去細胞異種神經復合同種異體脂肪干細胞修復獼猴周圍神經缺損的免疫反應研究_《中國修復重建外科雜志》

作者：

黃喜軍 ¹ , 朱慶棠 ¹ , 江麗 ² , 鄭燦鑌 ¹ , 朱昭煒 ¹ , 路慶森 ¹ , 許銀峰 ³ , 顧立強 ¹ , 劉小林 ¹

中山大學附屬第一醫院（廣州，510080）1顯微創傷骨科，2骨科研究所，3黃埔院區上肢骨科;

關鍵詞：

去細胞異種神經脂肪干細胞異種移植同種異體移植免疫反應獼猴豬

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目的觀察去細胞異種神經復合同種異體脂肪干細胞修復獼猴周圍神經缺損后的全身及局部免疫排斥反應，評價該修復材料的安全性。方法健康成年雄性長白豬1只，體重48 kg，取脛神經制備去細胞異種神經；健康成年雄性獼猴1只，體重4.5 kg，分離培養脂肪干細胞；健康成年雌性獼猴10只，體重3～5 kg，制備25 mm長橈神經缺損動物模型，隨機分為復合細胞組和無細胞組（n=5），前者采用去細胞異種神經復合第3代脂肪干細胞移植修復，后者采用去細胞異種神經移植修復。于術前及術后14、60、90 d抽取外周靜脈血行淋巴細胞分析；術后5個月取材觀察移植物組織免疫反應和神經再生情況，并與自體神經移植組作比較。結果復合細胞組與無細胞組術前及術后各時間點外周靜脈血淋巴細胞數量、T淋巴細胞百分率及其數量、CD8+ T淋巴細胞百分率、CD4+/CD8+ T淋巴細胞比值間比較，差異均無統計學意義（P gt; 0.05）；術后14 d，復合細胞組CD4+ T淋巴細胞百分率低于其余時間點，差異有統計學意義（P lt; 0.05）；同一時間點兩組間比較，除術后14 d復合細胞組CD4+ T淋巴細胞百分率低于無細胞組，差異有統計學意義（P lt; 0.05）外，其余各時間點各指標組間比較差異均無統計學意義（P gt; 0.05）。術后5個月，去細胞異種神經與周圍組織有輕度粘連，神經外膜較自體神經厚，未見組織壞死、纖維瘢痕形成，移植物內見再生神經纖維，復合細胞組、無細胞組均有稀疏的CD3+、CD4+、CD8+、CD68+、CD163+ T淋巴細胞散在分布，細胞浸潤情況與自體神經移植組類似。結論去細胞異種神經移植修復獼猴周圍神經缺損后未產生全身及局部免疫排斥反應；復合同種異體脂肪干細胞移植后也未發現免疫排斥反應，且術后早期可能抑制CD4+ T淋巴細胞增殖。

引用本文： 黃喜軍,朱慶棠,江麗,鄭燦鑌,朱昭煒,路慶森,許銀峰,顧立強,劉小林. 去細胞異種神經復合同種異體脂肪干細胞修復獼猴周圍神經缺損的免疫反應研究. 中國修復重建外科雜志, 2012, 26(8): 993-1000. doi: 復制

1.	Siemionow M, Bozkurt M, Zor F. Regeneration and repair of peripheral nerves with different biomaterials: review. Microsurgery, 2010, 30(7): 574-588.
2.	范偉杰, 丁志清. 不同方法制備的脫細胞神經組織學及組織相容性研究. 中國修復重建外科雜志, 2011, 25(2): 176-180.
3.	王冠軍, 盧世璧, 匡正達, 等. 化學去細胞異體神經移植促神經趨化性再生實驗研究. 中國修復重建外科雜志, 2010, 24(11): 1288-1292.
4.	趙喆, 王玉, 彭江, 等. 化學去細胞異體神經周圍復合BMSCs生物蛋白膠復合物促周圍神經缺損修復. 中國修復重建外科雜志, 2011, 25(4): 488-493.
5.	Moore AM, Macewan M, Santosa KB, et al. Acellular nerve allografts in peripheral nerve regeneration: a comparative study. Muscle Nerve, 2011, 44(2): 221-234.
6.	Johnson PJ, Newton P, Hunter DA, et al. Nerve endoneurial microstructure facilitates uniform distribution of regenerative fibers: a post hoc comparison of midgraft nerve fiber densities. J Reconstr Microsurg, 2011, 27(2): 83-90.
7.	丁小珩, 劉小林, 劉育杰, 等. 去細胞同種異體神經修復材料臨床應用初步報告. 中華顯微外科雜志, 2009, 32(6): 448-450.
8.	郭義柱, 王巖, 劉相成, 等. 同種異體神經移植治療銳性臂叢神經缺損5例臨床研究. 軍醫進修學院學報, 2011, 32(4): 317-318.
9.	Brooks DN, Weber RV, Chao JD, et al. Processed nerve allografts for peripheral nerve reconstruction: A multicenter study of utilization and outcomes in sensory, mixed, and motor nerve reconstructions. Microsurgery, 2012, 32(1): 1-14.
10.	何彩鳳, 朱慶棠, 江麗, 等. 豬與人去細胞神經結構與成分的對比研究. 實用手外科雜志, 2011, 25(1): 39-42, 46.
11.	趙喆, 趙斌, 王玉, 等. 化學去細胞異體神經添加不同組織來源雪旺細胞對周圍神經損傷修復的功能評價. 中國修復重建外科雜志, 2010, 24(11): 1281-1287.
12.	di Summa PG, Kingham PJ, Raffoul W, et al. Adipose-derived stem cells enhance peripheral nerve regeneration. J Plast Reconstr Aesthet Surg, 2010, 63(9): 1544-1552.
13.	Liu G, Cheng Y, Guo S, et al. Transplantation of adipose-derived stem cells for peripheral nerve repair. Int J Mol Med, 2011, 28(4): 565-572.
14.	朱慶棠, 朱家愷, 賴英榮, 等. 去細胞組織工程化神經支架的制備與形態學研究. 中華顯微外科雜志, 2004, 27(1): 35-37.
15.	Wang D, Liu XL, Zhu JK, et al. Repairing large radial nerve defects by acellular nerve allografts seeded with autologous bone marrow stromal cells in a monkey model. J Neurotrauma, 2010, 27(10): 1935-1943.
16.	金婷, 林星石, 楊若佳, 等. 小鼠同種異體及異種神經移植后脾臟T淋巴細胞亞群的變化. 實驗動物科學, 2007, 24(4): 1-4.
17.	Rovak JM, Bishop DK, Boxer LK, et al. Peripheral nerve transplantation: the role of chemical acellularization in eliminating allograft antigenicity. J Reconstr Microsurg, 2005, 21(3): 207-213.
18.	Kvist M, Sondell M, Kanje M, et al. Regeneration in, and properties of, extracted peripheral nerve allografts and xenografts. J Plast Surg Hand Surg, 2011, 45(3): 122-128.
19.	Zhang Y, Luo H, Zhang Z, et al. A nerve graft constructed with xenogeneic acellular nerve matrix and autologous adipose-derived mesenchymal stem cells. Biomaterials, 2010, 31(20): 5312-5324.
20.	Tse KH, Sun M, Mantovani C, et al. In vitro evaluation of polyester-based scaffolds seeded with adipose derived stem cells for peripheral nerve regeneration. J Biomed Mater Res A, 2010, 95(3): 701-708.
21.	Sun F, Zhou K, Mi WJ, et al. Combined use of decellularized allogeneic artery conduits with autologous transdifferentiated adipose-derived stem cells for facial nerve regeneration in rats. Biomaterials, 2011, 32(32): 8118-8128.
22.	Klyushnenkova E, Mosca JD, Zernetkina V, et al. T cell responses to allogeneic human mesenchymal stem cells: immunogenicity, tolerance, and suppression. J Biomed Sci, 2005, 12(1): 47-57.
23.	Sioud M, Mobergslien A, Boudabous A, et al. Evidence for the involvement of galectin-3 in mesenchymal stem cell suppression of allogeneic T-cell proliferation. Scand J Immunol, 2010, 71(4): 267-274.
24.	Comoli P, Ginevri F, Maccario R, et al. Human mesenchymal stem cells inhibit antibody production induced in vitro by allostimulation. Nephrol Dial Transplant, 2008, 23(4): 1196-1202.
25.	Sotiropoulou PA, Perez SA, Gritzapis AD, et al. Interactions between human mesenchymal stem cells and natural killer cells. Stem Cells, 2006, 24(1): 74-85.
26.	Kuo YR, Chen CC, Goto S, et al. Modulation of immune response and T-cell regulation by donor adipose-derived stem cells in a rodent hind-limb allotransplant model. Plast Reconstr Surg, 2011, 128(6): 661e-672e.

1. Siemionow M, Bozkurt M, Zor F. Regeneration and repair of peripheral nerves with different biomaterials: review. Microsurgery, 2010, 30(7): 574-588.
2. 范偉杰, 丁志清. 不同方法制備的脫細胞神經組織學及組織相容性研究. 中國修復重建外科雜志, 2011, 25(2): 176-180.
3. 王冠軍, 盧世璧, 匡正達, 等. 化學去細胞異體神經移植促神經趨化性再生實驗研究. 中國修復重建外科雜志, 2010, 24(11): 1288-1292.
4. 趙喆, 王玉, 彭江, 等. 化學去細胞異體神經周圍復合BMSCs生物蛋白膠復合物促周圍神經缺損修復. 中國修復重建外科雜志, 2011, 25(4): 488-493.
5. Moore AM, Macewan M, Santosa KB, et al. Acellular nerve allografts in peripheral nerve regeneration: a comparative study. Muscle Nerve, 2011, 44(2): 221-234.
6. Johnson PJ, Newton P, Hunter DA, et al. Nerve endoneurial microstructure facilitates uniform distribution of regenerative fibers: a post hoc comparison of midgraft nerve fiber densities. J Reconstr Microsurg, 2011, 27(2): 83-90.
7. 丁小珩, 劉小林, 劉育杰, 等. 去細胞同種異體神經修復材料臨床應用初步報告. 中華顯微外科雜志, 2009, 32(6): 448-450.
8. 郭義柱, 王巖, 劉相成, 等. 同種異體神經移植治療銳性臂叢神經缺損5例臨床研究. 軍醫進修學院學報, 2011, 32(4): 317-318.
9. Brooks DN, Weber RV, Chao JD, et al. Processed nerve allografts for peripheral nerve reconstruction: A multicenter study of utilization and outcomes in sensory, mixed, and motor nerve reconstructions. Microsurgery, 2012, 32(1): 1-14.
10. 何彩鳳, 朱慶棠, 江麗, 等. 豬與人去細胞神經結構與成分的對比研究. 實用手外科雜志, 2011, 25(1): 39-42, 46.
11. 趙喆, 趙斌, 王玉, 等. 化學去細胞異體神經添加不同組織來源雪旺細胞對周圍神經損傷修復的功能評價. 中國修復重建外科雜志, 2010, 24(11): 1281-1287.
12. di Summa PG, Kingham PJ, Raffoul W, et al. Adipose-derived stem cells enhance peripheral nerve regeneration. J Plast Reconstr Aesthet Surg, 2010, 63(9): 1544-1552.
13. Liu G, Cheng Y, Guo S, et al. Transplantation of adipose-derived stem cells for peripheral nerve repair. Int J Mol Med, 2011, 28(4): 565-572.
14. 朱慶棠, 朱家愷, 賴英榮, 等. 去細胞組織工程化神經支架的制備與形態學研究. 中華顯微外科雜志, 2004, 27(1): 35-37.
15. Wang D, Liu XL, Zhu JK, et al. Repairing large radial nerve defects by acellular nerve allografts seeded with autologous bone marrow stromal cells in a monkey model. J Neurotrauma, 2010, 27(10): 1935-1943.
16. 金婷, 林星石, 楊若佳, 等. 小鼠同種異體及異種神經移植后脾臟T淋巴細胞亞群的變化. 實驗動物科學, 2007, 24(4): 1-4.
17. Rovak JM, Bishop DK, Boxer LK, et al. Peripheral nerve transplantation: the role of chemical acellularization in eliminating allograft antigenicity. J Reconstr Microsurg, 2005, 21(3): 207-213.
18. Kvist M, Sondell M, Kanje M, et al. Regeneration in, and properties of, extracted peripheral nerve allografts and xenografts. J Plast Surg Hand Surg, 2011, 45(3): 122-128.
19. Zhang Y, Luo H, Zhang Z, et al. A nerve graft constructed with xenogeneic acellular nerve matrix and autologous adipose-derived mesenchymal stem cells. Biomaterials, 2010, 31(20): 5312-5324.
20. Tse KH, Sun M, Mantovani C, et al. In vitro evaluation of polyester-based scaffolds seeded with adipose derived stem cells for peripheral nerve regeneration. J Biomed Mater Res A, 2010, 95(3): 701-708.
21. Sun F, Zhou K, Mi WJ, et al. Combined use of decellularized allogeneic artery conduits with autologous transdifferentiated adipose-derived stem cells for facial nerve regeneration in rats. Biomaterials, 2011, 32(32): 8118-8128.
22. Klyushnenkova E, Mosca JD, Zernetkina V, et al. T cell responses to allogeneic human mesenchymal stem cells: immunogenicity, tolerance, and suppression. J Biomed Sci, 2005, 12(1): 47-57.
23. Sioud M, Mobergslien A, Boudabous A, et al. Evidence for the involvement of galectin-3 in mesenchymal stem cell suppression of allogeneic T-cell proliferation. Scand J Immunol, 2010, 71(4): 267-274.
24. Comoli P, Ginevri F, Maccario R, et al. Human mesenchymal stem cells inhibit antibody production induced in vitro by allostimulation. Nephrol Dial Transplant, 2008, 23(4): 1196-1202.
25. Sotiropoulou PA, Perez SA, Gritzapis AD, et al. Interactions between human mesenchymal stem cells and natural killer cells. Stem Cells, 2006, 24(1): 74-85.
26. Kuo YR, Chen CC, Goto S, et al. Modulation of immune response and T-cell regulation by donor adipose-derived stem cells in a rodent hind-limb allotransplant model. Plast Reconstr Surg, 2011, 128(6): 661e-672e.

《中國修復重建外科雜志》

去細胞異種神經復合同種異體脂肪干細胞修復獼猴周圍神經缺損的免疫反應研究

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《中國修復重建外科雜志》

去細胞異種神經復合同種異體脂肪干細胞修復獼猴周圍神經缺損的免疫反應研究

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