新型一体化纤维环-髓核双相支架的制备与评估_《中国修复重建外科杂志》

作者：

许海委 ^1,2 , 徐宝山 ¹ , 杨强 ¹ , 李秀兰 ³ , 马信龙 ⁴ , 夏群 ¹ , 张春秋 ⁵ , 伍耀宏 ^1,2

天津医院（天津，300211）1脊柱外科，3骨科研究所细胞工程室;;
2天津医科大学研究生院;;
4天津医科大学总医院骨科;;
5 天津理工大学机械工程学院;

关键词：

脱细胞脱钙骨基质脱细胞髓核基质组织工程椎间盘双相支架猪兔

DOI：

10.7507/1002-1892.20130108

视频：

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摘要 全文 图表 视频 参考文献 施引文献 补充材料

目的用脱细胞脱钙骨基质和脱细胞髓核基质构建新型一体化纤维环-髓核双相支架，并进行理化检测，探讨其作为组织工程椎间盘支架的可行性。方法取猪股骨近端松质骨，塑形成外径10 mm、内径5 mm、厚3 mm的骨环，经脱脂、脱钙、脱细胞处理制备成纤维环相支架；取猪尾髓核组织，Triton X-100和脱氧胆酸联合脱细胞后粉碎、离心，制备脱细胞髓核基质；将制备的脱细胞髓核基质注入纤维环相支架中央，冷冻干燥，联合应用紫外照射和碳化二亚胺进行交联，制备一体化纤维环-髓核双相支架。通过大体观察、HE染色、扫描电镜观察支架结构；测定支架的孔隙率、吸水率、压缩弹性模量；MTT法检测25%、50%、100%支架浸提液对新西兰大白兔脂肪来源干细胞（adipose-derived stem cells，ADSCs）增殖的影响；将ADSCs接种到支架上，活/死细胞染色观察细胞在支架上的活性。结果大体观察支架呈乳白色；HE染色可见支架均质红染，无细胞成分残留；扫描电镜可见外层纤维环相支架孔径大小均匀一致，孔隙相互贯通，中央脱细胞髓核基质微丝相互连接，形成均匀网状结构，交界处连接紧密。纤维环相支架孔径为（343.00± 88.25）µm，一体化纤维环-髓核双相支架孔隙率为82.98% ± 7.02%、吸水率为621.53% ± 53.31%，压缩弹性模量为（89.07± 8.73）kPa。经MTT测定支架浸提液对ADSCs增殖无影响；活/死细胞染色可见细胞在支架上生长良好，无死细胞。结论用脱细胞脱钙骨基质和脱细胞髓核基质构建的一体化纤维环-髓核双相支架具有合适的孔径和孔隙率且无毒，力学性能与人正常椎间盘接近，是构建组织工程椎间盘的理想支架载体。

引用本文： 许海委,徐宝山,杨强,李秀兰,马信龙,夏群,张春秋,伍耀宏. 新型一体化纤维环-髓核双相支架的制备与评估. 中国修复重建外科杂志, 2013, 27(4): 475-480. doi: 10.7507/1002-1892.20130108 复制

1.	Li CQ, Huang B, Luo G, et al. Construction of collagen II/hyaluronate/chondroitin-6-sulfate tri-copolymer scaffold for nucleus pulposus tissue engineering and preliminary analysis of its physico-chemical properties and biocompatibility. J Mater Sci Mater Med, 2010, 21(2): 741-751.
2.	Koepsell L, Zhang L, Neufeld D, et al. Electrospun nanofibrous polycaprolactone scaffolds for tissue engineering of annulus fibrosus. Macromol Biosci, 2011, 11(3): 391-399.
3.	Gilbert TW, Sellaro TL, Badylak SF. Decellularization of tissues and organs. Biomaterials, 2006, 27(19): 3675-3683.
4.	Wan Y, Feng G, Shen FH, et al. Biphasic scaffold for annulus fibrosus tissue regeneration. Biomaterials, 2008, 29(6): 643-652.
5.	Mercuri JJ, Gill SS, Simionescu DT. Novel tissue-derived biomimetic scaffold for regenerating the human nucleus pulposus. J Biomed Mater Res A, 2011, 96(2): 422-435.
6.	Zhang R, Ma PX. Biomimetic polymer/apatite composite scaffolds for mineralized tissue engineering. Macromol Biosci, 2004, 4(2): 100-111.
7.	许海委, 徐宝山, 杨强, 等. 细胞共培养法诱导兔脂肪来源干细胞向软骨细胞分化的实验研究. 中国修复重建外科杂志, 2013, 27(2): 193-198.
8.	杨强, 彭江, 卢世璧, 等. 软骨脱细胞基质多孔支架与骨髓基质干细胞体外构建组织工程软骨的研究. 中华医学杂志, 2011, 91(17): 1161-1166.
9.	李长青, 周跃. 椎间盘组织工程支架材料研究进展. 中国修复重建外科杂志, 2005, 19(12): 1033-1035.
10.	Calderon L, Collin E, Velasco-Bayon D, et al. Type II collagen-hyaluronan hydrogel—a step towards a scaffold for intervertebral disc tissue engineering. Eur Cell Mater, 2010, 20: 134-48.
11.	Wang B, Borazjani A, Tahai M, et al. Fabrication of cardiac patch with decellularized porcine myocardial scaffold and bone marrowmononuclear cells. J Biomed Mater Res A, 2010, 94(4): 1100-1110.
12.	Stapleton TW, Ingram J, Fisher J, et al. Investigation of the regenerative capacity of an acellular porcine medial meniscus for tissueengineering applications. Tissue Eng Part A, 2011, 17(1-2): 231-242.
13.	Zhao Y, Zhang Z, Wang J, et al. Abdominal hernia repair with a decellularized dermal scaffold seeded with autologous bone marrow-derived mesenchymal stem cells. Artif Organs, 2012, 36(3): 247-255.
14.	黄惠民, 马良龙, 任宏, 等. 骨髓间质干细胞和脱细胞基质构建组织工程血管的动物实验. 中华医学杂志, 2007, 87(48): 3440-3442.
15.	Du L, Wu X. Development and characterization of a full-thickness acellular porcine cornea matrix for tissueengineering. Artif Organs, 2011, 35(7): 691-705.
16.	Cheng HL, Loai Y, Beaumont M, et al. The acellular matrix (ACM) for bladder tissue engineering: A quantitative magnetic resonanceimaging study. Magn Reson Med, 2010, 64(2): 341-348.
17.	Xu CC, Chan RW, Weinberger DG, et al. A bovine acellular scaffold for vocal fold reconstruction in a rat model. J Biomed Mater Res A, 2010, 92(1): 18-32.
18.	Zhuang Y, Huang B, Li CQ, et al. Construction of tissue-engineered composite intervertebral disc and preliminary morphological and biochemical evaluation. Biochem Biophys Res Commun, 2011, 407(2): 327-332.
19.	潘勇, 周跃, 郝勇, 等. 脱矿脱细胞骨基质环支架体外构建组织工程化椎间盘纤维环. 中国脊柱脊髓杂志, 2009, 19(6): 451-457.
20.	Bowles RD, Gebhard HH, Härtl R, et al. Tissue-engineered intervertebral discs produce new matrix, maintain disc height, and restore biomechanical function to the rodent spine. Proc Natl Acad Sci U S A, 2011, 108(32): 13106-13111.

1. Li CQ, Huang B, Luo G, et al. Construction of collagen II/hyaluronate/chondroitin-6-sulfate tri-copolymer scaffold for nucleus pulposus tissue engineering and preliminary analysis of its physico-chemical properties and biocompatibility. J Mater Sci Mater Med, 2010, 21(2): 741-751.
2. Koepsell L, Zhang L, Neufeld D, et al. Electrospun nanofibrous polycaprolactone scaffolds for tissue engineering of annulus fibrosus. Macromol Biosci, 2011, 11(3): 391-399.
3. Gilbert TW, Sellaro TL, Badylak SF. Decellularization of tissues and organs. Biomaterials, 2006, 27(19): 3675-3683.
4. Wan Y, Feng G, Shen FH, et al. Biphasic scaffold for annulus fibrosus tissue regeneration. Biomaterials, 2008, 29(6): 643-652.
5. Mercuri JJ, Gill SS, Simionescu DT. Novel tissue-derived biomimetic scaffold for regenerating the human nucleus pulposus. J Biomed Mater Res A, 2011, 96(2): 422-435.
6. Zhang R, Ma PX. Biomimetic polymer/apatite composite scaffolds for mineralized tissue engineering. Macromol Biosci, 2004, 4(2): 100-111.
7. 许海委, 徐宝山, 杨强, 等. 细胞共培养法诱导兔脂肪来源干细胞向软骨细胞分化的实验研究. 中国修复重建外科杂志, 2013, 27(2): 193-198.
8. 杨强, 彭江, 卢世璧, 等. 软骨脱细胞基质多孔支架与骨髓基质干细胞体外构建组织工程软骨的研究. 中华医学杂志, 2011, 91(17): 1161-1166.
9. 李长青, 周跃. 椎间盘组织工程支架材料研究进展. 中国修复重建外科杂志, 2005, 19(12): 1033-1035.
10. Calderon L, Collin E, Velasco-Bayon D, et al. Type II collagen-hyaluronan hydrogel—a step towards a scaffold for intervertebral disc tissue engineering. Eur Cell Mater, 2010, 20: 134-48.
11. Wang B, Borazjani A, Tahai M, et al. Fabrication of cardiac patch with decellularized porcine myocardial scaffold and bone marrowmononuclear cells. J Biomed Mater Res A, 2010, 94(4): 1100-1110.
12. Stapleton TW, Ingram J, Fisher J, et al. Investigation of the regenerative capacity of an acellular porcine medial meniscus for tissueengineering applications. Tissue Eng Part A, 2011, 17(1-2): 231-242.
13. Zhao Y, Zhang Z, Wang J, et al. Abdominal hernia repair with a decellularized dermal scaffold seeded with autologous bone marrow-derived mesenchymal stem cells. Artif Organs, 2012, 36(3): 247-255.
14. 黄惠民, 马良龙, 任宏, 等. 骨髓间质干细胞和脱细胞基质构建组织工程血管的动物实验. 中华医学杂志, 2007, 87(48): 3440-3442.
15. Du L, Wu X. Development and characterization of a full-thickness acellular porcine cornea matrix for tissueengineering. Artif Organs, 2011, 35(7): 691-705.
16. Cheng HL, Loai Y, Beaumont M, et al. The acellular matrix (ACM) for bladder tissue engineering: A quantitative magnetic resonanceimaging study. Magn Reson Med, 2010, 64(2): 341-348.
17. Xu CC, Chan RW, Weinberger DG, et al. A bovine acellular scaffold for vocal fold reconstruction in a rat model. J Biomed Mater Res A, 2010, 92(1): 18-32.
18. Zhuang Y, Huang B, Li CQ, et al. Construction of tissue-engineered composite intervertebral disc and preliminary morphological and biochemical evaluation. Biochem Biophys Res Commun, 2011, 407(2): 327-332.
19. 潘勇, 周跃, 郝勇, 等. 脱矿脱细胞骨基质环支架体外构建组织工程化椎间盘纤维环. 中国脊柱脊髓杂志, 2009, 19(6): 451-457.
20. Bowles RD, Gebhard HH, Härtl R, et al. Tissue-engineered intervertebral discs produce new matrix, maintain disc height, and restore biomechanical function to the rodent spine. Proc Natl Acad Sci U S A, 2011, 108(32): 13106-13111.

《中国修复重建外科杂志》

新型一体化纤维环-髓核双相支架的制备与评估

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《中国修复重建外科杂志》

新型一体化纤维环-髓核双相支架的制备与评估

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