Experimental study on the repair of spinal cord injury by conducting hydrogel loaded with tetramethylpyrazine sustained-release microparticles_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

JIANG Shengyuan ¹ , DENG Bowen ¹ , LIU Gang ¹ , FAN Xiao ^1,2 , BAI Huizhong ¹ , ZHAO Yi ¹ , XU Lin ¹ ,  MU Xiaohong ¹

1. Department of Orthopedics, Dongzhimen Hospital of Beijing University of Chinese Medicine, Beijing, 100700, P. R. China;
2. Department of Orthopedics, Qingdao Municipal Hospital, Qingdao Shandong, 266000, P. R. China;

Corresponding author：

MU Xiaohong, Email: muxiaohong2006@126.com

Keywords：

Spinal cord injury; tetramethylpyrazine; hydrogels; inflammatory response; nerve repair; rat

DOI：

10.7507/1002-1892.202209013

Video：

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Objective To investigate the neuroprotective effect of conducting hydrogel loaded with tetramethylpyrazine sustained-release microparticles (hereinafter referred to as “TGTP hydrogel”) on spinal cord injury rats. Methods Forty-eight adult female Sprague Dawley rats were randomly divided into 4 groups: sham operation group (group A), model group (group B), conductive hydrogel group (group C), and TGTP hydrogel group (group D), with 12 rats in each group. Only laminectomy was performed in group A, and complete spinal cord transection was performed in groups B, C, and D. Basso-Bettie-Bresnahan (BBB) score was used to evaluate the recovery of hind limb motor function of each group before modeling and at 1, 3, 7, 14, and 28 days after modeling, respectively. At 28 days after modeling, the rats were sacrificed for luxol fast blue (LFB) staining to detect myelin regeneration. Nissl staining was used to detect the survival of neurons. Immunohistochemical staining was used to evaluate the expression of inflammation-related factors [nuclear factor кB (NF-кB), tumor necrosis factor α (TNF-α), and interleukin 10 (IL-10)]. Immunofluorescence staining and Western blot were used to evaluate the expression of neurofilament 200 (NF200). Rseults BBB scores of group A were significantly better than those of the other three groups at all time points after modeling (P<0.05); at 14 and 28 days after modeling, there was no significant difference in BBB scores between groups C and D (P>0.05), but the BBB score of group D was significantly better than that of group B (P<0.05). LFB staining and Nissl staining showed that the structure of neurons and myelin in group A was intact, and the myelin integrity and survival number of neurons in group D were significantly better than those in groups B and C. Immunohistochemical staining showed that the absorbency (A) value of NF-кB and TNF-α in group A were significantly lower than those in groups B and C (P<0.05), the A value of IL-10 was significantly higher than that in the other three groups (P<0.05); the A value of NF-κB in group D was significantly lower than that in groups B and C, the A value of TNF-α in group D was significantly lower than that in group B, while the A value of IL-10 in group D was significantly higher than that in group B (P<0.05). Immunofluorescence staining showed that the structure of neurons and nerve fibers in group A was clear and the fluorescence intensity was high. The fluorescence intensity of NF200 in group D was higher than that in groups B and C, and some nerve fibers could be seen. Western blot analysis showed that the relative expression of NF200 in group A was the highest, and the relative expression of NF200 in group D was significantly higher than that in groups B and C (P<0.05). Conclusion The TGTP hydrogel can effectively promote the recovery of motor function in rats with spinal cord injury, and its mechanism may be related to the regulation of inflammatory response.

Citation： JIANG Shengyuan, DENG Bowen, LIU Gang, FAN Xiao, BAI Huizhong, ZHAO Yi, XU Lin, MU Xiaohong. Experimental study on the repair of spinal cord injury by conducting hydrogel loaded with tetramethylpyrazine sustained-release microparticles. Chinese Journal of Reparative and Reconstructive Surgery, 2023, 37(1): 65-73. doi: 10.7507/1002-1892.202209013 Copy

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15.	Wang Y, Lv HQ, Chao X, et al. Multimodal therapy strategies based on hydrogels for the repair of spinal cord injury. Mil Med Res, 2022, 9(1): 16. doi: 10.1186/s40779-022-00376-1.
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18.	贺丰, 俞兴, 穆晓红, 等. 新型脊髓完全横断缺损模型大鼠的建立. 中国组织工程研究, 2016, 20(5): 635-639.
19.	Deng WS, Ma K, Liang B, et al. Collagen scaffold combined with human umbilical cord-mesenchymal stem cells transplantation for acute complete spinal cord injury. Neural Regen Res, 2020, 15(9): 1686-1700.
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21.	Basso DM, Beattie MS, Bresnahan JC. A sensitive and reliable locomotor rating scale for open field testing in rats. J Neurotrauma, 1995, 12(1): 1-21.
22.	Poplawski GH, Tuszynski MH. Regeneration of corticospinal axons into neural progenitor cell grafts after spinal cord injury. Neurosci Insights, 2020, 15: 2633105520974000. doi: 10.1177/2633105520974000.
23.	Yang S, Zhang N, Dong Y, et al. Research on polycaprolactone-gelatin composite scaffolds carrying nerve growth factor for the repair of spinal cord injury. Dis Markers, 2022, 2022: 3880687. doi: 10.1155/2022/3880687.
24.	赵书杰, 陈建, 凡进, 等. 创伤性脊髓损伤后脊髓微环境失衡的研究进展. 中国脊柱脊髓杂志, 2020, 30(10): 942-947.
25.	Lin S, Xu C, Lin J, et al. Regulation of inflammatory cytokines for spinal cord injury recovery. Histol Histopathol, 2021, 36(2): 137-142.
26.	许子星, 许卫红, 陈薛敏, 等. 急性脊髓损伤大鼠血管重构与炎症反应的相关性研究. 中国修复重建外科杂志, 2020, 34(11): 1429-1437.
27.	Freyermuth-Trujillo X, Segura-Uribe JJ, Salgado-Ceballos H, et al. Inflammation: A target for treatment in spinal cord injury. Cells, 2022, 11(17): 2692. doi: 10.3390/cells11172692.

1. Karsy M, Hawryluk G. Modern medical management of spinal cord injury. Curr Neurol Neurosci Rep, 2019, 19(9): 65. doi: 10.1007/s11910-019-0984-1.
2. Eli I, Lerner DP, Ghogawala Z. Acute traumatic spinal cord injury. Neurol Clin, 2021, 39(2): 471-488.
3. Quadri SA, Farooqui M, Ikram A, et al. Recent update on basic mechanisms of spinal cord injury. Neurosurg Rev, 2020, 43(2): 425-441.
4. 蒋昇源, 邓博文, 徐林, 等. 川芎嗪修复脊髓损伤的作用及机制. 中国组织工程研究, 2022, 26(11): 1799-1804.
5. Fan Y, Wu Y. Tetramethylpyrazine alleviates neural apoptosis in injured spinal cord via the downregulation of miR-214-3p. Biomed Pharmacother, 2017, 94: 827-833.
6. Li J, Wei J, Wan Y, et al. TAT-modified tetramethylpyrazine-loaded nanoparticles for targeted treatment of spinal cord injury. J Control Release, 2021, 335: 103-116.
7. Xia H, Cheng Z, Cheng Y, et al. Investigating the passage of tetramethylpyrazine-loaded liposomes across blood-brain barrier models in vitro and ex vivo. Mater Sci Eng C Mater Biol Appl, 2016, 69: 1010-1017.
8. Fan L, Liu C, Chen X, et al. Directing induced pluripotent stem cell derived neural stem cell fate with a three-dimensional biomimetic hydrogel for spinal cord injury repair. ACS Appl Mater Interfaces, 2018, 10(21): 17742-17755.
9. Anderson MA, O’Shea TM, Burda JE, et al. Required growth facilitators propel axon regeneration across complete spinal cord injury. Nature, 2018, 561(7723): 396-400.
10. 王健豪, 刘洋, 付玄昊, 等. 3D打印组织工程支架联合骨髓间充质干细胞移植修复脊髓损伤. 中华骨科杂志, 2021, 41(6): 376-385.
11. Luo Y, Fan L, Liu C, et al. An injectable, self-healing, electroconductive extracellular matrix-based hydrogel for enhancing tissue repair after traumatic spinal cord injury. Bioact Mater, 2021, 7: 98-111.
12. Zhou L, Fan L, Yi X, et al. Soft conducting polymer hydrogels cross-linked and doped by tannic acid for spinal cord injury repair. ACS Nano, 2018, 12(11): 10957-10967.
13. Pang QM, Chen SY, Xu QJ, et al. Effects of astrocytes and microglia on neuroinflammation after spinal cord injury and related immunomodulatory strategies. Int Immunopharmacol, 2022, 108: 108754. doi: 10.1016/j.intimp.2022.108754.
14. Pinelli F, Pizzetti F, Veneruso V, et al. Biomaterial-mediated factor delivery for spinal cord injury treatment. Biomedicines, 2022, 10(7): 1673. doi: 10.3390/biomedicines10071673.
15. Wang Y, Lv HQ, Chao X, et al. Multimodal therapy strategies based on hydrogels for the repair of spinal cord injury. Mil Med Res, 2022, 9(1): 16. doi: 10.1186/s40779-022-00376-1.
16. Lv Z, Dong C, Zhang T, et al. Hydrogels in spinal cord injury repair: A review. Front Bioeng Biotechnol, 2022, 10: 931800. doi: 10.3389/fbioe.2022.931800.
17. Liu X, Miller AL, Park S, et al. Functionalized carbon nanotube and graphene oxide embedded electrically conductive hydrogel synergistically stimulates nerve cell differentiation. ACS Appl Mater Interfaces, 2017, 9(17): 14677-14690.
18. 贺丰, 俞兴, 穆晓红, 等. 新型脊髓完全横断缺损模型大鼠的建立. 中国组织工程研究, 2016, 20(5): 635-639.
19. Deng WS, Ma K, Liang B, et al. Collagen scaffold combined with human umbilical cord-mesenchymal stem cells transplantation for acute complete spinal cord injury. Neural Regen Res, 2020, 15(9): 1686-1700.
20. Lu Y, Yang J, Wang X, et al. Research progress in use of traditional Chinese medicine for treatment of spinal cord injury. Biomed Pharmacother, 2020, 127: 110136. doi: 10.1016/j.biopha.2020.110136.
21. Basso DM, Beattie MS, Bresnahan JC. A sensitive and reliable locomotor rating scale for open field testing in rats. J Neurotrauma, 1995, 12(1): 1-21.
22. Poplawski GH, Tuszynski MH. Regeneration of corticospinal axons into neural progenitor cell grafts after spinal cord injury. Neurosci Insights, 2020, 15: 2633105520974000. doi: 10.1177/2633105520974000.
23. Yang S, Zhang N, Dong Y, et al. Research on polycaprolactone-gelatin composite scaffolds carrying nerve growth factor for the repair of spinal cord injury. Dis Markers, 2022, 2022: 3880687. doi: 10.1155/2022/3880687.
24. 赵书杰, 陈建, 凡进, 等. 创伤性脊髓损伤后脊髓微环境失衡的研究进展. 中国脊柱脊髓杂志, 2020, 30(10): 942-947.
25. Lin S, Xu C, Lin J, et al. Regulation of inflammatory cytokines for spinal cord injury recovery. Histol Histopathol, 2021, 36(2): 137-142.
26. 许子星, 许卫红, 陈薛敏, 等. 急性脊髓损伤大鼠血管重构与炎症反应的相关性研究. 中国修复重建外科杂志, 2020, 34(11): 1429-1437.
27. Freyermuth-Trujillo X, Segura-Uribe JJ, Salgado-Ceballos H, et al. Inflammation: A target for treatment in spinal cord injury. Cells, 2022, 11(17): 2692. doi: 10.3390/cells11172692.

Chinese Journal of Reparative and Reconstructive Surgery

Experimental study on the repair of spinal cord injury by conducting hydrogel loaded with tetramethylpyrazine sustained-release microparticles

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