Preparation and Characterization of a Brain Decellularized Scaffold
Zheng Fangping1*, Mao Kaili1, Zhao Yingzheng2
1Department of Pharmacy, People's Hospital of Quzhou, Quzhou 324000, Zhejiang, China 2School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou 325025, Zhejiang, China
Abstract：The aim of this work is to prepare a brain decellularized scaffold by chemical extraction combined with oscillation, and to perform preliminary characterization. Twenty SD rats were divided into decellularization and control group. In the decellularization group, the brain extracellular matrix (dBECM) was prepared by chemical extraction combined with oscillation, successively with 3% TritionX-100, 1%SDS,4% deoxycholic acid sodium and deionized water in 37℃. The microscopic morphology of dBECM was observed by scanning electron microscopy. The decellularization was analyzed by HE staining and DAPI staining. The components were identified by Masson staining and immunofluorescence staining. A lot of collagen fibers could be seen with HE and immunohistochemistry stain but no visible cell nuclei remained after decellularization. The degree of decellularization was about 99%. Masson staining and immunofluorescence staining revealed that dBECM retained elastin (4±1.1%), laminin (19±1.6%), fibronectin (9±2.1%) and collagen IV (16±1.9%). In conclusion, the method of chemical extraction combined with oscillation can effectively remove all cellular components while retain the extracellular matrix and three-dimensional structure. It is a convenient and ideal preparation method on decellularized brain scaffold for tissue engineering.
郑方平, 毛凯丽, 赵应征. 脑去细胞生物支架的制备与鉴定[J]. 中国生物医学工程学报, 2018, 37(2): 202-207.
Zheng Fangping, Mao Kaili, Zhao Yingzheng. Preparation and Characterization of a Brain Decellularized Scaffold. Chinese Journal of Biomedical Engineering, 2018, 37(2): 202-207.
[1] Wang Xiaoyan, Yu Tailong, Chen Guanghua, et al. Preparation and characterization of a chitosan/gelatin/extracellular matrix scaffold and its application in tissue engineering [J]. Tissue Engineering, 2017, 23(3): 169-179. [2] Xu Helin, Tian Furong, Lu Cuitao, et al. Thermo-sensitive hydrogels combined with decellularised matrix deliver bFGF for the functional recovery of rats after a spinal cord injury [J]. Scientific Reports, 2016, 6: 38332. [3] Wang RM, Johnson TD, He Jingjin, et al. Humanized mouse model for assessing the human immune response to xenogeneic and allogeneic decellularized biomaterials [J]. Biomaterials, 2017, 129: 98-110. [4] Hill RC, Calle EA, Dzieciatkowska M, et al. Quantification of extracellular matrix proteins from a rat lung scaffold to provide a molecular readout for tissue engineering [J]. Molecular & Cellular Proteomics: MCP, 2015, 14(4): 961-973. [5] Haykal S, Soleas JP, Salna M, et al. Evaluation of the structural integrity and extracellular matrix components of tracheal allografts following cyclical decellularization techniques: comparison of three protocols [J]. Tissue Engineering. Part C, Methods, 2012, 18(8): 614-623. [6] Baiguera S, Del Gaudio C, Lucatelli E, et al. Electrospun gelatin scaffolds incorporating rat decellularized brain extracellular matrix for neural tissue engineering [J]. Biomaterials, 2014, 35(4): 1205-1214. [7] Keane TJ, Swinehart IT, Badylak SF, Methods of tissue decellularization used for preparation of biologic scaffolds and in vivo relevance [J]. Methods (San Diego, Calif.), 2015, 84: 25-34. [8] Yi Sheng, Ding Fei, Gong L, et al. Extracellular matrix scaffolds for tissue engineering and regenerative medicine [J]. Current Stem Cell Research & Therapy, 2017,12(3): 233-246. [9] Polanco TO, Xylas J, Lantis JC, et al. Extracellular matrices (ECM) for tissue repair [J]. Surgical Technology International, 2016, 28: 43-57. [10] Vigier S, Gagnon H, Bourgade K, et al. Composition and organization of the pancreatic extracellular matrix by combined methods of immunohistochemistry, proteomics and scanning electron microscopy [J]. Current Research in Translational Medicine, 2017, 65(1): 31-39. [11] Almeida HV, Liu Y, Cunniffe GM, et al. Controlled release of transforming growth factor-beta3 from cartilage-extra-cellular-matrix-derived scaffolds to promote chondrogenesis of human-joint-tissue-derived stem cells [J]. Acta Biomaterialia, 2014, 10(10): 4400-4409. [12] Lin Qian, Wong HL, Tian Furong, et al. Enhanced neuroprotection with decellularized brain extracellular matrix containing bFGF after intracerebral transplantation in Parkinson's disease rat model [J]. International Journal of Pharmaceutics, 2017, 517(1-2): 383-394. [13] Xu Helin, Mao Kaili, Lu Cuitao, et al. An injectable acellular matrix scaffold with absorbable permeable nanoparticles improves the therapeutic effects of docetaxel on glioblastoma [J]. Biomaterials, 2016, 107: 44-60.
