦亡抑制大鼠肺纖維化研究. 中草藥, 2023, 54(3): 841-848.
10. 王力航, 姚書眈, 湯倩, 等. 亞低溫對SCI大鼠神經細胞凋亡、線粒體呼吸及TNF-α水平的影響. 中國老年學雜志, 2023, 43(9): 2220-2224.
11. 田珂, 趙瑞, 張昆龍, 等. 前扣帶回皮質參與小鼠脊髓損傷后疼痛和認知障礙共病. 神經解剖學雜志, 2025, 41(1): 9-17.
12. 朱強, 胡國良, 李全春, 等. 鼠神經生長因子、亞低溫聯合甲強龍在急性脊髓損傷中的應用及對氧化應激水平的影響. 中國現代醫學雜志, 2021, 31(23): 79-83.
13. Liu X, Zhang Y, Wang Y, et al. Inflammatory response to spinal cord injury and its treatment. World Neurosurg, 2021, 155: 19-31.
14. Chalif JI, Chavarro VS, Mensah E, et al. Epidural spinal cord stimulation for spinal cord injury in humans: A systematic review. J Clin Med, 2024, 13(4): 1090. doi: 10.3390/jcm13041090.
15. Xiao Q, Liu Y, Zhang X, et al. Mild hypothermia ameliorates hepatic ischemia reperfusion injury by inducing RBM3 expression. Apoptosis, 2022, 27(11-12): 899-912.
16. Ma Y, He Z, Wang J, et al. Mild hypothermia promotes neuronal differentiation of human neural stem cells via RBM3-SOX11 signaling pathway. iScience, 2024, 27(4): 109435. doi: 10.1016/j.isci.2024.109435.
17. Yao S, Wang L, Chen Q, et al. The effect of mild hypothermia plus rutin on the treatment of spinal cord injury and inflammatory factors by repressing TGF-β/smad pathway. Acta Cir Bras, 2021, 36(3): e360307. doi: 10.1590/ACB360307.
18. Chen MH, Fu QM. The roles of AMPK in revascularization. Cardiol Res Pract, 2020, 2020: 4028635. doi: 10.1155/2020/4028635.
19. Zhang J, Huang L, Shi X, et al. Metformin protects against myocardial ischemia-reperfusion injury and cell pyroptosis via AMPK/NLRP3 inflammasome pathway. Aging (Albany NY), 2020, 12(23): 24270-24287.
20. Zhou Z, Liu Z, Zhang C, et al. Mild hypothermia alleviates early brain injury after subarachnoid hemorrhage via suppressing pyroptosis through AMPK/NLRP3 inflammasome pathway in rats. Brain Res Bull, 2023, 193: 72-83.
21. Guo R, Gao S, Feng Y, et al. Ulinastatin attenuates spinal cord injury by targeting AMPK/NLRP3 signaling pathway. J Chem Neuroanat, 2022, 125: 102145. doi: 10.1016/j.jchemneu.2022.102145.
22. Cheng Z, Tu J, Wang K, et al. Wogonin alleviates NLRP3 inflammasome activation after cerebral ischemia-reperfusion injury by regulating AMPK/SIRT1. Brain Res Bull, 2024, 207: 110886. doi: 10.1016/j.brainresbull.2024.110886.
23. Luo X, Bao X, Weng X, et al. The protective effect of quercetin on macrophage pyroptosis via TLR2/Myd88/NF-κB and ROS/AMPK pathway. Life Sci, 2022, 291: 120064. doi: 10.1016/j.lfs.2021.120064.
24. He LL, Wang YC, Ai YT, et al. Qiangji decoction alleviates neurodegenerative changes and hippocampal neuron apoptosis induced by D-galactose via regulating AMPK/SIRT1/NF-κB signaling pathway. Front Pharmacol, 2021, 12: 735812. doi: 10.3389/fphar.2021.735812.
25. Song H, Ding Z, Chen J, et al. The AMPK-SIRT1-FoxO1-NF-κB signaling pathway participates in hesperetin-mediated neuroprotective effects against traumatic brain injury via the NLRP3 inflammasome. Immunopharmacol Immunotoxicol, 2022, 44(6): 970-983.

Chinese Journal of Reparative and Reconstructive Surgery

Latest ArticlesEffects of mild hypothermia on neurological function in rats with spinal cord injury based on adenosine monophosphate activated protein kinase/Nod-like receptor protein 3 pathway

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