去污剂-联合酶法行兔脱细胞组织工程气管制备及性能评价  

Preparation and performance evaluation of a novel detrgent-enzymatic tissue-engineered tracheal matrix in rabbits

作　　者：孙飞 潘枢 史宏灿 章方彪 张卫东 叶刚 刘星辰 

机构地区：[1]扬州大学临床医学院胸外科,225001

出　　处：《中华胸心血管外科杂志》2014年第1期38-41,55,共5页Chinese Journal of Thoracic and Cardiovascular Surgery

基　　金：国家自然科学基金（81370118、30672080、30972968、81170014）;江苏省“六大人才高峰”高层次人才基金（2009128）;江苏省“青蓝工程”学术带头人基金（20081215）

摘　　要：目的探讨兔气管支架最佳脱细胞周期，进行生物力学性能及细胞外基质结构完整性评价。方法选取40只健康新西兰兔，体质量2．5～3．0kg，雌雄不限。随机数字表法分为8组（n=5），A组为新鲜对照组，B、C、D、E、F、G、H组对应脱细胞1、3、5、6、7、8、9周期组。取同种气管，通过去污剂一联合酶法（DEM）分别行气管周期脱细胞处理，通过HE、Movat五色、DAPI染色显微镜观察、扫描电镜观察及生物力学性能测定，判断最佳脱细胞周期并行相关性能评价。结果与对照组相比，经7个DEM循环之后组织工程基质细胞核溶解、极少数细胞残留，结构体系未见明显破坏；组织学与分子生物学分析证实细胞成分及细胞核物质近乎全部去除；扫描电子显微镜（SEM）显示脱细胞基质依然保持完整的结构体系；生物力学证实脱细胞并未对基质形态学及机械性能产生明显影响；免疫荧光分析显示脱细胞气管基质细胞核物质显著减少（P〈0．05）。结论通过7周期改良DEM处理，能够有效获取长段无免疫原性脱细胞兔组织工程气管，具有与原生气管相似的结构体系及生物力学性能，是一种较为理想的支架材料。Objective To determine the detergent-enzymatic cycles and evaluate the biomechanical characteristics as well as extracellular matrix integrity of the decellularized tracheal scaffold in rabbit. Methods Forty tracheal segments were harvested from New Zealand white rabbits. Thirty-five of tracheas were subjected to a detergent-enzymatic method of decellu- larization for 1/3/5/6/7/8/9 cycles, respectively, and other five were stored in phosphate-buffered saline at 4℃ as a control. Comparative examinations were performed by the macroscopic view, histological view（ hematoxylin and eosin stain, Movat Pen- tachrome stain, 4：6-diamidino-2-phenylindole ）, scanning electron microscope （ SEM ） and biomechanical properties between decellularized groups and control group. Results After 7 detergent-enzymatic cycles, almost complete decellularized tracheae, retaining the hierarchical and mechanical properties of the native tissues, could be obtained. Histological and molecular biology analysis demonstrated that all cellular components and nuclear material were removed. SEM analysis revealed that the decellu-larized matrices retained the hierarchical structures of native trachea, and biomechanical tests showed that decellularization ap-proach did not led to any influence on tracheal morphological and mechanical properties. Immunofluorescence analysis show a significant reduction of nuclear material in decellularized tracheas （ P 〈 0.05 ）. Condusion In conclusion, this work suggests that 7 cycles of the modified DEM generates a bioengineered rabbit tracheal matrix that is structurally and mechanically similar to native trachea which could be a better selection for tracheal reconstruction with tissue engineering method.

关 键 词：兔 气管 组织工程 细胞外基质 

分 类 号：R318.08[医药卫生—生物医学工程]