摘要
目的应用不同方法对猪升主动脉进行脱细胞,从脱细胞有效性、细胞外基质破坏、生物力学和组织相容性等方面进行综合比较,以期获得适宜的组织工程血管支架。方法取市售猪升主动脉30根随机分成6组(n=5),分别行以下脱细胞处理:A组0.1%胰蛋白酶/0.02%EDTA/PBS,B组1%Triton X-100/0.02%EDTA/蒸馏水,C组1%脱氧胆酸钠/蒸馏水,D组0.5%脱氧胆酸钠/0.5%SDS/蒸馏水,E组1%脱氧胆酸/蒸馏水;F组未行脱细胞处理,作为对照。观察各组支架大体结构,行HE染色观察支架显微结构和脱细胞有效性,DNA定量分析评价脱细胞效率;免疫组织化学染色观察Ⅰ型胶原和弹性蛋白损伤;扫描电镜观察内膜表面结构,测定生物力学性能。取90只SD大鼠随机分为6组(n=15),于腹壁肌内分别植入各组支架,术后7、14、28 d行HE染色评价支架免疫原性和组织相容性。结果 HE染色及DNA定量分析示,仅A、D组支架完全去除细胞成分。A组Ⅰ型胶原表达量显著低于其余各组(P<0.05),基底膜破坏严重,生物力学性能下降,最大应力和抗张强度显著低于其余各组(P<0.05),断裂伸长率显著高于其余各组(P<0.05)。D组Ⅰ型胶原破坏较B、C、E、F组严重(P<0.05),但基底膜完整,生物力学性能接近于F组(P>0.05)。植入大鼠体内实验显示,D组支架免疫原性和组织相容性明显优于其余各组,炎性细胞浸润程度轻(P<0.05)。结论采用0.5%脱氧胆酸钠/0.5%SDS/蒸馏水对猪主动脉进行脱细胞,获得的细胞外基质支架具有用于构建组织工程血管的潜能。
Objective To prepare the aortic extracellular matrix （ECM） scaffold by using different methods to decellularize porcine ascending aorta and to comprehensively compare the efficiency of decellularization and the damage of ECM,evaluation of biomechanical property and biocompatibility. Methods Thirty specimens of fresh porcine ascending aorta were randomly divided into 6 groups （n=5）. The porcine ascending aorta was decellularized by 5 different protocols in groups A-E： 0.1% trypsin/0.02% ethylenediamine tetraacetic acid （EDTA）/PBS was used in group A, 1%Triton X-100/0.02% EDTA/distilled water in group B, 1% sodium deoxycholic acid/distilled water in group C, 0.5% sodium deoxycholic acid/0.5% sodium dodecyl sulfate/distilled water in group D, and 1% deoxycholic acid/distilled water in group E;and the porcine ascending aorta was not decellularized as control in group F. The ascending aorta scaffolds were investigated by gross examination, HE staining,DNA quantitative analysis, immunohistochemistry, and scanning electron microscopy were used to observe the efficiency of decellularization, microstructure of the ECM, the damage of collagen type I and elastin, the structure of intimal surface, and biomechanical property. The 90 Sprague Dawley rats were randomly divided into 6 groups （n=15）. Each scaffold was implanted in the abdominal muscles of rats respectively to evaluate the immunogenicity and biocompatibility. Results HE staining and quantitative analysis of DNA showed that the cells were completely removed only in groups A and D. The expression of collagen type I in group A was significantly lower than that in the other 5 groups （P〈0.05）, and serious damage of the basement membrane and decreased beomechanical property were observed. The maximum stress and tensile strength in group A was significantly lower than those in the other groups （P〈0.05）, and elongation at break was significantly higher than that in the other groups （P〈0.05）. The destruction of collagen type I was signifi
出处
《中国修复重建外科杂志》
CAS
CSCD
北大核心
2014年第11期1413-1421,共9页
Chinese Journal of Reparative and Reconstructive Surgery
基金
国家自然科学基金资助项目(31160230)
云南省自然科学基金资助项目(2010C119)
云南省科技厅-昆明医科大学联合专项基金资助项目(2011FB166
2013FB187
2013FB189)~~
关键词
血管组织工程
支架材料
细胞外基质
脱细胞
猪
大鼠
Vessel tissue engineering
Scaf old material
Extracellular matrix
Decellularization
Porcine
Rat
