人肝组织脱细胞支架的制备  

Preparation of a decellularized scaffold derived from human liver tissue

作　　者：郑幸龙[1,2,3] 向俊西[1,2,3] 吴万权 刘学民[1] 刘雯雁[1,2,3] 吕毅[1,2,3] 

机构地区：[1]西安交通大学第一附属医院肝胆外科,陕西西安710061 [2]西安交通大学先进外科技术与工程研究所,陕西西安710061 [3]陕西省再生医学与外科工程研究中心,陕西西安710061

出　　处：《南方医科大学学报》2015年第7期1028-1033,共6页Journal of Southern Medical University

基　　金：高等学校博士学科点专项科研基金(20110201130009);陕西省科学技术研究发展计划项目(2012KW-39-01)

摘　　要：目的探究人肝组织脱细胞支架的制备。方法本研究利用手术切除的人肝血管瘤左外叶组织,经反复冻融,0.01%SDS、0.1%SDS和1%Triton X-100循环灌注制备人肝组织脱细胞支架,并通过灌注过氧乙酸消毒。将L-02细胞通过门静脉插管种植于脱细胞支架内进行培养。结果 HE、DAPI染色和扫描电镜结果显示脱细胞支架内无细胞成分残留,残余DNA检测为25.3±14.6 ng/mg干质量,小于新鲜肝脏DNA含量的1%。免疫组化证实支架内保留了Ⅰ型胶原、Ⅳ型胶原、纤连蛋白、弹力蛋白成分。L-02细胞在支架上生长良好且有增殖,并表达白蛋白及葡萄糖-6-磷酸酶。结论利用手术切除的人肝标本行肝组织脱细胞支架的制备是可行的,且为构建更适用于临床的组织工程肝脏提供了新思路。Objective To develop a method for preparing a decellularized scaffold based on human liver tissue. Methods A surgical specimen of the left lateral lobe of the liver was obtained from a patients with hepatic hemangioma. The decellularization process was performed by repeated freezing-thawing, sequential perfusion with 0.01% SDS, 0.1% SDS and 1% Triton X-100 through the portal vein, and sterilization with peracetic acid. L-02 cells were then engrafted onto the decellularized liver scaffold. Results HE staining, DAPI staining and scanning electron microscopy all verified the absence of residual cellular components in the decellularized scaffold. The residual DNA content in the decellularized scaffolds was 25.3± 14.6 ng/mg （dry weight）, which was less than 1% of the total DNA content in a fresh human liver. Immunohistochemistry demonstrated that type I and IV collagens, fibronectin and elastin were all retained in the scaffold. The engrafted L-02 cells survived well on the scaffold with active proliferation and expressed albumin and G6pc. Conclusion It is feasible to prepare decellularized scaffolds using surgical specimens of human liver, which can be a new approach to constructing a tissue-engineered liver for clinical purposes.

关 键 词：肝组织工程 人肝组织脱细胞支架 细胞外基质 三维培养 

分 类 号：R657.3[医药卫生—外科学]