Department of Hepatobiliary Surgery, the Affiliated DrumTower Hospital of Nanjing University Medical School, Nanjing, China.
Liver Int. 2013 Mar;33(3):448-58. doi: 10.1111/liv.12088. Epub 2013 Jan 10.
Hepatic tissue engineering is considered as a possible alternative to liver transplantation for end-stage liver disease. Several methods of decellularization of xenogeneic liver are available to produce three-dimensional organ scaffolds for engineering liver tissues. However, rare studies have examined and compared the effectiveness of different methods on the structure and composition of intact decellularized liver extracellular matrix.
Two decellularization methods were adopted herein. Their effects on collagen, elastin, glycosaminoglycans (GAGs), hepatocyte growth factor (HGF) content and influence to the function of hepatocytes cultured in scaffolds were examined and compared.
The complete tissue decellularization was successfully achieved after treatment with sodium dodecyl sulphate (SDS) and Triton X-100. The total absence of nuclear structures and removal of viable cells were confirmed by haematoxylin-eosin staining and scanning electron microscopy. Collagen was preserved after both treatments. However, the elastin content decreased to about 20% and 60%, the GAGs content decreased to about 10% and 50% and the HGF content decreased to about 20% and 60% of the native liver level after SDS and Triton X-100 treatment respectively. The Triton X-100-treated scaffolds were much superior than SDS-treated scaffolds in supporting liver-specific function, including albumin secretion (P = 0.001), urea synthesis (P = 0.002), ammonia elimination (P = 0.007) and mRNA expression levels of drug metabolism enzymes.
This study suggested that liver extracellular matrix scaffolds constructed using perfusion of Triton X-100 as described herein might provide a more effective and ideal material for the usage in tissue engineering and regenerative medicine approaches.
肝组织工程被认为是治疗终末期肝病的肝移植的一种可能替代方法。有几种脱细胞化异种肝的方法可用于生产三维器官支架，用于工程化肝脏组织。然而，很少有研究检查和比较不同方法对完整脱细胞化肝细胞外基质的结构和组成的影响。
本文采用了两种脱细胞化方法。检查并比较了它们对胶原蛋白、弹性蛋白、糖胺聚糖（GAG）、肝细胞生长因子（HGF）含量的影响以及对支架中培养的肝细胞功能的影响。
用十二烷基硫酸钠（SDS）和 Triton X-100 处理后，成功地实现了组织的完全脱细胞化。苏木精-伊红染色和扫描电子显微镜证实了核结构的完全缺失和活细胞的去除。两种处理方法均保留了胶原蛋白。然而，弹性蛋白含量分别下降到约 20%和 60%，GAGs 含量分别下降到约 10%和 50%，HGF 含量分别下降到约 20%和 60%的天然肝水平。Triton X-100 处理的支架在支持肝脏特异性功能方面优于 SDS 处理的支架，包括白蛋白分泌（P = 0.001）、尿素合成（P = 0.002）、氨消除（P = 0.007）和药物代谢酶的 mRNA 表达水平。
本研究表明，本文所述的使用 Triton X-100 灌注构建的肝细胞外基质支架可能为组织工程和再生医学方法提供更有效和理想的材料。
