<i>In vivo</i> degradation and histocompatibility of modified chitosan based on conductive composite nerve conduit_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

 JIAO Haishan ¹ , SONG Yuening ¹ , HUANG Jian ¹ , LI Dongyin ¹ , HU Yi ²

1. Department of Clinical Medicine, Suzhou Vocational Health College, Suzhou Jiangsu, 215009, P.R.China;
2. Department of Pharmacy, Suzhou Vocational Health College, Suzhou Jiangsu, 215009, P.R.China;

Corresponding author：

JIAO Haishan, Email: jiaohaishan@szhct.edu.cn

Keywords：

Tissue engineered nerve; nanomaterial; polypyrrole; chitosan; in vivo degradation; histocompatibility; nerve conduit

DOI：

10.7507/1002-1892.202101088

Video：

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Objective To investigate the in vivo degradation and histocompatibility of modified chitosan based on conductive composite nerve conduit, so as to provide a new scaffold material for the construction of tissue engineered nerve.Methods The nano polypyrrole (PPy) was synthesized by microemulsion polymerization, blended with chitosan, and then formed conduit by injecting the mixed solution into a customized conduit formation model. After freeze-drying and deacidification, the nano PPy/chitosan composite conduit (CP conduit) was prepared. Then the CP conduits with different acetyl degree were resulted undergoing varying acetylation for 30, 60, and 90 minutes (CAP1, CAP2, CAP3 conduits). Fourier infrared absorption spectrum and scanning electron microscopy (SEM) were used to identify the conduits. And the conductivity was measured by four-probe conductometer. The above conduits were implanted after the subcutaneous fascial tunnels were made symmetrically on both sides of the back of 30 female Sprague Dawley rats. At 2, 4, 6, 8, 10, and 12 weeks after operation, the morphology, the microstructure, and the degradation rate were observed and measured to assess the in vivo degradation of conduits. HE staining and anti-macrophage immunofluorescence staining were performed to observe the histocompatibility in vivo.Results The characteristic peaks of the amide Ⅱ band around 1 562 cm⁻¹ appeared after being acetylated, indicating that the acetylation modification of chitosan was successful. There was no significant difference in conductivity between conduits (P>0.05). SEM observation showed that the surfaces of the conduits in all groups were similar with relatively smooth surface and compact structure. After the conduits were implanted into the rats, with the extension of time, all conduits were collapsed, especially on the CAP3 conduit. All conduits had different degrees of mass loss, and the higher the degree of acetylation, the greater the mass change (P<0.05). SEM observation showed that there were more pores at 12 weeks after implantation, and the pores showed an increasing trend as the degree of acetylation increased. Histological observation showed that there were more macrophages and lymphocytes infiltration in each group at the early stage. With the extension of implantation time, lymphocytes decreased, fibroblasts increased, and collagen fibers proliferated significantly. Conclusion The modified chitosan basedon conductive composite nerve conduit made of nano-PPy/chitosan composite with different acetylation degrees has good biocompatibility, conductivity, and biodegradability correlated with acetylation degree in vivo, which provide a new scaffold material for the construction of tissue engineered nerve.

Citation： JIAO Haishan, SONG Yuening, HUANG Jian, LI Dongyin, HU Yi. In vivo degradation and histocompatibility of modified chitosan based on conductive composite nerve conduit. Chinese Journal of Reparative and Reconstructive Surgery, 2021, 35(6): 769-775. doi: 10.7507/1002-1892.202101088 Copy

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11.	Wan Y, Wu H, Wen D. Porous-conductive chitosan scaffolds for tissue engineering, 1. Preparation and characterization. Macromol Biosci, 2004, 4(9): 882-890.
12.	Yang YM, Hu W, Wang XD, et al. The controlling biodegradation of chitosan fibers by N-acetylation in vitro and in vivo. J Mater Sci Mater Med, 2007, 18(11): 2117-2121.
13.	董炎明, 许聪义, 汪剑炜, 等. 红外光谱法测定N-酰化壳聚糖的取代度. 中国科学 (B 辑), 2001, 31(2): 153-160.
14.	Zhang PX, Han N, Kou YH, et al. Tissue engineering for the repair of peripheral nerve injury. Neural Regen Res, 2019, 14(1): 51-58.
15.	Park S, Liu CY, Ward PJ, et al. Effects of repeated 20-Hz electrical stimulation on functional recovery following peripheral nerve injury. Neurorehabil Neural Repair, 2019, 33(9): 775-784.
16.	Zuo KJ, Gordon T, Chan KM, et al. Electrical stimulation to enhance peripheral nerve regeneration: Update in molecular investigations and clinical translation. Exp Neurol, 2020, 332: 113397.
17.	王爱勤. 甲壳素化学. 北京: 科学出版社, 2008: 226-227.
18.	Shi G, Rouabhia M, Wang Z, et al. A novel electrically conductive and biodegradable composite made of polypyrrole nanoparticles and polylactide. Biomaterials, 2004, 25(13): 2477- 2488.
19.	Neisi Z, Ansari-Asl Z, Jafarinejad-Farsangi S, et al. Synthesis, characterization and biocompatibility of polypyrrole/Cu (Ⅱ) metal-organic framework nanocomposites. Colloids Surf B Biointerfaces, 2019, 178: 365-376.
20.	Freier T, Koh HS, Kazazian K, et al. Controlling cell adhesion and degradation of chitosan films by N-acetylation. Biomaterials, 2005, 26(29): 5872-5878.
21.	Younes I, Rinaudo M. Chitin and chitosan preparation from marine sources. Structure, properties and applications. Mar Drugs, 2015, 13(3): 1133-1174.

1. Wang X, Hu W, Cao Y, et al. Dog sciatic nerve regeneration across a 30-mm defect bridged by a chitosan/PGA artificial nerve graft. Brain, 2005, 128(Pt 8): 1897-1910.
2. Wang Y, Zhao Y, Sun C, et al. Chitosan degradation products promote nerve regeneration by dtimulating Schwann cell proliferation via miR-27a/FOXO1 axis. Mol Neurobiol, 2016, 53(1): 28-39.
3. Peng Y, Li KY, Chen YF, et al. Beagle sciatic nerve regeneration across a 30 mm defect bridged by chitosan/PGA artificial nerve grafts. Injury, 2018, 49(8): 1477-1484.
4. Jiao H, Yao J, Yang Y, et al. Chitosan/polyglycolic acid nerve grafts for axon regeneration from prolonged axotomized neurons to chronically denervated segments. Biomaterials, 2009, 30(28): 5004-5018.
5. 焦海山, 姚健, 任艳玲, 等. 甲壳素神经导管修复大鼠坐骨神经10 mm缺损的实验研究. 中国生物医学工程学报, 2008, 27(4): 597-602, 620.
6. Zhao Y, Liang Y, Ding S, et al. Application of conductive PPy/SF composite scaffold and electrical stimulation for neural tissue engineering. Biomaterials, 2020, 255: 120164. doi: 10.1016/j.biomaterials.2020.120164.
7. Huang J, Lu L, Zhang J, et al. Electrical stimulation to conductive scaffold promotes axonal regeneration and remyelination in a rat model of large nerve defect. PLoS One, 2012, 7(6): e39526. doi: 10.1371/journal.pone.0039526.
8. Baniasadi H, Ramazani SAA, Mashayekhan S. Fabrication and characterization of conductive chitosan/gelatin-based scaffolds for nerve tissue engineering. Int J Biol Macromol, 2015, 74: 360-366.
9. Wang Y, Zhang Y, Zhang Z, et al. An injectable high-conductive bimaterial scaffold for neural stimulation. Colloids Surf B Biointerfaces, 2020, 195: 111210. doi: 10.1016/j.colsurfb.2020.111210.
10. 焦海山, 曹萍, 陈颖, 等. 纳米聚吡咯/甲壳素复合膜的制备及其生物相容性观察. 中国修复重建外科杂志, 2018, 32(8): 1081-1087.
11. Wan Y, Wu H, Wen D. Porous-conductive chitosan scaffolds for tissue engineering, 1. Preparation and characterization. Macromol Biosci, 2004, 4(9): 882-890.
12. Yang YM, Hu W, Wang XD, et al. The controlling biodegradation of chitosan fibers by N-acetylation in vitro and in vivo. J Mater Sci Mater Med, 2007, 18(11): 2117-2121.
13. 董炎明, 许聪义, 汪剑炜, 等. 红外光谱法测定N-酰化壳聚糖的取代度. 中国科学 (B 辑), 2001, 31(2): 153-160.
14. Zhang PX, Han N, Kou YH, et al. Tissue engineering for the repair of peripheral nerve injury. Neural Regen Res, 2019, 14(1): 51-58.
15. Park S, Liu CY, Ward PJ, et al. Effects of repeated 20-Hz electrical stimulation on functional recovery following peripheral nerve injury. Neurorehabil Neural Repair, 2019, 33(9): 775-784.
16. Zuo KJ, Gordon T, Chan KM, et al. Electrical stimulation to enhance peripheral nerve regeneration: Update in molecular investigations and clinical translation. Exp Neurol, 2020, 332: 113397.
17. 王爱勤. 甲壳素化学. 北京: 科学出版社, 2008: 226-227.
18. Shi G, Rouabhia M, Wang Z, et al. A novel electrically conductive and biodegradable composite made of polypyrrole nanoparticles and polylactide. Biomaterials, 2004, 25(13): 2477- 2488.
19. Neisi Z, Ansari-Asl Z, Jafarinejad-Farsangi S, et al. Synthesis, characterization and biocompatibility of polypyrrole/Cu (Ⅱ) metal-organic framework nanocomposites. Colloids Surf B Biointerfaces, 2019, 178: 365-376.
20. Freier T, Koh HS, Kazazian K, et al. Controlling cell adhesion and degradation of chitosan films by N-acetylation. Biomaterials, 2005, 26(29): 5872-5878.
21. Younes I, Rinaudo M. Chitin and chitosan preparation from marine sources. Structure, properties and applications. Mar Drugs, 2015, 13(3): 1133-1174.

Chinese Journal of Reparative and Reconstructive Surgery

In vivo degradation and histocompatibility of modified chitosan based on conductive composite nerve conduit

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