骨骼肌無細胞基質的制備及其生物相容性研究_《中國修復重建外科雜志》

作者：

田春祥 ^1,2 , 范雪嬌 ^1,2 , 陳曉禾 ² , 鄧力 ² , 秦廷武 ² , 羅靜聰 ² , 李秀群 ² , 呂青 ¹

四川大學華西醫院（成都，610041） 1 甲狀腺乳腺外科，2 生物治療國家重點實驗室· 干細胞與組織工程研究室;

關鍵詞：

脂肪組織工程骨骼肌無細胞基質人脂肪干細胞支架材料

視頻：

導出 下載 收藏 掃碼 引用

摘要 全文 圖表 視頻 參考文獻 施引文獻 補充材料

目的細胞外基質是脂肪組織工程材料的研究熱點之一。通過探討骨骼肌無細胞基質的制作方法及生物相容性，為其在脂肪組織工程中的應用奠定基礎。方法取健康成年小香豬新鮮骨骼肌組織，橫切成厚2 ～ 3 mm 的組織塊，采用低滲- 去垢劑法脫細胞處理。處理后采用HE 染色、Masson 三色染色、免疫組織化學染色及掃描電鏡檢測骨骼肌無細胞基質是否有細胞成分殘留，并觀察其基本結構；應用MTT 法檢測骨骼肌無細胞基質細胞毒性。取乳腺癌患者自愿捐贈脂肪組織，分離培養人脂肪干細胞（human adipose-derived stem cells，hADSCs），從形態學、流式細胞學和成脂、成骨分化能力方面進行鑒定。將骨骼肌無細胞基質與第3 代hADSCs 共培養，于培養后第1、3、5、7 天通過細胞活性檢測材料上細胞黏附、擴散和增殖情況，了解其與細胞之間的相互作用。結果 HE、Masson、免疫組織化學染色及掃描電鏡觀察顯示骨骼肌無細胞基質肌纖維去除完全，無細胞核殘留，基質結構保留完整；大量連接成網狀的膠原纖維呈多孔隙樣結構，規則排列。MTT 檢測示骨骼肌無細胞基質細胞毒性為1 級，細胞相容性好。細胞活性檢測示hADSCs 在骨骼肌無細胞基質上能很好地伸展，且能與周圍基質黏附，進入基質內部并相互交織。結論經脫細胞處理的骨骼肌無細胞基質具有良好生物相容性，可能作為脂肪組織工程的支架材料。

引用本文： 田春祥,范雪嬌,陳曉禾,鄧力,秦廷武,羅靜聰,李秀群,呂青. 骨骼肌無細胞基質的制備及其生物相容性研究. 中國修復重建外科雜志, 2012, 26(6): 749-754. doi: 復制

1.	Lutolf MP, Hubbell JA. Synthetic biomaterials as instructive extracellular microenvironments for morphogenesis in tissue engineering. Nat Biotechnol, 2005, 23(1): 47-55.
2.	Gilbert TW, Sellaro TL, Badylak SF. Decellularization of tissues and organs. Biomaterials, 2006, 27(19): 3675-3683.
3.	Borschel GH, Dennis RG, Kuzon WM Jr. Contractile skeletal muscle tissue-engineered on an acellular scaffold. Plast Reconstr Surg, 2004, 113(2): 595-602.
4.	Valentin JE, Turner NJ, Gilbert TW, et al. Functional skeletal muscle formation with a biologic scaffold. Biomaterials, 2010, 31(29): 7475-7484.
5.	Penolazzi L, Mazzitelli S, Vecchiatini R, et al. Human mesenchymal stem cells seeded on extracellular matrix scaffold: Viability and osteogenic potential. J Cell Physiol, 2012, 227(2): 857-866.
6.	Zhu WD, Xu YM, Feng C, et al. Different bladder defects reconstructed with bladder acellular matrix grafts in a rabbit model. Urologe A, 2011, 50(11): 1420-1425.
7.	Zhang X, Yang J, Li Y, et al. Functional neovascularization in tissue engineering with porcine acellular dermal matrix and human umbilical vein endothelial cells. Tissue Eng Part C Methods, 2011, 17(4): 423-433.
8.	張建, 吳英鋒, 陳亮. 豬主動脈脫細胞基質的簡化制備及生物學評價. 中國修復重建外科雜志, 2008, 22(3): 364-369.
9.	Wainwright JM, Hashizume R, Fujimoto KL, et al. Right ventricular outflow tract repair with a cardiac biologic scaffold. Cells Tissues Organs, 2012, 195(1-2): 159-170.
10.	Soto-Gutierrez A, Zhang L, Medberry C, et al. A whole organ regenerative medicine approach for liver replacement. Tissue Eng Part C Methods, 2011, 17(6): 677-686.
11.	Uygun BE, Soto-Gutierrez A, Yagi H, et al. Organ reengineering through development of a transplantable recellularized liver graft using decellularized liver matrix. Nat Med, 2010, 16(7): 814-820.
12.	Petersen TH, Calle EA, Zhao L, et al. Tissue-engineered lungs for in vivo implantation. Science, 2010, 329(5991): 538-541.
13.	Karabekmez FE, Duymaz A, Moran SL. Early clinical outcomes with the use of decellularized nerve allograft for repair of sensory defects within the hand. Hand (N Y), 2009, 4(3): 245-249.
14.	Vindigni V, Mazzoleni F, Rossini K, et al. Reconstruction of ablated rat rectus abdominis by muscle regeneration. Plast Reconstr Surg, 2004, 114(6): 1509-1515.
15.	Conconi MT, De Coppi P, Bellini S, et al. Homologous muscle acellular matrix seeded with autologous myoblasts as a tissue-engineering approach to abdominal wall-defect repair. Biomaterials, 2005, 26(15): 2567-2574.
16.	薛輝, 陳東, 張秀英, 等. 化學去細胞肌肉組織工程支架與大鼠脊髓的生物相容性. 吉林大學學報: 醫學版, 2009, 35(5): 801-804.
17.	Zhang XY, Xue H, Liu JM, et al. Chemically extracted acellular muscle: A new potential scaffold for spinal cord injury repair. J Biomed Mater Res A, 2012, 100(3): 578-587.
18.	Wang B, Borazjani A, Tahai M, et al. Fabrication of cardiac patch with decellularized porcine myocardial scaffold and bone marrow mononuclear cells. J Biomed Mater Res A, 2010, 94(4): 1100-1110.
19.	Flynn L, Semple JL, Woodhouse KA. Decellularized placental matrices for adipose tissue engineering. J Biomed Mater Res A, 2006, 79(2): 359-369.
20.	Flynn L, Woodhouse KA. Adipose tissue engineering with cells in engineered matrices. Organogenesis, 2008, 4(4): 228-235.
21.	Gomillion CT, Burg KJ. Stem cells and adipose tissue engineering. Biomaterials, 2006, 27(36): 6052-6063.
22.	Cherubino M, Marra KG. Adipose-derived stem cells for soft tissue reconstruction. Regen Med, 2009, 4(1): 109-117.
23.	Huss FR, Kratz G. Adipose tissue processed for lipoinjection shows increased cellular survival in vitro when tissue engineering principles are applied. Scand J Plast Reconstr Surg Hand Surg, 2002, 36(3): 166-171.
24.	Beahm EK, Walton RL, Patrick CW Jr. Progress in adipose tissue construct development. Clin Plast Surg, 2003, 30(4): 547-558, viii.
25.	Patrick CW Jr. Tissue engineering strategies for adipose tissue repair. Anat Rec, 2001, 263(4): 361-366.
26.	Flynn LE. The use of decellularized adipose tissue to provide an inductive microenvironment for the adipogenic differentiation of human adipose-derived stem cells. Biomaterials, 2010, 31(17): 4715-4724.
27.	Crapo PM, Gilbert TW, Badylak SF. An overview of tissue and whole organ decellularization processes. Biomaterials, 2011, 32(12): 3233-3243.
28.	卿泉, 秦廷武. 大鼠骨骼肌脫細胞處理方法的優化研究. 中國修復重建外科雜志, 2009, 23(7): 836-839.
29.	Ciapetti G, Cenni E, Pratelli L, et al. In vitro evaluation of cell/biomaterial interaction by MTT assay. Biomaterials, 1993, 14(5): 359-364.
30.	Zuk PA, Zhu M, Mizuno H, et al. Multilineage cells from human adipose tissue: implications for cell-based therapies. Tissue Eng, 2001, 7(2): 211-228.
31.	Zuk PA, Zhu M, Ashjian P, et al. Human adipose tissue is a source of multipotent stem cells. Mol Biol Cell, 2002, 13(12): 4279-4295.
32.	Gimble JM, Guilak F. Adipose-derived adult stem cells: isolation, characterization, and differentiation potential. Cytotherapy, 2003, 5(5): 362-369.
33.	Gimble JM, Katz AJ, Bunnell BA. Adipose-derived stem cells for regenerative medicine. Circ Res, 2007, 100(9): 1249-1260.
34.	Dobson DE, Kambe A, Block E, et al. 1-Butyryl-glycerol: a novel angiogenesis factor secreted by differentiating adipocytes. Cell, 1990, 61(2): 223-230.
35.	Rodriguez AM, Pisani D, Dechesne CA, et al. Transplantation of a multipotent cell population from human adipose tissue induces dystrophin expression in the immunocompetent mdx mouse. J Exp Med, 2005, 201(9): 1397-1405.
36.	McIntosh K, Zvonic S, Garrett S, et al. The immunogenicity of human adipose-derived cells: temporal changes in vitro. Stem Cells, 2006, 24(5): 1246-1253.

1. Lutolf MP, Hubbell JA. Synthetic biomaterials as instructive extracellular microenvironments for morphogenesis in tissue engineering. Nat Biotechnol, 2005, 23(1): 47-55.
2. Gilbert TW, Sellaro TL, Badylak SF. Decellularization of tissues and organs. Biomaterials, 2006, 27(19): 3675-3683.
3. Borschel GH, Dennis RG, Kuzon WM Jr. Contractile skeletal muscle tissue-engineered on an acellular scaffold. Plast Reconstr Surg, 2004, 113(2): 595-602.
4. Valentin JE, Turner NJ, Gilbert TW, et al. Functional skeletal muscle formation with a biologic scaffold. Biomaterials, 2010, 31(29): 7475-7484.
5. Penolazzi L, Mazzitelli S, Vecchiatini R, et al. Human mesenchymal stem cells seeded on extracellular matrix scaffold: Viability and osteogenic potential. J Cell Physiol, 2012, 227(2): 857-866.
6. Zhu WD, Xu YM, Feng C, et al. Different bladder defects reconstructed with bladder acellular matrix grafts in a rabbit model. Urologe A, 2011, 50(11): 1420-1425.
7. Zhang X, Yang J, Li Y, et al. Functional neovascularization in tissue engineering with porcine acellular dermal matrix and human umbilical vein endothelial cells. Tissue Eng Part C Methods, 2011, 17(4): 423-433.
8. 張建, 吳英鋒, 陳亮. 豬主動脈脫細胞基質的簡化制備及生物學評價. 中國修復重建外科雜志, 2008, 22(3): 364-369.
9. Wainwright JM, Hashizume R, Fujimoto KL, et al. Right ventricular outflow tract repair with a cardiac biologic scaffold. Cells Tissues Organs, 2012, 195(1-2): 159-170.
10. Soto-Gutierrez A, Zhang L, Medberry C, et al. A whole organ regenerative medicine approach for liver replacement. Tissue Eng Part C Methods, 2011, 17(6): 677-686.
11. Uygun BE, Soto-Gutierrez A, Yagi H, et al. Organ reengineering through development of a transplantable recellularized liver graft using decellularized liver matrix. Nat Med, 2010, 16(7): 814-820.
12. Petersen TH, Calle EA, Zhao L, et al. Tissue-engineered lungs for in vivo implantation. Science, 2010, 329(5991): 538-541.
13. Karabekmez FE, Duymaz A, Moran SL. Early clinical outcomes with the use of decellularized nerve allograft for repair of sensory defects within the hand. Hand (N Y), 2009, 4(3): 245-249.
14. Vindigni V, Mazzoleni F, Rossini K, et al. Reconstruction of ablated rat rectus abdominis by muscle regeneration. Plast Reconstr Surg, 2004, 114(6): 1509-1515.
15. Conconi MT, De Coppi P, Bellini S, et al. Homologous muscle acellular matrix seeded with autologous myoblasts as a tissue-engineering approach to abdominal wall-defect repair. Biomaterials, 2005, 26(15): 2567-2574.
16. 薛輝, 陳東, 張秀英, 等. 化學去細胞肌肉組織工程支架與大鼠脊髓的生物相容性. 吉林大學學報: 醫學版, 2009, 35(5): 801-804.
17. Zhang XY, Xue H, Liu JM, et al. Chemically extracted acellular muscle: A new potential scaffold for spinal cord injury repair. J Biomed Mater Res A, 2012, 100(3): 578-587.
18. Wang B, Borazjani A, Tahai M, et al. Fabrication of cardiac patch with decellularized porcine myocardial scaffold and bone marrow mononuclear cells. J Biomed Mater Res A, 2010, 94(4): 1100-1110.
19. Flynn L, Semple JL, Woodhouse KA. Decellularized placental matrices for adipose tissue engineering. J Biomed Mater Res A, 2006, 79(2): 359-369.
20. Flynn L, Woodhouse KA. Adipose tissue engineering with cells in engineered matrices. Organogenesis, 2008, 4(4): 228-235.
21. Gomillion CT, Burg KJ. Stem cells and adipose tissue engineering. Biomaterials, 2006, 27(36): 6052-6063.
22. Cherubino M, Marra KG. Adipose-derived stem cells for soft tissue reconstruction. Regen Med, 2009, 4(1): 109-117.
23. Huss FR, Kratz G. Adipose tissue processed for lipoinjection shows increased cellular survival in vitro when tissue engineering principles are applied. Scand J Plast Reconstr Surg Hand Surg, 2002, 36(3): 166-171.
24. Beahm EK, Walton RL, Patrick CW Jr. Progress in adipose tissue construct development. Clin Plast Surg, 2003, 30(4): 547-558, viii.
25. Patrick CW Jr. Tissue engineering strategies for adipose tissue repair. Anat Rec, 2001, 263(4): 361-366.
26. Flynn LE. The use of decellularized adipose tissue to provide an inductive microenvironment for the adipogenic differentiation of human adipose-derived stem cells. Biomaterials, 2010, 31(17): 4715-4724.
27. Crapo PM, Gilbert TW, Badylak SF. An overview of tissue and whole organ decellularization processes. Biomaterials, 2011, 32(12): 3233-3243.
28. 卿泉, 秦廷武. 大鼠骨骼肌脫細胞處理方法的優化研究. 中國修復重建外科雜志, 2009, 23(7): 836-839.
29. Ciapetti G, Cenni E, Pratelli L, et al. In vitro evaluation of cell/biomaterial interaction by MTT assay. Biomaterials, 1993, 14(5): 359-364.
30. Zuk PA, Zhu M, Mizuno H, et al. Multilineage cells from human adipose tissue: implications for cell-based therapies. Tissue Eng, 2001, 7(2): 211-228.
31. Zuk PA, Zhu M, Ashjian P, et al. Human adipose tissue is a source of multipotent stem cells. Mol Biol Cell, 2002, 13(12): 4279-4295.
32. Gimble JM, Guilak F. Adipose-derived adult stem cells: isolation, characterization, and differentiation potential. Cytotherapy, 2003, 5(5): 362-369.
33. Gimble JM, Katz AJ, Bunnell BA. Adipose-derived stem cells for regenerative medicine. Circ Res, 2007, 100(9): 1249-1260.
34. Dobson DE, Kambe A, Block E, et al. 1-Butyryl-glycerol: a novel angiogenesis factor secreted by differentiating adipocytes. Cell, 1990, 61(2): 223-230.
35. Rodriguez AM, Pisani D, Dechesne CA, et al. Transplantation of a multipotent cell population from human adipose tissue induces dystrophin expression in the immunocompetent mdx mouse. J Exp Med, 2005, 201(9): 1397-1405.
36. McIntosh K, Zvonic S, Garrett S, et al. The immunogenicity of human adipose-derived cells: temporal changes in vitro. Stem Cells, 2006, 24(5): 1246-1253.

《中國修復重建外科雜志》

骨骼肌無細胞基質的制備及其生物相容性研究

摘要 全文 圖表 視頻 參考文獻 施引文獻 補充材料

Format

Content

《中國修復重建外科雜志》

骨骼肌無細胞基質的制備及其生物相容性研究

摘要 全文 圖表 視頻 參考文獻 施引文獻 補充材料

Format

Content

摘要全文圖表視頻參考文獻施引文獻補充材料